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622 Background: Colorectal small and large cell neuroendocrine carcinoma (SCNEC and LCNEC) comprise <1% of colorectal cancers and have poor prognosis. Detailed survival information is limited due to rarity. We analyzed survival outcomes using Survival, Epidemiology, and End Result (SEER) database (2000-2022) and compared with NETs and adenocarcinoma (AC). Methods: All patients with SCNEC, LCNEC, NETs or AC with colorectal primaries were extracted. AJCC stage was calculated with 8 th ed. TNM classification. Kaplan-Meier estimator, log-rank test, and Cox regression (including AJCC stage, surgery, radiation, and chemotherapy variables) were used to analyze survival outcomes. Results: Comparing colon and rectal primaries, the only significant mOS difference was in stage IV SCNEC (2 vs 7 months, p<0.0001). Comparing LCNEC and SCNEC, the only significant mOS difference was in stage IV colon SCNEC and LCNEC (2 vs 5 months, p<0.0001). Stage III and IV colorectal SCNEC and LCNEC had significantly shorter mOS than stage III and IV AC and NETs. In multivariate Cox regressions, surgery and chemotherapy were associated with significantly improved survival for both SCNEC and LCNEC at both primaries (p<0.001), while radiation was never significant. Conclusions: Most colorectal SCNEC and LCNEC patients present with stage IV disease. Stage IV colon SCNEC had significantly shorter mOS than both stage IV colon LCNEC and stage IV rectal SCNEC. Further research should explore early diagnosis and effective therapies for this rare, aggressive malignancy. Median overall survival (months) Colon SCNEC (n/mOS) Colon LCNEC (n/mOS) Colon NETs (n/mOS) Colon AC (n/mOS) SCNEC vs NETs in colon: P-value LCNEC vs NETs in colon: P-value SCNEC vs AC in colon: P-value LCNEC vs AC in colon: P-value SCNEC vs LCNEC in colon: P-value Rectal SCNEC (n/mOS) Rectal LCNEC (n/mOS) Rectal NETs (n/mOS) Rectal AC (n/mOS) SCNEC vs NETs in rectum: P-value LCNEC vs NETs in rectum: P-value SCNEC vs AC in rectum P-value LCNEC vs AC in rectum SCNEC vs LCNEC in rectum: P-value Colon SCNEC vs rectal SCNEC P-value Colon LCNEC vs rectal LCNEC: P-value Stage I 19/32 17/122 4496/NA 114,213/144 <0.0001 <0.0001 <0.0001 0.34 0.25 27/23 10/28 5185/NA 54,702/160 <0.0001 <0.0001 <0.0001 0.00047 0.88 0.99 0.24 Stage II 17/38 30/49 259/NA 121,757/106 0.0015 <0.0001 0.4 0.0068 0.65 17/16 3/NA 33/NA 38,417/106 <0.0001 0.57 <0.0001 0.87 0.3 0.065 0.55 Stage III 40/14 107/13 582/184 104,793/82 <0.0001 <0.0001 <0.0001 <0.0001 0.48 50/19 37/17 103/102 46,687/107 <0.0001 <0.0001 <0.0001 <0.0001 0.45 0.48 0.47 Stage IV 245/2 217/5 393/49 83,542/13 <0.0001 <0.0001 <0.0001 <0.0001 <0.0001 260/7 57/6 158/36 36,652/17 <0.0001 <0.0001 <0.0001 <0.0001 0.25 <0.0001 0.95

561 Background: For patients with aHCC, second-line treatment options are not well established. The primary aim of this study was to evaluate the efficacy of Futi, an FGFR 1-4 inhibitor, in combination with Pem as a second line therapy in patients with FGF19-expressing aHCC. Methods: Patients aged ≥ 18 years, with histological/cytological diagnosis or radiological diagnosis of HCC, Child-Pugh (CP) class A-B7, Barcelona Clinic Liver Cancer (BCLC) A, B or C, ECOG performance status of 0-2, and FGF19+, by in-situ hybridization or immunohistochemistry, received Futi 20 mg once daily plus i.v. Pem 200mg every 3 weeks. The primary endpoint was progression-free survival at 6 months (PFS6). Secondary endpoints include overall survival (OS). The study was designed to enroll 22 evaluable patients to detect PFS6 from 25% (historical control) to 47%, assuming a 1-sided alpha of 0.1 with 80% power. All patients provided written consent. Eligible patients with consent who started treatment are considered evaluable. Results: The study was closed to enrollment early at the request of funding agency due to poor accrual. Between 8/2021 and 3/2024, 14 patients enrolled with 1 patient withdrawing consent before beginning protocol treatment. Among 13 evaluable patients, median age was 72 (range: 60-87) years, 7 (53.8%) were male, 12 CP A (92.3%), 7 BCLC C (53.8%), with 7 (53.8%) having received prior liver directed therapy. Median PFS was 15 weeks (95% confidence interval [CI]: 9.0-Not Estimable) and PFS6 was 23.1% (95% CI: 8.6%, 62.3%). Objective response rate was 0% (95% CI: 0%, 24.71%) and stable disease occurred in 7 (53.9%) of patients. Median OS was 55.7 weeks (95% CI: 47.3-NE). Nine (69.2%) patients experienced at least 1 grade 3+ adverse event (AE), and 2 (15.4%) patients experienced at least a grade 4+ AE. One patient had a grade 5 AE unrelated to study treatment (acute kidney injury). Most commonly reported AEs (occurring in more than 31% of patients) were hyperphosphatemia (76.9%), fatigue (69.3%), diarrhea (61.6%), AST (53.9%) and ALT (46.2%), dry eye (38.5%), nail changes (38.5%), rash-maculo-papular (38.5%), and mucositis oral (38.5%). Conclusions: This prospective study evaluated the use of a targeted treatment in combination with immunotherapy following first line immune-based therapy. This combination had modest activity as a second line therapy for patients with aHCC but did not meet the primary endpoint. The safety profile was acceptable and consistent with prior reports. An optimal second-line regimen remains a critical need. Clinical trial information: NCT04828486 .

451 Background: In first-line E/GEJ/G adenocarcinoma, anti-PD-1 + chemotherapy improves outcomes, yet many patients do not respond. Beyond PD-L1 CPS, tumor-intrinsic biomarkers of primary resistance remain limited. We applied a validated IFN-γ–linked T cell-inflamed (Tinfl) framework to define exclusionary programs and nominate targets. Methods: Bulk RNAseq was analyzed from TCGA ESCA/STAD adenocarcinoma (n=79, discovery) and a de novo Mayo cohort of pretreatment samples (n=74, validation) with harmonized survival and response data. Tinfl scores (160-gene signature) defined quartiles (Q1 cold, Q4 hot). Cold versus hot tumors were compared by edgeR + limma-voom to identify differentially expressed genes (DEGs) (Bioconductor v3.20). Pathway and upstream-regulator/causal-network analyses used Ingenuity Pathway Analysis seeded by top DEGs (|log2FC|>1.5, p<0.05). Survival was assessed by Cox proportional hazards models (two-sided α=0.05) and KM log-rank testing (α=0.05). A 14-gene Tinfl-weighted cold signature produced a per-sample composite score. Results: In the Mayo cohort, median age was 62 years; 72% were stage IV at diagnosis; 24% were HER2-positive and 3% MSI-H; primary sites were GEJ/esophagus/gastric 50%/34%/16%; PD-L1 CPS <1/1–5/6–49/≥50 occurred in 18%/46%/27%/4%; patients received a mean of 2.4 lines of therapy. Across discovery and validation cohorts, Tinfl-cold tumors showed significantly higher expression of developmental and lineage transcription factors (HOXC12, HOXC13, ZIC1, DLX3, EVX1), an oncofetal post-transcriptional hub (IGF2BP1), cancer-testis and piRNA components (MAGEA4, TDRD12, THEG), and neuronal or transport genes (GABRQ, KCNF1, LDLRAD1, SLC6A10P) (logFC >2, p<0.05). Pathway analysis highlighted chromatin and lineage regulators (KAT6A, SIRT1, LEF1), TGF-β signaling (TGFBR2), and repressors of innate DNA/IFN sensing (TREX1, IRF2BP2) (Z-score >2, p<0.05). The 14-gene Tinfl-weighted cold signature was higher in cold versus hot tumors (p<0.05) and showed a directional association with inferior overall survival in the Mayo validation cohort. Among patients who received first-line anti-PD-1–containing regimens, several signature members and the composite score were higher in non-responders (p<0.05). Conclusions: Using a discovery–validation design, we define tumor-intrinsic expression pathways that characterize Tinfl-cold E/GEJ/G adenocarcinoma. We also derive a concise 14-gene Tinfl-weighted cold signature that is enriched in non-responders and associated with inferior overall survival. Targetable nodes such as IGF2BP1 and MAGEA4, together with chromatin and Wnt/TGF-β regulators, support rational combinations with PD-1 blockade. Planned validation in a third cohort, single-cell confirmation of tumor-cell expression, and in vivo cytokine profiling in knockdown models will refine mechanisms and nominate clinical candidates.

