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Yuk J, Kim J, Jung S, Um SH. Engineering Gizmos for Short Cancer Genetic Fragments Discrimination. Chembiochem 2025; 26:e202400867. [PMID: 39910951 DOI: 10.1002/cbic.202400867] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2024] [Revised: 02/04/2025] [Accepted: 02/06/2025] [Indexed: 02/07/2025]
Abstract
Currently, mankind is fiercely struggling with cancer. Recently, we have been winning the battle against cancer through precision medicine and accompanying diagnostic methods, and we are raising many hopes with blockbuster drugs. It would be even better if we could read the cancer nucleotide sequence, identify them in advance, and suggest treatments simultaneously. However, this may be an impossible dream because it takes a lot of time and effort to diagnose and ensure all the long gene sequences of cancer at once. Thus, victory will be even closer if a rapid and accurate diagnosis of the cancer-specific gene biomarkers that will soon be imprinted can be made. With the advent of nanotechnology, a new short cancer diagnostic toolkit has been proposed to achieve the goal. This review presents a small diagnostic device that detects certain cancers' genetic fragments (simply 'Gizmo'). The development of numerous diagnostic methods has focused on (1) directly detecting pre-selectively targeted genes using novel diagnostic systems, and (2) indirectly detecting substantial improvements in diagnostic sensitivity only through detection signal amplification without existing gene amplification steps. Our fight against cancer is not a dream, but the result of success, and it is assumed that victory will accelerate as soon as possible.
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Affiliation(s)
- Jisoo Yuk
- Department of Biological and Environmental Engineering, Cornell University, Ithaca, NY, 14853, USA
| | - Jeonghun Kim
- Progeneer Incorporation, #1002, 12, Digital-ro 31-gil, Guro-gu, Seoul, 08380, Korea
| | - Sunghwan Jung
- Department of Biological and Environmental Engineering, Cornell University, Ithaca, NY, 14853, USA
| | - Soong Ho Um
- School of Chemical Engineering, Sungkyunkwan University, 2066 Seobu-ro, Jangan-gu, Suwon-si, Gyeonggi-do, 16419, Korea
- Progeneer Incorporation, #1002, 12, Digital-ro 31-gil, Guro-gu, Seoul, 08380, Korea
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Vo DK, Trinh KTL. Polymerase Chain Reaction Chips for Biomarker Discovery and Validation in Drug Development. MICROMACHINES 2025; 16:243. [PMID: 40141854 PMCID: PMC11944077 DOI: 10.3390/mi16030243] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/02/2025] [Revised: 02/17/2025] [Accepted: 02/18/2025] [Indexed: 03/28/2025]
Abstract
Polymerase chain reaction (PCR) chips are advanced, microfluidic platforms that have revolutionized biomarker discovery and validation because of their high sensitivity, specificity, and throughput levels. These chips miniaturize traditional PCR processes for the speed and precision of nucleic acid biomarker detection relevant to advancing drug development. Biomarkers, which are useful in helping to explain disease mechanisms, patient stratification, and therapeutic monitoring, are hard to identify and validate due to the complexity of biological systems and the limitations of traditional techniques. The challenges to which PCR chips respond include high-throughput capabilities coupled with real-time quantitative analysis, enabling researchers to identify novel biomarkers with greater accuracy and reproducibility. More recent design improvements of PCR chips have further expanded their functionality to also include digital and multiplex PCR technologies. Digital PCR chips are ideal for quantifying rare biomarkers, which is essential in oncology and infectious disease research. In contrast, multiplex PCR chips enable simultaneous analysis of multiple targets, therefore simplifying biomarker validation. Furthermore, single-cell PCR chips have made it possible to detect biomarkers at unprecedented resolution, hence revealing heterogeneity within cell populations. PCR chips are transforming drug development, enabling target identification, patient stratification, and therapeutic efficacy assessment. They play a major role in the development of companion diagnostics and, therefore, pave the way for personalized medicine, ensuring that the right patient receives the right treatment. While this tremendously promising technology has exhibited many challenges regarding its scalability, integration with other omics technologies, and conformity with regulatory requirements, many still prevail. Future breakthroughs in chip manufacturing, the integration of artificial intelligence, and multi-omics applications will further expand PCR chip capabilities. PCR chips will not only be important for the acceleration of drug discovery and development but also in raising the bar in improving patient outcomes and, hence, global health care as these technologies continue to mature.
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Affiliation(s)
- Dang-Khoa Vo
- College of Pharmacy, Gachon University, 191 Hambakmoe-ro, Yeonsu-gu, Incheon 21936, Republic of Korea;
| | - Kieu The Loan Trinh
- Bionano Applications Research Center, Gachon University, 1342 Seongnam-daero, Sujeong-gu, Seongnam-si 13120, Gyeonggi-do, Republic of Korea
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Gu X, Minko T. Targeted Nanoparticle-Based Diagnostic and Treatment Options for Pancreatic Cancer. Cancers (Basel) 2024; 16:1589. [PMID: 38672671 PMCID: PMC11048786 DOI: 10.3390/cancers16081589] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/29/2024] [Revised: 04/17/2024] [Accepted: 04/19/2024] [Indexed: 04/28/2024] Open
Abstract
Pancreatic ductal adenocarcinoma (PDAC), one of the deadliest cancers, presents significant challenges in diagnosis and treatment due to its aggressive, metastatic nature and lack of early detection methods. A key obstacle in PDAC treatment is the highly complex tumor environment characterized by dense stroma surrounding the tumor, which hinders effective drug delivery. Nanotechnology can offer innovative solutions to these challenges, particularly in creating novel drug delivery systems for existing anticancer drugs for PDAC, such as gemcitabine and paclitaxel. By using customization methods such as incorporating conjugated targeting ligands, tumor-penetrating peptides, and therapeutic nucleic acids, these nanoparticle-based systems enhance drug solubility, extend circulation time, improve tumor targeting, and control drug release, thereby minimizing side effects and toxicity in healthy tissues. Moreover, nanoparticles have also shown potential in precise diagnostic methods for PDAC. This literature review will delve into targeted mechanisms, pathways, and approaches in treating pancreatic cancer. Additional emphasis is placed on the study of nanoparticle-based delivery systems, with a brief mention of those in clinical trials. Overall, the overview illustrates the significant advances in nanomedicine, underscoring its role in transcending the constraints of conventional PDAC therapies and diagnostics.
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Affiliation(s)
- Xin Gu
- Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, NJ 08554, USA
| | - Tamara Minko
- Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, Piscataway, NJ 08554, USA
- Rutgers Cancer Institute of New Jersey, Rutgers, The State University of New Jersey, New Brunswick, NJ 08901, USA
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Jain M, Atayan D, Rakhmatullin T, Dakhtler T, Popov P, Kim P, Viborniy M, Gontareva I, Samokhodskaya L, Egorov V. Cell-Free Tumor DNA Detection-Based Liquid Biopsy of Plasma and Bile in Patients with Various Pancreatic Neoplasms. Biomedicines 2024; 12:220. [PMID: 38255325 PMCID: PMC10813046 DOI: 10.3390/biomedicines12010220] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/25/2023] [Revised: 01/10/2024] [Accepted: 01/16/2024] [Indexed: 01/24/2024] Open
Abstract
The key challenge of cell-free tumor DNA (cftDNA) analysis in pancreatic ductal adenocarcinoma (PDAC) is overcoming its low detection rate, which is mainly explained by the overall scarcity of this biomarker in plasma. Obstructive jaundice is a frequent event in PDAC, which enables bile collection as a part of routine treatment. The aim of this study was to evaluate the performance of KRAS-mutated cftDNA detection-based liquid biopsy of plasma and bile in patients with pancreatic neoplasms using digital droplet PCR. The study included healthy volunteers (n = 38), patients with PDAC (n = 95, of which 20 had obstructive jaundice) and other pancreatic neoplasms (OPN) (n = 18). The sensitivity and specificity compared to the control group were 61% and 100% (AUC-ROC-0.805), and compared to the OPN group, they were 61% and 94% (AUC-ROC-0.794), respectively. Bile exhibited higher cftDNA levels than plasma (248.6 [6.743; 1068] vs. 3.26 [0; 19.225] copies/mL) and a two-fold higher detection rate (p < 0.01). Plasma cftDNA levels were associated with distant metastases, tumor size, and CA 19-9 (p < 0.05). The probability of survival was worse in patients with higher levels of cftDNA in plasma (hazard ratio-2.4; 95% CI: 1.3-4.6; p = 0.005) but not in bile (p > 0.05). Bile is a promising alternative to plasma in patients with obstructive jaundice, at least for the diagnostic purposes of liquid biopsy.
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Affiliation(s)
- Mark Jain
- Medical Research and Educational Center, Lomonosov Moscow State University, 119992 Moscow, Russia;
| | - David Atayan
- Joint Stock Company “Ilyinsky Hospital”, 143421 Moscow, Russia; (D.A.); (T.D.); (P.P.); (P.K.); (M.V.); (I.G.); (V.E.)
| | - Tagir Rakhmatullin
- Department of Fundamental Medicine, Lomonosov Moscow State University, 119991 Moscow, Russia;
| | - Tatyana Dakhtler
- Joint Stock Company “Ilyinsky Hospital”, 143421 Moscow, Russia; (D.A.); (T.D.); (P.P.); (P.K.); (M.V.); (I.G.); (V.E.)
| | - Pavel Popov
- Joint Stock Company “Ilyinsky Hospital”, 143421 Moscow, Russia; (D.A.); (T.D.); (P.P.); (P.K.); (M.V.); (I.G.); (V.E.)
| | - Pavel Kim
- Joint Stock Company “Ilyinsky Hospital”, 143421 Moscow, Russia; (D.A.); (T.D.); (P.P.); (P.K.); (M.V.); (I.G.); (V.E.)
| | - Mikhail Viborniy
- Joint Stock Company “Ilyinsky Hospital”, 143421 Moscow, Russia; (D.A.); (T.D.); (P.P.); (P.K.); (M.V.); (I.G.); (V.E.)
| | - Iuliia Gontareva
- Joint Stock Company “Ilyinsky Hospital”, 143421 Moscow, Russia; (D.A.); (T.D.); (P.P.); (P.K.); (M.V.); (I.G.); (V.E.)
| | - Larisa Samokhodskaya
- Medical Research and Educational Center, Lomonosov Moscow State University, 119992 Moscow, Russia;
| | - Vyacheslav Egorov
- Joint Stock Company “Ilyinsky Hospital”, 143421 Moscow, Russia; (D.A.); (T.D.); (P.P.); (P.K.); (M.V.); (I.G.); (V.E.)
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Radefeldt M, Stellmacher-Kaiser S, Krake S, Kragl B, Lemke S, Beetz C, Bauer P, Junghanß C, Al-Ali R. Basic ctDNA Panel Promises Affordable Clinical Validity in Colon Cancer Patients but Not in Pancreas Cancer Patients. Life (Basel) 2023; 13:2274. [PMID: 38137875 PMCID: PMC10744654 DOI: 10.3390/life13122274] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2023] [Revised: 11/10/2023] [Accepted: 11/19/2023] [Indexed: 12/24/2023] Open
Abstract
The potential of circulating tumor DNA (ctDNA) as a biomarker to assess the progression of various solid tumors has been explored extensively. In this study, we investigated the feasibility of utilizing a ctDNA sequencing panel specifically designed to target the most frequently mutated genomic regions in colon and pancreas cancers. Through somatic analysis of colon and pancreas tumors, we targeted 27 regions within eight genes. By employing PCR amplification and Illumina NGS, we ensured that each region was adequately covered with a minimum of 5000 reads (with an average of 12,000 reads). Our method exhibited reproducibility with repetition and dilutions. The positive detection threshold for ctDNA was set at a cutoff value of 0.5% ctDNA of the total reads using IGV. Among the samples analyzed, 71% of colon cancer cases displayed somatic mutations covered by the targeted regions. Within this group, detectable ctDNA was observed in 34% of the cases. Conversely, in pancreatic cancer, 55% of mutations were covered by the panel's regions, but only 13% of these cases exhibited detectable ctDNA. In follow-ups with the patients, changes in ctDNA percentages demonstrated complete concordance with changes in the clinical condition in 88% of the cases. Our findings suggest that employing a basic ctDNA-targeted panel can serve as a cost-effective and reliable approach for repeated monitoring of the efficacy of colon cancer therapy. However, in the case of pancreatic cancer, ctDNA showed limited utility, and alternative biomarkers may offer superior diagnostic value. Additionally, we found that a negative ctDNA test is not a guarantee for a relapse-free recovery; thus, we recommend a continuous follow-up with the patient on a long-term basis.
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Affiliation(s)
| | - Silke Stellmacher-Kaiser
- Clinical for Internal Medicine, Hematology, Oncology and Palliative Medicine, University Medicine Rostock, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany
| | - Susann Krake
- CENTOGENE GmbH, Am Strande 7, 18055 Rostock, Germany (S.K.)
| | - Brigitte Kragl
- Clinical for Internal Medicine, Hematology, Oncology and Palliative Medicine, University Medicine Rostock, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany
| | - Sabrina Lemke
- CENTOGENE GmbH, Am Strande 7, 18055 Rostock, Germany (S.K.)
| | | | - Peter Bauer
- CENTOGENE GmbH, Am Strande 7, 18055 Rostock, Germany (S.K.)
- Clinical for Internal Medicine, Hematology, Oncology and Palliative Medicine, University Medicine Rostock, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany
| | - Christian Junghanß
- Clinical for Internal Medicine, Hematology, Oncology and Palliative Medicine, University Medicine Rostock, Ernst-Heydemann-Str. 6, 18057 Rostock, Germany
| | - Ruslan Al-Ali
- CENTOGENE GmbH, Am Strande 7, 18055 Rostock, Germany (S.K.)
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Maloney S, Clarke SJ, Sahni S, Hudson A, Colvin E, Mittal A, Samra J, Pavlakis N. The role of diagnostic, prognostic, and predictive biomarkers in the management of early pancreatic cancer. J Cancer Res Clin Oncol 2023; 149:13437-13450. [PMID: 37460806 PMCID: PMC10587199 DOI: 10.1007/s00432-023-05149-4] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2023] [Accepted: 07/09/2023] [Indexed: 10/20/2023]
Abstract
Despite modern advances in cancer medicine, pancreatic cancer survival remains unchanged at just 12%. For the small proportion of patients diagnosed with 'early' (upfront or borderline resectable) disease, recurrences are common, and many recur soon after surgery. Whilst chemotherapy has been shown to increase survival in this cohort, the morbidity of surgery renders many candidates unsuitable for adjuvant treatment. Due to this, and the success of upfront chemotherapy in the advanced setting, use of neoadjuvant chemotherapy has been introduced in patients with upfront or borderline resectable disease. Randomized controlled trials have been conducted to compare upfront surgery to neoadjuvant chemotherapy in this patient cohort, opinions on the ideal upfront treatment approach are divided. This lack of consensus has highlighted the need for biomarkers to assist in clinical decision making. This review analyses the potential diagnostic, prognostic and predictive biomarkers that may assist in the diagnosis and management of early (upfront and borderline resectable) pancreatic cancer.
