Protein induced by vitamin K absence or antagonist-II in hepatocellular carcinoma with normal alpha-fetoprotein: Current advances and controversies

Abstract

A recent retrospective study evaluated the prognostic value of protein induced by vitamin K absence/antagonist-II (PIVKA-II) in patients with hepatocellular carcinoma (HCC) who have normal alpha-fetoprotein (AFP) levels. The study demonstrated that, among AFP-normal HCC patients, PIVKA-II exhibits a stratification threshold for invasive tumors and can serve as a supplementary indicator for risk stratification beyond AFP. Although this study suggests a potential correlation between elevated PIVKA-II levels and invasive tumor phenotypes, the predictive stability and generalizability of PIVKA-II for tumor recurrence still require further validation due to the limited clinical database and insufficient external verification. Future studies should be conducted in multiple prospective cohorts, with an expanded sample size and extended follow-up duration. In addition, the current study only included patients who had undergone PIVKA-II testing, which may introduce selection bias, thus limiting its general applicability. Finally, greater attention should be paid to the dynamic changes of PIVKA-II, and its predictive value for disease progression or treatment response should be evaluated by combining it with imaging and clinical variables. In this paper, we evaluate the strengths and limitations of this study and propose future research directions to refine the research model and deepen the understanding of the role of PIVKA-II in the invasive biology of AFP-normal HCC.

Key Words: Hepatocellular carcinoma; Alpha-fetoprotein; Protein induced by vitamin K absence/antagonist-II; Des-gamma carboxy prothrombin; Aggressive tumor

Core Tip: Alpha-fetoprotein (AFP)-normal hepatocellular carcinoma is a clinically challenging subgroup in which conventional biomarker-driven surveillance and risk assessment often underperform. Current evidence indicates that protein induced by vitamin K absence/antagonist-II (PIVKA-II) can supplement AFP for detection, provide information on tumor aggressiveness and microvascular invasion, and contribute to recurrence risk stratification after resection, locoregional therapy, and liver transplantation. At the same time, major controversies remain around assay harmonization, cut-off selection, regional generalizability, vitamin K-related confounding, and the incremental value of PIVKA-II relative to modern multi-marker panels and imaging-based models. Our perspective is that PIVKA-II is most promising when treated as a context-dependent component of integrated risk modelling rather than as a binary standalone test.

INTRODUCTION

Liver cancer is the fifth most common cancer and the second most frequent cause of cancer-related death globally, with 854000 new cases and 810000 deaths per year, accounting for 7% of all cancers[1,2]. Hepatocellular carcinoma (HCC), the most common type of primary liver cancer, accounting for 75%-86% of cases, remains a major global public health challenge[3]. The standard screening methods for the early diagnosis of HCC in high-risk populations include ultrasonography, computed tomography (CT), magnetic resonance imaging (MRI), and serum tumor markers (TMs)[4]. However, although CT and MRI can significantly improve the diagnostic accuracy of HCC, they are costly and therefore unsuitable for mass screening and surveillance[5]. Consequently, there is growing interest in the use of serum TMs for the early detection of HCC. Alpha-fetoprotein (AFP) levels above specific thresholds are widely used to predict HCC recurrence among clinicians for tumor staging, grading, and management[6]. However, several recent randomized trials have confirmed that AFP alone is not recommended for HCC surveillance, because nearly 80% of tumors smaller than 3 cm present with normal AFP levels, resulting in a low detection sensitivity of only 25% for these lesions[7,8]. Therefore, it is essential to search for new HCC-associated biomarkers, realize the combined detection of multiple indicators, improve the accuracy of early HCC diagnosis, and reduce the missed diagnosis rate. In this context, protein induced by vitamin K absence/antagonist II (PIVKA-II), also known as des-γ-carboxy prothrombin (DCP), as an abnormal form of prothrombin produced by liver tissues, was proven to improve HCC surveillance in high-risk populations[9]. Furthermore, the elevated degree of PIVKA-II is associated with high malignancy and poor prognosis, such as the presence of vascular invasion and poor differentiation of HCC cells[10]. Although various studies have demonstrated the utility of protein induced by vitamin K absence II (PIVKA-II) in surveillance, treatment monitoring, and predicting recurrence, it is still not recommended as a routine biomarker test[11,12]. Therefore, this article is to discuss the clinical usefulness and value of PIVKA-II for the surveillance and treatment monitoring of HCC, its benefits and limitations, and further steps required to improve its utility.

