Orme, Jacob J.

Background: Longitudinal genomic profiling through serial circulating tumor DNA (ctDNA) testing offers a noninvasive method to monitor clonal evolution in advanced prostate cancer. This study evaluated the frequency and nature of newly emergent and potentially actionable genomic alterations detected through serial testing in a real-world setting. Methods: We conducted a retrospective analysis of advanced prostate cancer patients who underwent multiple Guardant360 ctDNA tests between October 2020 and March 2023. The study focused on identifying new genomic alterations absent in the initial test, with particular attention to alterations relevant for on-label therapies, therapies approved in other oncologic indications (i.e., off-label), or a clinical trial. Results: Among 479 patients with at least two ctDNA tests, the median interval between the first and second evaluable tests was 207 days. New and potentially actionable alterations emerged in 57.8% of patients, including potential targets for on-label therapies (16.7%), off-label therapies (16.5%), and clinical trials (55.7%). Tumor mutational burden (TMB) increased from “low” to “high” in 11% of patients, although none had microsatellite instability or mismatch repair deficiency. In a Mayo Clinic subset, ten patients received olaparib based on treatment-emergent alterations, but none achieved a prostate-specific antigen (PSA) response. Two patients who transitioned from low to high TMB received pembrolizumab, both with progressive disease as best response. Conclusions: In a large real-world cohort, serial ctDNA testing frequently identified new alterations that were not detected at baseline and are potentially actionable therapeutic targets, highlighting the value of serial genomic profiling for capturing clonal dynamics. Additional research is needed to better establish a framework for retesting and to clarify how these results should influence subsequent treatment decisions.

Abstract The roles of nucleoplasm-residing nucleoporins (NUP) in solid tumors, including prostate cancer, remain unknown. In this study, we reveal the clinical significance and mechanistic role of the off-pore NUP, soluble POM121 (sPOM121), as a crucial transcriptional regulator that enhances the aggressiveness of metastatic prostate cancer. Using orthogonal methodologies in human samples, sPOM121 was identified as the predominantly expressed nucleoplasmic NUP in prostate cancer. Unbiased proteomic and epigenomic studies demonstrate that sPOM121, through its C-terminus, interacts with the chromatin remodeler SMARCA5 at gene promoter sites and localizes at nuclear condensates, reprogramming gene expression. Indeed, sPOM121 regulates a distinct oncogenic gene network, including β-catenin, leading to prostate cancer progression and immune evasion. Importantly, targeting the sPOM121/β-catenin axis in patient-derived preclinical and syngeneic mouse models halts prostate cancer aggressiveness and enhances antitumor immunity. Taken together, these findings reveal previously unknown actionable reprogramming functions of off-pore NUPs in solid tumors. Significance: This study uncovers how oncogenic signaling programs are transcriptionally heightened by the NUP sPOM121 in metastatic prostate cancer. Localization of sPOM121 at active transcriptional nuclear condensates propels disease progression and immune evasion, offering novel anticancer therapeutic opportunities.