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Affiliation(s)
- Sarah Maloney
- Faculty of Medicine and Health Sciences, Northern Clinical School, The University of Sydney, Sydney, 2065, Australia.
- Bill Walsh Translational Cancer Research Laboratory, Kolling Institute, The University of Sydney, Sydney, 2065, Australia.
- Department of Medical Oncology, Royal North Shore Hospital, St. Leonards, Sydney, NSW, 2065, Australia.
| | - Stephen J Clarke
- Faculty of Medicine and Health Sciences, Northern Clinical School, The University of Sydney, Sydney, 2065, Australia
- Bill Walsh Translational Cancer Research Laboratory, Kolling Institute, The University of Sydney, Sydney, 2065, Australia
- Department of Medical Oncology, Royal North Shore Hospital, St. Leonards, Sydney, NSW, 2065, Australia
| | - Sumit Sahni
- Faculty of Medicine and Health Sciences, Northern Clinical School, The University of Sydney, Sydney, 2065, Australia
- Bill Walsh Translational Cancer Research Laboratory, Kolling Institute, The University of Sydney, Sydney, 2065, Australia
| | - Amanda Hudson
- Faculty of Medicine and Health Sciences, Northern Clinical School, The University of Sydney, Sydney, 2065, Australia
- Bill Walsh Translational Cancer Research Laboratory, Kolling Institute, The University of Sydney, Sydney, 2065, Australia
| | - Emily Colvin
- Faculty of Medicine and Health Sciences, Northern Clinical School, The University of Sydney, Sydney, 2065, Australia
- Bill Walsh Translational Cancer Research Laboratory, Kolling Institute, The University of Sydney, Sydney, 2065, Australia
| | - Anubhav Mittal
- Faculty of Medicine and Health Sciences, Northern Clinical School, The University of Sydney, Sydney, 2065, Australia
- Bill Walsh Translational Cancer Research Laboratory, Kolling Institute, The University of Sydney, Sydney, 2065, Australia
- Upper Gastrointestinal Surgical Unit, Royal North Shore Hospital, St. Leonards, Sydney, NSW, 2065, Australia
| | - Jaswinder Samra
- Faculty of Medicine and Health Sciences, Northern Clinical School, The University of Sydney, Sydney, 2065, Australia
- Bill Walsh Translational Cancer Research Laboratory, Kolling Institute, The University of Sydney, Sydney, 2065, Australia
- Upper Gastrointestinal Surgical Unit, Royal North Shore Hospital, St. Leonards, Sydney, NSW, 2065, Australia
| | - Nick Pavlakis
- Faculty of Medicine and Health Sciences, Northern Clinical School, The University of Sydney, Sydney, 2065, Australia
- Bill Walsh Translational Cancer Research Laboratory, Kolling Institute, The University of Sydney, Sydney, 2065, Australia
- Department of Medical Oncology, Royal North Shore Hospital, St. Leonards, Sydney, NSW, 2065, Australia
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Takahashi K, Takeda Y, Ono Y, Isomoto H, Mizukami Y. Current status of molecular diagnostic approaches using liquid biopsy. J Gastroenterol 2023; 58:834-847. [PMID: 37470859 PMCID: PMC10423147 DOI: 10.1007/s00535-023-02024-4] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/21/2023] [Accepted: 07/08/2023] [Indexed: 07/21/2023]
Abstract
Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive and lethal cancers, and developing an efficient and reliable approach for its early-stage diagnosis is urgently needed. Precancerous lesions of PDAC, such as pancreatic intraepithelial neoplasia (PanIN) and intraductal papillary mucinous neoplasms (IPMN), arise through multiple steps of driver gene alterations in KRAS, TP53, CDKN2A, SMAD4, or GNAS. Hallmark mutations play a role in tumor initiation and progression, and their detection in bodily fluids is crucial for diagnosis. Recently, liquid biopsy has gained attention as an approach to complement pathological diagnosis, and in addition to mutation signatures in cell-free DNA, cell-free RNA, and extracellular vesicles have been investigated as potential diagnostic and prognostic markers. Integrating such molecular information to revise the diagnostic criteria for pancreatic cancer can enable a better understanding of the pathogenesis underlying inter-patient heterogeneity, such as sensitivity to chemotherapy and disease outcomes. This review discusses the current diagnostic approaches and clinical applications of genetic analysis in pancreatic cancer and diagnostic attempts by liquid biopsy and molecular analyses using pancreatic juice, duodenal fluid, and blood samples. Emerging knowledge in the rapidly advancing liquid biopsy field is promising for molecular profiling and diagnosing pancreatic diseases with significant diversity.
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Affiliation(s)
- Kenji Takahashi
- Division of Metabolism and Biosystemic Science, Gastroenterology, and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, 2-1 Midorigaoka Higashi, Asahikawa, Hokkaido, 078-8510, Japan.
| | - Yohei Takeda
- Division of Medicine and Clinical Science, Department of Multidisciplinary Internal Medicine, Faculty of Medicine, Tottori University, Tottori, Japan
| | - Yusuke Ono
- Division of Metabolism and Biosystemic Science, Gastroenterology, and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, 2-1 Midorigaoka Higashi, Asahikawa, Hokkaido, 078-8510, Japan
- Institute of Biomedical Research, Sapporo Higashi Tokushukai Hospital, Sapporo, Japan
| | - Hajime Isomoto
- Division of Medicine and Clinical Science, Department of Multidisciplinary Internal Medicine, Faculty of Medicine, Tottori University, Tottori, Japan
| | - Yusuke Mizukami
- Division of Metabolism and Biosystemic Science, Gastroenterology, and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, 2-1 Midorigaoka Higashi, Asahikawa, Hokkaido, 078-8510, Japan
- Institute of Biomedical Research, Sapporo Higashi Tokushukai Hospital, Sapporo, Japan
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Yang M, Zhang H, Ma W, Liu Q, Fu X, Fu Y. A triple-cycle amplification biosensor for colorimetric detection of mutant PIK3CA E545K based on cascade-driven DNA walker and branched hybridization strand reaction. Anal Chim Acta 2023; 1270:341452. [PMID: 37311611 DOI: 10.1016/j.aca.2023.341452] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2023] [Revised: 05/18/2023] [Accepted: 05/29/2023] [Indexed: 06/15/2023]
Abstract
Circulating tumor DNA (ctDNA) is an ideal candidate for liquid biopsy biomarkers. Therefore, detecting a low abundance of ctDNA is essential for early cancer diagnosis. Here, we developed a novel triple circulation amplification system integrating entropy and enzyme cascade-driven three-dimensional (3D) DNA walker and branched hybridization strand reaction (B-HCR) for ultrasensitive detection of breast cancer-related ctDNA. In this study, the 3D DNA walker was constructed by inner track probes (NH) and complex S on a microsphere. Once the DNA walker was triggered by the target, the strand replacement reaction ran first and kept circulating to rapidly displace the DNA walker containing 8-17 DNAzyme. Secondly, the DNA walker could repeatedly cleave NH autonomously along the inner track, generating numerous initiators, and then promoting B-HCR to activate the third cycle. Subsequently, the split G-rich fragments were brought in close to form the G-quadruplex/hemin DNAzyme by adding hemin, with the addition of H2O2 and ABTS, the target could be observed. Benefiting from triplex cycles, the PIK3CAE545K mutation detection possesses a good linear range from 1-103 fM, and the limit of detection was 0.65 fM. Due to the low cost and high sensitivity, the proposed strategy has great potential in early diagnosis of breast cancer.
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Affiliation(s)
- Mei Yang
- Hunan Provincial Key Laboratory of Environmental Catalysis and Waste Recycling, College of Materials and Chemical Engineering, Hunan Institute of Engineering, Xiangtan, 411104, China
| | - He Zhang
- Hunan Provincial Key Laboratory of Environmental Catalysis and Waste Recycling, College of Materials and Chemical Engineering, Hunan Institute of Engineering, Xiangtan, 411104, China.
| | - Wenjie Ma
- Hunan Provincial Key Laboratory of Environmental Catalysis and Waste Recycling, College of Materials and Chemical Engineering, Hunan Institute of Engineering, Xiangtan, 411104, China
| | - Qiong Liu
- Hunan Provincial Key Laboratory of Environmental Catalysis and Waste Recycling, College of Materials and Chemical Engineering, Hunan Institute of Engineering, Xiangtan, 411104, China
| | - Xin Fu
- Hunan Provincial Key Laboratory of Environmental Catalysis and Waste Recycling, College of Materials and Chemical Engineering, Hunan Institute of Engineering, Xiangtan, 411104, China
| | - Yu Fu
- Hunan Provincial Key Laboratory of Environmental Catalysis and Waste Recycling, College of Materials and Chemical Engineering, Hunan Institute of Engineering, Xiangtan, 411104, China
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9
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Cassese G, Han HS, Yoon YS, Lee JS, Lee B, Cubisino A, Panaro F, Troisi RI. Role of neoadjuvant therapy for nonmetastatic pancreatic cancer: Current evidence and future perspectives. World J Gastrointest Oncol 2023; 15:911-924. [PMID: 37389109 PMCID: PMC10302990 DOI: 10.4251/wjgo.v15.i6.911] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/28/2022] [Revised: 02/17/2023] [Accepted: 04/24/2023] [Indexed: 06/14/2023] Open
Abstract
Pancreatic adenocarcinoma (PDAC) is one of the most common and lethal human cancers worldwide. Surgery followed by adjuvant chemotherapy offers the best chance of a long-term survival for patients with PDAC, although only approximately 20% of the patients have resectable tumors when diagnosed. Neoadjuvant chemotherapy (NACT) is recommended for borderline resectable pancreatic cancer. Several studies have investigated the role of NACT in treating resectable tumors based on the recent advances in PDAC biology, as NACT provides the potential benefit of selecting patients with favorable tumor biology and controls potential micro-metastases in high-risk patients with resectable PDAC. In such challenging cases, new potential tools, such as ct-DNA and molecular targeted therapy, are emerging as novel therapeutic options that may improve old paradigms. This review aims to summarize the current evidence regarding the role of NACT in treating non-metastatic pancreatic cancer while focusing on future perspectives in light of recent evidence.
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Affiliation(s)
- Gianluca Cassese
- Department of Clinical Medicine and Surgery, Division of Minimally Invasive HPB Surgery and Transplantation Service, Federico II University Hospital, Naples 80131, Italy
| | - Ho-Seong Han
- Department of Surgery, Seoul National University College of Medicine, Seongnam 13620, Gyeonggi-do, South Korea
| | - Yoo-Seok Yoon
- Department of Surgery, Seoul National University College of Medicine, Seongnam 13620, Gyeonggi-do, South Korea
| | - Jun Suh Lee
- Department of Surgery, Seoul National University College of Medicine, Seongnam 13620, Gyeonggi-do, South Korea
| | - Boram Lee
- Department of Surgery, Seoul National University College of Medicine, Seongnam 13620, Gyeonggi-do, South Korea
| | - Antonio Cubisino
- Department of HPB Surgery and Transplantation, Beaujon Hospital, Clichy 92110, France
| | - Fabrizio Panaro
- Department of Digestive Surgery and Liver Transplantation, CHU Montpellier, Montpellier 34100, France
| | - Roberto Ivan Troisi
- Department of Clinical Medicine and Surgery, Division of Minimally Invasive HPB Surgery and Transplantation Service, Federico II University Hospital, Naples 80131, Italy
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10
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Li M, Luo N, Liao X, Zou L. Proximity hybridization-regulated CRISPR/Cas12a-based dual signal amplification strategy for sensitive detection of circulating tumor DNA. Talanta 2023; 257:124395. [PMID: 36858011 DOI: 10.1016/j.talanta.2023.124395] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/28/2023] [Revised: 02/20/2023] [Accepted: 02/23/2023] [Indexed: 02/27/2023]
Abstract
Circulating tumor DNA (ctDNA) is regarded as an ideal candidate biomarker for the non-invasive diagnosis of cancer. However, the lack of convenient and reliable detection methods for ctDNA restricts its clinical application. Herein, we developed a dual signal amplification strategy for sensitive detection of ctDNA based on hybridization chain reaction (HCR) and proximity hybridization-regulated CRISPR/Cas12a. The ctDNA initiates HCR through the continuous hybridization of two hairpin probes (H1 and H2), yielding long nicked double-stranded DNA nanowires composed of numerous split segments, which are successively connected to activate the trans-cleavage activity of CRISPR/Cas12a. In this case, the doubly labeled single-stranded DNA reporter can be cleaved to produce a strong fluorescent signal. Owing to the dual amplification of HCR and CRISPR/Cas12a, this strategy exhibits high sensitivity toward ctDNA with a low detection limit of 5.43 fM. Moreover, the proposed method was successfully applied for ctDNA detection in serum samples with satisfactory results, which has great potential in the clinical diagnosis of cancer.
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Affiliation(s)
- Mengyan Li
- School of Pharmacy, Guangdong Pharmaceutical University, Guangzhou, 510006, PR China
| | - Nian Luo
- School of Pharmacy, Guangdong Pharmaceutical University, Guangzhou, 510006, PR China
| | - Xiaofei Liao
- School of Pharmacy, Guangdong Pharmaceutical University, Guangzhou, 510006, PR China
| | - Li Zou
- School of Pharmacy, Guangdong Pharmaceutical University, Guangzhou, 510006, PR China; Key Specialty of Clinical Pharmacy, The First Affiliated Hospital of Guangdong Pharmaceutical University, Guangzhou, 510699, PR China.
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11
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Hu Q, Kanwal F, Lyu W, Zhang J, Liu X, Qin K, Shen F. Multiplex Digital Polymerase Chain Reaction on a Droplet Array SlipChip for Analysis of KRAS Mutations in Pancreatic Cancer. ACS Sens 2023; 8:114-121. [PMID: 36520653 DOI: 10.1021/acssensors.2c01776] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
Pancreatic cancer is a terminal disease with high mortality and very poor prognosis. A sensitive and quantitative analysis of KRAS mutations in pancreatic cancer provides a tool not only to understand the biological mechanisms of pancreatic cancer but also for diagnosis and treatment monitoring. Digital polymerase chain reaction (PCR) is a promising tool for KRAS mutation analysis, but current methods generally require a complex microfluidic handling system, which can be challenging to implement in routine research and point-of-care clinical diagnostics. Here, we present a droplet-array SlipChip (da-SlipChip) for the multiplex quantification of KRAS G12D, V, R, and C mutant genes with the wild-type (WT) gene background by dual color (FAM/ROX) fluorescence detection. This da-SlipChip is a high-density microwell array of 21,696 wells of 200 pL in 4 by 5424 microwell format with simple loading and slipping operation. It does not require the same precise alignment of microfeatures on the different plates that are acquired by the traditional digital PCR SlipChip. This device can provide accurate quantification of both mutant genes and the WT KRAS gene. We collected tumor tissue, paired normal pancreatic tissue, and other normal tissues from 18 pancreatic cancer patients and analyzed the mutation profiles of KRAS G12D, V, R, and C in these samples; the results from the multiplex digital PCR on da-SlipChip agree well with those of next-generation sequencing (NGS). This da-SlipChip moves digital PCR closer to the practical point-of-care applications not only for detecting KRAS mutations in pancreatic cancer but also for other applications that require precise nucleic acid quantification with high sensitivity.