HOW SHOULD WARFARIN/VITAMIN K STATUS, DYNAMIC MONITORING, AND CUT-OFF VALIDATION BE HANDLED TO IMPROVE THE CLINICAL APPLICATION OF PIVKA-II IN HCC?

Warfarin is a vitamin K antagonist, and both warfarin exposure and inadequate vitamin K intake can suppress vitamin K-dependent coagulation factor activity, potentially leading to non-tumoral elevations in DCP/PIVKA-II and thereby introducing interpretive bias[13]. Warfarin is an anticoagulant with a narrow therapeutic window, and its efficacy and safety are primarily monitored by the international normalized ratio (INR)[14]. Basit et al[15] demonstrated that warfarin therapy significantly reduced the level of fully carboxylated prothrombin, while the accumulation of PIVKA-II was positively correlated with an increase in the INR. Accordingly, anticoagulant exposure and vitamin K status should be prespecified as key confounders when evaluating PIVKA-II. For patients receiving warfarin, temporary interruption may be considered under clinician supervision in accordance with anticoagulation management standards, typically 5-7 days, which may allow partial recovery of vitamin K-dependent coagulation factors before PIVKA-II re-assessment[16,17].

In parallel, as an angiogenic factor, serum PIVKA-II was proposed as a predictor of microvascular invasion in HCC[18]. Therefore, studies should adhere to contemporary anticoagulation practice and implement an a priori analytic plan, including exclusion of patients receiving vitamin K antagonists, stratified reporting by anticoagulant exposure, cholestasis and nutritional status, or multivariable adjustment incorporating anticoagulant use, cholestasis-related indices, and a history of vitamin K supplementation as covariates. When anticoagulation exposure cannot be avoided, concurrent reporting of coagulation parameters and interpretation in conjunction with AFP and imaging features are recommended to mitigate false-positive PIVKA-II signals and reduce downstream inferential bias[19-21]. Longitudinal PIVKA-II monitoring should be aligned with imaging reassessment windows, with recommended sampling at baseline (pre-treatment), early after treatment initiation 2-4 weeks, and subsequently every 2-3 months during follow-up for at least 12-24 months[20]. Finally, candidate cut-offs should be evaluated through a minimal acceptable validation pathway, including external multicenter cohorts across assay platforms, use of prespecified thresholds to avoid re-derivation in validation datasets, anchoring to hard clinical endpoints such as recurrence and survival with aggressive phenotype as a secondary endpoint, and transparent reporting of transportability and decision utility[22,23]. Mechanistically and clinically, these findings are consistent with previous evidence linking PIVKA-II to invasive tumor behavior. In alcoholic cirrhosis, PIVKA-II has demonstrated utility not only for diagnosis but also for predicting vascular invasion, supporting its biological plausibility as a marker of aggressiveness. From a transplant perspective, serial PIVKA-II measurements have shown high sensitivity for monitoring post-transplant recurrence, including in patients with normal AFP, suggesting that PIVKA-II may be particularly informative where AFP underperforms[24].

HOW SHOULD ENDPOINTS, DEFINITIONS, AND ASSAY DETAILS BE STANDARDIZED WHEN INTERPRETING THE PREDICTIVE VALUE OF PIVKA-II IN AFP-NORMAL HCC?

