Kwon, Eugene D.

Abstract Introduction: Efforts to improve clear cell renal cell carcinoma (ccRCC) mortality have focused primarily on evaluating prognostic biomarkers in primary tumors in order to identify indicators of response to treatment and targets for novel therapies. To date little attention has been paid to the status of these biomarkers in the more therapeutically-relevant metastatic tumor. Thus, we evaluated concordance of expression of four key prognostic ccRCC biomarkers (BAP1, PBRM1, Ki-67 and TOPOIIa) in a large cohort of patient-matched primary ccRCC and metastatic tumors. Methods: We identified 114 patients treated surgically for ccRCC between 1990 and 2005 who had synchronous (M1) or metachronous (M0) metastases, underwent metastasectomy for at least one metastatic lesion and had archived tissue available from the primary and at least one metastatic tumor. We performed IHC on all primary and metastatic tumors for BAP1, PBRM1, Ki-67, and TOPOIIa using previously-published protocols. BAP1 and PBRM1 expression was categorized as positive or negative and Ki-67 and TOPOIIa was quantified as the number of positive cells per mm2. For BAP1 and PBRM1, we calculated the concordance between patient-matched primary and metastatic tumors as the percentage of pairs where the primary and metastatic tumor had the same designation (i.e., positive or negative). For TOPOIIa and Ki-67, we assessed concordance using Spearman correlation coefficient. Analysis of variance was used to evaluate if concordance between primary-metastatic tumor pairs for TOPOIIa and Ki-67 was associated with metastatic site, metastatic timing (M0/M1), metastatic tumor grade, metastatic tumor necrosis, and metastatic tumor sarcomatoid differentiation. Results: There were a total of 161 patient-matched metastatic tumors available for the 114 patients: 78 patients had only a single metastasis while 36 patients had two or more metastases. While sites of metastases varied, pulmonary metastases were the most common (44% of all metastases). Loss of BAP1 expression was observed in19% of the primary ccRCC and 98.6% of the tumor pairs had concordant BAP1 status between the paired primary and metastatic tumors. Loss of PBRM1 was observed in 56% of the primary ccRCC tumors and 89.3% of tumor pairs showed concordant PBRM1 status. By comparison, median expression of Ki-67 was 65.0mm2 in the primary ccRCC tumors and the Pearson correlation with the patient-matched metastatic tumors was 0.43. The difference in Ki-67 expression between primary-metastatic tumor pairs was associated with necrosis in the metastatic tumor (p=0.014). Median expression of TOPOIIa was 2.6mm2 in the primary ccRCC tumors and the Pearson correlation with the patient-matched metastatic tumors was 0.25. The difference in TOPOIIa expression between primary-metastatic tumor pairs was associated with metastatic site (p=0.0006) and metastases to the heart were significantly different than pulmonary metastases (p<0.0001). Additionally, synchronous metastases had larger differences from their patient-matched primary tumors in comparison to metachronous metastases (p=0.012), as did necrotic metastases (p=0.028). Conclusion: We observed high concordance for loss of BAP1 and PBRM1 expression between primary ccRCC and metastatic tumors in the same patient. Conversely, we noted high discordance in protein expression of Ki-67 and TOPOIIa between patient-matched primary-metastatic tumor pairs. Moreover, this discordance varied by key pathological features. Our results suggest that using the status of a biomarker in primary ccRCC as a guide for the status in metastatic tumors is not always appropriate and needs to be evaluated on a marker-by-marker basis. Citation Format: Jeanette E. Eckel-Passow, Daniel J. Serie, John C. Cheville, Thai H. Ho, Sue M. Harrington, Tracy Hilton, Christine Lohse, Amanda Shreders, Payal Kapur, James Brugarolas, R Houston Thompson, Bradley C. Leibovich, Eugene D. Kwon, Richard W. Joseph, Alexander S. Parker. Concordance of BAP1, PBRM1, Ki-67, and TOPOIIa protein expression in primary clear cell renal cell carcinoma tumors and patient-matched metastatic tumors. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Integrating Clinical Genomics and Cancer Therapy; Jun 13-16, 2015; Salt Lake City, UT. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(1_Suppl):Abstract nr 10.

