Biomarkers Of Response Research Articles

PIVKA-II as a surrogate biomarker for therapeutic response in Non-AFP-secreting hepatocellular carcinoma

BackgroundAlpha-fetoprotein (AFP) is a key biomarker for hepatocellular carcinoma (HCC), but 30–40% of cases are AFP-negative. Prothrombin induced by vitamin K absence II (PIVKA-II) is more sensitive for HCC detection, though its role in systemic therapy remains underexplored. This study aimed to evaluate PIVKA-II in non-AFP-secreting HCC treated with systemic therapy.MethodsPatients with unresectable HCC undergoing systemic therapy were enrolled. Baseline imaging and PIVKA-II levels were recorded. After 8–12 weeks of treatment, response was evaluated through imaging and repeat PIVKA-II measurements.ResultsA total of 116 treatment assessments from 61 cases were analyzed. Baseline PIVKA-II levels correlated with tumor size, but not tumor number or liver function. PIVKA-II regression (≥ 50% reduction) and progression (≥ 50% increase) were defined using ROC analysis. Imaging showed 71.0% objective response in the regression group, 50.0% stable disease in the stable group, and 83.7% progressive disease in the progression group (p < 0.001). This association held for targeted therapies, immune checkpoint inhibitors, and chemotherapy. Progression-free survival (PFS) for the regression, stable, and progression groups was non-reached, 6.7, and 3.2 months (p = 0.0002), and overall survival (OS) was non-reached, non-reached, and 18.5 months (p = 0.02).ConclusionsThis study is the first to establish the “50–50 rule” for PIVKA-II response in non-AFP-secreting HCC treated with systemic therapy, highlighting its value as a surrogate marker for radiological outcomes and prognosis.

The current status of tumor markers as biomarkers in the era of immunotherapy for hepatocellular carcinoma: alpha-fetoprotein alone is not sufficient.

Rapid development of systemic treatments has resulted in improved prognosis for unresectable hepatocellular carcinoma (uHCC) patients. Since immune therapy shows a favorable therapeutic efficacy, use of tumor markers as biomarkers for monitoring treatment response is necessary. This study aimed to elucidate changes in positive rates of 3 available tumor markers in Japan, including alpha-fetoprotein (AFP), des-gamma-carboxy prothrombin (DCP), and lens culinaris agglutinin-reactive AFP (AFP-L3) in uHCC patients treated with systemic therapies over time. From 2009 to 2023, 1470 uHCC patients with data of tumor markers before starting treatment were enrolled. The positivity cut-off value for AFP was 20 ng/mL, for AFP-L3 was 10%, and for DCP was 40 mAU/mL. After dividing the 15 years examined into three periods of five years each (period I, II, III), clinical features of the enrolled patients were evaluated, retrospectively. The percentage of Barcelona Clinic Liver Cancer stage B patients who received systemic therapy increased from period I to III (27.7%, 38.5%, 46.6%, respectively, P<0.001), which was also seen for HCC patients with a non-viral etiology (alcohol and others) (29.9%, 39.7%, 49.6%, respectively P<0.001). Positive rates for AFP (67.8%, 62.1%, 50.8%, respectively) and DCP (84.1%, 80.5%, 72.7%, respectively) were decreased (each P<0.001), while the AFP-L3 rate did not show a significant change (54.4%, 57.7%, 51.9%, respectively P=0.390). Among the AFP-negative patients, the rate of positive for DCP or AFP-L3 was increased (24.4%, 28.1%, 35.4%, respectively, P=0.002). Based on introduction of systemic treatment in an early stage and increasing numbers of HCC cases with a non-viral etiology, the positive rate of AFP level has been declining. Thus, determination of DCP and AFP-L3 in addition to AFP as markers should be more actively utilized in clinical practice, as well as clinical trials for monitoring and evaluating treatment response in this era of combination immunotherapy as a powerful treatment.

Photonic Nanomaterials for Wearable Health Solutions.

This review underscores the transformative potential of photonic nanomaterials in wearable health technologies, driven by increasing demands for personalized health monitoring. Their unique optical and physical properties enable rapid, precise, and sensitive real-time monitoring, outperforming conventional electrical-based sensors. Integrated into ultra-thin, flexible, and stretchable formats, these materials enhance compatibility with the human body, enabling prolonged wear, improved efficiency, and reduced power consumption. A comprehensive exploration is provided of the integration of photonic nanomaterials into wearable devices, addressing material selection, light-matter interaction principles, and device assembly strategies. The review highlights critical elements such as device form factors, sensing modalities, and power and data communication, with representative examples in skin patches and contact lenses. These devices enable precise monitoring and management of biomarkers of diseases or biological responses. Furthermore, advancements in materials and integration approaches have paved the way for continuum of care systems combining multifunctional sensors with therapeutic drug delivery mechanisms. To overcome existing barriers, this review outlines strategies of material design, device engineering, system integration, and machine learning to inspire innovation and accelerate the adoption of photonic nanomaterials for next-generation of wearable health, showcasing their versatility and transformative potential for digital health applications.

Biomarkers of response in patients with gastric/gastroesophageal junction adenocarcinoma (GEA) treated with telisotuzumab adizutecan.

