Epithelial-mesenchymal transition (EMT) has been extensively studied for its roles in tumor metastasis, the generation and maintenance of cancer stem cells and treatment resistance. Epithelial mesenchymal plasticity allows cells to switch between various states within the epithelial-mesenchymal spectrum, resulting in a mixed epithelial/mesenchymal phenotypic profile. This plasticity underlies the acquisition of multiple malignant features during cancer progression and poses challenges for EMT in tumors. MicroRNAs (miRNAs) in the microenvironment affect numerous signaling processes through diverse mechanisms, influencing physiological activities. This paper reviews recent advances in EMT, the role of different hybrid states in tumor progression, and the important role of miRNAs in EMT. Furthermore, it explores the relationship between miRNA-based EMT therapies and their implications for clinical practice, discussing how ongoing developments may enhance therapeutic outcomes.

The tumor budding and tumor cell nest size-based (TBNS) grading scheme is an emerging prognostic indicator for squamous cell carcinoma (SCC) across various organs; however, its significance in vulvar SCC (VSCC) remains poorly investigated. In this study, we applied the TBNS grading system to an institutional cohort of 62 consecutive surgically resected VSCC cases (39 HPV-independent and 23 HPV-associated), excluding patients with neoadjuvant chemotherapy or FIGO stage IA disease. High tumor budding activity, small cell nest size, and high TBNS grade were significantly associated with reduced overall survival (OS) and disease-free survival (DFS) in VSCC, as well as with adverse clinicopathologic features such as lymphovascular space invasion, perineural involvement, lymph node metastasis, and advanced FIGO stage (p < 0.05). Multivariate analysis revealed that TBNS grade 3 was independently associated with reduced DFS (p < 0.05). In contrast, among the two conventional grading systems, only the Gynecology Oncology Group grading system showed a significant association with OS in univariate analysis (p < 0.05), but not in multivariate analysis (p > 0.05). We conclude that the TBNS grading system is a promising prognostic indicator for VSCC, outperforming conventional grading systems. Further validation in larger cohorts is needed to expand the clinical applicability of this grading system in VSCC.

Microsatellite-stable colon cancer represents a heterogeneous group of tumors, where the identification of high-risk histopathological factors is particularly critical. In this study, we evaluate two morphological features associated with mesenchymal differentiation-tumor budding and tumor-associated stroma-with the aim of developing a robust recurrence risk stratification score and exploring its relationship with the tumor microenvironment composition in this clinical context. We retrospectively analyzed 254 formalin-fixed, paraffin-embedded colectomy specimens from patients with mismatch repair-proficient colon cancer (stages I to III). Tumor budding and tumor-associated stroma were assessed using two protocols: one focused on a single hotspot field and another on a broader tumor area. The tumor microenvironment composition, including the presence of tertiary lymphoid structures, was characterized using immunohistochemistry. We developed a three-tiered tumor budding-stroma (TBS) stratification score based on the evaluation of an extended tumor area. This score was independently associated with the disease-free survival probability (low-TBS: HR 0.12, 95 % CI 0.04-0.33, p < 0.001; intermediate-TBS: HR 0.26, 95 % CI 0.10-0.65, p = 0.003) and allowed the identification of a high-risk group characterized by an immune-depleted tumor microenvironment. The prognostic value of this approach outperformed that of each individual parameter and was superior to the stratification score obtained using the hotspot field-based assessment. In conclusion, the combined assessment of tumor budding and tumor-associated stroma over an extended tumor area provides a more comprehensive view of tumor heterogeneity. This approach may be suitable for routine evaluation of microsatellite-stable localized colon cancer patients.

Tumor budding (TB) and poorly differentiated clusters (PDCs) are well-established prognostic factors in various cancers. This study aimed to assess the independent prognostic role of these markers in endometrial carcinomas. Retrospective analysis of endometrial carcinoma resection specimens by examining traditional histologic prognostic parameters. TB and PDC were observed at 20× magnification in ten fields at the invasive front and categorized as present or absent. In addition, a count of ≥5 was stratified as "high." Clinical and follow-up details were extracted from Gynecologic Oncology records. Sixty-five endometrial carcinomas were studied and were predominantly endometrioid (n=47, 72.3%). TB was identified in 52.3% of cases, with high TB observed in 38.5%. PDC was evident in 44.6%, with high PDC seen in 29.2%. Associations were significant between the presence of TB/high TB and higher tumor grade ( P < 0.001), deep myometrial invasion ( P = 0.006/ P = 0.002), diffuse pattern of invasion ( P = 0.007/ P = 0.03), microcystic elongated and fragmented pattern ( P < 0.001), lymphovascular space invasion, lymph node metastasis ( P =<0.001) and International Federation of Gynecology and Obstetrics stage ( P = 0.000/ P = 0.002). PDC/high PDC showed similar associations, and, in addition, with nonendometrioid histologic type ( P = 0.02) and tumor location in a lower uterine segment (high PDC, P = 0.009). After adjusting for other significant parameters, both high TB ( P = 0.03) and high PDC ( P = 0.031) emerged as independent prognostic parameters for lymphovascular space invasion or Lymph node metastasis. No recorded deaths or significant events occurred, precluding commentary on overall survival status. High TB and PDC are independent predictors of Lymph node metastasis in endometrial carcinomas. Their association with the microcystic elongated and fragmented pattern makes them histologic predictors of epithelial-mesenchymal transition. Their simple application underscores their potential as valuable additional prognostic indicators for endometrial carcinomas.

Tumor budding (TB) is an independent predictor of adverse prognosis in colorectal cancer (CRC), defined as clusters of fewer than 5 tumor cells at the invasive margin of cancer. According to the international consensus criteria (ITBCC), TB should be evaluated from the non-mucinous regions. However, some tumors also contain tumor bud-like structures within extracellular mucin pools, and the prognostic impact of these structures remains unclear. To assess this, we defined a modified tumor budding variable (TB-Muc), representing the highest number of tumor buds/bud-like structures observed in a hotspot (0.785 mm2) at the invasive margin, including extracellular mucin regions. We analyzed the prognostic significance of TB (ITBCC criteria) and TB-Muc in two CRC cohorts (N = 1876). TB-ITBCC was associated with advanced stage and lymphovascular invasion (p < 0.001) but also with shorter cancer-specific survival independent of other prognostic factors (Cohort 1: HR for high vs. low 1.99, 95 % CI 1.32-3.01, ptrend = 0.0007; Cohort 2: HR 1.35, 95 % CI 0.98-1.85, ptrend = 0.037). TB-Muc had a comparable independent association with shorter cancer-specific survival (Cohort 1: HR for high vs. low 1.77, 95 % CI 1.18-2.65, ptrend = 0.006; Cohort 2: HR 1.39, 95 % CI 1.02-1.89, ptrend = 0.019). Our results indicate that tumor bud-like structures in mucin do not provide additional prognostic value and should not be included in TB evaluation.