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Affiliation(s)
- Qixin Hu
- School of Biomedical Engineering, Shanghai Jiao Tong University, 1954 Hua Shan Road, Shanghai 200030, China
| | - Fariha Kanwal
- School of Biomedical Engineering, Shanghai Jiao Tong University, 1954 Hua Shan Road, Shanghai 200030, China
| | - Weiyuan Lyu
- School of Biomedical Engineering, Shanghai Jiao Tong University, 1954 Hua Shan Road, Shanghai 200030, China
| | - Jiajie Zhang
- School of Biomedical Engineering, Shanghai Jiao Tong University, 1954 Hua Shan Road, Shanghai 200030, China
| | - Xu Liu
- School of Biomedical Engineering, Shanghai Jiao Tong University, 1954 Hua Shan Road, Shanghai 200030, China
| | - Kai Qin
- Department of General Surgery, Pancreatic Disease Center, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200031, China
| | - Feng Shen
- School of Biomedical Engineering, Shanghai Jiao Tong University, 1954 Hua Shan Road, Shanghai 200030, China
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12
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Pancreatic Cancer in Chronic Pancreatitis: Pathogenesis and Diagnostic Approach. Cancers (Basel) 2023; 15:cancers15030761. [PMID: 36765725 PMCID: PMC9913572 DOI: 10.3390/cancers15030761] [Citation(s) in RCA: 33] [Impact Index Per Article: 16.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2022] [Revised: 01/20/2023] [Accepted: 01/24/2023] [Indexed: 01/28/2023] Open
Abstract
Chronic pancreatitis is one of the main risk factors for pancreatic cancer, but it is a rare event. Inflammation and oncogenes work hand in hand as key promoters of this disease. Tobacco is another co-factor. During alcoholic chronic pancreatitis, the cumulative risk of cancer is estimated at 4% after 15 to 20 years. This cumulative risk is higher in hereditary pancreatitis: 19 and 12% in the case of PRSS1 and SPINK1 mutations, respectively, at an age of 60 years. The diagnosis is difficult due to: (i) clinical symptoms of cancer shared with those of chronic pancreatitis; (ii) the parenchymal and ductal remodeling of chronic pancreatitis rendering imaging analysis difficult; and (iii) differential diagnoses, such as pseudo-tumorous chronic pancreatitis and paraduodenal pancreatitis. Nevertheless, the occurrence of cancer during chronic pancreatitis must be suspected in the case of back pain, weight loss, unbalanced diabetes, and jaundice, despite alcohol withdrawal. Imaging must be systematically reviewed. Endoscopic ultrasound-guided fine-needle biopsy can contribute by targeting suspicious tissue areas with the help of molecular biology (search for KRAS, TP53, CDKN2A, DPC4 mutations). Short-term follow-up of patients is necessary at the clinical and paraclinical levels to try to diagnose cancer at a surgically curable stage. Pancreatic surgery is sometimes necessary if there is any doubt.
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13
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Dai M, Chen S, Teng X, Chen K, Cheng W. KRAS as a Key Oncogene in the Clinical Precision Diagnosis and Treatment of Pancreatic Cancer. J Cancer 2022; 13:3209-3220. [PMID: 36118526 PMCID: PMC9475360 DOI: 10.7150/jca.76695] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2022] [Accepted: 08/19/2022] [Indexed: 11/06/2022] Open
Abstract
Pancreatic ductal adenocarcinoma (PDAC) is one of the most malignant tumors, with a 5-year survival rate of less than 10%. At present, the comprehensive treatment based on surgery, radiotherapy and chemotherapy has encountered a bottleneck, and targeted immunotherapy turns to be the direction of future development. About 90% of PDAC patients have KRAS mutations, and KRAS has been widely used in the diagnosis, treatment, and prognosis of PDAC in recent years. With the development of liquid biopsy and gene testing, KRAS is expected to become a new biomarker to assist the stratification and prognosis of PDAC patients. An increasing number of small molecule inhibitors acting on the KRAS pathway are being developed and put into the clinic, providing more options for PDAC patients.
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Affiliation(s)
- Manxiong Dai
- Department of Hepatobiliary Surgery, Hunan Provincial People's Hospital, The First Affiliated Hospital of Hunan Normal University, Changsha, 410005 Hunan Province, China.,Translational Medicine Laboratory of Pancreas Disease of Hunan Normal University, Changsha 410005, China
| | - Shaofeng Chen
- Department of Hepatobiliary Surgery, Hunan Provincial People's Hospital, The First Affiliated Hospital of Hunan Normal University, Changsha, 410005 Hunan Province, China.,Translational Medicine Laboratory of Pancreas Disease of Hunan Normal University, Changsha 410005, China
| | - Xiong Teng
- Department of Hepatobiliary Surgery, Hunan Provincial People's Hospital, The First Affiliated Hospital of Hunan Normal University, Changsha, 410005 Hunan Province, China.,Translational Medicine Laboratory of Pancreas Disease of Hunan Normal University, Changsha 410005, China
| | - Kang Chen
- Department of Hepatobiliary Surgery, Hunan Provincial People's Hospital, The First Affiliated Hospital of Hunan Normal University, Changsha, 410005 Hunan Province, China.,Translational Medicine Laboratory of Pancreas Disease of Hunan Normal University, Changsha 410005, China
| | - Wei Cheng
- Department of Hepatobiliary Surgery, Hunan Provincial People's Hospital, The First Affiliated Hospital of Hunan Normal University, Changsha, 410005 Hunan Province, China.,Xiangyue Hospital Affiliated to Hunan Institute of Parasitic Diseases, National Clinical Center for Schistosomiasis Treatment, Yueyang 414000, Hunan Province, China.,Translational Medicine Laboratory of Pancreas Disease of Hunan Normal University, Changsha 410005, China
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14
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Sheel A, Addison S, Nuguru SP, Manne A. Is Cell-Free DNA Testing in Pancreatic Ductal Adenocarcinoma Ready for Prime Time? Cancers (Basel) 2022; 14:3453. [PMID: 35884515 PMCID: PMC9322623 DOI: 10.3390/cancers14143453] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/16/2022] [Revised: 07/03/2022] [Accepted: 07/13/2022] [Indexed: 11/17/2022] Open
Abstract
Cell-free DNA (cfDNA) testing currently does not have a significant role in PDA management: it is insufficient to diagnose PDA, and its use is primarily restricted to identifying targetable mutations (if tissue is insufficient or unavailable). cfDNA testing has the potential to address critical needs in PDA management, such as pre-operative risk stratification (POR), prognostication, and predicting (and monitoring) treatment response. Prior studies have focused primarily on somatic mutations, specifically KRAS variants, and have shown limited success in addressing prognosis and POR. Recent studies have demonstrated the importance of other less prevalent mutations (ERBB2 and TP53), but no studies have provided reliable mutation panels for clinical use. Methylation aberrations in cfDNA (epigenetic markers) in PDA have been relatively less explored. However, early evidence has suggested they offer diagnostic and, to some extent, prognostic value. The inclusion of epigenetic markers of cfDNA adds another dimension to genomic testing and may open new therapeutic avenues beyond addressing critical areas of need in PDA treatment. For cfDNA to substantially influence PDA management, concerted efforts are required to include less frequent mutations and epigenetic markers. Furthermore, relying on KRAS mutations for PDA management will always be inadequate.
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Affiliation(s)
- Ankur Sheel
- Department of Internal Medicine, The Ohio State University College of Medicine, Columbus, OH 432120, USA;
| | - Sarah Addison
- School of Medicine, The Ohio State University, Columbus, OH 432120, USA;
| | - Surya Pratik Nuguru
- Department of Internal Medicine, Kamineni Academy of Medical Sciences and Research Center, Hyderabad 500012, India;
| | - Ashish Manne
- Department of Internal Medicine, Division of Medical Oncology at the Arthur G. James Cancer Hospital and Richard J. Solove Research Institute, The Ohio State University Comprehensive Cancer Center, Columbus, OH 43210, USA
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15
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Ahmad E, Ali A, Nimisha, Kumar Sharma A, Apurva, Kumar A, Dar GM, Sumayya Abdul Sattar R, Verma R, Mahajan B, Singh Saluja S. Molecular markers in cancer. Clin Chim Acta 2022; 532:95-114. [DOI: https:/doi.org/10.1016/j.cca.2022.05.029] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 09/28/2023]
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16
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Zhang W, Zhao S, Xie Z, Chen S, Huang Y, Zhao Z, Yi G. The fluorescence amplification strategy based on 3D DNA walker and CRISPR/Cas12a for the rapid detection of BRAF V600E. ANAL SCI 2022; 38:1057-1066. [DOI: 10.1007/s44211-022-00131-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2022] [Accepted: 05/09/2022] [Indexed: 11/01/2022]
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17
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Ahmad E, Ali A, Nimisha, Kumar Sharma A, Apurva, Kumar A, Mehdi G, Sumayya Abdul Sattar R, Verma R, Mahajan B, Singh Saluja S. Molecular markers in cancer. Clin Chim Acta 2022; 532:95-114. [DOI: 10.1016/j.cca.2022.05.029] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/16/2022] [Revised: 05/31/2022] [Accepted: 05/31/2022] [Indexed: 12/01/2022]
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18
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Wang R, Zhao Y, Wang Y, Zhao Z, Chen Q, Duan Y, Xiong S, Luan Z, Wang J, Cheng B. Diagnostic and Prognostic Values of KRAS Mutations on EUS-FNA Specimens and Circulating Tumor DNA in Patients With Pancreatic Cancer. Clin Transl Gastroenterol 2022; 13:e00487. [PMID: 35351843 PMCID: PMC9132521 DOI: 10.14309/ctg.0000000000000487] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/14/2021] [Accepted: 03/25/2022] [Indexed: 11/17/2022] Open
Abstract
INTRODUCTION The ability of carbohydrate antigen 19-9 (CA19-9) to differentiate pancreatic cancer from other benign pancreatic lesions is unsatisfactory. This study explored the diagnostic value of KRAS gene mutations and plasma circulating tumor DNA (ctDNA) in patients with pancreatic cancer. METHODS The prospective cohort study comprised 149 consecutive patients with solid pancreatic lesions who underwent endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA). KRAS subtype mutations were analyzed by digital droplet PCR (ddPCR) in EUS-FNA histopathology tissue samples, and blood samples were sent for plasma ctDNA analysis. The final diagnosis was based on surgical resection pathology or follow-up for at least 2 years. RESULTS Adding KRAS mutation ddPCR increased the sensitivity and accuracy of EUS-FNA from 71.4% to 91.6% (P < 0.001) and 75.8% to 88.6% (P < 0.001), respectively. By comparison, the sensitivities of circulating biomarkers ctDNA and CA19-9 were only 35.2% and 71.2%. The area under the curve of the receiver operating characteristic curve (AUC) of EUS-FNA and KRAS ddPCR combination was >0.90 for distinguishing pancreatic cancer from benign lesions, whereas the AUC of EUS-FNA and CA19-9 combination was 0.83. The median survival time was significantly shorter in patients with G12D KRAS mutations than that in patients with other mutations (180 vs 240 days, P < 0.001). DISCUSSION FNA tissue sample KRAS mutation analysis in tissues significantly improves the diagnostic accuracy of cyto/histopathological evaluation in EUS-FNA samples. The combination of EUS-FNA and tissue sample KRAS ddPCR provided a more accurate method for pancreatic cancer diagnosis, superior to the combination of EUS-FNA and CA19-9/ctDNA. G12D KRAS mutations in pancreatic cancer were independently associated with poor overall survival.
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Affiliation(s)
- Ronghua Wang
- Department of Gastroenterology and Hepatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
- Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA
| | - Yuchong Zhao
- Department of Gastroenterology and Hepatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Yun Wang
- Department of Gastroenterology and Hepatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Zhenxiong Zhao
- Department of Gastroenterology and Hepatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Qian Chen
- Department of Gastroenterology and Hepatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Yaqi Duan
- Department of Pathology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Si Xiong
- Department of Gastroenterology and Hepatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Zhou Luan
- Department of Gastroenterology and Hepatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Jinlin Wang
- Department of Gastroenterology and Hepatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
- Department of Oncology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Bin Cheng
- Department of Gastroenterology and Hepatology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
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19
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Yan TB, Huang JQ, Huang SY, Ahir BK, Li LM, Mo ZN, Zhong JH. Advances in the Detection of Pancreatic Cancer Through Liquid Biopsy. Front Oncol 2021; 11:801173. [PMID: 34993149 PMCID: PMC8726483 DOI: 10.3389/fonc.2021.801173] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2021] [Accepted: 12/06/2021] [Indexed: 01/27/2023] Open
Abstract
Pancreatic cancer refers to the development of malignant tumors in the pancreas: it is associated with high mortality rates and mostly goes undetected in its early stages for lack of symptoms. Currently, surgical treatment is the only effective way to improve the survival of pancreatic cancer patients. Therefore, it is crucial to diagnose the disease as early as possible in order to improve the survival rate of patients with pancreatic cancer. Liquid biopsy is a unique in vitro diagnostic technique offering the advantage of earlier detection of tumors. Although liquid biopsies have shown promise for screening for certain cancers, whether they are effective for early diagnosis of pancreatic cancer is unclear. Therefore, we reviewed relevant literature indexed in PubMed and collated updates and information on advances in the field of liquid biopsy with respect to the early diagnosis of pancreatic cancer.
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Affiliation(s)
- Tian-Bao Yan
- Department of Hepatobiliary Surgery, Guangxi Medical University Cancer Hospital, Nanning, China
- Center for Genomics and Personalized Medicine, Guangxi Key Laboratory for Genomics and Personalized Medicine, Guangxi Collaborative Innovation Center for Genomics and Personalized Medicine, Guangxi Medical University, Nanning, China
| | - Jia-Qi Huang
- Department of Hepatobiliary Surgery, Guangxi Medical University Cancer Hospital, Nanning, China
- Center for Genomics and Personalized Medicine, Guangxi Key Laboratory for Genomics and Personalized Medicine, Guangxi Collaborative Innovation Center for Genomics and Personalized Medicine, Guangxi Medical University, Nanning, China
| | - Shi-Yun Huang
- Department of Hepatobiliary Surgery, Guangxi Medical University Cancer Hospital, Nanning, China
- Center for Genomics and Personalized Medicine, Guangxi Key Laboratory for Genomics and Personalized Medicine, Guangxi Collaborative Innovation Center for Genomics and Personalized Medicine, Guangxi Medical University, Nanning, China
| | - Bhavesh K. Ahir
- Section of Hematology and Oncology, Department of Medicine, University of Illinois at Chicago, Chicago, IL, United States
| | - Long-Man Li
- Center for Genomics and Personalized Medicine, Guangxi Key Laboratory for Genomics and Personalized Medicine, Guangxi Collaborative Innovation Center for Genomics and Personalized Medicine, Guangxi Medical University, Nanning, China
| | - Zeng-Nan Mo
- Center for Genomics and Personalized Medicine, Guangxi Key Laboratory for Genomics and Personalized Medicine, Guangxi Collaborative Innovation Center for Genomics and Personalized Medicine, Guangxi Medical University, Nanning, China
| | - Jian-Hong Zhong
- Department of Hepatobiliary Surgery, Guangxi Medical University Cancer Hospital, Nanning, China
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20
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Hipp J, Hussung S, Timme-Bronsert S, Boerries M, Biesel E, Fichtner-Feigl S, Fritsch R, Wittel UA. Perioperative cell-free mutant KRAS dynamics in patients with pancreatic cancer. Br J Surg 2021; 108:239-243. [PMID: 33793718 DOI: 10.1093/bjs/znaa116] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2020] [Revised: 06/22/2020] [Accepted: 11/03/2020] [Indexed: 12/24/2022]
Abstract
This prospective observational biomarker trial evaluated the diagnostic and prognostic value of circulating KRAS mutations (cmKRAS) and their perioperative dynamics in patients with resectable pancreatic ductal adenocarcinoma (PDAC). Plasma cmKRAS samples (G12D, G12V, G12R, and G12C) were analysed by droplet digital PCR in 51 patients with resectable PDAC, 20 with advanced PDAC, and 34 with non-malignant pancreatic pathology. Preoperative detection of cmKRAS alone did not correlate with poorer overall and disease-free survival in this patient cohort. However, a perioperative change in cmKRAS, particularly accurate when an intraoperative sample was included, was identified as a new and useful marker for prediction of prolonged survival.