490 Background: c-Met protein (MET protein) overexpression (OE) and MET amplification (amp) occur in several cancer types, including GEA, and are associated with poor prognosis. The c-Met–directed antibody-drug conjugate telisotuzumab adizutecan (ABBV-400) is being assessed in advanced solid tumors in an ongoing, first-in-human study (NCT05029882). Results showed promising antitumor activity and a tolerable safety profile in GEA (Strickler et al. ESMO 2024, 1439P). Here, we present our analysis on associations between c-Met OE, MET amp, and ctDNA-derived molecular response with clinical outcomes. Methods: c-Met expression was evaluated retrospectively in FFPE tissue (archival/fresh) collected at study screening using IHC (SP44 Ab clone) or whole transcriptomic RNA sequencing (Caris). Plasma ctDNA collected at baseline (BL) and cycle 3 day 1 (C3) was analyzed with the GuardantINFINITY assay. Molecular response (MR) was assessed using circulating tumor fraction (cTF), estimated on the basis of ctDNA variant allele frequencies in a 74-gene panel (Guardant360 CDx) and methylation signals (GuardantINFINITY methylome platform). MR was defined as a 50% decrease in cTF from BL. MET amp status was determined retrospectively by the Guardant Health algorithm. Radiographic response was assessed per RECIST v1.1. Results: Clinical data from 42 patients (pts) with advanced GEA were included in this biomarker analysis. There was a negative correlation between c-Met expression and tumor size change from BL (R=-0.59). Of 42 pts, 12 (29%) had tumors with MET amp per ctDNA analysis/local site reports, which was higher than the bioinformatic analysis of real-world data (Caris-ConcertAI linked RWD360 and PT360; ~3%). Concordance between c-Met expression per IHC and RNA was observed (R=0.75, p&lt;10 -6 ). There were 12 responders; 7 (58%) had MET amp, and 5 (42%) did not. Responders had lower ctDNA percentage change from BL vs nonresponders (1.5-fold and 1.6-fold difference based on 74-gene and methylation panel, respectively). Pts with MR had a greater decrease in tumor size from BL vs those without MR. A positive correlation between change in ctDNA levels (BL to C3) and change in tumor size was observed using the 74-gene panel and methylation panel (R=0.73 and 0.69, respectively). Additional biomarker analyses focused on the association between MET and other key biomarkers relevant in GEA will be discussed. Conclusions: In pts with advanced GEA treated with telisotuzumab adizutecan, c-Met OE/ MET amp and ctDNA-based MR were associated with enhanced antitumor activity. Compared with real-world data, MET amp was enriched in this study population. Phase 2 studies will continue to explore c-Met OE and MET amp prevalence and their potential predictive value with other biomarkers in a clinically representative population. Clinical trial information: NCT05029882 .

Single agent olaparib in advanced oesophago-gastric cancer: Primary results from the SOlar clinical phase II single arm trial.

427 Background: A subset of pts with advanced oesophago-gastric adenocarcinoma (OGA) are characterised by deficient DNA damage repair (DDR) providing a rationale for PARP inhibition (PARPi) in these tumours. Potential activity was observed in the phase 3 gastric cancer trial with Olaparib + paclitaxel, however the efficacy of Olaparib monotherapy and response biomarkers have not been established. Herein we report the primary clinical results from the SOlar trial. Methods: SOlar is a prospective, open label, single arm phase II trial using a Simon’s two stage optimal design, evaluating the efficacy and safety of Olaparib 300mg twice daily in pts with locally advanced/metastatic OGA who had previously progressed on ≥1 line of therapy in the advanced setting. Pts with measurable disease by RECIST v1.1 amenable to mandatory baseline biopsy were included. The primary end point was disease control rate (DCR) at 8 weeks from Olaparib initiation by RECIST v1.1. In this two-stage optimal design, if ≥5/27 pts had disease control in stage 1, a further 27 pts were to be recruited in stage 2. If ≥12/54 evaluable pts achieved disease control, Olaparib would warrant further investigation. Other key endpoints include; overall response rate, progression free survival and overall survival, and translational biomarker analyses (NCT03829345). Results: Between July 2019 - February 2024, 55 pts with locally advanced/metastatic OGA were enrolled. 51 pts received Olaparib and 50 were evaluable for the primary endpoint (5 withdrew). The median age was 61yrs (range: 54-67), 7 (14%) were HER2 -positive and 2 had a known germline BRCA mutation. 19 (38%) pts had received ≥3 lines of therapy and 22 (44%) pts had received prior immunotherapy for OGA. 30 (60%) pts had objective responses to prior platinum chemotherapy (CR + PR). The DCR at 8-weeks was 28% (91% one-sided confidence interval lower bound: 19.4) with 14/50 evaluable pts having stable disease. No objective response was observed. 36/51 (71%) treated pts experienced a treatment-related adverse event (TRAE) of any grade, and 9 (18%) experienced a grade ≥3 TRAE (most common being anaemia, fatigue, vomiting). Two serious adverse events were reported as treatment-related: definitely related grade 1 pneumonitis, and possibly related grade 4 lung infection. Conclusions: In SOlar, Olaparib met its primary endpoint with 28% DCR in this heavily pre-treated group. No new safety concerns were observed. Ongoing translational analyses include; homologous repair deficiency scoring, whole exome sequencing, RNA expression, serial ctDNA, and multiplex immunofluorescent assessment of the tumour micro-environment, with a view to identify a subgroup who might benefit from PARPi to support ongoing PARPi combinatorial studies in advanced OGA. Clinical trial information: NCT03829345 .