Abstract Addressing the limited responsiveness to immune checkpoint blockade (ICB) in human cancers remains a critical challenge. Efforts over the past two decades have primarily focused on enhancing CD8 T cell function and preventing their exhaustion. However, less attention has been given to understanding the molecular mechanisms that contribute to the immunosuppressive tumor microenvironment (TME), which impedes CD8 T cell infiltration and fosters ICB resistance. The TME in ICB-resistant tumors is characterized by Transforming Growth Factor beta (TGF-β) cytokine signaling, leading to a fibrotic extracellular matrix (ECM) and the accumulation of immunosuppressive fibroblasts and myeloid cells. These features hinder CD8 T cell penetration, thereby promoting immune escape. The immune checkpoint molecule B7-H3, overexpressed in ICB-resistant cancers, has been linked to impaired CD8 T cell infiltration and poor prognosis. However, the molecular mechanisms governing B7-H3 function remain poorly understood. We investigated the relationship between B7-H3 gene expression and ICB response using transcriptomic data from two randomized phase 3 trials (IMvigor010 and IMvigor210). Bulk RNAseq data, tissue staining, and spectral flow cytometry were used to characterize the TME composition of B7-H3 high and low tumors. Multi-omic profiling of tumor cell lines with manipulated B7-H3 expression was performed to identify downstream targets of B7-H3. Syngeneic mouse tumor models were used to assess the impact of B7-H3 on tumor growth and antitumor immunity. High B7-H3 expression was associated with shorter overall survival in both IMvigor010 (HR=0.48, 95% CI: 0.33-0.70, p<0.0001) and IMvigor210 trials (HR=0.68, 95% CI: 0.51-0.92, p=0.01). In ICB-refractory cancers, B7-H3 expression correlated with high levels of fibroblasts, collagen deposits, and M2 macrophages. Gene set enrichment analysis revealed a positive association between B7-H3 and TGF-β signaling. Genetic deletion of B7-H3 in tumor cells reduced the surface expression of MMP14, a metalloprotease that activates TGF-β. Pan-cancer analysis identified MMP14 as the gene most correlated with B7-H3 (Pearson r=0.69, p<0.0001). TGF-β activation was reduced by 70% in B7-H3 knockout cells compared to wild-type. Loss of B7-H3 led to MMP14 accumulation in lysosomes, resulting in its degradation. We generated a B7-H3 inhibitor, 5D10, which promotes MMP14 degradation and inhibits TGF-β activation. In humanized B7-H3 mouse models, 5D10 significantly reduced TGF-β activation, enhancing CD8 T cell infiltration and impairing tumor growth. Our research identifies B7-H3 as a driver of immune exclusion and ICB resistance. We describe a novel, receptor-independent mechanism where B7-H3 promotes TGF-β activation via stabilization of MMP14. Targeting B7-H3 represents a strategy to modulate the TME, transforming immune-excluded tumors into immune-inflamed states that may respond better to ICB. Citation Format: Fabrice Lucien, Roxane Lavoie, Jack Korleski, Yohan Kim, Bharath Wootla, Liguo Wang, Ava Farrell, Kelly Harper, Martine Charbonneau, Claire Dubois, Igor Frank, Sounak Gupta, John Cheville, Jacob Orme, Stephen Boorjian, Parash Shah, Eugene Kwon, Sean Park, Haidong Dong. The immune checkpoint molecule B7-H3 is a driver of tumor immune exclusion by mediating TGF-β activation [abstract]. In: Proceedings of the AACR IO Conference: Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2025 Feb 23-26; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2025;13(2 Suppl):Abstract nr B034.

261 Background: Clinicians use next-generation sequencing (NGS) to tailor targeted treatments for PCa patients. However, the results are often available in unstructured formats and are saturated with technical jargon, making them inaccessible for patients. Therefore, we aimed to assess the efficacy of LLMs to accurately extract genomic characteristics from unstructured reports and generate patient-friendly summaries for ease of variant interpretation in PCa patients. Methods: This retrospective study included patients with PCa who underwent NGS from 2023-2024. The GPT4 model was utilized using structured zero-shot prompts to extract genomic characteristics from unstructured reports in a non-decomposed manner. Prompt development was conducted iteratively using a 2% random sample of the dataset. Genomic characteristics ascertained by LLM were assessed against held out vendor-curated test dataset for evaluation. Performance was assessed using weighted evaluation metrics (precision, recall, and F1-score). Additionally, the extracted variables were organized using rule-based criterion and presented as a separate prompt to GPT4 to generate summaries for interpretation at the level of each biologically relevant variant. Mean Flesch-Kincaid scores and Vocd-D statistic with standard deviations were computed to assess readability and lexical diversity of LLM-generated variant interpretation summaries. Results: A total of 331 patients (370 NGS reports) were included in the study. Evaluation on the held-out test dataset (Table) showed that the weighted average precision ranged from 80% for extracting tumor mutational burden to 100% for extracting the CtDNA fraction from unstructured reports. The weighted average recall ranged from 85% for extracting CtDNA tumor fraction to 99% for ascertaining variant/alteration. The weighted average F1-scores varied from 0.83 for extracting tumor mutational burden to 1.0 for extracting specimen site from NGS reports. The mean Flesch-Kincaid score was 8.29 ± 1 indicating average/standard readability ease. The mean Vocd-D statistic was 7.55 ± 1 indicating focused summaries, however, with limited lexical diversity. Conclusions: This study suggests that large language models can effectively extract genomic characteristics from unstructured reports and can potentially improve genomic literacy among prostate cancer patients by providing easy-to-interpret variant summaries. However, targeted prompting may be required to increase lexical diversity for improved engagement. Performance across different genomic characteristics. Category Precision Recall F1-Score Altered gene 0.86 0.97 0.89 Variant/alteration type 0.98 0.99 0.98 Variant allele fraction 0.96 0.96 0.96 Test name/type 1 0.99 1 Specimen site 1 0.99 1 Tumor mutational burden 0.8 0.93 0.83 CtDNA tumor fraction 1 0.85 0.92 MSI status 0.99 0.97 0.98