Promising biomarker
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Affiliation(s)
- J Hipp
- Department of General and Visceral Surgery, Centre of Surgery, Medical Centre, Faculty of Medicine, University of Freiburg, Freiburg, Germany
| | - S Hussung
- Department of Medicine (Haematology, Oncology and Stem Cell Transplantation), Medical Centre, Faculty of Medicine, University of Freiburg, Freiburg, Germany.,German Cancer Consortium (DKTK) Partner Site Freiburg, Freiburg, Germany.,German Cancer Research Centre (DKFZ), Heidelberg, Germany
| | - S Timme-Bronsert
- Institute for Surgical Pathology, Medical Centre, Faculty of Medicine, University of Freiburg, Freiburg, Germany.,Tumorbank Comprehensive Cancer Centre Freiburg, Freiburg, Germany
| | - M Boerries
- German Cancer Consortium (DKTK) Partner Site Freiburg, Freiburg, Germany.,German Cancer Research Centre (DKFZ), Heidelberg, Germany.,Institute of Medical Bioinformatics and System Medicine, Medical Centre, Faculty of Medicine, University of Freiburg, Freiburg, Germany.,Comprehensive Cancer Centre Freiburg (CCCF), Freiburg, Germany
| | - E Biesel
- Department of General and Visceral Surgery, Centre of Surgery, Medical Centre, Faculty of Medicine, University of Freiburg, Freiburg, Germany
| | - S Fichtner-Feigl
- Department of General and Visceral Surgery, Centre of Surgery, Medical Centre, Faculty of Medicine, University of Freiburg, Freiburg, Germany
| | - R Fritsch
- Department of Medicine (Haematology, Oncology and Stem Cell Transplantation), Medical Centre, Faculty of Medicine, University of Freiburg, Freiburg, Germany.,German Cancer Consortium (DKTK) Partner Site Freiburg, Freiburg, Germany.,German Cancer Research Centre (DKFZ), Heidelberg, Germany.,Department of Medical Oncology and Haematology, Zurich University Hospital, Zurich, Switzerland
| | - U A Wittel
- Department of General and Visceral Surgery, Centre of Surgery, Medical Centre, Faculty of Medicine, University of Freiburg, Freiburg, Germany
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21
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Xue Z, You M, Peng P, Tong H, He W, Li A, Mao P, Xu T, Xu F, Yao C. Taqman-MGB nanoPCR for Highly Specific Detection of Single-Base Mutations. Int J Nanomedicine 2021; 16:3695-3705. [PMID: 34113098 PMCID: PMC8185130 DOI: 10.2147/ijn.s310254] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2021] [Accepted: 05/11/2021] [Indexed: 12/12/2022] Open
Abstract
PURPOSE Detection of single-base mutations is important for real-time monitoring of tumor progression, therapeutic effects, and drug resistance. However, the specific detection of single-base mutations from excessive wild-type background sequences with routine PCR technology remains challenging. Our objective is to develop a simple and highly specific qPCR-based single-base mutation detection method. METHODS Using EGRF T790M as a model, gold nanoparticles at different concentrations were separately added into the Taqman-MGB qPCR system to test specificity improvement, leading to the development of the optimal Taqman-MGB nanoPCR system. Then, these optimal conditions were used to test the range of improvement in the specificity of mutant-type and wild-type templates and the detection limit of mutation abundances in a spiked sample. RESULTS The Taqman-MGB nanoPCR was established based on the traditional qPCR, with significantly suppressed background noise and improved specificity for single-base mutation detection. With EGFR T790M as a template, we demonstrated that our Taqman-MGB nanoPCR system could improve specificity across a wide concentration range from 10-9 μM to 10 μM and detect as low as 0.95% mutation abundance in spiked samples, which is lower than what the traditional Taqman-MGB qPCR and existing PCR methods can detect. Moreover, we also proposed an experimentally validated barrier hypothesis for the mechanism of improved specificity. CONCLUSION The developed Taqman-MGB nanoPCR system could be a powerful tool for clinical single-base mutation detection.
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Affiliation(s)
- Zhenrui Xue
- Department of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University, Chongqing, 400038, People’s Republic of China
- The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
- Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
| | - Minli You
- The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
- Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
| | - Ping Peng
- Department of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University, Chongqing, 400038, People’s Republic of China
- The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
- Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
| | - Haoyang Tong
- The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
- Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
| | - Wanghong He
- The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
- Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
- Key Laboratory of Shaanxi Province for Craniofacial Precision Medicine Research, College of Stomatology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
| | - Ang Li
- Key Laboratory of Shaanxi Province for Craniofacial Precision Medicine Research, College of Stomatology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
| | - Ping Mao
- Department of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University, Chongqing, 400038, People’s Republic of China
- The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
- Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
| | - Ting Xu
- Department of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University, Chongqing, 400038, People’s Republic of China
| | - Feng Xu
- The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
- Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an, 710049, People’s Republic of China
| | - Chunyan Yao
- Department of Transfusion Medicine, Southwest Hospital, Third Military Medical University, Army Medical University, Chongqing, 400038, People’s Republic of China
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22
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Cazacu IM, Semaan A, Stephens B, Swartzlander DB, Guerrero PA, Singh BS, Lungulescu CV, Danciulescu MM, Cherciu Harbiyeli IF, Streata I, Popescu C, Saftoiu A, Roy-Chowdhuri S, Maitra A, Bhutani MS. Diagnostic value of digital droplet polymerase chain reaction and digital multiplexed detection of single-nucleotide variants in pancreatic cytology specimens collected by EUS-guided FNA. Gastrointest Endosc 2021; 93:1142-1151.e2. [PMID: 33058885 DOI: 10.1016/j.gie.2020.09.051] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/31/2020] [Accepted: 09/29/2020] [Indexed: 02/08/2023]
Abstract
BACKGROUND AND AIMS EUS-guided FNA is recommended as a first-line procedure for the histopathologic diagnosis of pancreatic cancer. Molecular analysis of EUS-FNA samples might be used as an auxiliary tool to strengthen the diagnosis. The current study aimed to evaluate the diagnostic performances of K-ras testing using droplet digital polymerase chain reaction (ddPCR) and a novel single-nucleotide variant (SNV) assay performed on pancreatic EUS-FNA samples. METHODS EUS-FNA specimens from 31 patients with pancreatic masses (22 pancreatic ductal adenocarcinomas, 7 chronic pancreatitis, and 2 pancreatic neuroendocrine tumors) were included in the study. K-ras testing was initially performed by ddPCR. In addition, mutational status was evaluated using an SNV assay by NanoString technology, using digital enumeration of unique barcoded probes to detect 97 SNVs from 24 genes of clinical significance. RESULTS The overall specificity and sensitivity of cytologic examination were 100% and 63%, respectively. K-ras mutation testing was performed using ddPCR, and the sensitivity increased to 87% with specificity 90%. The SNV assay detected at least 1 variant in 90% of pancreatic ductal adenocarcinoma samples; the test was able to detect 2 K-ras codon 61 mutations in 2 cases of pancreatic ductal adenocarcinoma, which were missed by ddPCR. The overall diagnostic accuracy of the cytologic examination alone was 74%, and it increased to 91% when the results of both molecular tests were considered for the cases with negative and inconclusive results. CONCLUSIONS The current study illustrated that integration of K-ras analysis with cytologic evaluation, especially in inconclusive cases, can enhance the diagnostic accuracy of EUS-FNA for pancreatic lesions.
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Affiliation(s)
- Irina M Cazacu
- Department of Gastroenterology, Hepatology and Nutrition, The University of Texas, MD Anderson Cancer Center, Houston, Texas, USA; Research Center of Gastroenterology and Hepatology, Craiova, Romania
| | - Alexander Semaan
- Sheikh Ahmed Pancreatic Cancer Research Center, The University of Texas, MD Anderson Cancer Center, Houston, Texas, USA
| | - Bret Stephens
- Sheikh Ahmed Pancreatic Cancer Research Center, The University of Texas, MD Anderson Cancer Center, Houston, Texas, USA
| | - Daniel B Swartzlander
- Sheikh Ahmed Pancreatic Cancer Research Center, The University of Texas, MD Anderson Cancer Center, Houston, Texas, USA
| | - Paola A Guerrero
- Sheikh Ahmed Pancreatic Cancer Research Center, The University of Texas, MD Anderson Cancer Center, Houston, Texas, USA
| | - Ben S Singh
- Department of Gastroenterology, Hepatology and Nutrition, The University of Texas, MD Anderson Cancer Center, Houston, Texas, USA
| | | | | | | | - Ioana Streata
- Research Center of Gastroenterology and Hepatology, Craiova, Romania
| | - Carmen Popescu
- Research Center of Gastroenterology and Hepatology, Craiova, Romania
| | - Adrian Saftoiu
- Research Center of Gastroenterology and Hepatology, Craiova, Romania
| | - Sinchita Roy-Chowdhuri
- Department of Pathology, The University of Texas MD Anderson Cancer Center, Houston, Texas, USA
| | - Anirban Maitra
- Sheikh Ahmed Pancreatic Cancer Research Center, The University of Texas, MD Anderson Cancer Center, Houston, Texas, USA
| | - Manoop S Bhutani
- Department of Gastroenterology, Hepatology and Nutrition, The University of Texas, MD Anderson Cancer Center, Houston, Texas, USA
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Heredia-Soto V, Rodríguez-Salas N, Feliu J. Liquid Biopsy in Pancreatic Cancer: Are We Ready to Apply It in the Clinical Practice? Cancers (Basel) 2021; 13:1986. [PMID: 33924143 PMCID: PMC8074327 DOI: 10.3390/cancers13081986] [Citation(s) in RCA: 41] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2021] [Revised: 04/14/2021] [Accepted: 04/16/2021] [Indexed: 12/11/2022] Open
Abstract
Pancreatic ductal adenocarcinoma (PDAC) exhibits the poorest prognosis of all solid tumors, with a 5-year survival of less than 10%. To improve the prognosis, it is necessary to advance in the development of tools that help us in the early diagnosis, treatment selection, disease monitoring, evaluation of the response and prognosis. Liquid biopsy (LB), in its different modalities, represents a particularly interesting tool for these purposes, since it is a minimally invasive and risk-free procedure that can detect both the presence of genetic material from the tumor and circulating tumor cells (CTCs) in the blood and therefore distantly reflect the global status of the disease. In this work we review the current status of the main LB modalities (ctDNA, exosomes, CTCs and cfRNAs) for detecting and monitoring PDAC.
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Affiliation(s)
- Victoria Heredia-Soto
- Translational Oncology Research Laboratory, Biomedical Research Institute, La Paz University Hospital, IdiPAZ, Paseo de la Castellana 261, 28046 Madrid, Spain; (V.H.-S.); (N.R.-S.)
- Centro de Investigación Biomédica en Red de Cáncer, CIBERONC, Instituto de Salud Carlos III, Monforte de Lemos 5, 28029 Madrid, Spain
| | - Nuria Rodríguez-Salas
- Translational Oncology Research Laboratory, Biomedical Research Institute, La Paz University Hospital, IdiPAZ, Paseo de la Castellana 261, 28046 Madrid, Spain; (V.H.-S.); (N.R.-S.)
- Centro de Investigación Biomédica en Red de Cáncer, CIBERONC, Instituto de Salud Carlos III, Monforte de Lemos 5, 28029 Madrid, Spain
- Cátedra UAM-AMGEN, Medical Oncology Department, La Paz University Hospital, Paseo de la Castellana 261, 28046 Madrid, Spain
| | - Jaime Feliu
- Translational Oncology Research Laboratory, Biomedical Research Institute, La Paz University Hospital, IdiPAZ, Paseo de la Castellana 261, 28046 Madrid, Spain; (V.H.-S.); (N.R.-S.)
- Centro de Investigación Biomédica en Red de Cáncer, CIBERONC, Instituto de Salud Carlos III, Monforte de Lemos 5, 28029 Madrid, Spain
- Cátedra UAM-AMGEN, Medical Oncology Department, La Paz University Hospital, Paseo de la Castellana 261, 28046 Madrid, Spain
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Sivapalan L, Kocher H, Ross-Adams H, Chelala C. Molecular profiling of ctDNA in pancreatic cancer: Opportunities and challenges for clinical application. Pancreatology 2021; 21:363-378. [PMID: 33451936 PMCID: PMC7994018 DOI: 10.1016/j.pan.2020.12.017] [Citation(s) in RCA: 33] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/05/2020] [Revised: 12/16/2020] [Accepted: 12/21/2020] [Indexed: 01/10/2023]
Abstract
Pancreatic ductal adenocarcinoma (PDAC) is predicted to become the second leading cause of cancer-related mortality within the next decade, with limited effective treatment options and a dismal long-term prognosis for patients. Genomic profiling has not yet manifested clinical benefits for diagnosis, treatment or prognosis in PDAC, due to the lack of available tissues for sequencing and the confounding effects of low tumour cellularity in many biopsy specimens. Increasing focus is now turning to the use of minimally invasive liquid biopsies to enhance the characterisation of actionable PDAC tumour genomes. Circulating tumour DNA (ctDNA) is the most comprehensively studied liquid biopsy analyte in blood and can provide insight into the molecular profile and biological characteristics of individual PDAC tumours, in real-time and in advance of traditional imaging modalities. This can pave the way for identification of new therapeutic targets, novel risk variants and markers of tumour response, to supplement diagnostic screening and provide enhanced scrutiny in treatment stratification. In the roadmap towards the application of precision medicine for clinical management in PDAC, ctDNA analyses may serve a leading role in streamlining candidate biomarkers for clinical integration. In this review, we highlight recent developments in the use of ctDNA-based liquid biopsies for PDAC and provide new insights into the technical, analytical and biological challenges that must be overcome for this potential to be realised.