Intratumoral heterogeneity, including epithelial-mesenchymal transition (EMT), is one major cause of therapeutic resistance. The induction of ferroptosis, an iron-dependent death, has the potential in overcoming this resistance to traditional treatment modalities. However, the roles of distinct EMT phenotypes in ferroptosis remain an enigma. This study reports that 3D soft fibrin microenvironment confers colorectal cancer (CRC) cells hybrid EMT phenotype and high level of resistance to ferroptosis. The activation of histone acetylation and WNT/β-catenin signaling drives this EMT phenotypic transition, which promotes the defense of 3D CRCs against ferroptosis via glutathione peroxidases/ferritin signaling axis. Unexpectedly, E-cadherin knockout in 3D but not 2D CRCs mediates an integrin β3 marked-late hybrid EMT state and further enhances the resistance to ferroptosis via integrin-mediated tension and mitochondrial reprogramming. The inhibition of integrin αvβ3-mediated tension and WNT/β-catenin-mediated hybrid EMT sensitizes 3D CRCs with and without E-cadherin deficiency to ferroptosis in vivo, respectively. Further, the EMT phenotype of patient-derived tumoroids is associated with CRC therapeutic resistance. In summary, this study uncovers previously unappreciated roles of hybrid EMT and cell membrane tension in ferroptosis, which not only predict the treatment efficacy but also potentiate the development of new ferroptosis-based targeted therapeutic strategies.

Invasion and metastasis are well-known hallmarks of cancer, with metastatic disease accounting for 60% to 90% of cancer-related deaths [...].

Stage II and III colorectal cancer (CRC) poses a significant challenge due to rising global incidence and mortality rates. Despite advancements in screening and treatment, there's a pressing need for reliable prognostic biomarkers. Tumor budding emerges as a promising marker associated with poor prognosis and higher recurrence. However, its incorporation into clinical guidelines differs when considering adjuvant treatment. This study assesses tumor budding's prognostic value for recurrence in stage II and III CRC, exploring its implications for risk stratification.

This retrospective study encompassed patients with stage II-III CRC at Hospital Universitario La Paz from October 2016 to January 2022. Tumor budding was assessed according to the 2016 ITBCC guidelines and categorized as low, intermediate, or high. The primary outcomes, time to recurrence (TTR) and overall survival (OS), were analyzed using Kaplan-Meier and Cox regression models.

A total of 390 patients were included in the final analysis. They were predominantly male (55%) with an average age of 75 years (range 35-95). Fifty percent of patients were stage II. Tumor budding was categorized as low, intermediate, and high in 186 (48%), 110 (28%), and 94 (24%) patients, respectively. After a median follow-up of 46.1 months, there were 71 recurrences and 96 deaths. Time to recurrence (TTR) was significantly shorter for patients with high tumor budding. At 24 months, freedom from recurrence was 92%, 84%, and 69% for low, intermediate, and high tumor budding groups, respectively. Median TTR was not reached in any group. Multivariate analysis revealed high-grade budding as an independent predictor of recurrence with a hazard ratio (HR) of 2.39 (P = .01; 95% CI, [1.42-4.04]).

Our study highlights the prognostic value of tumor budding in predicting recurrence in both stage II and III colorectal cancer patients, reinforcing its potential as an important biomarker beyond stage II CRC.

Breast cancer is a leading cause of morbidity and mortality among women. Advances in molecular biology have improved detection and treatment, but conventional histopathological factors remain crucial for prognosis. Tumour budding, defined as clusters of less than 5 tumour cells detached from the main tumour, has been linked to poor prognosis in several cancers. This study explores the association between intra-tumoral budding (ITB) and peripheral tumour budding (PTB) with known prognostic factors in Invasive Breast Carcinoma of no special type (IBC NST).

This retrospective study analysed 70 cases of IBC NST diagnosed at Kasturba Medical College, Manipal, between January 2020 and December 2021. Tumour budding was classified as high-grade or low-grade based on density, which denotes the number of buds per x20 field. Clinicopathological data, including hormone receptor status, Ki-67 index, lymphovascular invasion (LVI), perineural invasion (PNI), and axillary lymph node involvement, were obtained. Statistical analyses were performed to identify a significant association between tumour budding and these factors. Univariate and multivariate logistic regression analyses were also done to demonstrate the significance of association.

High-grade PTB showed significant associations with LVI (p = 0.046), PNI (p = 0.017), and axillary lymph node involvement (p = 0.021). In contrast, high-grade ITB was only significantly correlated with axillary lymph node involvement (p = 0.044). LVI (p-value = 0.240) and axillary lymph node involvement (p-value = 0.142) did not show any association with PTB on multivariate analysis and PNI (p-value = 0.074) near significant association with PTB). A significant inverse association was observed between PTB and Ki-67 (p = 0.012), which remained significant in univariate and multivariate analysis (p-value = 0.017). No significant associations were found between tumour budding and hormone receptor status or menopausal status.

Peripheral tumour budding (PTB) is significantly associated with several poor prognostic factors in IBC NST, while intra-tumoral budding (ITB) correlates primarily with axillary lymph node involvement. Tumor budding, particularly PTB, could serve as an important prognostic marker in breast cancer. Further research is needed to standardize tumour budding assessment in clinical practice.

Malic enzyme 1 (ME1) plays a key role in promoting malignant phenotypes in various types of cancer. ME1 promotes epithelial-mesenchymal transition (EMT) and enhances stemness via glutaminolysis, energy metabolism reprogramming from oxidative phosphorylation to glycolysis. As a result, ME1 promotes the malignant phenotypes of cancer cells and poor patient prognosis. In particular, ME1 expression is promoted in hypoxic environments associated with hypoxia-inducible factor (HIF1) α. ME1 is overexpressed in budding cells at the cancer invasive front, promoting cancer invasion and metastasis. ME1 also generates nicotinamide adenine dinucleotide (NADPH), which, together with glucose-6-phosphate dehydrogenase (G6PD) and isocitrate dehydrogenase (IDH1), expands the NADPH pool, maintaining the redox balance in cancer cells, suppressing cell death by neutralizing mitochondrial reactive oxygen species (ROS), and promoting stemness. This review summarizes the latest research insights into the mechanisms by which ME1 contributes to cancer progression. Because ME1 is involved in various aspects of cancer and promotes many of its malignant phenotypes, it is expected that ME1 will become a novel drug target in the near future.

Intratumor heterogeneity is considered a major cause of treatment failure in pancreatic ductal adenocarcinoma (PDAC). In recent years, marked heterogeneity at the genomic and transcriptional level has been revealed, but the spatial distribution of the heterogeneous cell populations has not been considered. Yet, it is assumed that cancer cells at the invasive front are endowed with enhanced migratory and invasive properties, although evidence is scanty, and cancer-associated fibroblasts (CAFs) in this location have not been characterized. In this study, digital spatial profiling was used to compare the transcriptional profiles of cancer cells and CAFs in the tumor center versus the invasive front of human PDAC. Four well-differentiated PDACs with conventional morphology were investigated with the GeoMx system (Nanostring). Regions of interest were analyzed in the tumor center and at the invasive front using a whole transcriptome assay in the cancer cell and CAF segments separately. Three of the PDACs harbored mutated KRAS, whereas the fourth case was confirmed wild-type KRAS. Substantial inter-regional heterogeneity was identified, with increased activity of pathways associated with cellular stress (including TNFα-signaling via NFκB, hypoxia, P53 pathway), proliferation (MYC targets, mitotic spindle), glycolysis, and epithelial-mesenchymal transition (EMT) at the invasive front in both the cancer cell and CAF segments compared with the center of the tumor. Immunohistochemical validation on 17 PDACs of well, moderate, and poor differentiation confirmed significant inter-regional heterogeneity in the expression level of markers of EMT and glycolysis. The results of this study show that in PDAC, transcriptional profiles of both cancer cells and CAFs differ between the center of the tumor and the invasive front.