Serum AFP is one of the main indicators of early screening of liver cancer, and it is recommended that high risk groups undergo liver ultrasound and AFP tests every 6 months; however, the sensitivity and specificity of AFP for HCC diagnosis are only 68.8% and 87.6%, respectively[25,26]. To interpret the predictive value of PIVKA-II in AFP-normal HCC, standardization should begin with the endpoint, not with the biomarker itself. Baseline aggressive phenotype, post-curative recurrence, and on-treatment response are different clinical questions. They should not be merged under a general “predictive” label. If recurrence is the intended endpoint, studies should include only patients treated with curative intent. These studies should also clearly define recurrence-free survival or time to recurrence, the start of follow-up, the surveillance schedule, the early-recurrence window, and censoring rules[27]. In contrast, studies linking PIVKA-II to vascular invasion, beyond-Milan status, or tumor burden are describing aggressive disease at presentation. Systemic-therapy studies are evaluating dynamic biomarker response at fixed post-treatment timepoints. Each of these settings needs a separate analytical framework and should be interpreted independently[28,29]. In the following sections, we highlight potential mechanisms of PIVKA-II in HCC, which are summarized in Table 1[10,30-33].

Table 1 Potential mechanisms of protein induced by vitamin K absence/antagonist-II in hepatocellular carcinoma.

Ref.	Year	Potential mechanism
Bhatti et al[30]	2021	Association with invasive tumor phenotype
Qian et al[31]	2023	Complementary value in AFP-normal HCC
Dong et al[10]	2023	Value for recurrence surveillance
Kudo[32]	2024	Abnormal vitamin K-dependent carboxylation
Chen et al[33]	2025	Dynamic marker of treatment response

AFP: Alpha-fetoprotein; HCC: Hepatocellular carcinoma.

Definitions and assay details should be standardized with equal rigor. AFP-normal should be defined numerically, preferably as a clear threshold like < 20 ng/mL or the assay-specific upper limit of normal[34]. Narrative descriptions should be avoided, as the proportion of AFP-normal HCC varies with the threshold. This may account for roughly 20%-40% of cases[35]. PIVKA-II should always be reported with the assay platform, manufacturer, unit of measurement, specimen type, timing of blood collection, and the cut-off value method. This is particularly important because recent studies show meaningful inter-platform and inter-study variability in PIVKA-II thresholds[36]. The interpretation of elevated PIVKA-II levels may be confounded by vitamin K deficiency, warfarin exposure, obstructive jaundice, and alcohol-related liver disease[37,38]. A practical standardization strategy should include baseline sampling in all patients and fixed longitudinal sampling points based on the clinical scenario. The assay-specific cut-off should be derived in a training cohort and validated externally with the same analytical platform before its routine clinical adoption can be proposed[39].

WHAT IS THE CURRENT EVIDENCE ON THE ROLE OF PIVKA-II IN HCC WITH NORMAL AFP LEVELS? AND WHAT CHALLENGES REMAIN FOR ITS CLINICAL APPLICATION?

Recently, the diagnostic role of PIVKA-II has been widely discussed. At present, the Japan Society of Hepatology lists PIVKA-II in its guidelines as an important biological indicator for liver cancer detection; the Chinese Guidelines for prevention and treatment of chronic hepatitis B recommend PIVKA-II as an important indicator for diagnosis of HCC, which can be combined with AFP to facilitate early diagnosis[40-42]. Feng et al[43] explored the relationship between PIVKA-II and survival and prognosis of HCC. In that study, a total of 3 cohorts encompassing 521 patients were included, and their results suggested that PIVKA-II was still valuable for the diagnosis of AFP-negative HCC and can be used as a supplement of AFP in the diagnosis of HCC. A prospective study by Si et al[44] of 433 participants showed that the serum levels of AFP and PIVKA-II were positively correlated with tumor differentiation and size, and high AFP and PIVKA-II expression was significantly associated with the presence of vascular invasion (P = 0.007 and 0.014, respectively). In addition, the AFP level > 64.4 ng/mL or PIVKA-II level > 957.61 mAU/mL was the best critical value to predict the presence of vascular invasion. In the following section, we discuss prospective studies in PIVKA-II-based risk stratification for AFP-normal HCC patients, and a summary of this section is provided in Table 2[33,45-54].

Table 2 Recent developments in protein induced by vitamin K absence/antagonist-II-based risk stratification for alpha-fetoprotein-normal hepatocellular carcinoma patients.