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Affiliation(s)
- L. Sivapalan
- Centre for Cancer Biomarkers and Biotherapeutics, Barts Cancer Institute, Queen Mary University of London, EC1M 6BQ, UK
| | - H.M. Kocher
- Centre for Tumour Biology, Barts Cancer Institute, Queen Mary University of London, EC1M 6BQ, UK
| | - H. Ross-Adams
- Centre for Cancer Biomarkers and Biotherapeutics, Barts Cancer Institute, Queen Mary University of London, EC1M 6BQ, UK
| | - C. Chelala
- Centre for Cancer Biomarkers and Biotherapeutics, Barts Cancer Institute, Queen Mary University of London, EC1M 6BQ, UK,Corresponding author.
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Jaworski JJ, Morgan RD, Sivakumar S. Circulating Cell-Free Tumour DNA for Early Detection of Pancreatic Cancer. Cancers (Basel) 2020; 12:E3704. [PMID: 33317202 PMCID: PMC7763954 DOI: 10.3390/cancers12123704] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2020] [Accepted: 12/04/2020] [Indexed: 01/11/2023] Open
Abstract
Pancreatic cancer is a lethal disease, with mortality rates negatively associated with the stage at which the disease is detected. Early detection is therefore critical to improving survival outcomes. A recent focus of research for early detection is the use of circulating cell-free tumour DNA (ctDNA). The detection of ctDNA offers potential as a relatively non-invasive method of diagnosing pancreatic cancer by using genetic sequencing technology to detect tumour-specific mutational signatures in blood samples before symptoms manifest. These technologies are limited by a number of factors that lower sensitivity and specificity, including low levels of detectable ctDNA in early stage disease and contamination with non-cancer circulating cell-free DNA. However, genetic and epigenetic analysis of ctDNA in combination with other standard diagnostic tests may improve early detection rates. In this review, we evaluate the genetic and epigenetic methods under investigation in diagnosing pancreatic cancer and provide a perspective for future developments.
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Affiliation(s)
- Jedrzej J. Jaworski
- MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH, UK;
| | - Robert D. Morgan
- Department of Medical Oncology, Christie NHS Foundation Trust, Manchester M20 4BX, UK;
- Division of Cancer Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester M13 9PL, UK
| | - Shivan Sivakumar
- Department of Oncology, University of Oxford, Old Road Campus Research Building, Roosevelt Drive, Oxford OX3 7DQ, UK
- Department of Medical Oncology, Oxford University Hospitals NHS Foundation Trust, Oxford OX3 7LE, UK
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Arshad J, Roberts A, Ahmed J, Cotta J, Pico BA, Kwon D, Trent JC. Utility of Circulating Tumor DNA in the Management of Patients With GI Stromal Tumor: Analysis of 243 Patients. JCO Precis Oncol 2020; 4:66-73. [DOI: 10.1200/po.19.00253] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
PURPOSE GI stromal tumor (GIST) is the most common sarcoma of the GI tract. Management of patients with GIST is determined by KIT, PDGFRA, or other genomic alterations. Tissue-based next-generation sequencing (NGS) analysis is the standard approach for diagnosis, prognosis, and treatment selection. However, circulating tumor DNA (ctDNA)–based NGS is a novel and noninvasive alternative. METHODS ctDNA sequencing results were evaluated in blood samples from 243 de-identified patients within the Guardant360 database. Under an approved institutional review board protocol, a retrospective analysis was performed on 45 single-institution patients. RESULTS Of 243 patients, 114 (47%) were women, and the median age was 59 years (range, 17-90 years). Patients with no alterations and variations of uncertain significance were excluded. Of the 162 patients with known pathogenic mutations, KIT was the most common (56%), followed by NF (7%), PDGFRA (6%), PI3KCA (6%), KRAS (5%), and others (6%). Most tumors harbored an actionable KIT or PDGFRA mutation. Our institutional cohort (n = 45) had 16 (35%) KIT exon 11 mutations, 3 (6%) KIT exon 9 mutations, and 1 (2%) PDGFRA mutation detected on ctDNA. Resistance mutations were observed in KIT exon 17 (8 patients), exon 13 (3 patients), and in both (3 patients). Our comparison of ctDNA with tissue NGS revealed a positive predictive value (PPV) of 100%. Failure of concordance was observed in patients with localized or low disease burden. From the time of ctDNA testing, the median overall survival was not reached, whereas the median progression-free survival was 7 months. CONCLUSION ctDNA provides a rapid, noninvasive analysis of current mutations with a high PPV for patients with metastatic GIST. ctDNA-based testing may help to define the optimal choice of therapy on the basis of resistance mutations and should be studied prospectively.
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Affiliation(s)
- Junaid Arshad
- University of Miami, Miller School of Medicine/Sylvester Comprehensive Cancer Center, Miami, FL
| | | | | | - Jared Cotta
- University of Miami, Miller School of Medicine/Sylvester Comprehensive Cancer Center, Miami, FL
| | - Brian A. Pico
- University of Miami, Miller School of Medicine/Sylvester Comprehensive Cancer Center, Miami, FL
| | - Deukoo Kwon
- University of Miami, Miller School of Medicine/Sylvester Comprehensive Cancer Center, Miami, FL
| | - Jonathan C. Trent
- University of Miami, Miller School of Medicine/Sylvester Comprehensive Cancer Center, Miami, FL
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Porta M, Pumarega J, Amaral AFS, Genkinger JM, Camargo J, Mucci L, Alguacil J, Gasull M, Zhang X, Morales E, Iglesias M, Ogino S, Engel LS. Influence of KRAS mutations, persistent organic pollutants, and trace elements on survival from pancreatic ductal adenocarcinoma. ENVIRONMENTAL RESEARCH 2020; 190:109781. [PMID: 32791343 PMCID: PMC7689512 DOI: 10.1016/j.envres.2020.109781] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/28/2020] [Revised: 05/02/2020] [Accepted: 06/02/2020] [Indexed: 05/09/2023]
Abstract
INTRODUCTION Reasons why pancreatic ductal adenocarcinoma (PDAC) continues to have poor survival are only partly known. No previous studies have analyzed the combined influence of KRAS mutations, persistent organic pollutants (POPs), and trace elements upon survival in PDAC or in any other human cancer. OBJECTIVE To analyze the individual and combined influence of KRAS mutations, POPs, and trace elements upon survival from PDAC. METHODS Incident cases of PDAC (n = 185) were prospectively identified in five hospitals in Eastern Spain in 1992-1995 and interviewed face-to-face during hospital admission. KRAS mutational status was determined from tumour tissue through polymerase chain reaction and artificial restriction fragment length polymorphism. Blood and toenail samples were obtained before treatment. Serum concentrations of POPs were analyzed by high-resolution gas chromatography with electron-capture detection. Concentrations of 12 trace elements were determined in toenail samples by inductively coupled plasma mass spectrometry. Multivariable Cox proportional hazards regression was used to assess prognostic associations. RESULTS Patients with a KRAS mutated tumor had a 70% higher risk of early death than patients with a KRAS wild-type PDAC (hazard ratio [HR] = 1.7, p = 0.026), adjusting for age, sex, and tumor stage. KRAS mutational status was only modestly and not statistically significantly associated with survival when further adjusting by treatment or by treatment intention. The beneficial effects of treatment remained unaltered when KRAS mutational status was taken into account, and treatment did not appear to be less effective in the subgroup of patients with a KRAS mutated tumor. POPs did not materially influence survival: the adjusted HR of the highest POP tertiles was near unity for all POPs. When considering the joint effect on survival of POPs and KRAS, patients with KRAS mutated tumors had modest and nonsignificant HRs (most HRs around 1.3 to 1.4). Higher concentrations of lead, cadmium, arsenic, vanadium, and aluminium were associated with better survival. When KRAS status, POPs, and trace elements were simultaneously considered along with treatment, only the latter was statistically significantly related to survival. CONCLUSIONS In this study based on molecular, clinical, and environmental epidemiology, KRAS mutational status, POPs, and trace elements were not adversely related to PDAC survival when treatment was simultaneously considered; only treatment was independently related to survival. The lack of adverse prognostic effects of POPs and metals measured at the time of diagnosis provide scientific and clinical reassurance on the effects of such exposures upon survival of patients with PDAC. The weak association with KRAS mutations contributes to the scant knowledge on the clinical implications of a genetic alteration highly frequent in PDAC.
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Affiliation(s)
- Miquel Porta
- School of Medicine, Universitat Autònoma de Barcelona, Barcelona, Spain; CIBER de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain; Hospital Del Mar Medical Research Institute (IMIM), Barcelona, Spain.
| | - José Pumarega
- CIBER de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain; Hospital Del Mar Medical Research Institute (IMIM), Barcelona, Spain
| | - André F S Amaral
- National Heart and Lung Institute, Imperial College London, London, United Kingdom
| | - Jeanine M Genkinger
- Department of Epidemiology, Columbia University, New York, USA; Herbert Irving Comprehensive Cancer Center, Columbia University Medical Center, New York, USA
| | - Judit Camargo
- School of Medicine, Universitat Autònoma de Barcelona, Barcelona, Spain; Hospital Del Mar Medical Research Institute (IMIM), Barcelona, Spain
| | - Lorelei Mucci
- Harvard Medical School, Harvard T. H. Chan School of Public Health, Brigham and Women's Hospital, Boston, USA
| | - Juan Alguacil
- CIBER de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain; Universidad de Huelva, Huelva, Spain
| | - Magda Gasull
- School of Medicine, Universitat Autònoma de Barcelona, Barcelona, Spain; CIBER de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain; Hospital Del Mar Medical Research Institute (IMIM), Barcelona, Spain
| | - Xuehong Zhang
- Harvard Medical School, Harvard T. H. Chan School of Public Health, Brigham and Women's Hospital, Boston, USA
| | - Eva Morales
- CIBER de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain; IMIB-Arrixaca, Department of Public Health Sciences, University of Murcia
| | - Mar Iglesias
- School of Medicine, Universitat Autònoma de Barcelona, Barcelona, Spain; Hospital Del Mar Medical Research Institute (IMIM), Barcelona, Spain
| | - Shuji Ogino
- Harvard Medical School, Harvard T. H. Chan School of Public Health, Brigham and Women's Hospital, Boston, USA
| | - Lawrence S Engel
- Department of Epidemiology, Gillings School of Global Public Health, University of North Carolina, Chapel Hill, USA
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Grunvald MW, Jacobson RA, Kuzel TM, Pappas SG, Masood A. Current Status of Circulating Tumor DNA Liquid Biopsy in Pancreatic Cancer. Int J Mol Sci 2020; 21:E7651. [PMID: 33081107 PMCID: PMC7589736 DOI: 10.3390/ijms21207651] [Citation(s) in RCA: 39] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/16/2020] [Revised: 10/12/2020] [Accepted: 10/14/2020] [Indexed: 02/06/2023] Open
Abstract
Pancreatic cancer is a challenging disease with a low 5-year survival rate. There are areas for improvement in the tools used for screening, diagnosis, prognosis, treatment selection, and assessing treatment response. Liquid biopsy, particularly cell free DNA liquid biopsy, has shown promise as an adjunct to our standard care for pancreatic cancer patients, but has not yet been universally adopted into regular use by clinicians. In this publication, we aim to review cfDNA liquid biopsy in pancreatic cancer with an emphasis on current techniques, clinical utility, and areas of active investigation. We feel that researchers and clinicians alike should be familiar with this exciting modality as it gains increasing importance in the care of cancer patients.
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Affiliation(s)
- Miles W. Grunvald
- Department of Surgery, Rush University Medical Center, Chicago, IL 60612, USA; (M.W.G.); (R.A.J.); (S.G.P.)
| | - Richard A. Jacobson
- Department of Surgery, Rush University Medical Center, Chicago, IL 60612, USA; (M.W.G.); (R.A.J.); (S.G.P.)
| | - Timothy M. Kuzel
- Division of Hematology/Oncology and Cell Therapy, Rush University Medical Center, Chicago, IL 60612, USA;
| | - Sam G. Pappas
- Department of Surgery, Rush University Medical Center, Chicago, IL 60612, USA; (M.W.G.); (R.A.J.); (S.G.P.)
| | - Ashiq Masood
- Division of Hematology/Oncology and Cell Therapy, Rush University Medical Center, Chicago, IL 60612, USA;
- Rush Precision Oncology Program, Rush University Medical Center, Chicago, IL 60612, USA
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Gaddes DE, Lee PW, Trick AY, Athamanolap P, O'Keefe CM, Puleo C, Hsieh K, Wang TH. Facile Coupling of Droplet Magnetofluidic-Enabled Automated Sample Preparation for Digital Nucleic Acid Amplification Testing and Analysis. Anal Chem 2020; 92:13254-13261. [PMID: 32869628 PMCID: PMC8549765 DOI: 10.1021/acs.analchem.0c02454] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
Digital nucleic acid amplification testing (dNAAT) and analysis techniques, such as digital polymerase chain reaction (PCR), have become useful clinical diagnostic tools. However, nucleic acid (NA) sample preparation preceding dNAAT is generally laborious and performed manually, thus creating the need for a simple sample preparation technique and a facile coupling strategy for dNAAT. Therefore, we demonstrate a simple workflow which automates magnetic bead-based extraction of NAs with a one-step transfer to dNAAT. Specifically, we leverage droplet magnetofluidics (DM) to automate the movement of magnetic beads between small volumes of reagents commonly employed for NA extraction and purification. Importantly, the buffer typically used to elute the NAs off the magnetic beads is replaced by a carefully selected PCR solution, enabling direct transfer from sample preparation to dNAAT. Moreover, we demonstrate the potential for multiplexing using a digital high-resolution melt (dHRM) after the digital PCR (dPCR). The utility of this workflow is demonstrated with duplexed detection of bacteria in a sample imitating a coinfection. We first purify the bacterial DNA into a PCR solution using our DM-based sample preparation. We then transfer the purified bacterial DNA to our microfluidic nanoarray to amplify 16S rRNA using dPCR and then perform dHRM to identify the two bacterial species.