Ovarian clear cell carcinoma (OCCC) is an endometriosis-related neoplasm, in which traditional histologic grading does not show prognostic significance. Tumor budding was associated with poorer outcomes in OCCC in previous studies. We aimed to evaluate the prognostic significance of tumor budding in OCCC in an independent cohort. Seventy patients diagnosed with OCCC were retrospectively identified. Slides from primary ovarian resections were reviewed by 2 pathologists blinded to outcomes. Tumor budding was defined as single or clusters of <5 tumor cells in peritumoral and/or intratumoral nonhyalinized stroma. Most patients were diagnosed at an early stage (stage I: 69%; II: 20%; III: 10%; IV: 1%). Twenty-one patients experienced recurrences (30%) and 2 progressive disease (3%). At the last follow-up, 52 patients had no evidence of disease, 6 were alive with disease, and 12 died of disease. The median follow-up time was 66.7 mo. Tumor budding was identified in 41 cases (59%) with a kappa coefficient of 0.60. On univariate analysis, tumor budding (P=0.022) and stage (P=0.0005) were associated with shorter progression-free survival (PFS), but only stage was independently associated with shorter PFS on multivariate analysis (P=0.003). Higher stage was the only variable associated with shorter overall survival (P=0.037). Tumor budding was associated with higher stage (P=0.039), absence of endometriosis (P=0.042) and adenofibroma (P=0.046), tumor-associated inflammation (P=0.002), and higher mitotic activity (P=0.022). There was no association between tumor budding and molecular characteristics in 32 cases with somatic tumor sequencing. Tumor budding was not independently associated with worse outcomes in this cohort of OCCC, although it was significantly associated with specific clinicopathologic features, including higher stage. Stage was the only independent variable predictive of poorer survival, which appears to drive the prognostic significance of tumor budding.

Breast cancer is a highly heterogenous tumor with different subtypes showing varying prognosis. Tumor budding is an unfavorable histological feature of many epithelial cancers. The purpose of this study is to analyze the association between tumor bud density with various histological and immunohistochemical characteristics and to explore its prognostic role in breast carcinoma.

A retrospective analysis was performed on 100 patients of breast cancer diagnosed in our institute from January to December 2017. Hematoxylin and eosin (H&E) stained slides from tumors and immunohistochemical slides were reviewed independently by two pathologists, and clinical data were acquired from computerized records. Patients on neoadjuvant chemotherapy were excluded from the study.

The study comprised 100 patients of invasive breast carcinoma. The median age was 52 years, and 96% were invasive ductal carcinoma. The median follow-up was 34 months. High tumor bud density was substantially correlated with primary tumor staging (T3, T4; 73% [11/15] cases) and lymph node staging (N2, N3; 68% [13/19] cases) with p -values of 0.017 and 0.023, respectively. Systemic metastasis (85% [6/7] cases) was significantly associated with high tumor bud density ( p =0.025) but lymphovascular invasion (LVI) and perineural invasion (PNI) were not significantly associated with tumor bud density ( p  = 0.762 and 0.862, respectively). Patients with N2 nodal stage had low event-free survival rate than N0/N1 nodal stage irrespective of tumor bud status. Grade 3 tumors with high tumor bud density had worse event-free survival than any other grades. There was no association of tumor bud density with tumor staging, necrosis, PNI, LVI, estrogen receptor (ER), progesterone receptor (PR) and Her2/neu , and event-free survival.

Strong relationships have been found between tumor bud density and poor prognostic variables such as primary tumor staging and lymph node staging. These results provide credence to the idea that tumor bud density can be an assessable prognostic feature that should be taken into account while reporting breast cancer cases. Tumor bud density evaluation has to be standardized nevertheless if it is to be widely adopted.

Gastric cancer remains the fifth most common malignant neoplasm in the world and ranks fifth among the causes associated with cancer. The TNM system remains the gold standard for predictive stratification of patients with gastric cancer, but the search for new sensitive, specific and reproducible biomarkers to develop a personalized approach to the management of patients with gastric cancer does not lose its relevance. The phenomenon of tumor budding is a well-established independent prognostic factor in colorectal cancer. In 2017, the first guideline on the method of calculating tumor budding for colorectal cancer was published. Despite the promising potential of using tumor budding in gastric cancer this parameter is still not evaluated in everyday practice. This lection provides data on various methods of counting tumor budding in gastric carcinomas, describes the molecular mechanisms of interaction between tumor cells and the immune microenvironment, and summarizes the available data on the relationship of clinical and morphological characteristics of gastric cancer with the degree of tumor budding. The relationship between the degree of tumor budding and the prognostic characteristics of gastric cancer and the prospects for its use is also described.

Breast cancer (BC) is the most prevalent cancer in women globally. Anti-cancer advancements have enabled the killing of BC cells through various therapies; however, cancer relapse is still a major limitation and decreases patient survival and quality of life. Epithelial-to-mesenchymal transition (EMT) is responsible for tumor relapse in several cancers. This highly regulated event causes phenotypic, genetic, and epigenetic changes in the tumor microenvironment (TME). This review summarizes the recent advancements regarding EMT using de-differentiation and partial EMT theories. We extensively review the mechanistic pathways, TME components, and various anti-cancer adjuvant and neo-adjuvant therapies responsible for triggering EMT in BC tumors. Information regarding essential clinical studies and trials is also discussed. Furthermore, we also highlight the recent strategies targeting various EMT pathways. This review provides a holistic picture of BC biology, molecular pathways, and recent advances in therapeutic strategies.

Cellular plasticity refers to the ability of cells to change their identity or behavior, which can be advantageous in some cases (e.g., tissue regeneration) but detrimental in others (e.g., cancer metastasis). With a better understanding of cellular plasticity, the complexity of cancer cells, their heterogeneity, and their role in metastasis is being unraveled. The plasticity of the cells could also prove as a nemesis to their characterization. In this review, we have attempted to highlight the possibilities and benefits of using multiomics approach in characterizing the plastic nature of cancer cells. There is a need to integrate fragmented evidence at different levels of cellular organization (DNA, RNA, protein, metabolite, epigenetics, etc.) to facilitate the characterization of different forms of plasticity and cell types. We have discussed the role of cellular plasticity in generating intra-tumor heterogeneity. Different omics level evidence is being provided to highlight the variety of molecular determinants discovered using different techniques. Attempts have been made to integrate some of this information to provide a quantitative assessment and scoring of the plastic nature of the cells. However, there is a huge gap in our understanding of mechanisms that lead to the observed heterogeneity. Understanding of these mechanism(s) is necessary for finding targets for early detection and effective therapeutic interventions in metastasis. Targeting cellular plasticity is akin to neutralizing a moving target. Along with the advancements in precision and personalized medicine, these efforts may translate into better clinical outcomes for cancer patients, especially in metastatic stages.