103 Background: Clinicians spend significant time extracting and synthesizing information to risk-stratify and make adjuvant chemotherapy (AC) recommendations for CRC. By automating parts of this process, LLMs could help reduce documentation burden and variability in decision-making. Methods: We selected 144 stage III, pMMR CRC patients treated at Mayo Clinic (2017–2021). HIPAA-compliant Gemini-2.0-Flash-001 and GPT-4o extracted clinical variables (age, neuropathy grade, pathological stage) from EHRs and generated AC recommendations. Two approaches were compared: (1) a rule-based algorithm with nested if/else statements translating variables into NCCN-based recommendations, and (2) a dynamic LLM agent generating recommendations using a structured prompt informed by NCCN guidelines. Concordance with treating physicians was assessed, with disagreements reviewed by a blinded GI oncologist. Results: The dataset included 144 patients with median age 67 (IQR 66–68), 48.6% female. The LLM pipeline extracted neuropathy grade with 100% accuracy and pathological T/N stages with 99.3% accuracy. Concordance with physician recommendations was 93.1% for the rule-based algorithm and 62.5% for the LLM agent. In discordant cases, blinded adjudication by a GI oncologist favored the algorithm and LLM agent over physicians in most cases (Table 1). Conclusions: This exploratory work shows LLM agents can reliably extract variables and generate evidence-based treatment recommendations for stage III CRC. The high rate of expert preference for LLM recommendations in discordant cases is intriguing but limited by small sample size and possible adjudicator bias. Discordant physician recommendations reflect the complexity of decision-making, including patient preferences, performance status, comorbidities, and subtle findings not fully captured in EHRs. Rather than replacing judgment, we envision LLMs as decision support tools—analogous to an advanced trainee reviewing charts and presenting evidence-based recommendations to the attending physician with ultimate authority. Larger multi-institutional studies with multiple adjudicators are needed to validate these findings and clarify AI’s role in oncology. Patient characteristics, LLM extraction accuracy, and concordance with physician recommendations. Metric Result Total patients 144 Median age (IQR) 67 (66-68) Female (%) 48.6% LLM extraction accuracy – Neuropathy grade 100% LLM extraction accuracy – Pathological T and N stages 99.3% Primary Concordance – Rule-based algorithm vs. physician 93.1% Adjudicator preference of rule-based algorithm over treating physician for discordant cases 100% Concordance – LLM agent vs. physician 62.5% Adjudicator preference of LLM agent over treating physician for discordant cases 82.0%

Abstract PAX5 is genetically altered in over 30% of B-ALL cases, with point mutations representing the second most common type of alteration. These point mutations are highly enriched in the DNA binding (paired) domain and have been recognized as founder events in PAX5 P80R and PAX5alt B-ALL subtypes, which account for over 12% of B-ALL. However, the molecular features and underlying mechanisms of these mutations in B-ALL remain largely undefined, limiting the development of targeted therapies and resulting in poor outcomes, especially in adult patients. We systematically investigated the functional roles and oncogenic mechanisms of PAX5 paired domain mutations. Integrated CRISPR screening and genomic profiling revealed that the paired domain is critical for PAX5 function. Disruptions in this region significantly reduced cell viability in