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Affiliation(s)
- David E Gaddes
- Department of Mechanical Engineering, Johns Hopkins University, Baltimore, Maryland 21218, United States
| | - Pei-Wei Lee
- Department of Mechanical Engineering, Johns Hopkins University, Baltimore, Maryland 21218, United States
| | - Alexander Y Trick
- Department of Biomedical Engineering, Johns Hopkins University, Baltimore, Maryland 21205, United States
| | - Pornpat Athamanolap
- Department of Biomedical Engineering, Johns Hopkins University, Baltimore, Maryland 21205, United States
- Department of Biomedical Engineering, Faculty of Engineering, Mahidol University, Nakorn Pathom 73170, Thailand
| | - Christine M O'Keefe
- Department of Biomedical Engineering, Johns Hopkins University, Baltimore, Maryland 21205, United States
| | - Chris Puleo
- Electronics Organization, GE Global Research Center, Niskayuna, New York 12309, United States
| | - Kuangwen Hsieh
- Department of Mechanical Engineering, Johns Hopkins University, Baltimore, Maryland 21218, United States
| | - Tza-Huei Wang
- Department of Mechanical Engineering, Johns Hopkins University, Baltimore, Maryland 21218, United States
- Department of Biomedical Engineering, Johns Hopkins University, Baltimore, Maryland 21205, United States
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Lee J, Kim JH, Kang SH, Yoo HM. Improvement of digital PCR conditions for direct detection of KRAS mutations. J Clin Lab Anal 2020; 34:e23344. [PMID: 32329932 PMCID: PMC7439326 DOI: 10.1002/jcla.23344] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2019] [Revised: 03/05/2020] [Accepted: 03/31/2020] [Indexed: 12/31/2022] Open
Abstract
BACKGROUND In standard analytical conditions, an isolation step is essential for circulating tumor DNA (ctDNA) analysis. The necessity of this step becomes unclear with the development of highly sensitive detection methods. The aim of this study was to evaluate ctDNA mimetic nDNA detection as reference materials (RMs) using dPCR technologies either directly from serum or without serum. METHODS To determine an absolute count of both mutation and wild-type bearing DNA molecules, genomic DNA (gDNA) and nucleosomal DNA (nDNA), which are similar in size to cell-free DNA, were evaluated. We tested 3 KRAS mutations in colorectal cancer cell lines. RESULTS We describe the recent progress in RMs. The short DNA fragments, such as sDNA and nDNA, exhibited higher quantitative values of dPCR compared to gDNA. The efficiency between Atlantis dsDNase (AD) and Micrococcal Nuclease (MN) affects DNA quantification. Moreover, there was a significant difference in dPCR output when spiking gDNA or nDNA containing KRAS mutations into FBS compared to the dPCR output under non-FBS conditions. CONCLUSION The matrix effect crucially affects the accuracy of gDNA and nDNA level estimation in the direct detection of mimic of patient samples. The form of reference material we proposed should be optimized for various conditions to develop reference materials that can accurately measure copy number and verify the detection of KRAS mutations in the matrix.
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Affiliation(s)
- Jina Lee
- Center for BioanalysisKorea Research Institute of Standards and Science (KRISS)DaejeonKorea
- College of PharmacyChungnam National UniversityDaejeonKorea
| | - Ji Hyun Kim
- Center for BioanalysisKorea Research Institute of Standards and Science (KRISS)DaejeonKorea
| | - Sun Hyung Kang
- Division of Gastroenterology and HepatologyDepartment of Internal MedicineChungnam National University School of MedicineDaejeonKorea
| | - Hee Min Yoo
- Center for BioanalysisKorea Research Institute of Standards and Science (KRISS)DaejeonKorea
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A self-powered bidirectional partition microfluidic chip with embedded microwells for highly sensitive detection of EGFR mutations in plasma of non-small cell lung cancer patients. Talanta 2020; 220:121426. [PMID: 32928434 DOI: 10.1016/j.talanta.2020.121426] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2020] [Revised: 07/12/2020] [Accepted: 07/15/2020] [Indexed: 02/07/2023]
Abstract
Circulating tumor DNA (ctDNA) is a promising biomarker for tumor genotyping and therapy monitoring. Herein, we developed a digital PCR chip with embedded microwell and bidirectional partition network for highly sensitive ctDNA analysis. The embedded microwell contributes to increasing microreaction density (up to 7000 microwells/cm2) and reducing evaporation during amplification. The bidirectional partition network can achieve fast and random distribution of targets, ensuring the precise quantification of nucleic acid. We used plasmids, artificial samples and 32 clinical blood samples from non-small cell lung cancer patients to test the performance of this platform. The results demonstrated that our chip has not only comparable quantification performance to commercial counterpart but also the ability to detect EGFR mutations with as low as 0.01% mutation rate and 20 alter molecules in 27 ng genomic DNA. The identification of EGFR mutations in plasma using developed chip exhibited 85.71% sensitivity and 94.44% specificity for L858R mutation and 100% sensitivity and 86.96% specificity for T790 M mutation. Moreover, the monitoring of mutant allele in plasma was accomplished in this work. In conclusion, the developed chip has a potential in lung tumor genotyping and therapy monitoring for precision medicine, even other tumors.
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32
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Leick KM, Kazarian AG, Rajput M, Tomanek-Chalkley A, Miller A, Shrader HR, McCarthy A, Coleman KL, Kasi PM, Chan CHF. Peritoneal Cell-Free Tumor DNA as Biomarker for Peritoneal Surface Malignancies. Ann Surg Oncol 2020; 27:5065-5071. [PMID: 32648179 DOI: 10.1245/s10434-020-08832-9] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2020] [Accepted: 06/27/2020] [Indexed: 12/11/2022]
Abstract
BACKGROUND Disease burden in patients with peritoneal carcinomatosis (PC) is difficult to estimate. We evaluate whether peritoneal cell-free tumor DNA can be used as a measure of disease burden. PATIENTS AND METHODS Malignant ascites or peritoneal lavage fluids were collected from patients with PC under approved IRB protocol. Cell-free DNA was extracted from peritoneal fluid. Droplet digital PCR (ddPCR) was performed using a commercially available KRAS G12/G13 screening kit. Mutant allele frequency (MAF) was calculated based on the numbers of KRAS wild-type and mutant droplets. Clinicopathological, treatment and outcome data were abstracted and correlated with MAF of cell-free KRAS mutant DNA. RESULTS Cell-free KRAS mutant DNA was detected in 15/37 (40%) malignant peritoneal fluids with a MAF of 0.1% to 26.2%. While peritoneal cell-free KRAS mutant DNA was detected in all the patients with KRAS mutant tumors (N = 10), 3/16 (19%) patients with KRAS wild-type tumors also had peritoneal cell-free KRAS mutant DNA. We also found that 71% (5/7) of patients with disease amenable to cytoreductive surgery (CRS) had a MAF of < 1% (median: 0.5%, range: 0.1-4.7%), while 75% (6/8) of patients with unresectable disease had a MAF of > 1% (median: 4.4%, range: 0.1-26.2%). CONCLUSIONS This pilot proof-of-principle study suggests that peritoneal cell-free tumor DNA detected by ddPCR may enable prediction of disease burden and a measure of disease amenable to CRS in patients with PC.
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Affiliation(s)
- Katie M Leick
- Department of Surgery, University of Iowa, Iowa City, IA, USA
| | | | - Maheen Rajput
- Department of Radiology, University of Iowa, Iowa City, IA, USA
| | | | - Ann Miller
- Department of Surgery, University of Iowa, Iowa City, IA, USA
| | | | - Ashley McCarthy
- Holden Comprehensive Cancer Center, University of Iowa, Iowa City, IA, USA
| | - Kristen L Coleman
- Holden Comprehensive Cancer Center, University of Iowa, Iowa City, IA, USA
| | - Pashtoon M Kasi
- Holden Comprehensive Cancer Center, University of Iowa, Iowa City, IA, USA.,Internal Medicine, University of Iowa, Iowa City, IA, USA
| | - Carlos H F Chan
- Department of Surgery, University of Iowa, Iowa City, IA, USA. .,Holden Comprehensive Cancer Center, University of Iowa, Iowa City, IA, USA.
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Yuanfeng P, Ruiyi L, Xiulan S, Guangli W, Zaijun L. Highly sensitive electrochemical detection of circulating tumor DNA in human blood based on urchin-like gold nanocrystal-multiple graphene aerogel and target DNA-induced recycling double amplification strategy. Anal Chim Acta 2020; 1121:17-25. [DOI: 10.1016/j.aca.2020.04.077] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/10/2020] [Revised: 04/26/2020] [Accepted: 04/30/2020] [Indexed: 12/12/2022]
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34
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Abdallah R, Taly V, Zhao S, Pietrasz D, Bachet JB, Basile D, Mas L, Zaanan A, Laurent-Puig P, Taieb J. Plasma circulating tumor DNA in pancreatic adenocarcinoma for screening, diagnosis, prognosis, treatment and follow-up: A systematic review. Cancer Treat Rev 2020; 87:102028. [PMID: 32485509 DOI: 10.1016/j.ctrv.2020.102028] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2020] [Revised: 04/27/2020] [Accepted: 04/29/2020] [Indexed: 12/19/2022]
Abstract
While no biomarker is currently recommended for the management of pancreatic adenocarcinoma (PA), circulating tumor DNA (ctDNA) seems promising but little is known on how it may help to manage our patients in the near future. This systematic review of literature was designed to explore the current knowledge on ctDNA as a screening, diagnostic, prognostic, predictive and theranostic biomarker in the management of PA. We retrieved 62 full-text articles, 3 meta-analyses, 2 clinical trials, 1 abstract and 13 ongoing trials. Results were categorized into sections about screening, diagnosis, prognosis and follow-up of localized and advanced PA together with possible theranostics applications. Although its specificity is excellent, the current sensitivity of ctDNA remains a limitation especially in patients without metastatic disease. Therefore, this biomarker cannot be currently used as a screening or diagnostic tool. Increasing evidence suggests that ctDNA is a relevant candidate biomarker to assess minimal residual disease after radical surgery, but also a strong independent biomarker linked to a poor prognosis in advanced PA. Some recent data also indicates that ctDNA is an attractive biomarker for longitudinal follow-up and possibly early treatment adaptation. Its role in tumor profiling in advanced disease to decide targeted treatments remains to be explored. Altogether, ctDNA appears to be a reliable prognostic tool. Though promising results have been reported, further studies are still needed to define exactly how ctDNA can help physicians in the screening, diagnosis and treatment, as PA is expected to become a major cause of cancer-related deaths in the forthcoming decade.
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Affiliation(s)
- Raëf Abdallah
- Université de Paris, Department of Hepatogastroenterology and GI Oncology, Georges Pompidou European Hospital, APHP Centre, Paris, France; Centre de Recherche des Cordeliers, INSERM, CNRS, Sorbonne Université, USPC, Université Paris Descartes, Université Paris Diderot, Equipe labellisée Ligue Nationale contre le cancer, Paris, France
| | - Valérie Taly
- Centre de Recherche des Cordeliers, INSERM, CNRS, Sorbonne Université, USPC, Université Paris Descartes, Université Paris Diderot, Equipe labellisée Ligue Nationale contre le cancer, Paris, France
| | - Shulin Zhao
- Centre de Recherche des Cordeliers, INSERM, CNRS, Sorbonne Université, USPC, Université Paris Descartes, Université Paris Diderot, Equipe labellisée Ligue Nationale contre le cancer, Paris, France
| | - Daniel Pietrasz
- Centre de Recherche des Cordeliers, INSERM, CNRS, Sorbonne Université, USPC, Université Paris Descartes, Université Paris Diderot, Equipe labellisée Ligue Nationale contre le cancer, Paris, France
| | - Jean-Baptiste Bachet
- Department of Hepatogastroenterology and GI Oncology, La Pitié-Salpêtrière Hospital, Paris, INSERM UMRS 1138, Université de Paris, Paris, France
| | - Debora Basile
- Université de Paris, Department of Hepatogastroenterology and GI Oncology, Georges Pompidou European Hospital, APHP Centre, Paris, France; Department of Medicine (DAME), University of Udine, Italy
| | - Léo Mas
- Department of Hepatogastroenterology and GI Oncology, La Pitié-Salpêtrière Hospital, Paris, INSERM UMRS 1138, Université de Paris, Paris, France
| | - Aziz Zaanan
- Université de Paris, Department of Hepatogastroenterology and GI Oncology, Georges Pompidou European Hospital, APHP Centre, Paris, France; Centre de Recherche des Cordeliers, INSERM, CNRS, Sorbonne Université, USPC, Université Paris Descartes, Université Paris Diderot, Equipe labellisée Ligue Nationale contre le cancer, Paris, France
| | - Pierre Laurent-Puig
- Université de Paris, Department of Hepatogastroenterology and GI Oncology, Georges Pompidou European Hospital, APHP Centre, Paris, France
| | - Julien Taieb
- Université de Paris, Department of Hepatogastroenterology and GI Oncology, Georges Pompidou European Hospital, APHP Centre, Paris, France; Centre de Recherche des Cordeliers, INSERM, CNRS, Sorbonne Université, USPC, Université Paris Descartes, Université Paris Diderot, Equipe labellisée Ligue Nationale contre le cancer, Paris, France.
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35
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Macgregor-Das A, Yu J, Tamura K, Abe T, Suenaga M, Shindo K, Borges M, Koi C, Kohi S, Sadakari Y, Dal Molin M, Almario JA, Ford M, Chuidian M, Burkhart R, He J, Hruban RH, Eshleman JR, Klein AP, Wolfgang CL, Canto MI, Goggins M. Detection of Circulating Tumor DNA in Patients with Pancreatic Cancer Using Digital Next-Generation Sequencing. J Mol Diagn 2020; 22:748-756. [PMID: 32205290 DOI: 10.1016/j.jmoldx.2020.02.010] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2019] [Revised: 01/08/2020] [Accepted: 02/26/2020] [Indexed: 12/19/2022] Open
Abstract
Circulating tumor DNA (ctDNA) measurements can be used to estimate tumor burden, but avoiding false-positive results is challenging. Herein, digital next-generation sequencing (NGS) is evaluated as a ctDNA detection method. Plasma KRAS and GNAS hotspot mutation levels were measured in 140 subjects, including 67 with pancreatic ductal adenocarcinoma and 73 healthy and disease controls. To limit chemical modifications of DNA that yield false-positive mutation calls, plasma DNA was enzymatically pretreated, after which DNA was aliquoted for digital detection of mutations (up to 384 aliquots/sample) by PCR and NGS. A digital NGS score of two SDs above the mean in controls was considered positive. Thirty-seven percent of patients with pancreatic cancer, including 31% of patients with stages I/II disease, had positive KRAS codon 12 ctDNA scores; only one patient had a positive GNAS mutation score. Two disease control patients had positive ctDNA scores. Low-normal-range digital NGS scores at mutation hotspots were found at similar levels in healthy and disease controls, usually at sites of cytosine deamination, and were likely the result of chemical modification of plasma DNA and NGS error rather than true mutations. Digital NGS detects mutated ctDNA in patients with pancreatic cancer with similar yield to other methods. Detection of low-level, true-positive ctDNA is limited by frequent low-level detection of false-positive mutation calls in plasma DNA from controls.
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Affiliation(s)
- Anne Macgregor-Das
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Jun Yu
- Department of Surgery, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Koji Tamura
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Toshiya Abe
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Masaya Suenaga
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Koji Shindo
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Michael Borges
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Chiho Koi
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Shiro Kohi
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Yoshihiko Sadakari
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Marco Dal Molin
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Jose A Almario
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Madeline Ford
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Miguel Chuidian
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Richard Burkhart
- Department of Surgery, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Jin He
- Department of Surgery, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Ralph H Hruban
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland; Department of Oncology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - James R Eshleman
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland; Department of Oncology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Alison P Klein
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland; Department of Oncology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Christopher L Wolfgang
- Department of Surgery, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Marcia I Canto
- Department of Oncology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland; Department of Medicine, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland
| | - Michael Goggins
- Department of Pathology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland; Department of Oncology, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland; Department of Medicine, The Sol Goldman Pancreatic Cancer Research Center, Johns Hopkins Medical Institutions, Baltimore, Maryland.
