Citations

Abstract Introduction: Immune checkpoint inhibitors (ICI) are an essential systemic therapy for advanced melanoma. However, most melanomas develop resistance to ICI. Tumor-derived soluble PD-L1 (sPD-L1) and other soluble immunosuppressive factors drive checkpoint inhibitor resistance and correlate with inferior survival. We previously showed that therapeutic plasma exchange (TPE) removes sPD-L1 from circulation. Thus, we hypothesized that TPE-mediated removal of sPD-L1 and other immunosuppressive factors could overcome immunotherapy resistance in refractory melanoma. Methods: In this clinical trial (NCT04581382), we prospectively enrolled eighteen (18) patients with widely metastatic melanoma with progression despite anti-PD-1 ICI and elevated sPD-L1 by ELISA (≥1.7ng/mL). Each patient received radiotherapy to between one and three metastatic lesions (at least two unirradiated lesions) followed by three TPE sessions on consecutive days and re-challenge with checkpoint inhibitor. The primary safety and efficacy endpoints of the study were adverse events (AEs) and sPD-L1 reduction by TPE, respectively. Secondary endpoints included RECIST-based response in unirradiated lesions and overall survival. Correlative studies included kinetics of sPD-L1 and soluble immonsuppressive factors and dynamics of peripheral immune cell phenotypes. Results: Mean age was 62 (SD 13) and seven of eighteen (39%) were female. Mean baseline sPD-L1 was 26.31 ng/mL (40.01). The treatment was well-tolerated with one (6%) patient with grade 3 and 4 adverse events from a central line infection after TPE. Levels of sPD-L1 were significantly reduced by TPE (mean 80.2% reduction, p<0.0001). Two patients (11.1%) experienced complete response (CR), one (5.6%) partial response, three (16.7%) stable disease, and 12 (66.7%) progressive disease. In one case, immunotherapy was discontinued after two years due to no detectable lesions. Changes in tumor-reactive (TTR) GZMB+/CX3CR1+/CD11ahigh and other peripheral immune cell populations predicted overall survival in this cohort. In addition to sPD-L1, other soluble mediators of ICI resistance were also reduced by TPE and predicted overall survival in this cohort. Summary/Conclusion: sPD-L1 and other soluble immunoregulatory signaling molecules are important mediators of ICI resistance. SBRT and TPE can resensitize ICI-refractory melanoma by removing these factors. Patients with persistently elevated or rapid rebound of sPD-L1 following TPE experienced poor response and overall survival. Serial monitoring of sPD-L1 may predict response to ICI and multiple courses of TPE may be necessary. Our findings may apply in other ICI-resistant cancers with elevated sPD-L1. ClinicalTrials.gov registration: NCT04581382, ReCIPE-M1 (Rescuing Cancer Immunotherapy with Plasma Exchange in Melanoma 1).

Abstract Purpose: Metastatic castration-resistant prostate cancer (mCRPC) resistant to androgen receptor signaling inhibitors (ARSI) is often lethal. Liquid biopsy biomarkers for this deadly form of disease remain under investigation, and underpinning mechanisms remain ill-understood. Experimental Design: We applied targeted cell-free DNA (cfDNA) sequencing to 126 patients with mCRPC from three academic cancer centers and separately performed genome-wide cfDNA methylation sequencing on 43 plasma samples collected prior to the initiation of first-line ARSI treatment. To analyze the genome-wide sequencing data, we performed nucleosome positioning and differential methylated region analysis. We additionally analyzed single-cell and bulk RNA sequencing data from 14 and 80 patients with mCRPC, respectively, to develop and validate a stem-like signature, which we inferred from cfDNA. Results: Targeted cfDNA sequencing detected AR/enhancer alterations prior to first-line ARSIs that correlated with significantly worse progression-free survival (P = 0.01; HR = 2.12) and overall survival (P = 0.02; HR = 2.48). Plasma methylome analysis revealed that AR/enhancer lethal mCRPC patients have significantly higher promoter-level hypomethylation than AR/enhancer wild-type mCRPC patients (P &lt; 0.0001). Moreover, gene ontology and CytoTRACE analysis of nucleosomally more accessible transcription factors in cfDNA revealed enrichment for stemness-associated transcription factors in patients with lethal mCRPC. The resulting stemness signature was then validated in a completely held-out cohort of 80 patients with mCRPC profiled by tumor RNA sequencing. Conclusions: We analyzed a total of 220 patients with mCRPC, validated the importance of cell-free AR/enhancer alterations as a prognostic biomarker in lethal mCRPC, and showed that the underlying mechanism for lethality involves reprogramming developmental states toward increased stemness. See related commentary by Nawfal et al., p. 7