Assessing Torquetenovirus (TTV) as a Biomarker for Immune Responses to SARS-CoV-2 mRNA Vaccines in People Living with HIV and Healthy Individuals

Background: Torquetenovirus (TTV) viremia is increasingly recognized as a marker of immune competence. In the context of COVID-19, TTV viral load (VL) has been shown to predict anti-Spike antibody levels in severely immunocompromised patients. This study aimed to evaluate whether pre-vaccine TTV VL could predict humoral and cellular immune responses to SARS-CoV-2 mRNA vaccines in people living with HIV (PLWH) and healthy individuals (HP). Methods: TTV VL was measured via real-time PCR in serum samples collected before the second and third doses of mRNA vaccines in 93 PLWH and 48 HP (second dose) and 255 PLWH and 48 HP (third dose). Immune responses were assessed through anti-SARS-CoV-2 receptor-binding domain (RBD) IgG, neutralizing antibodies, and IFN-γ release. Statistical analyses included correlation studies between TTV VL and vaccine-induced immune responses. Results: TTV VL did not significantly correlate with anti-RBD IgG or neutralizing antibody levels in either cohort; highlighting its limited predictive value for humoral responses in relatively immunocompetent populations. However, a strong inverse correlation was observed between TTV VL and IFN-γ release after the third, but not the second, vaccine dose. These findings suggest that higher TTV VL, indicative of reduced immune competence, may impair T-cell-mediated immunity to vaccines. Conclusions: In virologically suppressed PLWH and HP, TTV VL is not a reliable predictor of humoral immune responses to COVID-19 vaccines. However, its inverse relationship with cellular responses warrants further investigation in more immunosuppressed populations. These results reinforce the continuum model of TTV VL as a biomarker, with predictive utility increasing alongside the degree of immunosuppression

Tumor infiltrating T-cells and loss of expression of SWI/SNF genes in varying stages of clear cell renal cell carcinoma.

Patients with clear cell renal cell carcinoma (ccRCC) metastases face poor prognoses, even with adjuvant therapies. Tumor-infiltrating T-cells and macrophages are critical in targeting tumor cells within the renal microenvironment. Beyond VHL mutations, loss-of-function mutations in SWI/SNF complex genes, including PBRM1, BAP1, ARID1A, SETD2, SMARCA4 (BRG1), and SMARCA2 (BRM), have been implicated in ccRCC progression. A tissue microarray of 160 ccRCC cases was analyzed via immunohistochemistry (IHC) for SWI/SNF protein expression and CD4 + /CD8 + T-cell infiltration. Clinical and pathologic features were obtained through electronic medical records. Statistical analyses included one-way ANOVA, two-way ANOVA, Pearson's chi-square and t-tests. Loss of SWI/SNF protein expression was observed in PBRM1 (31 %), ARID1A (51 %), SETD2 (14 %), BRG1 (15 %), BRM (38 %), and BAP1 (40 %). T-cell counts varied significantly with stage: CD4 + counts peaked at Stage 3, while CD8 + counts increased through Stage 4 (p < 0.001). Loss of PBRM1 was more frequent in advanced stages (29.4 %, p < 0.001), while BRM and BRG1 losses were more common in earlier stages (p < 0.001, p = 0.006). ARID1A and BRM losses correlated with reduced CD8 + counts (p = 0.016, p = 0.032) and stage-specific CD4 + variations (p < 0.001, p = 0.042). Loss of PBRM1, SMARCA2/BRM, and SMARCA4/BRG1 expression is significantly associated with ccRCC progression, with PBRM1 loss prevalent in advanced stages and SMARCA2/BRM and SMARCA4/BRG1 in earlier stages. ARID1A and SMARCA2/BRM losses correlate with reduced CD8 + counts and stage-specific CD4 + infiltration, highlighting their potential as biomarkers for disease progression and immunotherapeutic response.

Fueling metabolic disruption via FMD to boost chemotherapy in TNBC.

Triple-negative breast cancer (TNBC) is highly glycolytic and lacks effective biomarkers for therapy response. The BREAKFAST trial showed that a fasting-mimicking diet (FMD) improved pathological complete response(pCR) rates to 56.6% compared to historical chemotherapy averages (30%-40%), with minimal severe adverse events. FMD's metabolic and immune-modulating effects warrant further study with immunotherapy.

Evaluation of EpiSwitch in predicting immunotherapy response in hepatocellular carcinoma and gastrointestinal tumors.