524 Background: Antiangiogenic tyrosine kinase inhibitors (TKIs) play a key role in treating renal cell carcinoma and other solid organ malignancies, but TKIs cause bothersome side effects that frequently necessitate dose reductions or treatment holds. Olanzapine has been useful for palliating symptoms from classical chemotherapies and advanced cancer, but its effect on TKI-related symptoms is largely unknown. This retrospective study aimed to estimate the efficacy of olanzapine for TKI-induced nausea, vomiting, anorexia, weight loss, and insomnia. Methods: All patients prescribed olanzapine with lenvatinib, cabozantinib, axitinib, or tivozanib at Mayo Clinic between January 1, 2018, and June 30, 2024 were assessed for inclusion. For baseline symptom assessment, clinical care team notes documenting the presence or absence of symptoms and indication(s) for starting olanzapine were reviewed. For response assessment, clinical notes and patient portal messages from the first three months after starting olanzapine were evaluated for qualitative descriptions of change in symptom burden. Relevant data were extracted as direct quotations, which were reviewed by the research team and categorized as “improved,” “worsened,” “stable,” or “missing data.” Each symptom domain was analyzed independently when olanzapine was prescribed for multiple interrelated symptoms. Results: A total of 60 patients who received olanzapine for the treatment of TKI-related adverse effects were included. The most common documented indication for olanzapine was nausea without vomiting (n=35), followed by anorexia (n=25), nausea with vomiting (n=16), weight loss (n=16), and insomnia (n=11). In 32 cases (53%), olanzapine was prescribed for multiple, simultaneous symptoms. Response rates for each symptom domain are shown in the table. In addition, prior to olanzapine, 34 patients (57%) had documented weight loss. Following olanzapine, 50% (n=17) gained weight (median increase: 6.1 kg; range: 2.0 - 10.7 kg), 26% (n=9) stabilized their weight (±1 kg), and 24% (n=8) continued to lose weight. Clinicians utilized a TKI dose reduction within one week of olanzapine initiation in 5 cases (8.3%), and 3 of these patients could resume the higher dose of TKI after starting olanzapine. Only 4 patients (7%) discontinued olanzapine within 3 months due to perceived side effects. Conclusions: Olanzapine appears to be effective in treating TKI-induced nausea, vomiting, anorexia, insomnia, and weight loss. Prospective randomized placebo-controlled studies are needed to confirm these findings. Improvedn (%) Worsened n (%) Stablen (%) Missing Datan (%) Nausea without emesis 32 (84) 3 (8) 1 (3) 2 (5) Nausea with emesis 13 (93) 0 (0) 0 (0) 1 (7) Anorexia 26 (74) 2 (6) 4 (11) 3 (9) Insomnia 11 (85) 0 (0) 2 (15) 0 (0)

745 Background: The combination of EV-P therapy has recently been approved as first-line (1L) therapy for patients with LA/mUC based on the results of the EV-302 trial. Herein, we report the updated results from a cohort of patients with LA/mUC who received EV-P in real world clinical setting. Methods: This retrospective study included patients with LA/mUC (07/2022-08/2024) at the Mayo Clinic who completed at least one cycle of EV-P. The best overall response (BOR) was evaluated using radiographic imaging and was adjudicated as complete response (CR), partial response (PR), stable disease (SD), mixed response (MR), or progressive disease (PD). The median progression-free survival (PFS), defined as the time from treatment initiation to disease progression or death, was estimated using the Kaplan-Meier method. Patients who had not experienced disease progression by their last follow-up were censored for the PFS analysis. Results: A total of 120 patients were included; 79 (65.8%) were males, 41 (34.1%) females, 109 (91%) White, and 119 (99%) were non-Hispanic/Latino. The median age was 72 years (IQR: 65-77). Of these, only 22 (18.3%) had locally advanced disease while the remaining had mUC; 62 (51.6%) had lower tract urothelial carcinoma (LUTC), 36 (30%) had upper tract urothelial carcinoma (UTUC). The median follow-up time was 7.1 months (IQR: 4.93-9.26). In terms of BOR, 56 patients (46.6%) experienced PR, 34 (28.3%) CR, 12 (10%) PD, 7 (5.8%) MR, and 8 (6.6%) experienced SD. 3 (2.5%) patients were not evaluable. The BOR across subgroups, LA and mUC, including LUTC and UTUC, is summarized in the table. Among the responders, the median number of treatment cycles to first response was 3 (IQR: 2.25–4). Within the follow-up, 28 patients (23.3%) experienced eventual radiographic disease progression; death was observed in 25 patients (20.8%). The median PFS was 12.7 months (95% CI: 9.8-NE), and OS was 25.1 months (16.3-NE). Conclusions: In real world setting, the combination of EV-P in LA/mUC demonstrated clinical efficacy compatible with the findings from the EV-302 trial. Our results showed comparable efficacy with EV-P in both UTUC and LUTC, highlighting it's broad therapeutic potential. However, clinical data at longer follow-up is required to fully evaluate overall survival and the durability of response. mUC LA Overall LUTC UTUC Total patients 98 62 36 22 BOR CR 23 (23.4%) 13 (20.9%) 10 (28.5%) 11 (50%) PR 49 (52.6%) 31 (50%) 18 (50%) 7 (31.8%) SD 6 (6.4%) 2 (3.2%) 4 (11.4%) 2 (9.0%) PD 10 (10.7%) 9 (15.5%) 1 (2.8%) 2 (9.0%) MR 7 (7.5%) 5 (8.6%) 2 (5.7%) 0 (0%)