Abstract Metastasis-ablative radiotherapy is a new therapeutic approach that prolongs survival in patients with advanced cancers. Tumour irradiation can elicit an immunogenic cell death resulting in the expansion of tumour-reactive T cells and eradication of both irradiated and distant non-irradiated metastases, referred to as the “abscopal effect”. This abscopal response, nevertheless, is observed in a minority of patients suggesting uncharacterized mechanisms that prevent robust systemic anti-tumor immune response. Tumour-derived extracellular vesicles (TdEVs) have recently emerged as mediators of antitumor immunity but their immunomodulatory role in response to radiotherapy remains largely unknown. Recent clinical studies by our lab have demonstrated that circulating levels of tumour-derived extracellular vesicles (tdEVs) are elevated in response to radiotherapy and high tdEV concentrations are associated with reduced levels of peripheral cytotoxic CD8+ T cells and higher risk of disease recurrence in advanced prostate cancer. Based on our findings, we hypothesize that radiotherapy induces the release of tumor-derived EVs that suppress anti-tumor immunity. Prostate cancer cells (PC3 and DU145) were irradiated with clinically used doses of ionizing radiation (20Gy x 1 or 10Gy x 3 fractions) and tdEVs were co-cultured with human CD8+ T cells to measure their proliferation and cytotoxic activity. Increased levels of tdEVs were released by tumour cells treated with radiotherapy. A higher frequency of impaired T cell cytotoxic activity and a reduction in CD8 T cell proliferation was found in the 10Gy x 3 radiation regimen. Proteomic analysis and western blot showed an increased expression of the immune checkpoint molecule B7-H3 on the tumour cell surface and tdEVs in response to radiotherapy. While coculture of CD8 T cells with B7-H3 depleted tdEVs led to recovery of T-cell cytotoxic ability, overexpression of the B7-H3 in tdEVs inhibited cytotoxic activity. This study provides a rationale for future studies in which we will test the immune-mediated effects of radiotherapy-induced tdEVs in immunocompetent mouse models and investigate B7-H3 blockade as a therapeutic approach to boost abscopal response in the treatment of advanced prostate cancer. Citation Format: Chisom J. Uzendu, Yohan Kim, Roxane R. Lavoie, Haidong Dong, Sean Park, Fabrice Lucien-Matteoni. Impact of radiotherapy on extracellular vesicle mediated anti-tumor immunity in oligometastatic prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4995.