623 Background: The EpiSwitch Checkpoint Inhibitor Response Test (CiRT) is a blood-based assay that analyzes DNA conformations in immune cells to predict responses to immune checkpoint inhibitors (ICIs) targeting PD-L1/PD-1. CiRT has demonstrated superior predictive accuracy compared to traditional biomarkers, such as tumor mutational burden (TMB) and PD-L1 immunohistochemistry (IHC), for predicting responses in urothelial cancer (ESMO 2022). However, its role in hepatocellular carcinoma (HCC) and other gastrointestinal (GI) tumors remains unclear. As immunotherapy becomes more common in these cancers, identifying reliable predictive biomarkers is essential to optimize treatment outcomes. Methods: This retrospective study evaluates CiRT-predicted immunotherapy responses and clinical outcomes in patients with HCC and GI tumors, including cholangiocarcinoma, pancreatic adenocarcinoma, and gastric cancer. All patients received one or more lines of immunotherapy. CiRT responses were categorized into high probability (HP) and low probability (LP). Treatment response was assessed per RECIST 1.1 criteria. Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), overall survival (OS), and progression-free survival (PFS) were analyzed. Results: 43 patients (24 HP, 19 LP) were included. HCC patients (n = 33) received varied immunotherapy regimens, such as combinations of bevacizumab with atezolizumab, tremelimumab with durvalumab, and nivolumab with ipilimumab. Non-HCC patients (n = 10) received combinations of immunotherapy and chemotherapy such as durvalumab with cisplatin and gemcitabine and nivolumab with oxaliplatin and capecitabine. Across all patients, CiRT revealed a sensitivity of 73.91%, a specificity of 65%, a PPV of 70.83%, and an NPV of 68.42%. In HCC, sensitivity was 70.59% and specificity was 68.75%. In non-HCC, sensitivity was 83.33% and specificity 50%. HP responders showed better treatment outcomes compared to LP responders, with higher rates of complete response (CR: 12.5% vs 0%), partial response (PR: 29.2% vs 21.1%), and stable disease (SD: 29.2% vs 10.5%), and a lower rate of progressive disease (PD: 29.2% vs 68.4%) (p = 0.0467). Overall, 70.83% of HP achieved CR, PR, or SD vs. 31.58% of LP (p = 0.0098). PFS was significantly better in HP (p = 0.044, Log-rank test; median PFS = 2.0 months for the LP group, mPFS not reached for the HP group). No statistically significant OS difference was noted due to the lack of events in the HP group. Conclusions: CiRT HP status was associated with improved response rates to immunotherapy, longer PFS, and better treatment outcomes in HCC and GI tumors. HP responders experienced notably better outcomes, suggesting that CiRT could serve as a promising predictive biomarker for immunotherapy response, addressing a critical unmet need for reliable biomarkers to treat HCC and other GI tumors.

Histological and genetic features and therapeutic responses of lung cancers explored via the global analysis of their metabolome profile.

Lung cancer is the deadliest disease globally, with more than 120,000 diagnosed cases and more than 75,000 deaths annually in Japan. Several treatment options for advanced lung cancer are available, and the discovery of biomarkers will be useful for personalized medicine. Using metabolome analysis, we aimed to identify biomarkers for diagnosis and treatment response by examining the changes in metabolites associated with lung cancer progression. Plasma samples from patients with recurrent or metastatic non-small cell lung carcinomas diagnosed at Tohoku University Hospital between 2019 and 2024 were used in this study. Metabolomic analysis was performed using the Biocrates Life Sciences MxP Quant 500 kit. Multivariate, principal component, and orthogonal partial least squares discriminant analyses were performed. The triglyceride and phosphatidylcholine concentrations were higher in the patients with early than in those with advanced lung adenocarcinomas. However, the cholesterol ester concentrations were higher for the patients with advanced lung cancer. The concentrations of hexosylceramide were higher in patients with early lung adenocarcinoma than in those with squamous cell carcinoma. Relative to epidermal growth factor receptor (EGFR)-mutation negative cases, the EGFR-mutation positive cases showed marked differences between the ceramide and triglyceride concentrations. For the best therapeutic effect of EGFR-TKI treatment, the hexosylceramide (HexCer) (d18:1/24:0), ceramide (Cer) (d18:2/22:0), and ceramide (Cer) (d18:2/24:0) concentrations were higher for the stable and progressive disease groups. The concentrations of phosphatidylcholine (PC) ae C42:2, sphingomyelin (SM) C24:1, and lysophosphatidylcholine (lysoPC) a C18:2 were higher in the partial response group treated with immune checkpoint inhibitors and chemotherapy. Metabolomic analysis may be useful for the diagnosis and treatment of lung cancer and may provide clues for new therapeutic strategies. PC ae C42:2, SM C24:1, and lysoPC a C18:2 can serve as predictive biomarkers for monitoring the therapeutic effects of the combination of immune checkpoint inhibitors and chemotherapy.

Alteration of gut microbiota in patients with advanced hepatocellular carcinoma.

