Prostate cancer is one of the most common and heritable human cancers. Our aim was to find germline biomarkers that can predict disease outcome. We previously detected predisposing signals at 2q37, the location of the prostate specific ANO7 gene. To investigate, in detail, the associations between the ANO7 gene and PrCa risk and disease aggressiveness, ANO7 was sequenced in castration resistant tumors together with samples from unselected PrCa patients and unaffected males. Two pathogenic variants were discovered and genotyped in 1769 patients and 1711 unaffected males. Expression of ANO7 vs. PrCa aggressiveness was investigated. Different databases along with Swedish and Norwegian cohorts were used for validation. Case-control and aggressive vs. nonaggressive association analyses were performed against risk and/or cancer aggressiveness. The ANO7 mRNA level and patient survival were analyzed using expression data from databases. Variant rs77559646 showed both risk (OR 1.40; p = 0.009, 95% CI 1.09-1.78) and association with aggressive PrCa (Genotype test p = 0.04). It was found to be an eQTL for ANO7 (Linear model p-values for Finnish patients p = 0.009; Camcap prostate tumor p = 2.53E-06; Stockholm prostate tumor cohort p = 1.53E-13). rs148609049 was not associated with risk, but was related to shorter survival (HR 1.56; 95% CI 1.03-2.36). High ANO7 expression was independently linked to poor survival (HR 18.4; 95% CI 1.43-237). ANO7 genotypes correlate with expression and biochemical relapse, suggesting that ANO7 is a potential PrCa susceptibility gene and that its elevated expression correlates with disease severity and outcome.

BACKGROUND: We have previously shown that TILs are predictive of benefit to trastuzumab and chemotherapy in HER2+ BC in the FinHER study, an adjuvant, phase III study in early-stage BC, where HER2+ patients were randomized to 9 weeks of trastuzumab or no trastuzumab in addition to chemotherapy (Loi et al, ASCO 2012). We sought to further confirm this positive association as well as to understand the composition of TILs. METHODS: The association between TILs and response to trastuzumab with chemotherapy (epirubicin/cyclophosphamide with docetaxel with or without capecitabine) was evaluated in 156 HER2+ patients from the neoadjuvant GeparQuattro trial. The primary correlative endpoint was the association between TILs (quantified using the same method as previously published) with pathological complete response rates (pCR), adjusted for clinicopathological characteristics. To understand the composition of TILs, correlations between TILs and gene expression levels of 13 pre-defined immune markers were evaluated from 202 HER2+ samples from the FinHER study. These represented T and B cell infiltration (CD3D, IGKC), Th1 (IFNG, CD8A), chemoattractants (CXCL9, CXCL13), immunosuppression (VEGFA, FOXP3, IDO1) and T-cell checkpoint receptors and ligands (PD-1, PD-L1, CTLA-4, CD80). Prognostic associations and interactions with trastuzumab were studied in Cox regression models for distant-disease free survival (DDFS). Preclinical mice models of HER2 mammary cancer were used to investigate combination therapies. RESULTS: In the GeparQuattro trial data, each 10% increment in TILs was associated with higher rates of pCR (adjusted OR:1.14 95%CI:1.01-1.29;P = 0.037) after neoadjuvant trastuzumab and chemotherapy supporting the findings from the FinHER study. Gene expression analyses using the FinHER samples revealed IDO1 and CXCL13 were most highly correlated with TILs (R = 0.58 and 0.51;p<0.001 respectively). Whilst no immune genes were significantly associated with prognosis in HER2+ BC, high expression of PD-1 and IDO1 were significantly associated with greater trastuzumab benefit for DDFS (Interaction p values: PD-1 = 0.029; IDO1 = 0.039) suggesting that trastuzumab modulates the immune microenvironment. We next postulated that enhancing T-cell responses would be synergistic with trastuzumab. Trastuzumab in combination with several inhibitors of T-cell negative regulation (anti-CTLA4, anti-PD-1, anti-PD-L1) resulted in higher tumor regressions compared with monotherapy in a mouse transplant model of HER2+ mammary cancer. In particular, anti-PD1 (P = 0.02) and anti-PD-L1 (P = 0.008) were most effective. In a BALB/c-MMTV-neu transgenic mice mouse model (i.e. immune tolerant to HER2 antigen), the combination of trastuzumab and anti-PD1 antibody could significantly delay mammary gland tumor formation (p<0.001). CONCLUSIONS: We confirm that TILs are associated with higher responses to trastuzumab and chemotherapy. Furthermore, tumor-mediated immunosuppression is evident in the lymphocytic infiltrate with PD-1 and IDO1 significantly predictive of trastuzumab benefit. The addition of a T-cell checkpoint inhibitor in a preclinical model significantly enhanced trastuzumab responses. These combinations warrant evaluation in the clinical setting. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr S1-05.