Abstract PO-046: Single-cell profiling of tumor evolution and complex immune infiltrate induced by ADT and PD-1 blockade in metastatic hormone-sensitive prostate cancer
Authors: Obradovic Aleksandar , Hawley Jessica , Guo Xinzheng , Chaimowitz Matthew , Dallos Matthew , Stein Mark , Califano Andrea , Drake Charles 
Published: December 2020

Abstract
Patients presenting with de novo metastatic, hormone-sensitive prostate cancer (mHSPC), while initially responsive to androgen deprivation therapy (ADT), invariably progress to castration-resistant prostate cancer. We and others have shown that ADT induces a complex immune infiltrate in localized prostate cancer, in both animal models and humans, leading to trials of combination ADT with immunotherapy, but the effect of ADT alone or in combination with checkpoint blockade have not been well-described in metastatic tumor sites. We are conducting a clinical trial (NCT03951831) to test the hypothesis that ADT-induced immune infiltrate can be further augmented with anti-PD-1 inhibition in men with mHSPC. Here, we present single-cell RNA-sequencing analysis of paired metastatic tumor biopsies at baseline and on-treatment. In our phase 2, open-label, single-center trial, eligible patients with mHSPC are treated with phased administration of ADT, cemiplimab (anti-PD-1 inhibitor), and docetaxel. All patients undergo baseline metastatic tumor biopsies as well as on-treatment biopsies after either ADT alone or ADT in combination with cemiplimab. Biopsies are collected from bone, lymph node, and liver metastases, and are dissociated by gentleMACS protocol for sequencing using the 10X Chromium platform. Cells are first clustered at the transcriptional level by unsupervised Louvain algorithm with optimization of the resolution parameter by a novel silhouette-score subsampling technique. Cells are further sub-clustered following inference of protein activity by the VIPER algorithm. Cell types are assigned by singleR and tumor cells verified by inference of Copy Number Variations (InferCNV). Baselines are compared to their paired on-treatment sample. Differences in treatment-induced immune cell fold-change are compared for patients profiled following ADT only (n=3) and patients profiled following ADT in combination with checkpoint blockade (n=2). Furthermore, transcriptional and proteomic profiles of tumor cells persisting after treatment are compared to those present before treatment to identify consistent treatment-induced changes in tumor cell state. Preliminary results showed significant increase in the fold-change of CD8+ T cells in combination-treated samples relative to baseline (P &lt; 0.001), which was not observed in patients treated with ADT only. There was also a dramatic shift in other immune cell types with treatment, including macrophages and Treg. In addition, we found a consistent up-regulation of antigen-presentation in tumor cells following ADT, and identified that tumor cells expressing KLK2/KLK3 significantly dissipate with treatment (mean fold-change 0.56, P &lt; 0.001). Taken together, these data demonstrate the feasibility of scRNA-seq on prostate cancer metastatic lesions and provide initial support for the hypothesis that ADT in combination with checkpoint blockade induces complex immunological changes in the metastatic tumor microenvironment.
Citation Format: Aleksandar Obradovic, Jessica Hawley, Xinzheng Guo, Matthew Chaimowitz, Matthew Dallos, Mark Stein, Andrea Califano, Charles Drake. Single-cell profiling of tumor evolution and complex immune infiltrate induced by ADT and PD-1 blockade in metastatic hormone-sensitive prostate cancer [abstract]. In: Proceedings of the AACR Virtual Special Conference on Tumor Heterogeneity: From Single Cells to Clinical Impact; 2020 Sep 17-18. Philadelphia (PA): AACR; Cancer Res 2020;80(21 Suppl):Abstract nr PO-046.