641 Background: Hepatocellular carcinoma (HCC) is the most common primary liver cancer and the third leading cause of cancer-related death worldwide. Despite advancements in treatment, such as novel combinations of immunotherapy and targeted therapy or dual immunotherapy, the prognosis remains poor due to a lack of predictive biomarkers for treatment response. Gut microbiota alterations have been proposed as both potential diagnostic biomarker and predictive biomarker. However, data in this field are not well-established and are mostly derived from studies conducted in China and Western countries, which may not fully reflect the situation in Thailand. Methods: Fecal samples were collected from pre-treated advanced HCC patients who visited medical oncology outpatient clinic at Srinagarind hospital, Khon Kaen University (HCC group = 27) and analyzed using 16S rRNA sequencing of gut microbiota. Additional sequenced data from a healthy population (Control group = 31) who had no underlying disease and had normal liver ultrasonography were retrieved from a previous study conducted by Khon Kaen University (Cholangiocarcinoma Screening and Care Program, CASCAP). The datasets were compared using the Wilcoxon rank-sum test to analyze the alteration of gut microbiota between HCC group and control group. Results: The HCC group exhibited significantly lower biodiversity index (p&lt; 0.001) than control group in terms of richness, Shannon diversity index and Simpson’s index. The HCC group had significantly higher relative abundance of the phylum Proteobacteria (p&lt; 0.001), Firmicutes (p&lt; 0.001) and a lower abundance of the phylum Actinobacteria (p&lt; 0.001) compared to the control group. Additionally, the HCC group had significantly higher relative abundance of Stenotrophomonas, Granulicatella, Ruminococcus, Blautia, Phascolarctobacterium, Butyricoccus, Streptococcus, Escherichia-Shigella, Flavonifractor, Haemophilus and Lachnospira at the genus level. Further analysis of relative abundance of gut microbiota at the genus level and clinical prognostic parameters showed positive correlation between Streptococcus and 6-month survival status. Conclusions: Gut microbiota profile in patients with advanced HCC was significantly altered from healthy control group. This could be the evidence of microbial dysbiosis and may potentially be biomarker for prognosis.

Unraveling the role of interferon-stimulated neutrophils: A promising biomarker for immunotherapy efficacy and prognosis in gastric adenocarcinoma.

472 Background: Despite significant advancements in immunotherapy, the identification of effective predictive biomarkers for treatment response in gastric adenocarcinoma (GAC) remains a critical challenge. Discovering such biomarkers could enhance patient stratification and improve therapeutic outcomes. Methods: We conducted a comprehensive analysis of 18 specimens obtained from 10 GAC patients, both prior to and following immunotherapy. Utilizing single-cell RNA sequencing, we compared the cellular composition and functional phenotypes of these specimens, with particular emphasis on tumor-infiltrating immune cells and the identification of previously uncharacterized neutrophil subclusters. Results: Our analysis identified eight distinct neutrophil subclusters, including Neu01_S100A12, Neu02_VEGF, Neu03_CCL3_CCL4, Neu04_IL1β, Neu05_CXCR4, Neu06_IFN-Stimulated, Neu07_Proliferative, and Neu08_B-cell_related. Notably, the presence of IFN-Stimulated neutrophils was positively correlated with the efficacy of immunotherapy in GAC, a finding that was further validated through analyses of public datasets and in-house cohort studies employing multiplex immunohistochemistry. Additionally, IFN-Stimulated neutrophils served as prognostic indicators across multiple datasets. Investigations revealed that these cells engage in significant cellular communication with other immune components within the tumor microenvironment, including CD8+ T cells, macrophages, and B cells, thereby promoting a pro-inflammatory and anti-tumor phenotype. Importantly, pan-cancer analyses demonstrated the consistent prognostic and immunotherapy efficacy-predicting potential of IFN-Stimulated neutrophils across various cancer types. Conclusions: IFN-Stimulated neutrophils represent a promising biomarker for predicting immunotherapy response and patient prognosis in GAC. Our findings offer novel insights into the underlying mechanisms of immunotherapy and highlight potential avenues for personalized treatment strategies in the management of GAC.

Use of immune repertoire sequencing analysis to detect differences in treatment responses for advanced esophageal squamous cell carcinoma treated with camrelizumab and platinum-based chemotherapy.

488 Background: The study evaluated the efficacy and safety of camrelizumab combined with platinum-based chemotherapy (platinum plus paclitaxel [TP] or fluorouracil [FP] agents) as first-line treatment in patients with advanced esophageal squamous cell carcinoma (ESCC), and explored potential biomarkers for treatment response using immune repertoire (IR) sequencing. Methods: In this multi-center, prospective cohort study, advanced ESCC patients were treated with camrelizumab and TP or FP chemotherapy. Primary outcome was 1-year progression free survival (PFS). Secondary outcomes included 1-year overall survival (OS), objective response rate (ORR), disease control rate (DCR) and safety. Patients were categorized based on their treatment responses into the non-ORR and ORR groups. IR sequencing was exploratorily performed on the peripheral blood mononuclear cells from 15 patients in each group. Results: From June 2020 to April 2023, 88 out of 98 screened patients were enrolled, with a median follow-up of 15.9 months. The 1-year PFS was 56.8%, OS was 68.2%, ORR was 64.8%, and DCR was 91.1%. Most treatment-related adverse events were grade 1-2, though 12.5% experienced grade ≥3 toxicities. The FP regimen's efficacy was comparable to the TP regimen. The analysis of the complementarity-determining region 3 polypeptide sequences revealed significant differences in amino acid composition between the non-ORR and ORR groups in T-cell receptor beta-chain (TRB) and immunoglobulin heavy chain (IGH) repertoire. Furthermore, the ORR group exhibited a more concentrated distribution of clones within TRB. Two V genes (TRBV29-1 and TRBV4-1) and one J gene (TRBJ1-3) in the TRB region, along with six V genes (IGHV1-45, IGHV3-20, IGHV3-48, IGHV3-49, IGHV4-4, and IGHV5-51) in the IGH region, were differentially expressed between the two groups. Conclusions: Camrelizumab with platinum-based chemotherapy is effective and well-tolerated as first-line treatment in advanced ESCC. IR sequencing may provide insights into the underlying mechanisms influencing treatment response. Clinical trial information: ChiCTR2000037942. Objective response and disease response. Efficacy evaluation Total (N=88) Camrelizumab plus T（n=69） Camrelizumab plus FP (n=19) p 1-year PFS rate 50/88 (56.8%) 39/69 (56.5%) 11/19 (57.9%) 1.000 1-year OS rate 60/88 (68.2%) 47/69 (68.1%) 13/19 (68.4%) 1.000 ORR 57/88 (64.8%) 43/69 (62.3%) 11/19 (57.9%) 0.793 DCR 81/88 (91.1%) 63/69 (91.3%) 15/19 (79.0%) 0.213 ORR group, n (%) 54/88 (61.4%) 43/69 (62.3%) 11/19 (57.89%) 0.793 Non-ORR Group, n (%) 34/88 (38.6%) 26/69 (37.7%) 8/19 (42.1%) 1.000 2-year PFS rate 38/88 (43.2%) 30/69 (43.5%) 8/19 (42.1%) 1.000 2-year OS rate 42/88 (47.7%) 33/69 (47.8%) 9/19 (47.4%) 1.000 *p&lt;0.05, a statistically difference.

