Serie, Daniel

530 Background: Therapeutic efficacy of H involves activation of the immune system.Age-dependent progressive deterioration of the immune response is referred as immunosenescence. In HER2+ breast cancer, the effects of aging and the ability of H to activate endogenous anti-tumor immunity is unknown. A previous report from HERA trial showed no significant increase in risk of early recurrence with age (≤ or > 40). We evaluated the long-term outcome of HER2+ patients (pt) related to age and immune landscape. Methods: 1,392 pt from N9831 trial were evaluated. Stromal tumor infiltrating lymphocytes (sTIL) were evaluated in H&E slides. Lymphocyte predominant breast cancer (LPBC) was defined as sTIL ≥ 50%. Molecular TIL (mTIL) and immune subset signatures were evaluated using NanoString research CodeSets. Cox proportional hazard ratio (HR) was used for analysis. Results: There were 1,111 (79.8%) pt > 40 years old (yo) and 281 (20.2%) pt ≤ 40 yo. Younger age was significantly associated with hormone receptor positivity (p 0.00011) and luminal B subtype (p 0.011). With a median follow up of 10.6 years, there was no significant difference in long-term outcome among pt ≤ 40 vs. > 40 yo who received H (HR = 0.88 (0.62-1.24), p = 0.45). Similar findings were observed when age was dichotomized at 50 and 60 yo. While there was no association between sTIL and age, a small but significant increase in mTIL CD45 expression with age (p 0.003) was observed. Similar small increases in cytotoxic cell (p 0.007) and T cell (p 0.015) immune scores were also observed with increasing age. Among pt who received chemotherapy alone, pt > 40 yo with LPBC (n 55) had excellent outcome with 92.4% recurrence free survival (RFS) at 10 years. However, there was no significant difference in RFS among pt ≤ 40 yo with or without LPBC. Conclusions: Among pt treated with H, there was no significant difference in outcome related to age. In contrast to a decline expected from immunosenescence, we observed small but significant increases in mTIL signatures for total lymphocytes, cytotoxic, and T cell subsets. Among patients who received chemotherapy alone, our data suggested that pt > 40 yo with LPBC had excellent prognosis, compared to pt > 40 yo without LPBC.

Anthracycline-induced cardiotoxicity (ACT) is a key limiting factor in setting optimal chemotherapy regimes, with almost half of patients expected to develop congestive heart failure given high doses. However, the genetic basis of sensitivity to anthracyclines remains unclear. We created a panel of iPSC-derived cardiomyocytes from 45 individuals and performed RNA-seq after 24 hr exposure to varying doxorubicin dosages. The transcriptomic response is substantial: the majority of genes are differentially expressed and over 6000 genes show evidence of differential splicing, the later driven by reduced splicing fidelity in the presence of doxorubicin. We show that inter-individual variation in transcriptional response is predictive of in vitro cell damage, which in turn is associated with in vivo ACT risk. We detect 447 response-expression quantitative trait loci (QTLs) and 42 response-splicing QTLs, which are enriched in lower ACT GWAS p -values, supporting the in vivo relevance of our map of genetic regulation of cellular response to anthracyclines.

Abstract Introduction Comparative transcriptome analyses in Alzheimer's disease (AD) and other neurodegenerative proteinopathies can uncover both shared and distinct disease pathways. Methods We analyzed 940 brain transcriptomes including patients with AD, progressive supranuclear palsy (PSP; a primary tauopathy), and control subjects. Results We identified transcriptional coexpression networks implicated in myelination, which were lower in PSP temporal cortex (TCX) compared with AD. Some of these associations were retained even after adjustments for brain cell population changes. These TCX myelination network structures were preserved in cerebellum but they were not differentially expressed in cerebellum between AD and PSP. Myelination networks were downregulated in both AD and PSP, when compared with control TCX samples. Discussion Downregulation of myelination networks may underlie both PSP and AD pathophysiology, but may be more pronounced in PSP. These data also highlight conservation of transcriptional networks across brain regions and the influence of cell type changes on these networks.