Abstract Objective . An algorithm was developed for automated positioning of lattice points within volumetric modulated arc lattice radiation therapy (VMAT LRT) planning. These points are strategically placed within the gross tumor volume (GTV) to receive high doses, adhering to specific separation rules from adjacent organs at risk (OARs). The study goals included enhancing planning safety, consistency, and efficiency while emulating human performance. Approach . A Monte Carlo-based algorithm was designed to optimize the number and arrangement of lattice points within the GTV while considering placement constraints and objectives. These constraints encompassed minimum spacing between points, distance from OARs, and longitudinal separation along the z -axis. Additionally, the algorithm included an objective to permit, at the user’s discretion, solutions with more centrally placed lattice points within the GTV. To validate its effectiveness, the automated approach was compared with manually planned treatments for 24 previous patients. Prior to clinical implementation, a failure mode and effects analysis (FMEA) was conducted to identify potential shortcomings. Main results. The automated program successfully met all placement constraints with an average execution time (over 24 plans) of 0.29 ±0.07 min per lattice point. The average lattice point density (# points per 100 c.c. of GTV) was similar for automated (0.725) compared to manual placement (0.704). The dosimetric differences between the automated and manual plans were minimal, with statistically significant differences in certain metrics like minimum dose (1.9% versus 1.4%), D5% (52.8% versus 49.4%), D95% (7.1% versus 6.2%), and Body-GTV V30% (20.7 c.c. versus 19.7 c.c.). Significance. This study underscores the feasibility of employing a straightforward Monte Carlo-based algorithm to automate the creation of spherical target structures for VMAT LRT planning. The automated method yields similar dose metrics, enhances inter-planner consistency for larger targets, and requires fewer resources and less time compared to manual placement. This approach holds promise for standardizing treatment planning in prospective patient trials and facilitating its adoption across centers seeking to implement VMAT LRT techniques.

191 Background: Two randomized clinical trials have demonstrated that SABR can prolong time to progression in oligometastatic castration-sensitive prostate cancer (omCSPC) patients. However, there is a need for predictive tools to identify patients who will benefit from SABR alone or with systemic therapy. We investigated the prognostic value of PSMA-positive extracellular vesicles (PSMA+EVs) in a blinded observational study using blood samples from three independent patient cohorts. Methods: We obtained plasma samples from 157 omCSPC patients treated with SABR from the ORIOLE cohort (N=30, JHU) and STOMP-like cohorts (N=80, UGhent; N=47, Iridium Net). Baseline PSMA+EV levels (EV/ml) were measured by nanoscale flow cytometry. Primary clinical endpoints were biochemical progression (bPFS) and radiographic progression (rPFS). Optimal cutoffs for PSA (ng/ml) and PSMA+EV (EVs/ml) were defined as the point with the most significant log-rank test split. Kaplan-Meier curves and univariate Cox regression models were used to determine the association of PSMA+EV levels with clinical outcomes. Results: Twenty percent of patients (N=157 total) were diagnosed with bone scan, 79% with PET imaging, and 1% with other modalities. Ninety-four percent of patients presented with ≤3 metastases on imaging. In the pooled STOMP-like cohorts, median bPFS was 27.9 and 18.0 months for PSMA+EV low and high groups, respectively (p=0.039). Median rPFS was 36.0 versus 27.0 months (p=0.029). In the ORIOLE cohort, median bPFS was 24.3 and 5.9 months in PSMA+EV low and high, respectively (p=0.021). Median rPFS was 36.0 versus 11.1 months (p=0.019). High baseline PSMA+EV levels were associated with higher risk of both biochemical and radiographic progression (Table). Combination of baseline PSA low and PSMA+EV low outperformed either biomarker alone and better predicted risk of disease progression after SABR. Conclusions: While prospective biomarker-guided trials are warranted, a PSMA+EV blood test can aid patient selection by identifying omCSPC patients who may have durable responses to SABR without ADT. [Table: see text]

Patients with advanced cancers who either do not experience initial response to or progress while on immune checkpoint inhibitors (ICIs) receive salvage radiotherapy to reduce tumor burden and tumor-related symptoms. Occasionally, some patients experience substantial global tumor regression with a rebound of cytotoxic CD8 + T cells. We have termed the rebound of cytotoxic CD8 + T cells in response to salvage therapy as T cell resilience and examined the underlying mechanisms of resilience. Resilient T cells are enriched for CX3CR1 + CD8 + T cells with low mitochondrial membrane potential, accumulate less reactive oxygen species (ROS), and express more malic enzyme 1 (ME1). ME1 overexpression enhanced the cytotoxicity and expansion of effector CD8 + T cells partially via the type I interferon pathway. ME1 also increased mitochondrial respiration while maintaining the redox state balance. ME1 increased the cytotoxicity of peripheral lymphocytes from patients with advanced cancers. Thus, preserved resilient T cells in patients rebound after salvage therapy and ME1 enhances their resiliency.

