Citations

5546 Background: CHFR, an E3 ubiquitin ligase that regulates Aurora A and Pololike Kinase 1, plays a critical role in the cellular response to mitotic stress. In particular, cells containing CHFR arrest in G2 when microtubule dynamics are disrupted. Conversely, low CHFR expression is associated with hypersensitivity to mitotic spindle poisons in breast cancer cell lines. Despite the extensive use of taxanes in front line treatment of epithelial ovarian cancer (EOC), little is known about CHFR expression in this disease. We examined CHFR expression and its relationship with clinical characteristics and outcome in women with EOC. Methods: Pretreatment EOC samples from women enrolled in the Mayo Clinic Biospecimen Resource for Ovarian Cancer Research who underwent initial surgical debulking between 1999-2009 were stained in triplicate with anti-CHFR antibodies. Samples were scored in a blinded fashion for CHFR expression. Associations between CHFR staining and histology, grade, and stage were assessed using chi-squared tests; associations between CHFR and overall survival (OS) or time to disease recurrence (TTR) were assessed using Cox proportional hazards models. Results: Samples from 354 women were stained; median age at diagnosis was 60.5 (range 21-93). At a median follow-up of 67 months, 221 women (62%) had died. Across all patients, moderate/strong CHFR staining was associated with poor OS (HR = 1.39, 95% CI: 1.07-1.81, p = 0.015). Moreover, moderate/strong CHFR expression was strongly associated with serous histology (p < 0.0001), high grade (p < 0.0001) and advanced stage (p = 0.0002). To assess whether the association between CHFR expression and OS was related to taxane sensitivity, a subgroup analysis was performed in the subset of patients (N=131) with high-grade serous EOC who received initial platinum/taxane chemotherapy. Moderate/strong CHFR expression failed to correlate with OS (HR=0.91, 95% CI: 0.57-1.45, p = 0.68) or TTR from the start of chemotherapy (HR = 0.85, 95% CI: 0.55-1.31, p = 0.55) in this subset. Conclusions: CHFR expression is associated with poor OS and adverse clinical characteristics in patients with EOC but does not appear to be related to taxane sensitivity in high-grade serous tumors.

5510 Background: Transcriptional profiling of ovarian cancers has proven to be complex. As such it has been difficult to validate existing signatures across studies. Cancer Genome Atlas (TCGA) researchers have identified four molecular subtypes of high-grade serous ovarian cancer (HGSOC). However, survival duration did not differ significantly for the TCGA subtypes. Potential limitations of the TCGA data include short clinical follow-up (45% were alive at the time of last follow-up) and the need to unify gene expression measures from multiple platforms. Methods: Clinically annotated stage-II–IV HGSOC samples (n = 175) with >70% tumor cell content were profiled using the Agilent Whole Human Genome 4 x 44K chip. To identify subtypes non-negative matrix factorization (NMF) of mRNA expression was performed using ~2000 genes with the highest variability across patients. In parallel differentially expressed genes were identified using the Rosetta Similarity Search Tool as well as analysis of variance based on genes known to be involved in epithelial to mesenchymal transition. Results: Median follow-up time was 35 months (range, 0–202 months, 12% were alive at the time of last follow-up). NMF clustering confirmed four HGSOC subtypes (immunoreactive, differentiated, proliferative, and mesenchymal) on the basis of gene content in the clusters. Pathway signatures with therapeutic potential were identified for individual or multiple subtypes. Survival differed significantly between the four molecular subgroups in univariate (Hazard Ratio [HR] 2.4, 95% CI 1.5-4.1, p = 0.007) and multivariate (HR 2.3, 95% CI 1.3-4.0, p = 0.003) analyses when accounting for age, stage, grade, and postoperative residual tumor. Using the supervised clustering approach two distinct molecular subtypes of HGSOC based on epithelial and mesenchymal gene expression signatures were identified with significantly different survival outcomes. Conclusions: Here we independently validate and expand upon the molecular TCGA classification of HGSOC. The potential of these two prognostic classifiers may lie in their ability to recognize categories of patients that are more likely to respond to particular therapies.