Alcohol consumption has been associated inversely with renal cell carcinoma (RCC) risk; however, no study has examined effect modification by germline variation in alcohol‐metabolizing genes. We investigated whether the association between alcohol intake and RCC risk is modulated by germline variants in alcohol dehydrogenase genes in a large case–control study. Data from 652 RCC cases and 1,366 non‐cancer controls were analyzed. Alcohol intake was assessed using a standardized risk factor questionnaire. Three previously genotyped polymorphisms in ADH6 and ADH7 with the TaqMan assay were examined. Odds ratios (ORs) and 95% confidence interval (CI) were calculated using logistic regression, adjusting for covariates. Compared to non‐drinkers, ever consumption of alcohol was associated with lower RCC risk (OR = 0.52, 95% CI = 0.42–0.65). Analysis with cubic spline regression curve showed a “J‐shaped” relationship between alcohol drinks/day and RCC risk, such that there was no added benefit against RCC for consumption of more than two drinks/day. We observed effect modification by variation in rs1154454 (ADH7) (pinteraction = 0.007); a per unit increase in alcohol drink/day was associated with 35% lower RCC risk among non‐minor allele carriers, a 27% lower risk among those who carry one copy of the minor allele, but no association was observed among those with two copies of the minor allele. These findings indicate that alcohol consumption is associated with lower RCC risk. Consuming more than two drinks a day does not confer additional protection against RCC. The association between alcohol intake and RCC risk appears to be modulated by inter‐individual germline variation in alcohol‐metabolizing genes.

Purpose Enhancer of zeste homolog 2 (EZH2), a chromatin remodeler, is implicated in the pathogenesis of clear cell renal cell carcinoma (ccRCC). However, the effect of EZH2 on outcomes in localized ccRCC is unclear, and molecular biomarkers are not currently integrated into prognostic models or adjuvant therapy trials. Methods We performed Cox regression to evaluate the association of tumor-based EZH2 gene and protein expression with survival in three independent cohorts: a cohort from The Cancer Genome Atlas (n = 532), a cohort from University of Texas Southwestern Medical Center (n = 122), and a cohort from Mayo Clinic (n = 1,338). Analyses were adjusted for the prognostic stage, size, grade, and necrosis (SSIGN) score as well as within low-, intermediate-, and high-risk SSIGN groups. Results Patients in The Cancer Genome Atlas cohort with EZH2-high gene expression were 1.5 times more likely to experience overall death than patients with EZH2-low expression (95% CI, 1.1 to 2.3; P = .028). Patients in the University of Texas Southwestern Medical Center cohort with EZH2-high protein expression were two times more likely to experience overall death than patients with EZH2-low expression (95% CI, 1.1 to 4.4; P = .034). Similarly, patients in the Mayo Clinic cohort with EZH2-high protein expression were 1.4 times more likely to experience overall death (95% CI, 1.2 to 1.7; P < .001). Patients in the Mayo Clinic cohort with EZH2-high protein expression were nearly two times more likely to experience RCC-specific death (95% CI, 1.5 to 2.6; P < .001); EZH2 protein expression was particularly prognostic among patients with low-risk SSIGN tumors (HR, 6.1; 95% CI, 3.4 to 11.1; P < .001). Conclusion EZH2 expression accurately predicts risk of RCC death beyond existing clinicopathologic models, particularly in low- and intermediate-risk SSIGN tumors. Further studies are required to incorporate molecular biomarkers into surveillance guidelines and adjuvant clinical trials.

Abstract Background: Understanding heterogeneity within individual breast tumors is key to the ability to predict therapeutic outcome. Molecular heterogeneity is commonly evaluated based on genomic features, including mRNA abundance, gene copy number events, and somatic mutations. The expression profile and activation state of key proteins is widely recognized as another key element in defining tumor heterogeneity. We have taken advantage of NanoString 3D Biology™ technology (for research use only) and curated nCounter Vantage 3DTM Solid Tumor Assay to interrogate a survey panel of HER2-positive breast tumors with the ultimate goal of determining key relationships between multiple genomic and proteomic profiles in individual tumors. Methods: We analyzed samples from 24 HER2+ breast cancer patients using NanoString technology to quantify the expression profile for over 25 total and phospho signaling proteins, including PI3K/MAPK/EGFR/HER2, 770 mRNA corresponding to 13 canonical cancer pathways, and 104 somatic mutations and small INDELS that are commonly associated with cancer, including 8 known PIK3CA mutations. These analyses were carried out in a matched fresh frozen and FFPE samples on the nCounter paltform. Data were analyzed by nSolver to identify genotype specific expression profiles across the 24 samples. Results: In our proof-of-concept data set, we successfully demonstrate that NanoString’s 3D biology Technology shows concordance across both FFPE and fresh frozen sample types for DNA, RNA, and protein. NanoString analysis also showed high concordance to gold-standard techniques used to assess genotype and RNA expression profiles. The combination of digital DNA, RNA, and protein data from our HER2+ breast cancer samples yielded potentially actionable data based on mapping of mutational status as the driver of key differences in protein expression and mRNA abundance of the signaling targets profiled. This work sheds new light on HER2+ breast cancer biology and the interplay between genomic and proteomic profiles while setting the stage for future studies that further probe the differences observed in this sample set. Conclusions: Simultaneous analysis of mutational status (SNV) and expression at the level of both mRNA and protein promises to enable a more detailed view of the relationship between genotype and the biological and clinical behavior of key tumor types. The NanoString Vantage 3DTM Solid Tumor platform provides a rapid, reliable, and economic means of assessing these analytes simultaneously. The application of these analytes to models that make clinically actionable predictions will require additional analyses of large sample cohorts, but such analysis is quite feasible using a variety of sample types. Acknowledgements: Supported in part by grants from the Breast Cancer Research Foundation and the 26.2 with Donna Foundation. Citation Format: Sarayna Chumsri, Daniel J. Serie, Brian M. Necela, Jennifer M. Kachergus, Bianca C. Axenfeld, Gokhan Demirkan, Gavin Meredith, P. Martin Ross, Anisha Kharkia, Erin Piazza, Afshin Mashadi-Hossein, Sarah Warren, Sarah A. McLaughlin, Joseph Beechem, Gary Geiss, E. Aubrey Thompson. Simultaneous analysis of the mutational landscape and RNA and protein expression profile of HER2-positive breast cancer using 3D BiologyTM [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3377. doi:10.1158/1538-7445.AM2017-3377