B-ALL cell lines, and analysis of 2,955 B-ALL samples showed that recurrent PAX5 missense mutations are highly enriched in this domain, further underscoring its functional importance. Functional analysis of knock-in mice with PAX5 mutations revealed that R38H and R140L severely impair DNA binding and block B-cell differentiation, while P34L and P80R retain binding ability but alter binding specificity and gene regulation. Mice carrying homozygous (P34L, R38H, P80R, R140L) or heterozygous (R38H, P80R) mutations developed spontaneous B-ALL within one year. To model co-occurring mutations observed in patients, Pax5 R38H/P34L and R38H/R140L mice were generated. These compound mutations exhibited greater leukemogenic potential than single mutants. Transcriptomic analysis of leukemia samples showed consistent up-regulation of metabolic pathways and Myc target genes. Notably, biallelic PAX5 alterations and additional genetic lesions in signaling pathways were detected in all PAX5R38H/+ and PAX5P80R/+ leukemias, faithfully recapitulating the genetic alteration features of patient B-ALL driven by PAX5 mutations. Among Pax5-mutant mouse strains, Pax5P80R/+ mice develop B-ALL with short latency and complete penetrance, making them an ideal model to study the mutational evolution and leukemogenesis of B-ALL. To characterize the stepwise leukemogenesis process, we performed serial bone marrow aspirations in the same Pax5P80R/+ mice. In all 17 mice analyzed, we identified early preleukemic clones with only Pax5 WT deletions, followed by the emergence of leukemic clones harboring Jak1/3 mutations. Single-cell RNA/BCR-seq revealed that Pax5 WT allele deletions relieve metabolic constraints, thereby promoting clonal expansion of preleukemic B cells blocked at the pre-B stage. These changes coincide with downregulation of metabolic checkpoint genes and upregulation of glycolytic enzymes, promoting genetic instability and facilitating malignant transformation. Clinically, PAX5 P80R patients have better outcomes than PAX5alt cases. Using our mouse models, we observed that leukemic cells driven by the PAX5 P80R mutation exhibit greater sensitivity to dexamethasone compared to the ones carrying other PAX5 mutations in PAX5alt subtype, potentially due to their heightened dependence on glucose uptake and glycolysis, suggesting that metabolic rewiring driven by PAX5 mutants may influence therapeutic response. Interestingly, we found that an aberrant isoform of MEGF10 (aMEGF10) is highly and specifically expressed in PAX5 P80R B-ALL. This N-terminal truncated isoform encodes a protein that retains the intracellular immunoreceptor tyrosine-based activation motifs (ITAMs). Using CUT&Tag, we showed that PAX5 P80R directly binds to the aMEGF10 promoter. By engineering human CD34⁺ cells, we established a humanized PAX5 P80R leukemia model that recapitulates the gene expression profile of patient samples, including robust activation of aMEGF10. Functionally, aMEGF10 binds to and activates SYK, thereby promoting leukemic cell proliferation. Importantly, this oncogenic effect depends on intact ITAM motifs and SYK signaling, revealing a novel targetable vulnerability in PAX5 P80R leukemias. Overall, the study establishes that PAX5 DNA binding domain mutations contribute to leukemogenesis through stepwise loss of PAX5 WT alleles and acquisition of signaling pathway mutations, involving impaired differentiation, metabolic deregulation, and novel oncogene activation. These findings not only deepen the understanding of PAX5-driven leukemias but also highlight aMEGF10–SYK signaling as a promising therapeutic target.