Abstract Purpose: Immunotherapeutic strategies to treat patients with renal cell carcinoma (RCC) offer new opportunities for disease management. Further improvements to immunotherapy will require additional understanding of the host response to RCC development. Experimental Design: Using a novel approach to understanding the immune status of cancer patients, we previously showed that patients with a certain immune profile had decreased overall survival. Here, we examine in more detail the phenotypic changes in peripheral blood and the potential consequences of these changes in RCC patients. Results: We found that CD14+HLA-DRlo/neg monocytes were the most predominant phenotypic change in peripheral blood of RCC patients, elevated nearly 5-fold above the average levels measured in healthy volunteers. Intratumoral and peritumoral presence of CD14 cells was an independent prognostic factor for decreased survival in a cohort of 375 RCC patients. The amount of peripheral blood CD14+HLA-DRlo/neg monocytes was found to correlate with the intensity of CD14 staining in tumors, suggesting that the measurement of these cells in blood may be a suitable surrogate for monitoring patient prognosis. The interaction of monocytes and tumor cells triggers changes in both cell types with a loss of HLA-DR expression in monocytes, increases of monocyte survival factors such as GM-CSF in tumors, and increased production of angiogenic factors, including FGF2. Conclusions: Our results suggest a model of mutually beneficial interactions between tumor cells and monocytes that adversely affect patient outcome. Clin Cancer Res; 21(18); 4224–33. ©2015 AACR.

Abstract We investigated the influence of PD-1 expression on the systemic antitumor response (abscopal effect) induced by stereotactic ablative radiotherapy (SABR) in preclinical melanoma and renal cell carcinoma models. We compared the SABR-induced antitumor response in PD-1–expressing wild-type (WT) and PD-1–deficient knockout (KO) mice and found that PD-1 expression compromises the survival of tumor-bearing mice treated with SABR. None of the PD-1 WT mice survived beyond 25 days, whereas 20% of the PD-1 KO mice survived beyond 40 days. Similarly, PD-1–blocking antibody in WT mice was able to recapitulate SABR-induced antitumor responses observed in PD-1 KO mice and led to increased survival. The combination of SABR plus PD-1 blockade induced near complete regression of the irradiated primary tumor (synergistic effect), as opposed to SABR alone or SABR plus control antibody. The combination of SABR plus PD-1 blockade therapy elicited a 66% reduction in size of nonirradiated, secondary tumors outside the SABR radiation field (abscopal effect). The observed abscopal effect was tumor specific and was not dependent on tumor histology or host genetic background. The CD11ahigh CD8+ T-cell phenotype identifies a tumor-reactive population, which was associated in frequency and function with a SABR-induced antitumor immune response in PD-1 KO mice. We conclude that SABR induces an abscopal tumor-specific immune response in both the irradiated and nonirradiated tumors, which is potentiated by PD-1 blockade. The combination of SABR and PD-1 blockade has the potential to translate into a potent immunotherapy strategy in the management of patients with metastatic cancer. Cancer Immunol Res; 3(6); 610–9. ©2015 AACR.