5076 Background: Advanced OVCA should be managed aggressively, and extensive surgery has been the most accepted initial treatment. Medically unfit patients or those with extensive disease, in which complete cytoreduction is unlikely, may not benefit from upfront radical surgery, and neoadjuvant chemotherapy might be an appropriate alternative. Thus, reliable preoperative indicators of surgical outcome are necessary for considering primary surgery vs. neoadjuvant chemotherapy. Our aim is to determine if C-reactive protein (CRP), IL-6, albumin and Glasgow Prognostic Score (GPS – score based on CRP and albumin) correlate with overall survival (OS), length of hospital stay (LOS), surgical morbidity, and suboptimal cytoreduction. Methods: We randomly selected 50 stages III/IV OVCA who underwent surgery as a primary treatment between July 2002 and June 2009 at Mayo Clinic with serum albumin levels and frozen serum available. CRP and IL-6 were measured in stored serum. Univariate and multivariate regression models were fit to evaluate associations with each of the outcomes. Results: Among the 50 patients, the mean age was 67.7 years. 34% had pretreatment albumin <3.5 g/ml, 22.4% had CRP level ≥10 mg/l, 26.5% had IL-6 ≥24 pg/ml and 45% had abnormal GPS score. At 1, 3 and 5 years following surgery, the OS was 75.6%, 49.8% and 36.9%, respectively. RD (0, <1, ≥1cm; p<0.001) was the only independent predictor of OS. Also, IL-6 (p=0.028) and stage (p=0.046) were independently associated with LOS, but no inflammatory or nutritional markers were significant associated with post surgical complications. Stage IV (p=0.019) and elevated CRP (p=0.044) were independent predictors of suboptimal surgery (RD ≥ 1cm). Conclusions: One-third of the patients in our series had low serum albumin at the time of the OVCA diagnosis, and at least one-fourth had elevated inflammatory markers. Advanced stage and elevated inflammatory markers (CRP and IL-6) were independent predictors of longer hospital stay and suboptimal debulking. These pilot data, if confirmed in a larger population, may help in the selection of candidates for neoadjuvant chemotherapy.

20043 Background: Studies have demonstrated that the generation of endogenous immunity to specific tumor antigens is associated with improved prognosis for many cancers. Thus, there has been increased efforts to identify tumor-associated antigens to which there is immunity, as these may be tumor rejection antigens. A candidate antigen is the folate receptor alpha (FRα) which is overexpressed in malignancies, especially breast and ovarian cancer. Prior studies involving ovarian and breast cancer patients have demonstrated the presence of FRα-specific cytotoxic T cells in the tumor-associated lymphocyte population. Methods: Using the RANKPEP CD4 T cell epitope prediction algorithm, we predicted promiscuous immunogenic regions of FRα, and tested for immunity in 30 breast or ovarian cancer patients and 18 healthy volunteer donors in order to attain a better understanding of the levels and extent of the endogenous FRα immune response. Immunity was examined using IFN-γ ELIspot analysis and IgG ELISAs. Results: Fourteen peptides were predicted as potential epitopes to which T cells may respond, seven each from the carboxy- and amino-terminus halves of the protein. Greater than 70% of patients demonstrated T cell immunity to at least one of the fourteen peptides. Patients responded to an average of 3 ± 0.7 peptides while healthy donors responded to only 1 ± 0.5 peptides (p = 0.02). Five peptides were recognized by >25% of patients. Responses to three of these peptides were higher (p > 0.05) in the patients than in healthy donors, suggesting that patients generated immunity upon cancer development. Compared to healthy individuals, patients developed higher immunity to the amino-terminus half of the receptor, suggesting natural epitope spreading (p = 0.03). There was no difference between patients and healthy donors in the immune responses to either non-specific stimuli (p = 0.2) or viral peptides (p = 0.5). Patients had detectable FRα-specific IgG antibody consistent with active FRα-specific helper T cell immunity. Conclusions: These findings demonstrate that the FRα is a target of the immune system in patients with breast or ovarian cancer. Understanding the antigens that are naturally targeted by the immune system may be important for diagnosis, prognosis, and immune-based therapies. No significant financial relationships to disclose.

BACKGROUNDThe prevention of excessive allogeneic red blood cell (RBC) transfusion is an important component of patient blood management initiatives. In this investigation, changes in transfusion behaviors following modification of computerized physician order entry (CPOE) procedures for RBC transfusions to a single‐unit default quantity were assessed.STUDY DESIGN AND METHODSThis is an observational cohort study of adults for whom nonemergency allogeneic RBC transfusions were ordered in the 2 years before and 2 years after the date of modification of the CPOE system to a single‐unit default (June 18, 2015). Changes in the frequency of single‐ versus multiunit RBC transfusion orders and other transfusion metrics were compared between preintervention and postintervention cohorts.RESULTSA total of 52,773 unique transfusion orders for 61,989 RBC units were included, of which 60,045 (96.9%) were transfused. Single‐unit orders increased annually, from 10,404 (74.1%) in the first year to 11,645 (88.6%) in the last year, while multiunit orders decreased by more than half (p < 0.0001). The number of RBC units transfused decreased by 13.9% from 32,528 in the preintervention cohort to 27,497 in the post intervention cohort (p < 0.0001) with an estimated reduction in transfusion‐related expenditures of nearly $4 million. The percentage of transfusions associated with a posttransfusion hemoglobin of10 g/dL or greater decreased by 34.5% (p < 0.0001).CONCLUSIONModification of the CPOE process such that nonemergency RBC transfusion orders were defaulted to a single unit was associated with decreased rates of multiunit RBC transfusion orders, lower transfusion volumes, and substantial cost savings.