Circulating tumor DNA for predicting complete response to total neoadjuvant therapy in locally advanced rectal cancer: ENSEMBLE-2.

284 Background: Total neoadjuvant therapy (TNT) has dramatically shifted the paradigm in the treatment of locally advanced rectal cancer (LARC), prolonging survival with high rates of pathologic complete response (pCR) and introducing non-operative management (NOM). However, no predictive biomarkers of TNT efficacy, the regrowth of NOM, or prognosis have been developed. Circulating tumor DNA (ctDNA) is a minimally invasive biomarker used to detect molecular residual disease (MRD) and predict recurrence in colon cancer after curative resection. The effectiveness of ctDNA MRD status as a predictive biomarker for TNT was evaluated in the Phase II TNT study, ENSEMBLE-2 (jRCTs071210143), conducted in Japan. Methods: Patients with LARC undergoing TNT were enrolled in ENSEMBLE-2. Protocol treatment was defined as total mesorectal excision (TME) following long course chemoradiotherapy (LCCRT: 50.4Gy, capecitabine) plus four cycles of CAPOX. NOM was allowed if a clinical complete response (cCR) was achieved in the evaluation after TNT. ctDNA MRD was measured by Signatera (Natera, Inc.) at the following time points: baseline, after LCCRT, after TNT, post operative 4w, 12w, 24, 36 and 48w in the GALAXY trial (UMIN000039205). Results: A total of 28 patients were enrolled in the study. Treatment was discontinued at the patient's request in one case. After completing TNT, TME and NOM were performed in 21 (77.8%) and 6 (21.4%) patients, respectively. ctDNA positivity rates were 96.4. % (27/28) at baseline, 14.8% (4/27) after LCCRT, and 34.6 % (9/26) after TNT. Post-LCCRT ctDNA status was not significantly associated with cCR + near CR (nCR) vs. incomplete clinical response (iCR), pCR vs. non-pCR (p=0.065 and p=0.539, respectively). In contrast, ctDNA status after TNT was significantly associated with cCR + nCR vs. iCR (p=0.028), as well as pCR vs. non-pCR (p=0.038). Conclusions: Our study indicates that post TNT ctDNA status may be a predictive biomarker for TNT response in LARC patients. ctDNA negativity upon completion of neoadjuvant treatment may indicate a favorable response. Clinical trial information: jRCTs071210143 . Correlation with ctDNA, clinical response, treatment after TNT and pathological response. After LCCRT p value After TNT p value Clinical ResponsecCR + nCR vs. iCR (ctDNA -/+) cCR + nCR 18 / 1 0.065 cCR + nCR 15 / 4 0.028 iCR 5 / 3 iCR 2 / 5 Treatment after TNTNOM vs. TME (ctDNA -/+) TME 17 / 3 0.438 TME 11 / 9 0.063 NOM 6 / 0 NOM 6 / 0 Pathological response after TMEpCR vs. non pCR (ctDNA -/+) pCR 5 / 0 0.539 pCR 5 / 0 0.038 non pCR 12 / 3 non pCR 6 / 9 Fisher's exact test was used to statistically examine the correlation between ctDNA MRD status and the factors at each timing.

Highly accurate detection of early-stage colorectal cancer using tumor and immune extracellular vesicles biomarkers.

