Immunomodulatory response to neoadjuvant nivolumab in non-metastatic clear cell renal cell carcinoma.

Novel perioperative strategies are needed to reduce recurrence rates in patients undergoing nephrectomy for high-risk, non-metastatic clear cell renal cell carcinoma (ccRCC). We conducted a prospective, phase I trial of neoadjuvant nivolumab prior to nephrectomy in 15 evaluable patients with non-metastatic ccRCC. We leveraged tissue from that cohort to elucidate the effects of PD-1 inhibition on immune cell populations in ccRCC and correlate the evolving immune milieu with anti-PD-1 response. We found that nivolumab durably induces a pro-inflammatory state within the primary tumor, and baseline immune infiltration within the primary tumor correlates with nivolumab responsiveness. Nivolumab increases CTLA-4 expression in the primary tumor, and subsequent nephrectomy increases circulating concentrations of sPD-L1, sPD-L3 (sB7-H3), and s4-1BB. These findings form the basis to consider neoadjuvant immune checkpoint inhibition (ICI) for high-risk ccRCC while the tumor remains in situ and provide the rationale for perioperative strategies of novel ICI combinations.

Li-Fraumeni Syndrome-Associated Dimer-Forming Mutant p53 Promotes Transactivation-Independent Mitochondrial Cell Death.

Cancer-relevant mutations in the oligomerization domain (OD) of the p53 tumor suppressor protein, unlike those in the DNA binding domain, have not been well elucidated. Here, we characterized the germline OD mutant p53(A347D), which occurs in cancer-prone Li-Fraumeni syndrome (LFS) patients. Unlike wild-type p53, mutant p53(A347D) cannot form tetramers and exists as a hyperstable dimeric protein. Further, p53(A347D) cannot bind or transactivate the majority of canonical p53 target genes. Isogenic cell lines harboring either p53(A347D) or no p53 yield comparable tumorigenic properties, yet p53(A347D) displays remarkable neomorphic activities. Cells bearing p53(A347D) possess a distinct transcriptional profile and undergo metabolic reprogramming. Further, p53(A347D) induces striking mitochondrial network aberration and associates with mitochondria to drive apoptotic cell death upon topoisomerase II inhibition in the absence of transcription. Thus, dimer-forming p53 demonstrates both loss-of-function (LOF) and gain-of-function (GOF) properties compared with the wild-type form of the protein. SIGNIFICANCE: A mutant p53 (A347D), which can only form dimers, is associated with increased cancer susceptibility in LFS individuals. We found that this mutant wields a double-edged sword, driving tumorigenesis through LOF while gaining enhanced apoptogenic activity as a new GOF, thereby yielding a potential vulnerability to select therapeutic approaches. See related commentary by Stieg et al., p. 1046. See related article by Gencel-Augusto et al., p. 1230. This article is highlighted in the In This Issue feature, p. 1027.

Anti-PD-1 immunotherapy with androgen deprivation therapy induces robust immune infiltration in metastatic castration-sensitive prostate cancer.

When compared to other malignancies, the tumor microenvironment (TME) of primary and castration-resistant prostate cancer (CRPC) is relatively devoid of immune infiltrates. While androgen deprivation therapy (ADT) induces a complex immune infiltrate in localized prostate cancer, the composition of the TME in metastatic castration-sensitive prostate cancer (mCSPC), and the effects of ADT and other treatments in this context are poorly understood. Here, we perform a comprehensive single-cell RNA sequencing (scRNA-seq) profiling of metastatic sites from patients participating in a phase 2 clinical trial (NCT03951831) that evaluated standard-of-care chemo-hormonal therapy combined with anti-PD-1 immunotherapy. We perform a longitudinal, protein activity-based analysis of TME subpopulations, revealing immune subpopulations conserved across multiple metastatic sites. We also observe dynamic changes in these immune subpopulations in response to treatment and a correlation with clinical outcomes. Our study uncovers a therapy-resistant, transcriptionally distinct tumor subpopulation that expands in cell number in treatment-refractory patients.

Longitudinal Immune Profiling Reveals Unique Myeloid and T-cell Phenotypes Associated with Spontaneous Immunoediting in a Prostate Tumor Model.