AbstractBackgroundProstate cancer (PCa) parenchymal brain metastases are uncommon and troubling observations in the course of the disease. Our study aims to evaluate the prevalence of brain metastases among PCa patients while reporting various therapeutic modalities, clinical features, and oncological outcomes.MethodsWe retrospectively identified 34 patients with parenchymal brain metastasis out of 4575 patients using a prospectively maintained database that contains clinicopathologic characteristics of PCa patients between January 2012 and December 2021. Based on the three treatment modalities used, the patients were divided into three groups: stereotactic radiosurgery (SRS), whole brain radiotherapy (WBRT), and systemic therapy alone. The Kaplan–Meier curve was used to calculate overall survival [OS] probability and the Cox proportional hazards regression model was used to compare between groups.ResultsAt the time of brain metastasis diagnosis, the median age was 66 years, the median (interquartile range [IQR]) prostate‐specific antigen (PSA) was 2.2 (0.1–26.6) ng/ml and the median (IQR) months from initial PCa diagnosis to brain metastasis development was 70.8 (27.6–100.9). The median (IQR) primary Gleason score was 8 (7–9) and over a median (IQR) follow‐up time of 2.2 (1.2–16.5) months, 76.5% (n = 26) of the patients died. Thirteen (38.2%) patients had solitary lesion, whereas 21 (61.8%) had ≥2 lesions. The lesions were supratentorial in 19 (55.9%) patients, infratentorial in six (17.6%), and both sides in nine (26.5%). Among all 34 patients, 10 (29.4%) were treated with SRS, seven (20.6%) with WBRT, and 17 (50%) with systemic therapy alone. OS varied greatly between the three treatment modalities (log‐rank test, p = 0.049). Those who were treated with SRS and WBRT had better OS compared with patients who were treated with systemic therapy alone (hazard ratio: 0.37, 95% confidence interval: 0.16–0.86, p = 0.022).ConclusionsIn our single‐institutional study, we confirmed that PCa brain metastasis is associated with poor survival outcomes and more advanced metastatic disease. Furthermore, we found that SRS and WBRT for brain metastasis in patients with recurrent PCa appear to be associated with improved OS as compared with systemic therapy alone and are likely secondary to selection bias.

Abstract Cytotoxic T lymphocytes (CTL) are indispensable in anti-tumor immunity. Although CTLs are prone to exhaustion in patients with advanced cancer, T cell resiliency explains the presence of tumor-reactive CTLs that are less exhausted, capable of cytolytic function, expansion, and rebound in response to immunotherapy to reject metastatic malignancies. However, the features of resilient T cells have not been clearly defined. In this report, we demonstrate that peripheral CX3CR1+ CD8+ T cells with low mitochondrial membrane potential rebounded CTL function quickly after radiation therapy in patients with large tumor burden portraying their functional resiliency. Furthermore, CX3CR1+ CD8+ T cell with low, but not high, mitochondrial membrane potential are highly cytotoxic, accumulate less reactive oxygen species (ROS), and express more Malic enzyme 1 (ME1). ME1 overexpression increases ATP production in a glycolysis-independent manner while concurrently curtailing excessive ROS in activated CD8+ T cells; and expands CX3CR1+NKG7+ effector CD8+ T cells with enhanced cytotoxicity. Importantly, transfection of ME1 mRNA promotes tumoricidal activity in CD8+ T cells from patients with advanced cancers. Our study reveals a mechanism used by CTLs to balance excessive ROS via ME1 to maintain a metabolic and functional resiliency. Modification of ME1 expression in CTLs may be a novel method to improve the efficacy of cancer immunotherapy by preventing T cell exhaustion. Supported by grants from Mayo Clinic Foundation and NIH (R01CA 256927-2)