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36
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Buscail L, Bournet B, Cordelier P. Role of oncogenic KRAS in the diagnosis, prognosis and treatment of pancreatic cancer. Nat Rev Gastroenterol Hepatol 2020; 17:153-168. [PMID: 32005945 DOI: 10.1038/s41575-019-0245-4] [Citation(s) in RCA: 449] [Impact Index Per Article: 89.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 11/21/2019] [Indexed: 02/08/2023]
Abstract
Pancreatic ductal adenocarcinoma (PDAC) is predicted to be the second most common cause of death within the next 10 years. The prognosis for this disease is poor despite diagnostic progress and new chemotherapeutic regimens. The oncogenic KRAS mutation is the major event in pancreatic cancer; it confers permanent activation of the KRAS protein, which acts as a molecular switch to activate various intracellular signalling pathways and transcription factors inducing cell proliferation, migration, transformation and survival. Several laboratory methods have been developed to detect KRAS mutations in biological samples, including digital droplet PCR (which displays high sensitivity). Clinical studies have revealed that a KRAS mutation assay in fine-needle aspiration material combined with cytopathology increases the sensitivity, accuracy and negative predictive value of cytopathology for a positive diagnosis of pancreatic cancer. In addition, the presence of KRAS mutations in serum and plasma (liquid biopsies) correlates with a worse prognosis. The presence of mutated KRAS can also have therapeutic implications, whether at the gene level per se, during its post-translational maturation, interaction with nucleotides and after activation of the various oncogenic signals. Further pharmacokinetic and toxicological studies on new molecules are required, especially small synthetic molecules, before they can be used in the therapeutic arsenal for pancreatic ductal adenocarcinoma.
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Affiliation(s)
- Louis Buscail
- Department of Gastroenterology, University of Toulouse III, Rangueil Hospital, Toulouse, France. .,INSERM UMR 1037, Toulouse Centre for Cancer Research, University of Toulouse III, Toulouse, France.
| | - Barbara Bournet
- Department of Gastroenterology, University of Toulouse III, Rangueil Hospital, Toulouse, France.,INSERM UMR 1037, Toulouse Centre for Cancer Research, University of Toulouse III, Toulouse, France
| | - Pierre Cordelier
- INSERM UMR 1037, Toulouse Centre for Cancer Research, University of Toulouse III, Toulouse, France
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37
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Wu C, Maley AM, Walt DR. Single-molecule measurements in microwells for clinical applications. Crit Rev Clin Lab Sci 2019:1-21. [PMID: 31865834 DOI: 10.1080/10408363.2019.1700903] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
The ability to detect and analyze proteins, nucleic acids, and other biomolecules is critical for clinical diagnostics and for understanding the underlying mechanisms of disease. Current detection methods in clinical and research laboratories rely upon bulk measurement techniques such as immunoassays, polymerase chain reaction, and mass spectrometry to detect these biomarkers. However, many potentially useful protein or nucleic acid biomarkers in blood, saliva, or other biofluids exist at concentrations well below the detection limits of current methods, necessitating the development of more sensitive technologies. Single-molecule measurements are poised to address this challenge, vastly improving sensitivity for detecting low abundance biomarkers and rare events within a population. Microwell arrays have emerged as a powerful tool for single-molecule measurements, enabling ultrasensitive detection of disease-relevant biomolecules in easily accessible biofluids. This review discusses the development, fundamentals, and clinical applications of microwell-based single-molecule methods, as well as challenges and future directions for translating these methods to the clinic.
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Affiliation(s)
- Connie Wu
- Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.,Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA, USA
| | - Adam M Maley
- Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.,Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA, USA
| | - David R Walt
- Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.,Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA, USA
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38
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Zhou X, Ravichandran GC, Zhang P, Yang Y, Zeng Y. A microfluidic alternating-pull-push active digitization method for sample-loss-free digital PCR. LAB ON A CHIP 2019; 19:4104-4116. [PMID: 31720646 PMCID: PMC6894176 DOI: 10.1039/c9lc00932a] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/04/2023]
Abstract
Digital polymerase chain reaction (dPCR) is a powerful tool for genetic analysis, providing superior sensitivity and accuracy. In many applications that demand minuscule reaction volumes, such as single cell analysis, efficient and reproducible sample handling and digitization is pivotal for accurate absolute quantification of targets, but remains a significant technical challenge. In this paper, we described a robust and flexible microfluidic alternating-pull-push active digitization (μAPPAD) strategy that confers close to 100% sample digitization efficiency for microwell-based dPCR. Our strategy employs pneumatic valve control to periodically manipulate air pressure inside the chip to greatly facilitate the vacuum-driven partition of solution into microwells, enabling efficient digitization of a small-volume solution with significantly reduced volume variability. The μAPPAD method was evaluated on both tandem-channel and parallel-channel chips, which achieved a digitization efficiency of 99.5 ± 0.3% and 94.6 ± 0.9% within 10.5 min and 2 min, respectively. To assess the analytical performance of the μAPPAD chip, we calibrated it for absolution dPCR quantitation of λDNA across a range of concentrations. The results obtained with our chip matched well with the theoretical curve computed from Poisson statistics. Compared to the existing methods for highly efficient sample digitization, not only does our technology greatly reduce the constraints on microwell geometries and channel design, but also benefits from the intrinsic amenability of the pneumatic valve technique with device integration and automation. Thus we envision that the μAPPAD technology will provide a scalable and widely adaptable platform to promote the development of advanced lab-on-a-chip systems integrating microscale sample processing with dPCR for a broad scope of applications, such as single cell analysis of tumor heterogeneity and genetic profiling of circulating exosomes directly in clinical samples.
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Affiliation(s)
- Xin Zhou
- Department of Chemistry, University of Kansas, Lawrence, KS 66045, USA.
| | | | - Peng Zhang
- Department of Chemistry, University of Kansas, Lawrence, KS 66045, USA.
| | - Yang Yang
- Department of Chemistry, University of Kansas, Lawrence, KS 66045, USA.
| | - Yong Zeng
- Department of Chemistry, University of Kansas, Lawrence, KS 66045, USA. and University of Kansas Cancer Center, Kansas City, KS 66160, USA
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39
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Affiliation(s)
- Yun Ding
- Institute for Chemical and Bioengineering, Department of Chemistry and Applied Biosciences, ETH Zurich, 8093 Zürich, Switzerland
| | - Philip D. Howes
- Institute for Chemical and Bioengineering, Department of Chemistry and Applied Biosciences, ETH Zurich, 8093 Zürich, Switzerland
| | - Andrew J. deMello
- Institute for Chemical and Bioengineering, Department of Chemistry and Applied Biosciences, ETH Zurich, 8093 Zürich, Switzerland
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40
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Hank T, Strobel O. Conversion Surgery for Advanced Pancreatic Cancer. J Clin Med 2019; 8:jcm8111945. [PMID: 31718103 PMCID: PMC6912686 DOI: 10.3390/jcm8111945] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2019] [Revised: 11/03/2019] [Accepted: 11/06/2019] [Indexed: 12/24/2022] Open
Abstract
While primarily unresectable locally advanced pancreatic cancer (LAPC) used to be an indication for palliative therapy, a strategy of neoadjuvant therapy (NAT) and conversion surgery is being increasingly used after more effective chemotherapy regimens have become available for pancreatic ductal adenocarcinoma. While high-level evidence from prospective studies is still sparse, several large retrospective studies have recently reported their experience with NAT and conversion surgery for LAPC. This review aims to provide a current overview about different NAT regimens, conversion rates, survival outcomes and determinants of post-resection outcomes, as well as surgical strategies in the context of conversion surgery after NAT. FOLFIRINOX is the predominant regimen used and associated with the highest reported conversion rates. Conversion rates considerably vary between less than 5% and more than half of the study population with heterogeneous long-term outcomes, owing to a lack of intention-to-treat analyses in most studies and a high heterogeneity in resectability criteria, treatment strategies, and reporting among studies. Since radiological criteria of local resectability are no longer applicable after NAT, patients without progressive disease should undergo surgical exploration. Surgery after NAT has to be aimed at local radicality around the peripancreatic vessels and should be performed in expert centers. Future studies in this rapidly evolving field need to be prospective, analyze intention-to-treat populations, report stringent and objective inclusion criteria and criteria for resection. Innovative regimens for NAT in combination with a radical surgical approach hold high promise for patients with LAPC in the future.
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41
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Gall TMH, Belete S, Khanderia E, Frampton AE, Jiao LR. Circulating Tumor Cells and Cell-Free DNA in Pancreatic Ductal Adenocarcinoma. THE AMERICAN JOURNAL OF PATHOLOGY 2019; 189:71-81. [PMID: 30558725 DOI: 10.1016/j.ajpath.2018.03.020] [Citation(s) in RCA: 53] [Impact Index Per Article: 8.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/14/2017] [Revised: 03/06/2018] [Accepted: 03/26/2018] [Indexed: 12/21/2022]
Abstract
Pancreatic cancer is detected late in the disease process and has an extremely poor prognosis. A blood-based biomarker that can enable early detection of disease, monitor response to treatment, and potentially allow for personalized treatment would be of great benefit. This review analyzes the literature regarding two potential biomarkers, circulating tumor cells (CTCs) and cell-free DNA (cfDNA), with regard to pancreatic ductal adenocarcinoma. The origin of CTCs and the methods of detection are discussed and a decade of research examining CTCs in pancreatic cancer is summarized, including both levels of CTCs and analyzing their molecular characteristics and how they may affect survival in both advanced and early disease and allow for treatment monitoring. The origin of cfDNA is discussed, and the literature over the past 15 years is summarized. This includes analyzing cfDNA for genetic mutations and methylation abnormalities, which have the potential to be used for the detection and prognosis of pancreatic ductal adenocarcinoma. However, the research certainly remains in the experimental stage, warranting future large trials in these areas.
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Affiliation(s)
- Tamara M H Gall
- Hepato-Pancreato-Biliary Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, United Kingdom.
| | - Samuel Belete
- Hepato-Pancreato-Biliary Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, United Kingdom
| | - Esha Khanderia
- Hepato-Pancreato-Biliary Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, United Kingdom
| | - Adam E Frampton
- Hepato-Pancreato-Biliary Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, United Kingdom
| | - Long R Jiao
- Hepato-Pancreato-Biliary Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, United Kingdom
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42
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Gómez-Tomás Á, Pumarega J, Alguacil J, Amaral AF, Malats N, Pallarès N, Gasull M, Porta M, PANKRAS II Study Group. Concentrations of trace elements and KRAS mutations in pancreatic ductal adenocarcinoma. ENVIRONMENTAL AND MOLECULAR MUTAGENESIS 2019; 60:693-703. [PMID: 31066938 PMCID: PMC6786909 DOI: 10.1002/em.22296] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/07/2018] [Revised: 04/10/2019] [Accepted: 04/26/2019] [Indexed: 05/04/2023]
Abstract
Trace elements are a possible risk factor for pancreatic ductal adenocarcinoma (PDAC). However, their role in the occurrence and persistence of KRAS mutations remains unstudied. There appear to be no studies analyzing biomarkers of trace elements and KRAS mutations in any human cancer. We aimed to determine whether patients with KRAS mutated and nonmutated tumors exhibit differences in concentrations of trace elements. Incident cases of PDAC were prospectively identified in five hospitals in Spain. KRAS mutational status was determined through polymerase chain reaction from tumor tissue. Concentrations of 12 trace elements were determined in toenail samples by inductively coupled plasma mass spectrometry. Concentrations of trace elements were compared in 78 PDAC cases and 416 hospital-based controls (case-control analyses), and between 17 KRAS wild-type tumors and 61 KRAS mutated tumors (case-case analyses). Higher levels of iron, arsenic, and vanadium were associated with a statistically nonsignificant increased risk of a KRAS wild-type PDAC (OR for higher tertile of arsenic = 3.37, 95% CI 0.98-11.57). Lower levels of nickel and manganese were associated with a statistically significant higher risk of a KRAS mutated PDAC (OR for manganese = 0.34, 95% CI 0.14-0.80). Higher levels of selenium appeared protective for both mutated and KRAS wild-type PDAC. Higher levels of cadmium and lead were clear risk factors for both KRAS mutated and wild-type cases. This is the first study analyzing biomarkers of trace elements and KRAS mutations in any human cancer. Concentrations of trace elements differed markedly between PDAC cases with and without mutations in codon 12 of the KRAS oncogene, thus suggesting a role for trace elements in pancreatic and perhaps other cancers with such mutations. Environ. Mol. Mutagen., 60:693-703, 2019. © 2019 Wiley Periodicals, Inc.
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Affiliation(s)
- Álvaro Gómez-Tomás
- School of Medicine, Universitat Autònoma de Barcelona, Barcelona, Spain
- Faculty of Health and Life Sciences, Universitat Pompeu Fabra, Barcelona, Spain
| | - José Pumarega
- CIBER de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain
- Hospital del Mar Medical Research Institute (IMIM), Barcelona, Spain
| | - Juan Alguacil
- CIBER de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain
- Universidad de Huelva, Huelva, Spain
| | - André F.S. Amaral
- Population Health and Occupational Disease, National Heart and Lung Institute, Imperial College London, London, United Kingdom
| | - Núria Malats
- Genetic and Molecular Epidemiology Group, Spanish National Cancer Research Centre (CNIO), Madrid, Spain
- Centro de Investigación Biomédica en Red de Cáncer (CIBERONC), Madrid, Spain
| | - Natàlia Pallarès
- School of Medicine, Universitat Autònoma de Barcelona, Barcelona, Spain
- Hospital del Mar Medical Research Institute (IMIM), Barcelona, Spain
| | - Magda Gasull
- School of Medicine, Universitat Autònoma de Barcelona, Barcelona, Spain
- CIBER de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain
- Hospital del Mar Medical Research Institute (IMIM), Barcelona, Spain
| | - Miquel Porta
- School of Medicine, Universitat Autònoma de Barcelona, Barcelona, Spain
- CIBER de Epidemiología y Salud Pública (CIBERESP), Madrid, Spain
- Hospital del Mar Medical Research Institute (IMIM), Barcelona, Spain
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43
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Luchini C, Veronese N, Nottegar A, Cappelletti V, Daidone MG, Smith L, Parris C, Brosens LAA, Caruso MG, Cheng L, Wolfgang CL, Wood LD, Milella M, Salvia R, Scarpa A. Liquid Biopsy as Surrogate for Tissue for Molecular Profiling in Pancreatic Cancer: A Meta-Analysis Towards Precision Medicine. Cancers (Basel) 2019; 11:1152. [PMID: 31405192 PMCID: PMC6721631 DOI: 10.3390/cancers11081152] [Citation(s) in RCA: 37] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2019] [Revised: 08/05/2019] [Accepted: 08/07/2019] [Indexed: 12/24/2022] Open
Abstract
Liquid biopsy (LB) is a non-invasive approach representing a promising tool for new precision medicine strategies for cancer treatment. However, a comprehensive analysis of its reliability for pancreatic cancer (PC) is lacking. To this aim, we performed the first meta-analysis on this topic. We calculated the pooled sensitivity, specificity, positive (LR+) and negative (LR-) likelihood ratio, and diagnostic odds ratio (DOR). A summary receiver operating characteristic curve (SROC) and area under curve (AUC) were used to evaluate the overall accuracy. We finally assessed the concordance rate of all mutations detected by multi-genes panels. Fourteen eligible studies involving 369 patients were included. The overall pooled sensitivity and specificity were 0.70 and 0.86, respectively. The LR+ was 3.85, the LR- was 0.34 and DOR was 15.84. The SROC curve with an AUC of 0.88 indicated a relatively high accuracy of LB for molecular characterization of PC. The concordance rate of all mutations detected by multi-genes panels was 31.9%. LB can serve as surrogate for tissue in the molecular profiling of PC, because of its relatively high sensitivity, specificity and accuracy. It represents a unique opportunity to be further explored towards its introduction in clinical practice and for developing new precision medicine approaches against PC.