Ref.	Year	Number of subjects	Primary role of PIVKA-II
Wang et al[45]	2022	46	PIVKA-II may be used as a marker for predicting transarterial chemoembolization response
Devillers et al[46]	2023	203	PIVKA-II’s better prediction of microvascular invasion suggests its role in identifying high-risk HCC recurrence post LT
Yu et al[47]	2023	809	PIVKA-II contributed to the development of prediction and diagnostic models for AFP-negative populations
Zhu et al[48]	2024	258	Serum AFP and PIVKA-II were evaluated for the detection of recurrent HCC, with PIVKA-II showing superior diagnostic performance
Lin et al[49]	2025	639	Serum glypican-3 and PIVKA-II were evaluated for detection and prognostic assessment in AFP-negative HCC
Gongor et al[50]	2025	707	Preoperative PIVKA-II, alone or in combination with AFP, may serve as an accessible long-term risk assessment marker for HCC recurrence and mortality following LDLT
Chen et al[33]	2025	61	PIVKA-II response was evaluated as an early marker of radiological and clinical outcomes in non-AFP-secreting HCC undergoing systemic therapy
Bhatti et al[51]	2025	400	Pre-transplant AFP and PIVKA-II were evaluated as biomarkers for recurrence risk assessment and selection of liver transplantation candidates with HCC
Zhang et al[52]	2025	751	PIVKA-II and AFP were evaluated for HCC diagnosis, with PIVKA-II showing superior diagnostic accuracy and complementary value for early detection
Abbas et al[53]	2026	113	PIVKA-II was evaluated for identifying aggressive disease and prognostic risk in HCC patients with normal AFP levels
Gao et al[54]	2026	117	PIVKA-II combined with MRI features, including intratumoral arteries and tumor ADC values, was evaluated for predicting microvascular invasion in HCC

PIVKA-II: Protein induced by vitamin K absence/antagonist-II; AFP: Alpha-fetoprotein; HCC: Hepatocellular carcinoma; LT: Liver transplantation; LDLT: Living donor liver transplantation; MRI: Magnetic resonance imaging; ADC: Apparent diffusion coefficient.

PIVKA-II is a well-known tumor marker of HCC[55]. Previous studies on the relationship between serum PIVKA-II level and various clinicopathologic factors of HCC have shown that elevation of PIVKA-II may be related to worse tumor behavior and prognosis in HCC patients[56]. However, there is still no consensus on the superior diagnostic value of PIVKA-II when compared with AFP. Furthermore, several unresolved issues require further discussion. First, although PIVKA-II/DCP is widely used across parts of the Asia-Pacific region and is increasingly positioned as complementary to AFP, substantial heterogeneity in proposed cut-offs remains a major implementation barrier, especially across assays, patient populations, disease etiologies, and treatment contexts[57,58]. Relevant Asia-Pacific consensus statements underscore both the clinical value of PIVKA-II in AFP-negative settings and the practical challenges of harmonizing thresholds and defining evidence-graded indications[59]. Furthermore, because PIVKA-II is biologically linked to vitamin K-dependent pathways, rigorous control of vitamin K-related confounding is essential. Vitamin K deficiency and exposure to vitamin K antagonists can increase DCP/PIVKA-II, potentially biasing interpretation if not appropriately addressed[60,61].

CONCLUSION

In summary, PIVKA-II is a promising adjunctive biomarker for AFP-normal HCC, particularly in improving detection, reflecting tumor aggressiveness, and supporting recurrence risk assessment across different treatment settings. Nevertheless, its routine clinical adoption remains constrained by unresolved issues, including endpoint inconsistency, assay heterogeneity, variable cut-off definitions, regional differences in evidence, and confounding from vitamin K-related factors. The next stage of research should move beyond isolated assessments of biomarker performance and focus on standardized, clinically stratified, and externally validated study designs. PIVKA-II is most likely to achieve meaningful clinical utility when incorporated into integrated models alongside AFP, imaging findings, and treatment context, rather than being interpreted as a universal stand-alone marker.

ACKNOWLEDGEMENTS

We thank all members of Xu Laboratory for helpful suggestions about the manuscript.