PURPOSE Immunotherapy has demonstrated limited efficacy in the treatment of microsatellite-stable (MSS) metastatic colorectal cancer (mCRC) to date. We evaluated whether an off-the-shelf, multipeptide vaccine, PolyPEPI1018, in conjunction with standard chemotherapy can induce broad antitumor immune responses, potentially improving disease control in refractory mCRC. PATIENTS AND METHODS This single-arm, open-label study enrolled adults with MSS mCRC who had progressed after ≤2 previous lines of therapy for metastatic disease. PolyPEPI1018 was administered subcutaneously to four anatomical sites on days 1 and 15 and TAS-102 orally twice a day on days 1-5 and 8-12 of a 28-day cycle. Patients were treated for seven cycles or until disease progression. The primary end point was safety. Secondary end points were overall response rate, progression-free survival (PFS), overall survival (OS), and immune and genetic correlates. RESULTS Fifteen patients were enrolled; 80% had liver metastases and 67% had KRAS mutations. Adverse events (AEs) related to PolyPEPI1018 were grade 1 and 2 local skin reactions (n = 14). Grade 3 AEs (n = 15) at least possibly related to treatment were fatigue, decreased lymphocyte and neutrophil counts, and diarrhea. None had objective tumor response; the disease control rate was 53.3%. The median PFS (mPFS) was 4.0 (95% CI, 2.2 to 6.1) months and the median OS (mOS) was 8.7 (95% CI, 6.4 to 15.2) months. In post hoc subgroup analyses, patients without active liver metastasis had improved mPFS of 9.7 months and mOS of 14.8 months. In addition, patients who were predicted to have broader immune responses to vaccine based on human leukocyte antigen genotype had longer mOS (10.3 v 4.6 months, hazard ratio, 0.29 [95% CI, 0.07 to 1.17]). CONCLUSION The addition of PolyPEPI1018 to TAS-102 was safe and induced immunological responses that were associated with improved clinical outcomes. Further investigation of this chemo-immunotherapeutic is warranted.

426 Background: Testing for DPYD gene variations prior to administration of fluoropyrimidines ([FP] e.g. 5-FU or capecitabine) can determine if patients are at risk of significant toxicity and may benefit from dose reduction or elimination of chemotherapy. Barriers to testing include lack of recommendations in US guidelines for routine DPYD testing, absence of insurance coverage, and limitations in provider support. We designed an electronic health record (EHR)-based intervention to support shared decision-making (SDM) between providers and patients regarding DPYD testing. Methods: We built an EHR alert identifying all patients initiating FP without DPYD testing. The alert fired to remind providers upon signing FP. Clinical decision support (CDS) was included in the alert to help with DPYD testing selection along with an educational note template containing the incidence of DPYD deficiency, rationale for testing, test costs, and turnaround time. Interactions with the EHR were recorded to determine why providers declined ordering pharmacogenomic testing. Testing rates and results for patients initiating FP were collected from the EHR. Finally, providers were surveyed to determine the impact of the alert on conversations with patients as well as the utility of the embedded CDS and educational note template. Results: Within the first 8 months after alert activation, 34 patients completed DPYD testing of 444 starting FP (testing rate of 7.7%) vs the baseline testing rate in the 3 months prior to alert go-live of 6.8% (p = 0.86 via chi-squared analysis, two-tailed). Of the 34 patients, 5 were intermediate DPYD metabolizers. Reasons for alert dismissal per provider selection were: “Patient already tolerated prior fluoropyrimidines” (15.7%), “Patient declines” (12.9%), “Will order separately” (5.7%), “Not discussed as not appropriate” (5.3%), and “Not discussed, deferred” (60.4%). Thirty-nine providers (n = 77 eligible) responded to the survey; 22 prescribed FP in their practice and had encountered the alert. 21 (95.5%) of these reported the alert was “Very Useful” or “Somewhat Useful” and 18 (81.8%) used the DPYD testing options available in the alert. Sixteen (72.7%) discussed DPYD testing more frequently with patients as a result of the alert. Barriers to DPYD testing were “Concerned about the impact of lowering FP dosing” and “Lack of knowledge about DPYD testing”, while “Cost of DPYD testing” was scored lowest. Conclusions: Utilizing an EHR alert with embedded CDS had a positive impact on provider awareness and led to more discussions regarding DPYD pharmacogenomic testing with patients in line with current FDA recommendations. Limitations in provider knowledge regarding DPYD testing and concerns about lowering chemotherapy doses posed barriers to ordering testing. Further interventions including additional education are required.