Abstract Purpose: High-dose aldesleukin (HD IL2) received FDA approval for the treatment of metastatic renal cell carcinoma (MRCC) in 1992, producing a 14% objective response rate (ORR) and durable remissions. Retrospective studies suggested that clinical and pathologic features could predict for benefit. The Cytokine Working Group conducted this prospective trial to validate proposed predictive markers of response to HD IL2. Experimental Design: Standard HD IL2 was administered to prospectively evaluate whether the ORR of patients with mRCC with “good” predictive pathologic features based on an “integrated selection” model [ISM (e.g., clear-cell histology subclassification and carbonic anhydrase-9 (CA-9) IHC staining] was significantly higher than the ORR of a historical, unselected population. Archived tumor was collected for pathologic analysis including tumor programmed death-ligand 1 (PD-L1) expression. Results: One hundred and twenty eligible patients were enrolled between June 11 and September 7; 70% were Memorial Sloan Kettering Cancer Center (New York, NY) intermediate risk, 96% had clear cell RCC, and 99% had prior nephrectomy. The independently assessed ORR was 25% (30/120, 95% CI, 17.5%–33.7%, P = 0.0014; 3 complete responses, 27 partial responses) and was higher than a historical ORR. Thirteen patients (11%) remained progression free at 3 years and the median overall survival was 42.8 months. ORR was not statistically different by ISM classification (“good-risk” 23% vs. “poor-risk” 30%; P = 0.39). ORR was positively associated with tumor PD-L1 expression (P = 0.01) by IHC. Conclusions: In this prospective, biomarker validation study, HD IL2 produced durable remissions and prolonged survival in both “good” and “poor-risk” patients. The proposed ISM was unable to improve the selection criteria. Novel markers (e.g., tumor PD L1 expression) appeared useful, but require independent validation. Clin Cancer Res; 21(3); 561–8. ©2014 AACR.

Abstract Androgen has been implicated as a negative regulator of host immune function and a factor contributing to the gender dimorphism of autoimmunity. Conversely, androgen deprivation has been suggested to potentiate male host immunity. Studies have shown that removal of androgen in postpubertal male mice produces an increase in size and cellularity of primary and peripheral lymphoid organs, and enhances a variety of immune responses. Yet, few details are known about the effect of androgen removal on T cell-mediated immunity. In this study, we demonstrate two pronounced and independent alterations in T cell immunity that occur in response to androgen deprivation, provided by castration, in postpubertal male mice. First, we show that levels of T cells in peripheral lymphoid tissues of mice are increased by androgen deprivation. Second, T cells from these mice transiently proliferate more vigorously to TCR- and CD28-mediated costimulation as well as to Ag-specific activation. In addition, androgen deprivation accelerates normalization of host T and B cell levels following chemotherapy-induced lymphocyte depletion. Such alterations induced by androgen deprivation may have implications for enhancing immune responses to immunotherapy and for accelerating the recovery of the immune system following chemotherapy.

Abstract Adoptive γδ T cell immunotherapy has moved briskly into clinical trials prompted by several small studies suggesting abundant accumulation of γδ T cells within renal cell carcinoma (RCC). In this study, we re-examined levels of γδ T cells within RCC tumors and correlated levels of these cells with pathologic features and outcome associated with this form of cancer. Tissues from 248 consecutive clear cell RCC tumors obtained from 2000 to 2003 were stained and quantified for total CD3+ and γδ T cells per mm2. Wilcoxon rank sum and Kruskal-Wallis tests were used to evaluate associations between T cell amounts and prognostic factors (age, gender, tumor size, stage, grade, tumor necrosis). Cox models were used to assess associations with RCC-specific death. Median numbers of total CD3+ and γδ T cells were 281/mm2 (interquartile range (IQR): 149–536) and 2.6/mm2 (IQR: 1.3–4.6), respectively. The median percentage of CD3+ T cells that were γδ T cells was 1.0% (IQR: 0.4–1.9). This low percentage of intratumoral γδ T cells was diluted even further with rising CD3+ T cell infiltration. Percentages of γδ T cells were not associated with even one single clinicopathologic feature examined. Median follow-up for this study was 3.1 years (48 patients died of RCC) and Cox analysis failed to demonstrate that γδ T cells (hazard ratio = 1.02, p = 0.25) were predictive of RCC-specific death. γδ T cells are rare and not recruited nor expanded within RCC tumors. Percentages of γδ T cells fail to correlate with any prognostic features of RCC nor specific death. As such, the role of γδ T cells in RCC immunobiology remains questionable.