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Affiliation(s)
- Claudio Luchini
- Department of Diagnostics and Public Health, Section of Pathology, University of Verona, 37134 Verona, Italy
| | - Nicola Veronese
- National Institute of Gastroenterology-Research Hospital, IRCCS "S. de Bellis", Castellana Grotte, 70013 Bari, Italy
| | - Alessia Nottegar
- Department of Surgery, Section of Pathology, San Bortolo Hospital, 36100 Vicenza, Italy
| | - Vera Cappelletti
- Applied Research and Technological Development Department, Fondazione IRCCS Istituto Nazionale dei Tumori di Milano, 20133 Milano, Italy
| | - Maria G Daidone
- Applied Research and Technological Development Department, Fondazione IRCCS Istituto Nazionale dei Tumori di Milano, 20133 Milano, Italy
| | - Lee Smith
- Faculty of Science and Engineering, Anglia Ruskin University, Cambridge CB1 1PT, UK
| | - Christopher Parris
- Faculty of Science and Engineering, Anglia Ruskin University, Cambridge CB1 1PT, UK
| | - Lodewijk A A Brosens
- Department of Diagnostics and Public Health, Section of Pathology, University of Verona, 37134 Verona, Italy
- Department of Pathology, University Medical Center Utrecht, Utrecht University, 3584CX Utrecht, The Netherlands
- Department of Pathology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, 6526GA Nijmegen, The Netherlands
| | - Maria G Caruso
- National Institute of Gastroenterology-Research Hospital, IRCCS "S. de Bellis", Castellana Grotte, 70013 Bari, Italy
| | - Liang Cheng
- Department of Pathology and Laboratory Medicine, Indiana University School of Medicine, Indianapolis, IN 46202, USA
| | - Christopher L Wolfgang
- Department of Surgery, Sol Goldman Pancreatic Cancer Research Center, The Johns Hopkins University School of Medicine, Baltimore, MD 21211, USA
| | - Laura D Wood
- Department of Pathology, Sol Goldman Pancreatic Cancer Research Center, The Johns Hopkins University School of Medicine, Baltimore, MD 21211, USA
- Department of Oncology, Sol Goldman Pancreatic Cancer Research Center, The Johns Hopkins University School of Medicine, Baltimore, MD 21211, USA
| | - Michele Milella
- Department of Medicine, Section of Medical Oncology, University and Hospital Trust of Verona, 37134 Verona, Italy
| | - Roberto Salvia
- Department of General and Visceral Surgery, The Pancreas Institute, University and Hospital Trust of Verona, 37134 Verona, Italy
| | - Aldo Scarpa
- Department of Diagnostics and Public Health, Section of Pathology, University of Verona, 37134 Verona, Italy.
- ARC-Net Research Center, University of Verona, 37134 Verona, Italy.
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Enrichment technique to allow early detection and monitor emergence of KRAS mutation in response to treatment. Sci Rep 2019; 9:11346. [PMID: 31383871 PMCID: PMC6683117 DOI: 10.1038/s41598-019-47700-9] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2019] [Accepted: 07/18/2019] [Indexed: 02/08/2023] Open
Abstract
Sensitivity of cell-free circulating tumour DNA (ctDNA) assays is often hampered by the limited quantity of intact mutant nucleotide fragments. To overcome the issue of substrate limitation in clinical applications, we developed an enrichment method utilizing pyrrole-imidazole (PI) polyamides and their ability to bind the minor groove of B-DNA. We present here a proof-of-concept experiment to enrich specific mutant KRAS alleles with biotinylated PI polyamides. We investigated the clinical feasibility of incorporating PI polyamides to detect KRAS mutations in ctDNA from 40 colorectal cancer (CRC) patients, of whom 17 carried mutations in KRAS. After enriching ctDNA with those polyamides, we used digital PCR to detect several common KRAS codon 12 mutations. Enrichment by biotinylated PI polyamides improved the sensitivity of ctDNA analysis (88.9% vs. 11.1%, P < 0.01) in 9 non-metastatic mutation-positive patients. We observed no differences in performance for the 8 metastatic subjects (100% vs. 75%, P = 0.47). In the remaining 23/40 patients with wild type KRAS codon 12, no mutant alleles were detected with or without polyamide-facilitated enrichment. Enriching B-form of ctDNA with PI polyamides significantly improved the assay sensitivity in detecting KRAS mutations in non-metastatic CRC patient samples.
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Lee J, Park SS, Lee YK, Norton JA, Jeffrey SS. Liquid biopsy in pancreatic ductal adenocarcinoma: current status of circulating tumor cells and circulating tumor DNA. Mol Oncol 2019; 13:1623-1650. [PMID: 31243883 PMCID: PMC6670020 DOI: 10.1002/1878-0261.12537] [Citation(s) in RCA: 62] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2018] [Revised: 06/07/2019] [Accepted: 06/25/2019] [Indexed: 12/22/2022] Open
Abstract
Reliable biomarkers are required to evaluate and manage pancreatic ductal adenocarcinoma. Circulating tumor cells and circulating tumor DNA are shed into blood and can be relatively easily obtained from minimally invasive liquid biopsies for serial assays and characterization, thereby providing a unique potential for early diagnosis, forecasting disease prognosis, and monitoring of therapeutic response. In this review, we provide an overview of current technologies used to detect circulating tumor cells and circulating tumor DNA and describe recent advances regarding the multiple clinical applications of liquid biopsy in pancreatic ductal adenocarcinoma.
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Affiliation(s)
- Jee‐Soo Lee
- Department of Laboratory MedicineHallym University Sacred Heart HospitalAnyangKorea
- Department of Laboratory MedicineSeoul National University College of MedicineSeoulKorea
| | - Sung Sup Park
- Department of Laboratory MedicineSeoul National University College of MedicineSeoulKorea
| | - Young Kyung Lee
- Department of Laboratory MedicineHallym University Sacred Heart HospitalAnyangKorea
- Department of Laboratory MedicineHallym University College of MedicineAnyangKorea
| | - Jeffrey A. Norton
- Department of SurgeryStanford University School of MedicineStanfordCAUSA
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Circulating Hybrid Cells Join the Fray of Circulating Cellular Biomarkers. Cell Mol Gastroenterol Hepatol 2019; 8:595-607. [PMID: 31319228 PMCID: PMC6889578 DOI: 10.1016/j.jcmgh.2019.07.002] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/06/2019] [Revised: 07/03/2019] [Accepted: 07/05/2019] [Indexed: 12/11/2022]
Abstract
Gastrointestinal cancers account for more cancer-related deaths than any other organ system, owing in part to difficulties in early detection, treatment response assessment, and post-treatment surveillance. Circulating biomarkers hold the promise for noninvasive liquid biopsy platforms to overcome these obstacles. Although tumors shed detectable levels of degraded genetic material and cellular debris into peripheral blood, identifying reproducible and clinically relevant information from these analytes (eg, cell-free nucleotides, exosomes, proteins) has proven difficult. Cell-based circulating biomarkers also present challenges, but have multiple advantages including allowing for a more comprehensive tumor analysis, and communicating the risk of metastatic spread. Circulating tumor cells have dominated the cancer cell biomarker field with robust evidence in extraintestinal cancers; however, establishing their clinical utility beyond that of prognostication in colorectal and pancreatic cancers has remained elusive. Recently identified novel populations of tumor-derived cells bring renewed potential to this area of investigation. Cancer-associated macrophage-like cells, immune cells with phagocytosed tumor material, also show utility in prognostication and assessing treatment responsiveness. In addition, circulating hybrid cells are the result of tumor-macrophage fusion, with mounting evidence for a role in the metastatic cascade. Because of their relative abundance in circulation, circulating hybrid cells have great potential as a liquid biomarker for early detection, prognostication, and surveillance. In all, the power of the cell reaches beyond enumeration by providing a cellular source of tumor DNA, RNA, and protein, which can be harnessed to impact overall survival.
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Cervena K, Vodicka P, Vymetalkova V. Diagnostic and prognostic impact of cell-free DNA in human cancers: Systematic review. MUTATION RESEARCH-REVIEWS IN MUTATION RESEARCH 2019; 781:100-129. [PMID: 31416571 DOI: 10.1016/j.mrrev.2019.05.002] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/15/2019] [Revised: 05/03/2019] [Accepted: 05/07/2019] [Indexed: 02/06/2023]
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Pratt ED, Cowan RW, Manning SL, Qiao E, Cameron H, Schradle K, Simeone DM, Zhen DB. Multiplex Enrichment and Detection of Rare KRAS Mutations in Liquid Biopsy Samples using Digital Droplet Pre-Amplification. Anal Chem 2019; 91:7516-7523. [DOI: 10.1021/acs.analchem.8b01605] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/06/2023]
Affiliation(s)
- Erica D. Pratt
- Ahmed Center for Pancreatic Cancer Research, Department of Gastroenterology, Hepatology and Nutrition, University of Texas MD Anderson Cancer Center, Houston, Texas, United States
| | - Robert W. Cowan
- Ahmed Center for Pancreatic Cancer Research, Department of Gastroenterology, Hepatology and Nutrition, University of Texas MD Anderson Cancer Center, Houston, Texas, United States
| | - Sara L. Manning
- Ahmed Center for Pancreatic Cancer Research, Department of Gastroenterology, Hepatology and Nutrition, University of Texas MD Anderson Cancer Center, Houston, Texas, United States
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Liu X, Liu L, Ji Y, Li C, Wei T, Yang X, Zhang Y, Cai X, Gao Y, Xu W, Rao S, Jin D, Lou W, Qiu Z, Wang X. Enrichment of short mutant cell-free DNA fragments enhanced detection of pancreatic cancer. EBioMedicine 2019; 41:345-356. [PMID: 30857943 PMCID: PMC6442234 DOI: 10.1016/j.ebiom.2019.02.010] [Citation(s) in RCA: 60] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2018] [Revised: 01/30/2019] [Accepted: 02/06/2019] [Indexed: 02/08/2023] Open
Abstract
Background Analysis of cell-free DNA (cfDNA) is promising for broad applications in clinical settings, but with significant bias towards late-stage cancers. Although recent studies have discussed the diverse and degraded nature of cfDNA molecules, little is known about its impact on the practice of cfDNA analysis. Methods We developed single-strand library preparation and hybrid-capture-based cfDNA sequencing (SLHC-seq) to analysis degraded cfDNA fragments. Next we used SLHC-seq to perform cfDNA profiling in 112 pancreatic cancer patients, and the results were compared with 13 previous reports. Extensive analysis was performed in terms of cfDNA fragments to explore the reasons for higher detection rate of KRAS mutations in the circulation of pancreatic cancers. Findings By applying the new approach, we achieved higher efficiency in analysis of mutations than previously reported using other detection assays. 791 cancer-specific mutations were detected in plasma of 88% patients with KRAS hotspots detected in 70% of all patients. Only 8 mutations were detected in 28 healthy controls without any known oncogenic or truncating alleles. cfDNA profiling by SLHC-seq was largely consistent with results of tissue-based sequencing. SLHC-seq rescued short or damaged cfDNA fragments along to increase the sensitivity and accuracy of circulating-tumour DNA detection. Interpretation We found that the small mutant fragments are prevalent in early-stage patients, which provides strong evidence for fragment size-based detection of pancreatic cancer. The new pipeline enhanced our understanding of cfDNA biology and provide new insights for liquid biopsy.
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Affiliation(s)
- Xiaoyu Liu
- Department of Interventional Radiology, Zhongshan Hospital Fudan University, Shanghai, China; Shanghai Institute of Medical Imaging, Shanghai, China; Institute of Neuroscience, State Kay Laboratory of Neuroscience, CAS Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Shanghai, China
| | - Lingxiao Liu
- Department of Interventional Radiology, Zhongshan Hospital Fudan University, Shanghai, China; Shanghai Institute of Medical Imaging, Shanghai, China
| | - Yuan Ji
- Department of Pathology, Zhongshan Hospital Fudan University, Shanghai, China
| | - Changyu Li
- Department of Interventional Radiology, Zhongshan Hospital Fudan University, Shanghai, China; Shanghai Institute of Medical Imaging, Shanghai, China
| | - Tao Wei
- Department of Hepatobiliary and Pancreatic Surgery, School of Medicine, The Second Affiliated Hospital of Zhejiang University, Hangzhou, China
| | - Xuerong Yang
- Department of Radiology, Shuguang Hospital Shanghai University of Traditional Chinese Medicine, Shanghai, China
| | - Yuefang Zhang
- Institute of Neuroscience, State Kay Laboratory of Neuroscience, CAS Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Shanghai, China
| | - Xuyu Cai
- Precision Medicine Center, West China Hospital, Sichuan University, Chengdu, Sichuan, China
| | | | - Weihong Xu
- Stanford Genome Technology Center, Palo Alto, CA, USA
| | - Shengxiang Rao
- Department of Radiology, Zhongshan Hospital Fudan University, Shanghai, China
| | - Dayong Jin
- Department of Pancreatic Surgery, Zhongshan Hospital Fudan University, Shanghai, China; Department of General Surgery, Zhongshan Hospital Fudan University, Shanghai, China
| | - Wenhui Lou
- Department of Pancreatic Surgery, Zhongshan Hospital Fudan University, Shanghai, China; Department of General Surgery, Zhongshan Hospital Fudan University, Shanghai, China.
| | - Zilong Qiu
- Institute of Neuroscience, State Kay Laboratory of Neuroscience, CAS Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Shanghai, China.
| | - Xiaolin Wang
- Department of Interventional Radiology, Zhongshan Hospital Fudan University, Shanghai, China; Shanghai Institute of Medical Imaging, Shanghai, China.
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The cornerstone of integrating circulating tumor DNA into cancer management. Biochim Biophys Acta Rev Cancer 2018; 1871:1-11. [PMID: 30419316 DOI: 10.1016/j.bbcan.2018.11.002] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2018] [Revised: 08/23/2018] [Accepted: 11/07/2018] [Indexed: 12/26/2022]
Abstract
Recent circulating tumor DNA (ctDNA) research has demonstrated its potential as a non-invasive biomarker for cancer. However, the deployment of ctDNA assays in routine clinical practice remains challenging owing to variability in analytical approaches and the assessment of clinical significance. A well-developed, analytically valid ctDNA assay is a prerequisite for integrating ctDNA into cancer management, and an appropriate analytical technology is crucial for the development of a ctDNA assay. Other determinants including pre-analytical procedures, test validation, internal quality control (IQC), and continual proficiency testing (PT) are also important for the accuracy of ctDNA assays. In the present review, we will focus on the most widely used ctDNA detection technologies and the key quality management measures used to assure the accuracy of ctDNA assays. The aim of this review is to provide useful information for technology selection during ctDNA assay development and assure a reliable test result in clinical practice.
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