Inflammatory breast cancer (IBC) is characterized by numerous tumor emboli within lymphatics. In a recent study, we observed tumor embolic budding both in vitro and in vivo within lymphovascular spaces and proposed this to account for the plethora of tumor emboli seen in IBC. These observations did not address, however, how lymphovascular invasion is initiated or the mechanisms involved. In the present study, using the well-characterized patient-derived xenograft (PDX), Mary-X, which exhibited florid lymphovascular invasion (LVI) in athymic mice (LVI) as defined by E-cadherin-positive tumor emboli within lymphatic channels distinguished by podoplanin and LYVE1 membrane and Prox1 nuclear immunoreactivities and spontaneous spheroidgenesis in vitro and human cases of IBC which showed similar LVI, we compared laser-captured microdissected emboli from Mary-X and from the cases of human IBC to non-embolic areas. Mary-X and IBC emboli expressed high levels of E-cadherin and no evidence of epithelial-mesenchymal transition (EMT). Mary-X spheroids expressed high levels of VEGF, especially VEGF-C, and stimulated both vascular and lymphatic endothelial haptotaxis. We then transplanted Mary-X serially into green, cyano, red, and nestin-green fluorescing protein (GFP-, CFP-, RFP-, and nestin-GFP) transgenic reporter mice in various combinations. Multicolor murine imaging studies indicated that reporter-labeled stroma initially encircled clumps of tumor cells and then served as a scaffold that supported nestin-GFP-labeled endothelial haptotaxis resulting in encircling lymphangiogenesis, confirmed by dual LYVE1 immunofluorescence. The present studies demonstrate a possible mechanism of a critical step of the tumor emboli formation of IBC.

It is of great importance to study the detachment/attachment behaviors of cells (cancer cell, immune cell, and epithelial cell), as they are closely related with tumor metastasis, immunoreaction, and tissue development at variety scales. To characterize the detachment/attachment during the interaction between cells and substrate, some researchers proposed using cell traction force (CTF) as the indicator. To date, various strategies have been developed to measure the CTF. However, these methods only realize the measurements of cell passive forces on flat cases. To quantify the active CTF on nonflat surfaces, which can better mimic the in vivo case, we employed elastic hydrogel microspheres as a force sensor. The microspheres were fabricated by microfluidic chips with controllable size and mechanical properties to mimic substrate. Cells were cultured on microsphere and the CTF led to the deformation of microsphere. By detecting the morphology information, the CTF exerted by attached cells can be calculated by the in-house numerical code. Using these microspheres, the CTF of various cells (including tumor cell, immunological cell, and epithelium cell) were successfully obtained on nonflat surfaces with different curvature radii. The proposed method provides a versatile platform to measure the CTF with high precision and to understand the detachment/attachment behaviors during physiology processes.

Laryngeal squamous cell carcinoma (LSCC) is a commonly occurring malignancy in the head and neck region. However, due to the heterogeneity of primary tumor sites, tumor behavior, and molecular mechanisms, there is currently no consensus on the accuracy of clinicopathological prognostic factors for individual cases. Tumor histopathologic behavior remains a crucial factor in predicting aggressiveness. Recent studies have shown that peritumoral tumor budding (TB) combined with cell nest size (CNS) is a reliable marker for predicting lymph node metastasis, advanced cancer prognosis, and therapeutic response in SCCs of different origins.

This study aims to investigate the relationship between TB and CNS in the context of nodal metastasis and overall prognosis in patients diagnosed with LSCC. Our objective is to establish the significance of TB and CNS status as a cost-effective, easily assessed, and highly reliable prognostic factor among this patient population.

In this retrospective cross-sectional study, we analyzed 128 LSCC cases that underwent total laryngectomy at Amir Alam Hospital. We evaluated TB and CNS based on the Boxberg et al.

Our study demonstrated a significant correlation between TB, and nodal involvement (p = 0.015), vascular invasion (p = 0.035), and mortality rate (p = 0.001), as well as a significant statistical correlation between high TB and extra-laryngeal extension (p = 0.006), clinical stage (p = 0.011), and mortality rate (p = 0.001). Moreover, small nest size was also associated with the clinical stage (p = 0.047), extra-laryngeal extension (p = 0.015), and mortality rate (p < 0.001). Based on our results, TB, CNS, and clinical stage are independent prognostic factors for mortality rate and are correlated with disease-specific survival.

Given the effect of TB and CNS on the overall prognosis and survival of patients with LSCC, evaluating these two factors on routine H&E microscopic examination of LSCC specimens is recommended to facilitate individualized risk assessment and treatment planning.

Serous ovarian carcinoma (SOC) is the most common subtype of epithelial ovarian cancer, with high-grade (HGSOC) and low-grade (LGSOC) subtypes presenting distinct clinical behaviours. This study aimed to evaluate histopathologic features in SOC, correlating these with prognostic outcomes, and explore the potential clinical implications.

We analysed 51 SOC cases for lymphovascular space invasion (LVSI), tumour border configuration (TBC), microvessel density (MVD), tumour budding (TB), the tumour-stroma ratio (TSR), the stromal type, tumour-infiltrating lymphocytes (TILs), and tertiary lymphoid structures (TLSs). A validation cohort of 54 SOC cases from The Cancer Genome Atlas (TCGA) was used for comparison.

In the discovery set, significant predictors of aggressive behaviour included LVSI, high MVD, high TB, and low TILs. These findings were validated in the validation set where the absence of TLSs, lower peritumoural TILs, immature stromal type, and low TSR were associated with worse survival outcomes. The stromal type was identified as an independent prognostic predictor in SOC across both datasets. Inter-observer variability analysis demonstrated substantial to almost perfect agreement for these features, ensuring the reproducibility of the findings.

The histopathological evaluation of immune and stromal features, such as TILs, TLSs, TB, TSR, and stromal type, provides critical prognostic information for SOC. Incorporating these markers into routine pathological assessments could enhance risk stratification and guide treatment, offering practical utility, particularly in low-resource settings when molecular testing is not feasible.

Tumor budding has been proposed as a potential prognostic marker in various cancers, but its association with survival outcomes in breast cancer (BC) remains unclear. This meta-analysis aimed to clarify the relationship between tumor budding and survival outcomes in patients with BC. A comprehensive literature search was conducted in PubMed, EMBASE, and Web of Science. Cohort studies examining the association between tumor budding and overall survival (OS) and progression-free survival (PFS) in BC patients were included. Hazard ratios (HRs) and 95% confidence intervals (CIs) were pooled using a random-effects model to account for potential heterogeneity. Eleven cohort studies, including 2,828 patients, met the inclusion criteria. High tumor budding was significantly associated with poorer OS (HR = 1.89, 95% CI = 1.37-2.60, P < 0.001) and PFS (HR = 1.89, 95% CI = 1.32-2.71, P < 0.001). Subgroup analyses revealed a stronger association in studies where high tumor budding was defined as ≥ 10 buds / high-power field (HPF) compared to those with lower cutoffs. Sensitivity analyses confirmed the robustness of the findings. This meta-analysis demonstrates that high tumor budding is associated with significantly worse OS and PFS in BC patients, underscoring its prognostic significance. These findings suggest tumor budding could be a valuable marker in clinical assessments, and further research is needed to standardize its evaluation criteria in BC.