The theory of cancer immunoediting, which describes the dynamic interactions between tumors and host immune cells that shape the character of each compartment, is foundational for understanding cancer immunotherapy. Few models exist that facilitate in-depth study of each of the three canonical phases of immunoediting: elimination, equilibrium, and escape. Here, we utilized NPK-C1, a transplantable prostate tumor model that we found recapitulated the three phases of immunoediting spontaneously in immunocompetent animals. Given that a significant portion of NPK-C1 tumors reliably progressed to the escape phase, we were able to delineate cell types and mechanisms differentially prevalent in equilibrium versus escape phases. Using high-dimensional flow cytometry, we found that activated CD4+ effector T cells were enriched in regressing tumors, highlighting a role for CD4+ T cells in antitumor immunity. CD8+ T cells were also important for NPK-C1 control, specifically, central memory-like cytotoxic CD8+ T cells. Regulatory T cells (Treg), as a whole, were counterintuitively enriched in regressing tumors; however, high-dimensional analysis revealed their significant phenotypic diversity, with a number of Treg subpopulations enriched in progressing tumors. In the myeloid compartment, we found that iNOS+ dendritic cell (DC)-like cells are enriched in regressing tumors, whereas CD103+ DCs were associated with late-stage tumor progression. In total, these analyses of the NPK-C1 model provide novel insights into the roles of lymphoid and myeloid populations throughout the cancer immunoediting process and highlight a role for multidimensional, flow-based analyses to more deeply understand immune cell dynamics in the tumor microenvironment.

Blocking IL1 Beta Promotes Tumor Regression and Remodeling of the Myeloid Compartment in a Renal Cell Carcinoma Model: Multidimensional Analyses.

PURPOSE: Intratumoral immunosuppression mediated by myeloid-derived suppressor cells (MDSC) and tumor-associated macrophages (TAM) represents a potential mechanism of immune checkpoint inhibitor (ICI) resistance in solid tumors. By promoting TAM and MDSC infiltration, IL1ß may drive adaptive and innate immune resistance in renal cell carcinoma (RCC) and in other tumor types. EXPERIMENTAL DESIGN: Using the RENCA model of RCC, we evaluated clinically relevant combinations of anti-IL1ß plus either anti-PD-1 or the multitargeted tyrosine kinase inhibitor (TKI), cabozantinib. We performed comprehensive immune profiling of established RENCA tumors via multiparameter flow cytometry, tumor cytokine profiling, and single-cell RNA sequencing (RNA-seq). Similar analyses were extended to the MC38 tumor model. RESULTS: Analyses via multiparameter flow cytometry, tumor cytokine profiling, and single-cell RNA-seq showed that anti-IL1ß reduces infiltration of polymorphonuclear MDSCs and TAMs. Combination treatment with anti-IL1ß plus anti-PD-1 or cabozantinib showed increased antitumor activity that was associated with decreases in immunosuppressive MDSCs and increases in M1-like TAMs. CONCLUSIONS: Single-cell RNA-seq analyses show that IL1ß blockade and ICI or TKI remodel the myeloid compartment through nonredundant, relatively T-cell-independent mechanisms. IL1ß is an upstream mediator of adaptive myeloid resistance and represents a potential target for kidney cancer immunotherapy.

T-Cell Infiltration and Adaptive Treg Resistance in Response to Androgen Deprivation With or Without Vaccination in Localized Prostate Cancer.

PURPOSE: Previous studies suggest that androgen deprivation therapy (ADT) promotes antitumor immunity in prostate cancer. Whether a vaccine-based approach can augment this effect remains unknown. PATIENTS AND METHODS: We conducted a neoadjuvant, randomized study to quantify the immunologic effects of a GM-CSF-secreting allogeneic cellular vaccine in combination with low-dose cyclophosphamide (Cy/GVAX) followed by degarelix versus degarelix alone in patients with high-risk localized prostate adenocarcinoma who were planned for radical prostatectomy. RESULTS: Both Cy/GVAX plus degarelix and degarelix alone led to significant increases in intratumoral CD8+ T-cell infiltration and PD-L1 expression as compared with a cohort of untreated, matched controls. However, the CD8+ T-cell infiltrate was accompanied by a proportional increase in regulatory T cells (Treg), suggesting that adaptive Treg resistance may dampen the immunogenicity of ADT. Although Cy/GVAX followed by degarelix was associated with a modest improvement in time-to-PSA progression and time-to-next treatment, as well as an increase in PD-L1, there was no difference in the CD8+ T-cell infiltrate as compared with degarelix alone. Gene expression profiling demonstrated that CHIT1, a macrophage marker, was differentially upregulated with Cy/GVAX plus degarelix compared with degarelix alone. CONCLUSIONS: Our results highlight that ADT with or without Cy/GVAX induces a complex immune response within the prostate tumor microenvironment. These data have important implications for combining ADT with immunotherapy. In particular, our finding that ADT increases both CD8+ T cells and Tregs supports the development of regimens combining ADT with Treg-depleting agents in the treatment of prostate cancer.