262 Background: Colorectal cancer (CRC) is one of the most common cancers worldwide, and early detection is critical for successful treatment and improved survival rates. However, precancerous and early-stage CRC present significant diagnostic challenges due to the small size of lesions and the very low expression of tumor-specific biomarkers in the bloodstream. Current genomics-based diagnostic methods struggle to detect these early-stage and precancerous lesions, making it imperative to develop more sensitive and specific approaches. Circulating extracellular vesicles (EVs) are emerging as a promising solution for early-stage CRC detection. Since EVs are produced by tumor cells as well as tumor microenvironment and host immune response cells, EVs offer the means to detect and monitor small early-stage tumors through both direct detection of tumor-associated biomarkers as well as tumor specific host response and tumor microenvironment biomarkers. Methods: To identify novel early-stage CRC specific biomarkers, we performed proteomics analysis on EVs purified from patient plasma using size exclusion chromatography and a proprietary buffer system that enhances EV and corona protein recovery. TrueDiscovery Data-independent acquisition (DIA) mass spectrometry (MS) analysis was conducted on 24 pre-cancer/stage 0 and 25 stage 1 CRC patients and 75 normal patient plasma samples. An in-house developed machine learning pipeline was used to identify differentially expressed proteins and model candidate multiplexes to identify those with extremely high diagnostic accuracy (&gt; 0.99). Results: An average of ~2,500 proteins were identified per sample and included in bioinformatics analysis. Comparative analysis between control patient EVs and either pre-cancerous/Stage 0, or Stage 1 colon cancer EVs using an in-house Machine Learning pipeline identified 336 and 493 differentially expressed proteins for each group (FDR adjusted p-value &lt; 0.001). Of these, the best 17 pre/stage 0 proteins and 53 stage 1 proteins were trained and tested using Support Vector Machine to assess all potential 3-plexes in order to identify those with mean diagnostic accuracy &gt; 99%. This yielded 5 3-plexes in precancer/stage 0 and 34 3-plexes in stage 1 with near perfect diagnostic accuracy. These plexes are currently being assessed in single analyte and multiplex immunoassays with the goal of translating candidate 3-plexes to the clinical. Conclusions: Key biomarkers from CRC patient EVs indicate the potential to detect and diagnose early-stage and low tumor burden cancers using our methods. Interestingly, the most accurate 3-plexes consist of proteins from immune, inflammatory and metabolic processes, suggesting that for the detection of early stage cancer it may be critical to include both tumor and host specific biomarkers.

A prospective multi-site translational study investigating the association of gut microbiome (GM) diversity with pathological complete response (pCR) after neoadjuvant treatment in early stage rectal and esophageal cancer.

TPS849 Background: The diversity of the GM is defined as the number and relative abundance distribution of distinct types of microorganisms colonizing within the gut. Studies have suggested that dysbiosis of the GM confers a predisposition to certain malignancies and impacts response to immunomodulating therapies. The influence of the GM diversity on the pathological response after neoadjuvant chemotherapy and radiotherapy is unclear. Some studies have suggested that the GM may offer predictive biomarkers for response to chemoradiation in rectal cancer. Other studies in early-stage rectal cancer patients indicated an association between GM diversity and pathological outcomes following neo-adjuvant therapy (NAT). We hypothesize that a more diverse GM constitution at baseline leads to an improved pathological response at the time of definitive surgery. Methods: We designed a cross-institutional translational study investigating the impact of the GM diversity on the efficacy of NAT in GI cancers by assessing its association with pathological response. The study population includes patients with early-stage rectal or esophageal cancer due to commence NAT (including chemotherapy and chemoradiation) who are planned for definitive surgery. Exclusion criteria includes prior allogenic tissue/solid organ transplantation and prior receipt of anti-cancer therapy. The study assessments include fecal sampling of the GM prior to NAT, upon completion and again six months post completion of therapy. Fecal samples are analysed by 16S RNA sequencing. Pathological response will be examined at time of surgery and patients will be classified as responders (complete pathological response) or non-responders. The primary endpoint of the study is to examine the association between the GM diversity and pathological response. Exploratory analysis will include the assessment of the association between cf-DNA and the GM diversity as well as an assessment of the association between cf-DNA at baseline and pCR. Species richness (Alpha Diversity) will be analysed using the Shannon diversity index and Jaccard similarity index will be used to calculate beta diversity. Following planned study recruitment, classification and clustering analysis will be performed with Principal Component Analysis (PCA) and Random Forest analysis. Logistic regression analysis adjusting for potential confounding factors will be employed to assess the primary endpoint of the association between GM and complete pCR in the final statistical analysis. Adjusted odds ratios (OR) and 95% confidence intervals will be presented. This trial accrued 11 patients between May 2023 and Sept 2024. Out of the 11 patients enrolled, 9 patients have undergone their planned surgery. We are expecting to have 30 patients accrued prior to Jan 2025.

BRCA1 and BRCA2 mutations testing in prostate cancer: Detection in formalin fixed paraffin embedded (FFPE) and blood samples.

Prostate cancer (PC) represents one of the leading causes of cancer-related morbidity and mortality in men, requiring further understanding to improve diagnosis and treatment. Germline BRCA1/2 mutations, primarily identified in other hereditary cancers, confer an increased risk of developing PC; thus, testing is essential to assess cancer risk, guiding preventive strategies and screening. Recently, somatic BRCA1/2 mutations have emerged as pivotal predictive biomarkers of responsiveness to the poly ADP-ribose polymerase (PARP) inhibitors. This review provides a comprehensive overview of BRCA1/2 mutations testing in PC, focusing on the germline and somatic mutations frequencies and the technical approach for their identification. A revision of the main data reported in the literature regarding BRCA1/2 mutations identification will be presented, highlighting the critical issue for the detection both in formalin fixed paraffin embedded (FFPE) and blood samples.

A phase II double window of opportunity trial evaluating the oral TGF-beta receptor I inhibitor vactosertib in patients undergoing standard of care preoperative therapy for locally advanced esophageal adenocarcinoma.