Some colorectal cancers (CRCs) are clinically diagnosed as cT4a with　serosal invasion (SI). However, the cT4a is most often underdiagnosed pathologically as pT3 without SI by hematoxylin-eosin (H&E) staining alone. Using Elastica van Gieson (EVG) staining, some pT3 tumors invade the elastic lamina (EL), which extends just below the serosal layer. Recently, EL invasion (ELI) has been described as a poor prognostic factor for disease-free survival (DFS) and overall survival (OS) in patients with pStage II CRC. However, its clinicopathological significance remains unclear due to the limited number of studies and poor understanding of ELI.

This study investigated the association between the ELI and patient prognosis.

After 1982, pathological diagnosis was routinely performed using H&E and EVG staining methods, and long-term follow up was performed until 2016. All clinicopathological features including ELI were prospectively registered into our computer and 569 patients with pStage II CRC were collected from the database. Based on the ELI status, pT3 was divided into three pathological categories: pT3ELI - was defined as pT3a, pT3ELI + as pT3b and unidentified EL (pT3EL -) as pT3u.

Using H&E staining alone, gross cT4a was most often pathologically underdiagnosed as pT3 (93.8%) and very rarely as pT4a, resulting in a large diagnostic discrepancy. Using EVG staining, 60.7% of the cT4a tumors were diagnosed as pT3b. The 10-year DFS and OS rates were similar for pT3a and pT3u patients. However, the 10-year DFS and OS rates of pT3b patients were significantly lower than those of pT3a patients (75.6% vs. 95.6%, p < 0.0001 and 58.4% vs. 70.6%, p = 0.0024, respectively) but did not differ from those of pT4a patients (70.6%, p = 0.5799 and 52.0%, p = 0.1116, respectively). Multivariate analysis revealed that the ELI was the strongest independent risk factor for recurrence and CRC-specific death (p < 0.0001).

A better understanding of the ELI allows us to reconsider the diagnostic discrepancy of serosal invasion, i.e., pT3b should be considered pT4a. The ELI-based subclassification of pT3 is expected to be incorporated into the TNM staging system in the future. The ELI is a notable prognostic indicator in patients with pStage II CRC.

Squamous cell carcinoma (SQCC) represents a significant proportion of human malignancies affecting various anatomical sites, including the lung. Understanding the prognostic factors is crucial for establishing effective risk stratification in these patients, as multiple critical aspects significantly impact overall survival.

A retrospective study was conducted on 99 patients with operable lung SQCC treated at a tertiary center. The exclusion criteria included patients under 18, those with in situ or metastatic SQCC, and those who received neoadjuvant therapy. The surgical specimens were re-analyzed, and data were collected on multiple variables, including pTNM staging, tumor characteristics, and overall survival (OS). The Kaplan-Meier survival analysis and Cox regression models were used to identify significant prognostic factors.

The Kaplan-Meier analysis showed a median survival of 36 months with a 65.65% mortality rate. Significant factors influencing survival included keratinization, histological grading, tumor size and stage, pleural invasion, tumor cell arrangement, tumor budding, spread through air space (STAS), and mitotic index. A multiple Cox regression highlighted the nonkeratinizing tumors, advanced pT stages, single-cell invasion, and high mitotic index as key predictors of poorer outcomes. The nonkeratinizing tumors showed higher mortality and shorter median survival rates compared to keratinizing tumors. The tumor staging, cell arrangement, and tumor budding significantly impacted the survival curves.

The study underscores the importance of detailed histopathological evaluations in lung SQCC. The nonkeratinizing tumors, advanced pT stage, single-cell invasion, and high mitotic index were associated with higher hazard rates, emphasizing the need for a comprehensive grading system incorporating these factors to improve prognostic accuracy and guide treatment strategies.

Cancer dormancy followed by recurrence remains an enigma in cancer biology. Since both local and systemic recurrences are thought to emanate from dormant micrometastasis which take origin from lymphovascular tumor emboli we wondered whether the process of dormancy might initiate within lymphovascular emboli. This study combines experimental studies with a patient-derived xenograft (PDX) of inflammatory breast cancer (Mary-X) that spontaneously forms spheroids in vitro and budding lymphovascular tumor emboli in vivo with observational studies utilizing tissue microarrays (TMAs) of human breast cancers. In the experimental studies, Mary-X during both lymphovascular emboli formation in vivo and spheroidgenesis in vitro exhibited decreased proliferation, a G0/G1 cell cycle arrest and decreased mTOR signaling. This induction of dormancy required calpain-mediated E-cadherin proteolysis and was mediated by decreased P13K signaling, resulting in decreased mTOR activity. In observational human breast cancer studies, increased E-cadherin immunoreactivity due to increased E-cad/NTF-1 but both decreased Ki-67 and mTOR activity was observed selectively and differentially within the lymphovascular tumor emboli. Both our experimental as well as observational studies indicate that in vivo lymphovascular tumor emboli and their in vitro spheroid equivalent initiate dormancy through these pathways.

Tumor microenvironment (TME) characteristics including tumor stroma ratio (TSR), tumor budding (TB), and tumor-infiltrating lymphocytes (TILs) were examined in resected gastric cancer. These TME features have been shown to indicate metastatic potential in colon cancer, and intestinal-type gastric cancer (IGC) has pathological similarities with that malignancy.

TSR, TB, and TILs were quantified in routine histological sections from 493 patients with IGC who underwent radical resection at 2 university hospitals in China from 2010 to 2016. TME variables were dichotomized as follows: TSR (50%), TILs (median), TB per international guidelines (4 buds/0.785mm2), and platelet-lymphocyte ratio (PLR) per survival ROC. Association of TME features with patient clinicopathological characteristics, time-to-recurrence (TTR), and cancer-specific-survival (CSS) were examined using univariate and multivariate analysis, including a relative contribution analysis by Cox regression.

Patients whose tumors showed high TSR or high TB or low TILs were each significantly associated with increased T and N stage, higher histological grade, and poorer TTR and CSS at 5 years. Only TSR and N stage were independently associated with TTR and CSS after adjustment for covariates. PLR was only independently associated with TTR after adjustment for covariates. Among the variables examined, only TSR was significantly associated with both TTR (HR 1.72, 95% CI, 1.14-2.60, P = .01) and CSS (HR 1.62, 95% CI, 1.05-2.51, P = .03) multivariately. Relative contribution to TTR revealed that the top 3 contributors were N stage (45.1%), TSR (22.5%), and PLR (12.9%), while the top 3 contributors to CSS were N stage (59.9%), TSR (14.7%), and PLR (10.9%).

Among the examined TME features, TSR was the most robust for prognostication and was significantly associated with both TTR and CSS. Furthermore, the relative contribution of TSR to patient TTR and CSS was second only to nodal status.