114 Background: Oncologists have traditionally relied upon serial assessments of PSA and periodic imaging (i.e., every 2 to 3 cycles) to monitor treatment response. Early identification of disease progression allows for faster transitions to alternative therapies that might be more effective. In this study, we evaluated the utility of post-treatment SPECT/CT for earlier detection of treatment failure in patients receiving Lutetium-177–PSMA-617 (LuPSMA). Methods: We queried an IRB-approved registry including all patients starting LuPSMA at Mayo Clinic, MN, in the interval of March 2022 to March 2023. Those receiving fewer than 2 cycles of treatment and missing SPECT/CT data were excluded. We reviewed clinical notes to document the date and reason(s) for treatment discontinuation. The assembled cohort consists of patients stopping LuPSMA for progressive disease ([PD], categorized as biochemical, radiographic, or both). Results of the prior SPECT/CT imaging reports were reviewed. At our institution SPECT/CT images are typically acquired on the day after each infusion. Metrics collected from the radiology reports included: new foci of PSMA localization since the prior PSMA PET or SPECT/CT, suspicion for new non-PSMA avid disease, and a visual determination of PSMA-avid tumor volume. Results: A total of 256 patients received an initial cycle of LuPSMA. SPECT/CT imaging data for at least 2 consecutive treatment cycles was available for 68 patients who developed PD after a median (IQR) of 3 (2-4) cycles. There were 55 patients (81%) with both biochemical and radiographic evidence of PD, 7 (10%) with biochemical only progression, and 6 (9%) with radiographic only progression. The most used imaging modality for response assessment was PSMA PET/CT (n=46, 67%), followed by conventional imaging (n=9, 13%), and choline PET/CT (n=5, 7%). The median time between most recent post-therapy SPECT/CT and determination of PD was 39 days. The prior SPECT/CT had detected new foci of PSMA avid disease in 12 (18%) cases and new non-PSMA avid disease in 5 (7%) cases. Two patients had both new PSMA-avid and non-PSMA avid lesions. The most common sites of new PSMA-avid disease on SPECT/CT were bone (n=10) and liver (n=2). A total of 20 (29%) patients had visually increased tumor volume on SPECT/CT, 9 (45%) of which had no new lesions. In total, 24 of 68 patients (35%) had early detection of treatment futility detected by SPECT/CT. At a median (IQR) follow-up time of 8 (6.4-12) mo, the median OS [95% CI] was similar for patients with PD detected on SPECT/CT (7.8 [6.6 – 9] mo) compared to those without clear evidence of PD on SPECT/CT(8.4 [6.4-10.3] mo, p=0.885). Conclusions: Post-treatment SPECT/CT imaging may complement traditional forms of response assessment and provide an earlier warning of treatment failure through identification of new PSMA-avid lesions, non-PSMA avid disease, and increased tumor volume.