Abstract Background Unresectable hepatocellular carcinoma (uHCC) is a leading cause of cancer death. FDA-approved first-line systemic therapies include atezolizumab/bevacizumab (atezo/bev) and durvalumab/tremelimumab (durva/treme); however, there is a lack of comparative data. Methods We reviewed outcomes of patients with uHCC who initiated atezo/bev or durva/treme between 2017 and 2024, across six institutions. Overall survival (OS) and time to treatment discontinuation (TTD) were analyzed using the Kaplan–Meier and Cox models, adjusting for baseline characteristics. Results Four hundred fifty-two uHCC pts were included. Median age: 68 years; 77% male; 81% white. Most common etiologies were viral hepatitis (38.9%) and metabolic dysfunction-associated steatohepatitis (19.5%). Disease progression was the primary reason for treatment discontinuation, atezo/bev (56%) and durva/treme (42%). Outcomes were not statistically significant (median OS [month, m]: 14.0 vs 14.6 [P = .66]; median TTD [m]: 4.9 vs 3.9 [P = .42] for atezo/bev vs durva/treme). Outcomes were significantly different between Child-Pugh classes (CP: A, B7, B8/9, C) respectively, median OS(m): 19.0, 6.1, 5.1, 2.0 (P &lt; .001); median TTD(m): 6.1, 2.3, 3.0, 1.3 (P &lt; .001). Conclusions In this real-world study of uHCC, no significant difference in clinical outcomes was observed between atezo/bev and durva/treme in the first-line setting. CP scores were a key prognostic variable with both regimens.

This cross-sectional study analyzes patient-reported outcome findings from the Enhanced, Electronic Health Record–Facilitated Cancer Symptom Control (E2C2) trial to investigate level of symptom burden in adolescents and young adults with cancer.

PURPOSE Colorectal cancer (CRC) incidence and mortality have decreased since the 1970s, but the incidence in young adults (<50 years, named early-onset CRC [eoCRC]) has been increasing. PATIENTS AND METHODS Individual patient data on 13,365 patients with metastatic CRC enrolled between 2000 and 2012 in 17 first-line randomized trials in the Aide et Recherche en Cancérologie Digestive database were pooled. The distribution of demographics, clinicopathologic features, biomarkers, and outcome data were summarized and compared by age groups. Progression-free survival (PFS) and overall survival (OS) were assessed by Kaplan-Meier curves and Cox models stratified by treatment arms within studies, adjusting for potential confounders. Predictive value of age group on clinical outcomes was evaluated by testing interaction effect between treatment and age variables. RESULTS Overall, 2,045 patients with eoCRC (median age, 42.5) and 11,320 patients with average-onset CRC (aoCRC; median age, 63.8) were included in this analysis. Within the eoCRC population, treatment with bevacizumab in addition to chemotherapy improved PFS (9.9 v 6.8 months; hazard ratio [HR], 0.66 [95% CI, 0.54 to 0.80]; P < .0001), which was similar to the findings in aoCRC population (9.4 v 7.3 months; HR, 0.73 [95% CI, 0.67 to 0.80]; P < .001; interaction P = .5415). However, epithelial growth factor receptor inhibitor (EGFRi) did not improve PFS in RAS wild-type (WT) patients with eoCRC who had left-sided primary tumors (8.3 v 8.9 months; HR, 1.20 [95% CI, 0.81 to 1.77]; P = .36), whereas EGFRi significantly improved PFS in the aoCRC population (9.9 v 8.5 months; HR, 0.74 [95% CI, 0.64 to 0.86]; P < .0001; interaction P = .083). CONCLUSION Treatment-naïve patients with metastatic eoCRC appear to derive similar benefit from bevacizumab as patients with aoCRC. However, patients with eoCRC with left-sided RAS/RAF WT tumors did not appear to derive benefit from first-line EGFRi.