The liver's unique cellular structure makes it a frequent site for metastatic cancer. In colorectal liver metastasis (CRLM), surgical resection is essential for long-term survival. Histopathological growth patterns (HGPs) in CRLM, including desmoplastic and nondesmoplastic patterns, provide critical prognostic information. Tumor budding (TB) and poorly differentiated clusters (PDCs), indicators of aggressive cancer behavior, are evaluated using standardized histological scoring systems and are linked to epithelial-mesenchymal transition. This study explored the correlation between HGPs, TB, and PDCs in CRLM. Archived data from Thammasat University Hospital, including resected CRLM specimens, were analyzed. This study evaluated 51 CRLM resection specimens treated with preoperative systemic therapy, finding most to be nondesmoplastic with low TB and grade 1 PDC. Desmoplastic growth was significantly more prevalent in cases receiving preoperative chemotherapy than those that did not. Higher 3-year mortality was noted in nondesmoplastic groups and those with higher TB and tumor regression grade (TRG) scores. Significant correlations were observed between HGPs, TB, and PDCs, despite challenges in assessing these parameters due to issues with noncancer cells, extracellular mucin, bile ductular proliferation, and retraction artifacts. This study underscores the prognostic significance of HGPs, TB, PDCs, and TRG scores in CRLM, highlighting the need for precise histopathological evaluation for more accurate prognostic implications.

The long-term prognosis of colon cancer patients remains little changed with relatively high mortality and morbidity. Since the most widely used prognostic parameter TNM staging system is less satisfactory in predicting prognosis in early-stage cancers, numerous clinicopathological factors, including tumor necrosis, have been proposed for prognosis stratification, but substantial evidences are still lacking for early-stage colon cancer.

In the retrospective study, a total of eligible 173 stage I-II colon cancer patients, who received tumor radical resection and lymphadenectomy in the local hospital between January 1, 2010, and December 31, 2018, were enrolled for analyzing the prognostic role of tumor necrosis. The primary endpoints included 5-year overall survival (OS) and progression-free survival (PFS).

The median follow-up of enrolled early-stage colon cancer patients was 58.3 months. The 2-year and 5-year OS rates were 88.3% and 68.2%, respectively, and the 2-year and 5-year PFS rates were 85.6% and 62.7%, respectively. Seventy-eight patients (45.1%) were diagnosed with tumor necrosis by pathological examination. Demographic analysis revealed a significant association of tumor necrosis with larger tumor size and a marginal association with vascular invasion. Kaplan-Meier survival curves demonstrated that tumor necrosis was associated with worse OS (log-rank P = 0.003) and PFS (log-rank P = 0.002). The independent unfavorable prognostic effect of tumor necrosis was further validated in univariate and multivariate Cox regression analysis (hazard ratio = 1.91 (1.52-2.40), P = 0.004).

The current study confirmed the independent prognostic role of tumor necrosis from pathological review in early-stage colon cancer patients. This pathological criterion promises to help in identifying high-risk subgroup from early-stage colon cancer patients, who may benefit from strict follow-up and adjuvant therapy.

Pancreatic ductal adenocarcinoma (PDAC) is considered a low-immunogenic (LI) tumor with a "cold" tumor microenvironment and is mostly unresponsive to immune checkpoint blockade therapies. In this study, we decipher the impact of intratumoral heterogeneity of immune determinants on antitumor responses.

We performed spatial proteomic and transcriptomic analyses and multiplex immunofluorescence on multiple tumor regions, including tumor center (TC) and invasive front (IF), from 220 patients with PDAC, classified according to their transcriptomic immune signaling into high-immunogenic PDAC (HI-PDAC, n = 54) and LI PDAC (LI-PDAC, n = 166). Spatial compartments (tumor: pancytokeratin+/CD45- and leukocytes: pancytokeratin-/CD45+) were defined by fluorescence imaging.

HI-PDAC exhibited higher densities of cytotoxic T lymphocytes with upregulation of T-cell priming-associated immune determinants, including CD40, ITGAM, glucocorticoid-induced TNF-related receptor, CXCL10, granzyme B, IFNG, and HLA-DR, which were significantly more prominent at the IF than at the TC. In contrast, LI-PDAC exhibited immune-evasive tumor microenvironments with downregulation of immune determinants and a negative gradient from TC to IF. Patients with HI-PDAC had significantly better outcomes but showed more frequently exhausted immune phenotypes.

Our results indicate strategic differences in the regulation of immune determinants, leading to different levels of effectiveness of antitumor responses between HI and LI tumors and dynamic spatial changes, which affect the evolution of immune evasion and patient outcomes. This finding supports the coevolution of tumor and immune cells and may help define therapeutic vulnerabilities to improve antitumor immunity and harness the responsiveness to immune checkpoint inhibitors in patients with PDAC.

Background Triple-negative breast cancer (TNBC) is a highly aggressive subtype of breast cancer characterized by the lack of expression of estrogen and progesterone receptors and the absence of HER2 protein overexpression or gene amplification. How TNBC becomes so aggressive at the molecular level is not yet fully understood. The epithelial-mesenchymal transition (EMT) has been increasingly recognized as playing a pivotal role in cancer progression and metastasis. This study aimed to elucidate the connection between TNBC progression with EMT-related markers, including vimentin, beta-catenin, and E-cadherin. Methodology Rigorous immunohistochemical analysis was employed to assess the expression of vimentin, beta-catenin, and E-cadherin in primary tumors, tumor buds, and lymph node metastases (LNMs) from 137 cases with an invasive ductal carcinoma triple-negative phenotype diagnosed between 2018 and 2024. The EMT index, which was especially important in our work, is the sum of vimentin and beta-catenin expression divided by that of E-cadherin. Estimated Pearson correlation, multiple linear regression, and Kruskal-Wallis tests were used to determine the relationships of the EMT index with tumor buds and tumor-infiltrating lymphocytes (TILs). Results Vimentin highly correlated within separate regions of interest with Pearson correlation ranging from 0.90 to 0.92 (p < 0.001). Strong negative correlations between E-cadherin and vimentin (r = -0.81 to - 0.89, p < 0.001) showed its role in preserving the epithelial phenotype. The presence of tumor buds, aggregates, or clusters of cancer cells shed from the primary tumor mass invading the connective tissue showed very strong associations with the EMT index (r = 0.91, p < 0.001). Its presence is suggestive of aggressive disease and may identify a high-risk subpopulation that may benefit from more active surveillance or adjuvant treatment. Similarly, TILs correlated inversely with the EMT index (r = -0.90, p < 0.001). The most significant predictor of the EMT index, i.e., vimentin, had a model R-squared value of 1.000 in the regression analysis. Conclusions This study brings to light the importance of EMT-related markers in TNBC progression, with special emphasis on tumor buds as possible prognostic indicators for aggressive disease. The negative correlation of TILs with the EMT index indicates that an effective immune response could antagonize EMT-mediated tumor progression. These results suggest that EMT-based treatments in TNBC should be designed from a multimarker perspective by including interactions among several markers to optimize predictions and therapeutics. The results hold the potential to set future research directions and actionable outcomes that could influence clinical utility in the battle against TNBC.