Abstract Background: Trastuzumab (H), a monoclonal antibody against HER2, has revolutionized the treatment for HER2-positive breast cancer. Besides inhibiting downstream signaling of HER2, several studies showed that H also exerts its anti-tumor activity via immune-related mechanisms. While H is quite effective in preventing recurrence, significant numbers of patients still develop recurrence despite adjuvant H-based chemotherapy. In this study, we analyzed immune-related genes that were associated with poor outcome in N9831. Methods: NanoStringTM technology was used to quantify mRNA of immune-related genes in baseline samples from 1,280 patients in N9831. In N9831, patients in arm A were treated with chemotherapy alone (AC-T), arm B received chemotherapy followed by sequential H (AC-T-H), and arm C received H concurrently with chemotherapy (AC-TH). Cox proportional hazard ratio (HR) was used to determine the association of each immune-related gene with recurrence free survival (RFS). Different immune subset signatures, including CD45, CD8, cytotoxic-cells, and T-cells were analyzed using algorithms developed by NanoString. Results: With the median follow up of 10.6 years, we identified a total 77 genes that were associated with improved outcome in arm C. Among these 77 genes, there were 20 tumor necrosis factor (TNF)-related genes. Of those, only OX40 (TNFRSF4) and its ligand TNFSF4 have interaction p &lt; 0.10. Interestingly, we found uneven distribution of OX40 expression in the N9831 specimens. Approximately, 9.5% of HER2-positive breast cancer expressed OX40 at distinctly low level. Low expression of OX40 was significantly associated with HR positivity (OX40 low 61% vs. OX40 high 51%, p 0.003) and larger tumor size but not patients’ age, tumor grade, and lymph node status. Low expression of OX40 was significantly associated with low mTIL-CD45, CD8, cytotoxic-cells, and T-cells immune signature scores. Tumors with low OX40 expression had significantly lower levels of CTLA4 (p 9.11e-71) and PD-L1 expressions (p 2.49e-92). Low expression of OX40 is associated with poorer outcome among patients treated with H in both sequential (HR 0.88, 95% CI 0.79-0.98, p 0.022) and concurrent arms (HR 0.86, 95%CI 0.76-0.98, p 0.027) but not in chemotherapy only arm (HR 0.9, 95%CI 0.90-1.09, p 0.8). Similar findings were observed with its ligand TNFSF4 in arm A (HR 1.02, 95%CI 0.92-1.15, p 0.68), arm B (HR 0.87, 95%CI 0.76-0.99, p 0.04), and arm C (HR 0.81, 95%CI 0.68-0.95, p 0.01). Conclusion: Our study suggests that pre-existing expression of OX40 and its ligand TNFSF4 are prognostic and may also be predictive of adjuvant H benefit. Patients with distinctly low level of OX40 had poor outcome despite adjuvant H-based adjuvant chemotherapy. Our study provides a rationale to further evaluate the strategy to increase immune activation to improve outcome in this group of patients. Citation Format: Saranya Chumsri, Daniel J. Serie, Afshin Mashadi-Hossein, Sarah Warren, Alvaro Moreno-Aspitia, Geraldo Colon-Otero, Keith L. Knutson, Edith A. Perez, E. Aubrey Thompson. Correlation of immune co-stimulatory molecule OX40 and outcome in trastuzumab treated HER2-positive breast cancer patients in the NCCTG-N9831 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1682. doi:10.1158/1538-7445.AM2017-1682