TPS516 Background: Esophageal adenocarcinoma (EAC) is the seventh most common cancer globally and ranks sixth in overall mortality. Despite improvements in pre-operative therapy, EAC has a high recurrence rate, especially among patients whose tumors do not achieve pathologic complete response (pCR), thus there is a critical need to enhance pCR rate and improve long-term outcomes. We recently discovered that a subset of EACs are driven by tumor-intrinsic oncogenic TGFβ signaling and have identified two potential biomarkers to predict response to anti-TGFβ therapy. One of these biomarkers is enriched in poorly differentiated EAC. We hypothesize that TGFβ signaling blockade will exert a tumor-inhibitory effect in patients with EAC and propose a double window-of-opportunity trial to test if TGFβ receptor I inhibitor vactosertib can improve pCR and induce tumor metabolic response in patients undergoing neoadjuvant therapy for localized EAC. Methods: This phase II double window of opportunity study assesses the efficacy and pharmacodynamics of the oral TGFβ Receptor I inhibitor vactosertib in patients with locally advanced esophageal adenocarcinoma (EAC). Patients will be treated with single-agent vactosertib in two windows of opportunity: the first will be prior to initiation of standard of care neoadjuvant therapy and the second after completion of neoadjuvant therapy prior to surgery. Patients will be eligible if they have poorly differentiated EAC and are appropriate for neoadjuvant therapy followed by resection as per standard of care. The primary objectives are improvement in pCR compared to historical controls and metabolic response (decrease in fluorodeoxyglucose uptake by PET CT) in the primary tumor after the first window. Secondary objectives will evaluate biomarkers of response identified in preclinical studies. Pre- and post-first window of opportunity treatment biopsies will be obtained for correlative and exploratory analyses. Clinical trial information: NCT06044311 .

The impact of bisphenol A on gill health: A focus on mitochondrial dysfunction induced disorders of energy metabolism and apoptosis in Meretrix petechialis.

Bisphenol A (BPA), a well-known chemical compound used in various daily goods, has been associated with adverse effects on animal metabolic processes. However, the specific impacts of BPA exposure on clam gills remain largely unexplored. To investigate the effects of BPA on energy metabolism and apoptosis in Meretrix petechialis gills, clams were exposed to varying concentrations of BPA (1, 10, and 100 μg/L) for 21 days. Results showed that BPA exposure induced gill histopathological injuries and inhibited filtration rates. Transmission electron microscopy (TEM) analysis revealed mitochondrial injury and dysfunction as potential mechanisms of gill damage. Transcriptome analysis identified differentially expressed genes (DEGs) primarily enriched in energy metabolism and apoptosis pathways. BPA-induced changes in ATP content, ATPase, and lactate dehydrogenase (LDH) activities suggested dysregulation of energy metabolism. TUNEL staining demonstrated enhanced apoptotic signals with increasing BPA concentrations. Activation of the caspase-3/9 pathway indicated a concentration-dependent, mitochondria-dependent apoptotic process. Additionally, the expression of genes associated with mitochondria (NNT, TOMM40, and SLC25A11), energy metabolism (PCK1 and pdhC), inducing mitochondria-dependent apoptosis (NFKB1, RAC1, and TRAF2), and oxidative stress (GSTT1) was affected by BPA exposure. Integrated biomarker response version 2 (IBRv2) values further confirmed a concentration-dependent gill toxicity of BPA via the mitochondrial pathway. These findings provide a deeper understanding of the toxicological mechanisms underlying BPA-induced toxicity in bivalves and contribute to assessing the risks posed by BPA in benthic ecosystems.

The diving beetle, Cybister lateralimarginalis (De Geer, 1774), as a bioindicator for subcellular changes affected by heavy metal(loid) pollution in freshwater ecosystems.

The present study assessed Cybister lateralimarginalis as a model species for ecotoxicological studies of metal(loid) bioaccumulation in freshwater ecosystems, by optimising protocols and analysing biomarker activities and metal(loid) concentrations from different sites and investigating their association. To accomplish this, levels of arsenic (As), selenium (Se), cadmium (Cd), mercury (Hg), and lead (Pb) were analysed, as well as a set of biomarkers were evaluated, esterase activity and oxidative stress biomarkers: fluorescence- and glutathione-based. C. lateralimarginalis were sampled in the spring/summer of 2023 along the Sava, Drava and Danube floodplains in Croatia. C. lateralimarginalis samples from Sava floodplain (Mužilovčica) showed altered carboxylesterase (CES) activity, lower glutathione (GSH) levels, and elevated Hg and As concentrations. The Weighted Pollution Index (WPI) indicated moderate pollution at the Sava floodplain, possibly due to agricultural or petrochemical impacts. At the Drava floodplain (Podravlje), higher glutathione S-transferase (GST) activity and elevated Cd, Hg, and As concentrations were observed, likely from soil accumulation, while Stara Drava beetles had lower GSH levels, suggesting agricultural impacts. Despite this, the Drava floodplain showed the lowest WPI. Higher GST activity was noted at the Danube floodplain (Kopački rit and Topoljski Dunavac), with reactive oxygen species (ROS) concentrations (Kopački rit and Podunavlje) possibly reflecting agricultural practices or illegal hunting. The present research successfully applied C. lateralimarginalis as a bioindicator species for assessing metal(loid)s in different environments, highlighting its reliability as an abundant apex predator, emphasizing the significance of integrating biomarker response evaluation into pollutant monitoring for efficient environmental risk assessment in freshwater ecosystems.