 Tumor budding, defined as small clusters of tumor cells at the invasive front of carcinomas, has gained attention as a potential prognostic marker in various cancers. This study aimed to evaluate the utility of tumor budding as a histopathological marker in breast cancer and compare it to traditional prognostic markers such as histological grading, tumor-node-metastasis (TNM) staging, and molecular subtypes.

 A prospective, cross-sectional study was conducted over two years (June 2022 to May 2024) in the Department of Pathology at Jawaharlal Nehru Medical College, Wardha. Seventy-two female patients diagnosed with breast carcinoma who underwent modified radical mastectomy were included. Tumor budding was assessed through histopathological analysis and categorized as high or low. Statistical correlations were established between tumor budding and tumor size, histological grade, lymph node involvement, vascular invasion, and molecular subtypes (estrogen receptor, progesterone receptor, human epidermal growth factor receptor 2, and triple-negative breast cancer, TNBC). The chi-square test and multiple regression analyses were applied to assess significance.

 High tumor budding was observed in 68.1% of patients and was significantly associated with larger tumor size (p = 0.040), higher histological grade (p = 0.009), lymph node metastasis (p = 0.002), and vascular invasion (p = 0.003). The postmenopausal age group (>55 years) demonstrated a higher prevalence of high budding (p = 0.010). No significant correlation was found between tumor budding and molecular subtypes (p = 0.39), although high budding was more frequent in TNBC cases.

 Tumor budding is significantly associated with more aggressive tumor characteristics in breast cancer. Incorporating tumor budding into routine pathological assessments alongside TNM staging and histological grading may enhance the ability to identify high-risk patients and guide treatment strategies. Further large-scale, multicenter studies are warranted to confirm its prognostic value.

The aim of this study was to analyze the expression of Snail in the colorectal adenocarcinoma.

This study used a cross-sectional design. Seventy four paraffin embedded block of Colorectal Adenocarcinoma were assessed using Snail rabbit polyclonal antibody and their expression were performed using Olympus CX-43 light microscope. The relationship between Snail expression with histopathological grading, tumor budding grading, lymphovascular invasion and metastases of colorectal adenocarcinoma ability were statistically analyzed by Mann Whitney tests and presented in tables using SPSS 27.

From 74 samples examined, in samples with low grade tumor budding (n=11), there were 9 samples (81.8%) with weak expression, while those with strong expression were 2 samples (18.2%). In samples with intermediate grade tumor budding (n=28), there were 17 samples (60.7%) with weak expression, while those with strong expression were 11 samples (39.3%). In samples with high grade tumor budding (n=35), there were 13 samples (37.1%) with weak expression, while those with strong expression were 22 samples (62.9%). In samples with lymphovascular invasion (n=14), there were 10 samples (71.4%) with strong expression, while those with weak expression were 4 samples (28.6%). In samples with metastases (n=23), there were 16 samples (69.6%) with strong expression, while those with weak expression were 7 samples (30.4%). There was a significant relationship between the expression of Snail with tumor budding grade (p=0.003), lymphovascular invasion and metastases (p=<0.001), but there was no significant relationship with histopathological grade (p=0.942).

The Snail expression can be used as a prognostic factor in colorectal adenocarcinoma.

Epithelial-mesenchymal transition (EMT) is defined as a cellular process during which epithelial cells acquire mesenchymal phenotypes and behavior following the downregulation of epithelial features. EMT and its reversed process, the mesenchymal-epithelial transition (MET), and the special form of EMT, the endothelial-mesenchymal transition (EndMT), have been considered as mainstream concepts and general rules driving developmental and pathological processes, particularly cancer. However, discrepancies and disputes over EMT and EMT research have also grown over time. EMT is defined as transition between two cellular states, but it is unanimously agreed by EMT researchers that (1) neither the epithelial and mesenchymal states nor their regulatory networks have been clearly defined, (2) no EMT markers or factors can represent universally epithelial and mesenchymal states, and thus (3) EMT cannot be assessed on the basis of one or a few EMT markers. In contrast to definition and proposed roles of EMT, loss of epithelial feature does not cause mesenchymal phenotype, and EMT does not contribute to embryonic mesenchyme and neural crest formation, the key developmental events from which the EMT concept was derived. EMT and MET, represented by change in cell shapes or adhesiveness, or symbolized by EMT factors, are biased interpretation of the overall change in cellular property and regulatory networks during development and cancer progression. Moreover, EMT and MET are consequences rather than driving factors of developmental and pathological processes. The true meaning of EMT in some developmental and pathological processes, such as fibrosis, needs re-evaluation. EMT is believed to endow malignant features, such as migration, stemness, etc., to cancer cells. However, the core property of cancer (tumorigenic) cells is neural stemness, and the core EMT factors are components of the regulatory networks of neural stemness. Thus, EMT in cancer progression is misattribution of the roles of neural stemness to the unknown mesenchymal state. Similarly, neural crest EMT is misattribution of intrinsic property of neural crest cells to the unknown mesenchymal state. Lack of basic rationale in EMT and related concepts urges re-evaluation of their significance as general rules for understanding developmental and pathological processes, and re-evaluation of their significance in scientific research.

The progression of cancer cell migration, invasion and subsequent metastasis is the main cause of mortality in cancer patients. Through creating more accurate cancer models, we can achieve more precise results, which will lead to a better understanding of the invasion process. This holds promise for more effective prevention and treatment strategies. Although numerous 2D and 3D cell culture systems have been developed, they poorly reflect the in vivo situation and many questions have remained unanswered. This work describes a novel dynamic 3D cell culture system aimed at advancing our comprehension of cancer cell migration. With the newly designed cultivation chamber, 3D tumor spheroids were cultivated within a collagen I matrix in the presence of fluid flow to study the migration of cancer cells from spheroids in the matrix. Using light sheet microscopy and histology, we demonstrated that the morphology of spheroids is influenced by dynamic culture and that, in contrast to static culture, spheroids in dynamic culture are characterized by the absence of a large necrotic core. Additionally, this influence extends to an increase in the size of migration area, coupled with an increase in expression of some genes related to epithelial-mesenchymal transition (EMT). The results here highlight the importance of dynamic culture in cancer research. Although the dynamic 3D cell culture system in this study was used to investigate migration of one cell type into a matrix, it has the potential to be further developed and used for more complex models consisting of different cell types or to analyze other steps of metastasis development such as transendothelial migration or extravasation.

Epithelial-mesenchymal transition (EMT) is a cellular process in embryonic development, wound healing, organ fibrosis, and cancer metastasis. Previously, we and others have reported that proinflammatory cytokine interleukin-1β (IL-1β) induces EMT. However, the exact mechanisms, especially the signal transduction pathways, underlying IL-1β-mediated EMT are not yet completely understood. Here, we found that IL-1β stimulation leads to the partial EMT-like phenotype in human lung epithelial A549 cells, including the gain of mesenchymal marker (vimentin) and high migratory potential, without the complete loss of epithelial marker (E-cadherin). IL-1β-mediated partial EMT induction was repressed by PI3K inhibitor LY294002, indicating that the PI3K/AKT pathway plays a significant role in the induction. In addition, ERK1/2 inhibitor FR180204 markedly inhibited the IL-1β-mediated partial EMT induction, demonstrating that the MEK/ERK pathway was also involved in the induction. Furthermore, we found that the activation of the PI3K/AKT and MEK/ERK pathways occurred downstream of the epidermal growth factor receptor (EGFR) pathway and the IL-1 receptor (IL-1R) pathway, respectively. Our findings suggest that the PI3K/AKT and MEK/ERK pathways coordinately promote the IL-1β-mediated partial EMT induction. The inhibition of not one but both pathways is expected yield clinical benefits by preventing partial EMT-related disorders such as organ fibrosis and cancer metastasis.