112 Background: We now have more than a decade of experience using Androgen Receptor Pathway Inhibitors (ARPIs) across various disease states of prostate cancer, and the safety of their combination with Lutetium-177–PSMA-617 has been demonstrated in VISION and other large randomized clinical trials. However, real-world data on treatment patterns and outcomes involving combination therapies with 177 Lu-PSMA-617 remain limited. Methods: For this analysis, we utilized the Mayo Clinic Rochester radiopharmaceutical database, a prospectively maintained retrospective database containing all patients who received 177 Lu-PSMA-617 at our institution. We focused on patients who started treatment from March 2022 to March 2023. Patients receiving an ARPI (abiraterone acetate, enzalutamide, apalutamide, or darolutamide) during the course of 177 Lu-PSMA-617 treatment were identified, inclusive of patients who continued using an ARPI prescribed as a prior line of treatment and those who started (i.e., switched) to a new ARPI. Baseline clinicopathologic and imaging characteristics were abstracted and compared using the Mann-Whitney U and Chi-square tests. Best PSA response during treatment was reported as a percent decline from baseline. Survival was calculated from the date of the first cycle of 177 Lu-PSMA-617. PSA50 response and overall survival (OS) outcomes for the two groups (concurrent use of ARPI vs. 177 Lu-PSMA-617 alone) were compared using the Chi-square test and Kaplan-Meier method, respectively. A multivariate Cox regression analysis was performed, including established prognostic factors. Results: An ARPI was prescribed to 106 of the 256 patients (41.4%) starting 177 Lu-PSMA-617 in the interval of March 2022 to March 2023. With regards to baseline characteristics, those receiving an ARPI concurrently with 177 Lu-PSMA-617 had a lower PSA (3.4 vs 29.7 ng/mL, p < 0.001) and a lower frequency of bone (77.4% vs. 87.4%, p = 0.035) and visceral metastases (18.9 vs 34%, p = 0.008) at start of treatment. Median follow-up for the overall cohort (IQR) was 19.1 months (8.7 – 23.6). Patients receiving an ARPI plus 177 Lu-PSMA-617, as compared to 177 Lu-PSMA-617 alone, were more likely to complete all 6 planned doses of treatment (63.2% vs. 48.7%, p < 0.001), though the PSA50 response rate was similar (49.1% vs 47.3%, p = 0.786). Median OS [95% CI] for the overall cohort was 21.3 [16.7 – 25.9] months and significantly longer for those receiving ARPI with 177 Lu-PSMA-617 (NR [NE – NE]) as compared to 177 Lu-PSMA-617 alone (15.3 mo [11.6 – 19.1], p < 0.001). However, on multivariate analysis including known prognostic variables, use of ARPI was not independently associated with improved survival (HR = 1.03 [CI: 0.68 to 1.55], p = 0.891). Conclusions: Patients receiving an ARPI with 177 Lu-PSMA-617 were more likely to complete all 6 cycles of treatment, but no clear differences in survival were observed on multivariate analysis.

136 Background: The ECOG performance status (PS) is a critical measure for evaluating functional capacity and determining clinical trial eligibility in patients with advanced prostate cancer. We aim to assess the capability of large language models (LLMs) to accurately infer ECOG PS from unstructured oncology notes. Methods: This retrospective study included patients with metastatic castration-resistant prostate cancer (mCRPC) receiving Lutetium-177–PSMA-617 ( 177 Lu) at the Mayo Clinic (2022-2023). Baseline ECOG PS scores were manually curated by a trained clinician from clinical notes recorded within four weeks prior to 177 Lu initiation. Structured zero-shot prompts were then used to prompt GPT-4 to infer ECOG PS scores from unstructured oncology notes. Total dataset was randomly sampled to prompt development (20%) and held-out test set (80%). Three experimental setups were employed to assess model performance: (i) utilizing unmasked clinical notes, (ii) clinical notes with only numerical ECOG PS scores masked while retaining narrative descriptive/assessment by oncologist of ECOG score, and (iii) clinical notes with both numerical scores and narrative ECOG descriptors masked. Final performance was assessed using evaluation metrics (precision, recall, F1 score) on the held-out test set. Results: A total of 240 patients (n: 50 prompt development; n: 190 test set) were included in the analysis. The median age was 70 (IQR: 66-76); a majority of patients were White (n: 229; 95%) and non-Hispanic (n: 233; 97%). The most prevalent ECOG PS score was 0 (n: 140; 58%), followed by 1 (n: 88; 37%), and 2 (n: 12; 5%). The performance was numerically higher when using unmasked notes (weighted average precision: 0.89; recall: 0.89; F1: 0.89), compared to using notes with masked scores only (weighted average precision: 0.76; recall: 0.79; F1: 0.75) and notes with both scores and narrative masked (weighted average precision: 0.57; recall: 0.53; F1: 0.48). Metrics by each ECOG PS score is outlined (Table). Conclusions: Large language models demonstrate significant potential in precisely extracting ECOG performance status from unstructured oncology notes, offering valuable support for treatment decision-making and clinical trial enrollment. However, model performance declines as reliance on implicit reporting within clinical notes increases. Future efforts focusing on prediction of ECOG using a series of notes over time will be the next step. ECOG Unmasked oncology notes ECOG Score masked only ECOG Score + ECOG narrative marked Precision Recall F1 Score Precision Recall F1 Score Precision Recall F1 Score 0 0.89 0.92 0.91 0.75 0.93 0.83 0.59 0.74 0.66 1 0.88 0.86 0.87 0.76 0.74 0.75 0.19 0.47 0.27 2 1.00 0.82 0.90 0.78 0.33 0.47 0.78 0.16 0.26