Clinical outcome for patients suffering from HPV-negative head and neck squamous cell carcinoma (HNSCC) remains poor. This is mostly due to highly invasive tumors that cause loco-regional relapses after initial therapeutic intervention and metastatic outgrowth. The molecular pathways governing the detrimental invasive growth modes in HNSCC remain however understudied. Here, we have established HNSCC patient derived organoid (PDO) models that recapitulate 3-dimensional invasion in vitro. Single cell mRNA sequencing was applied to study the differences between non-invasive and invasive conditions, and in a collective versus single cell invading PDO model. Differential expression analysis under invasive conditions in Collagen gels reveals an overall upregulation of a YAP-centered transcriptional program, irrespective of the invasion mode. However, we find that collectively invading HNSCC PDO cells show elevated levels of YAP transcription targets when compared to single cell invasion. Also, collectively invading cells are characterized by increased nuclear translocation of YAP within the invasive strands, which coincides with Collagen-I matrix alignment at the invasive front. Using gene set enrichment analysis, we identify immune cell-like migratory pathways in the single cell invading HNSCC PDO, while collective invasion is characterized by overt upregulation of adhesion and migratory pathways. Lastly, based on clinical head and neck cancer cohorts, we demonstrate that the identified collective invasion signature provides a candidate prognostic platform for survival in HNSCC. By uncoupling collective and single cell invasive programs, we have established invasion signatures that may guide new therapeutic options.

Tumor budding (TB) has shown promising results as a prognostic marker in several cancers such as colorectal carcinoma, breast carcinoma etc. It has been co-related to aggressiveness of the tumor and can also predict the metastasis to the lymph nodes. This systematic review evaluates the prognostic potential of TB in predicting lymph node metastasis (LNM) in OSCC.

Systematic search was carried out in the electronic data-bases i.e. PubMed, Cochrane and Google scholar for original studies related to TB in OSCC. The assessment of risk bias was done using QUIPS tool. Meta-analysis was done using STATA software.

A total of 25 articles were included. A significant association was noted for overall survival and prognosis but not for TB LNM in OSCC. Meta-analysis revealed a pooled estimate i.e odds ratio of 2.10 (CI - 0.00 - 4.20) for TB and LNM while for overall survival, it was 2.29 (CI-1.81-2.76).

Tumor budding though is strongly associated with LNM in OSCC did not show significant relationship in this systematic review but demonstrated a higher correlation with overall survival. It highlights that TB is an important parameter for prognosis of oral cancer but its potential in prediction of LNM needs further validation.

Sinonasal intestinal-type adenocarcinoma (ITAC) is a very rare, closely occupational-related tumor with strong histological similarities to colorectal cancer (CRC). In the latter, tumor budding (TB) is widely recognized as a negative prognostic parameter. The aim of this study was to evaluate the prognostic role of TB in ITAC and to correlate it with other established or emerging biomarkers of the disease, such as p53 and deficient DNA mismatch repair (MMR) system status/microsatellite instability (MSI). We retrospectively analyzed 32 consecutive specimens of patients with ITAC diagnosis treated in two institutions in Northern Italy. We reviewed surgical specimens for TB evaluation (low-intermediate/high); p53 expression and MMR proteins were evaluated via immunohistochemistry. Results were retrospectively stratified using clinical data and patients' outcomes. According to bud counts, patients were stratified into two groups: intermediate/high budding (>4 TB) and low budding (≤4 TB). Patients with high TB (>4) have an increased risk of recurrence and death compared to those with low TB, with a median survival of 13 and 54 months, respectively. On multivariate analysis, considering TB, therapy, and stage as covariates, TB emerged as an independent prognostic factor net of the stage of disease or type of therapy received. No impact of p53 status as a biomarker of prognosis was observed and no alterations regarding MMR proteins were identified. The results of the present work provide further significant evidence on the prognostic role of TB in ITAC and underline the need for larger multicenter studies to implement the use of TB in clinical practice.

At present, there is limited research on the mechanisms underlying moyamoya disease (MMD). Herein, we aimed to determine the role of glutamine in MMD pathogenesis, and 360 adult patients were prospectively enrolled. Human brain microvascular endothelial cells (HBMECs) were subjected to Integrin Subunit Beta 4 (ITGB4) overexpression or knockdown and atorvastatin. We assessed factors associated with various signaling pathways in the context of the endothelial-to-mesenchymal transition (EndMT), and the expression level of related proteins was validated in the superficial temporal arteries of patients. We found glutamine levels were positively associated with a greater risk of stroke (OR = 1.599, p = 0.022). After treatment with glutamine, HBMECs exhibited enhanced proliferation, migration, and EndMT, all reversed by ITGB4 knockdown. In ITGB4-transfected HBMECs, the MAPK-ERK-TGF-β/BMP pathway was activated, with Smad4 knockdown reversing the EndMT. Furthermore, atorvastatin suppressed the EndMT by inhibiting Smad1/5 phosphorylation and promoting Smad4 ubiquitination in ITGB4-transfected HBMECs. We also found the protein level of ITGB4 was upregulated in the superficial temporal arteries of patients with MMD. In conclusion, our study suggests that glutamine may be an independent risk factor for hemorrhage or infarction in patients with MMD and targeting ITGB4 could potentially be therapeutic approaches for MMD.

Epidermal growth factor receptor (EGFR)-mutant non-small-cell lung cancer (NSCLC) patients treated with EGFR-tyrosine kinase inhibitors (TKIs) inevitably develop resistance through several biological mechanisms. However, little is known on the molecular mechanisms underlying acquired resistance to suboptimal EGFR-TKI doses, due to pharmacodynamics leading to inadequate drug exposure. To evaluate the effects of suboptimal EGFR-TKI exposure on resistance in NSCLC, we obtained HCC827 and PC9 cell lines resistant to suboptimal fixed and intermittent doses of gefitinib and compared them to cells exposed to higher doses of the drug. We analyzed the differences in terms of EGFR signaling activation and the expression of epithelial-mesenchymal transition (EMT) markers, whole transcriptomes byRNA sequencing, and cell motility. We observed that the exposure to low doses of gefitinib more frequently induced a partial EMT associated with an induced migratory ability, and an enhanced transcription of cancer stem cell markers, particularly in the HCC827 gefitinib-resistant cells. Finally, the HCC827 gefitinib-resistant cells showed increased secretion of the EMT inducer transforming growth factor (TGF)-β1, whose inhibition was able to partially restore gefitinib sensitivity. These data provide evidence that different levels of exposure to EGFR-TKIs in tumor masses might promote different mechanisms of acquired resistance.