Significance Provision of sufficient activating signals can drive self-tolerant T cells to cross-react on self-peptide-MHC in a breakdown of tolerance. This approach has been applied in the development of heteroclitic peptide vaccines, which retain sufficient similarity to self to allow reactivation of effector cells on unaltered self-epitopes but require detailed knowledge about the behavior of individual altered self-peptide antigens. We designed altered-self MHC mutant antigen-presenting molecules to drive CTL capable of cross-reacting on self-peptide-MHC and demonstrate the breaking of self-tolerance to defined and undefined self-antigens. These findings bear particular importance in the field of cancer immunotherapy in which breaking tolerance to weakly antigenic cancer-associated antigens is fundamental to vaccination strategies.

Abstract Background: High Grade Serous Ovarian Cancer (HGSOC) is a highly aggressive form of ovarian cancer accounting for up to 70% of all cases. The Cancer Genome Atlas (TCGA) has further defined 4 molecular subtypes within HGSOC: differentiated (DIF), immunoreactive (IMM), mesenchymal (MES), and proliferative (PRO). The MES subtype is associated with the worst prognosis and the least likely to be completely resectable when compared with other subtypes. A recent study suggested the existence of a 5-th, so-called anti-mesenchymal (ANM), subtype which is characterized by down-regulation of genes typically upregulated in the MES subtype. Importantly, previous molecular subtype studies have been performed using tumor samples consisting of both cancer cells and stroma. However, the origin of MES biomarkers in OC is still largely unknown and several studies suggest that many of the MES biomarkers could arise from cancer-associated stroma rather than cancer cells. Methods: Specimens from 20 patients (MES subtype, n=15; ANM subtype, n=5) diagnosed with HGSOC between 2010 and 2013 were obtained from Pathology Research Core in Mayo Clinic. A total of 60 FFPE blocks from 1 primary tumor and 2 metastatic sites of each patient were used to make the slides. Five MES-subtype biomarkers, including ACTA2, FAP, POSTN, COL5A1, and VCAN, were selected for immunohistochemistry (IHC) to determine the protein localization and intensities. Two researchers scored tumor sections for intensity as 0 (negative), 1 (weak), 2 (moderate), and 3 (strong), and for localization (stroma/tumor, cytoplasm/nuclear). Statistical analysis was performed using GraphPad Prism 7 software and statistical differences were evaluated using t-test. Results: All 5 proteins evaluated exclusively localized in stroma rather than cancer cells (p&lt;0.001 for all 5 proteins). Only FAP showed significantly different staining intensity between MES and ANM subtype of OC patient samples (p= 0.0114). When comparing primary vs. metastatic sites, only COL5A1 showed significant difference (p=0.019). Conclusion: We report that 5 biomarkers traditionally associated with the MES subtype HGSOC arise from cancer-associated stroma instead of cancer cells. Considering the potential roles of these proteins in mediating HGSOC behavior, it may suggest “stroma targeting” as an alternative therapy to “cancer targeting” to improve OC patient outcomes. Although all 5 genes were reported as MES biomarkers in mRNA-based analysis, we found that FAP was the only protein differentially expressed between MES and ANM subtype suggesting it is a better characteristic biomarker for MES subtype. COL5A1 showed significance in staining between primary and metastatic cancer which may indicates the potential role of COL5A1 in metastasis in OC. Citation Format: Qing Zhang, Kimberly R. Kalli, Chen Wang, William Cliby. The mesenchymal subtype biomarkers arise from stroma rather than cancer cells in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4491.

Abstract Background: Endometrioid carcinoma (EC) and clear cell carcinoma (CC) histotypes of epithelial ovarian cancer are understudied compared with the more common high-grade serous carcinomas (HGSC). We therefore sought to characterize EC and CC transcriptomes in relation to HGSC. Methods: Following bioinformatics processing and gene abundance normalization, differential expression analysis of RNA sequence data collected on fresh-frozen tumors was completed with nonparametric statistical analysis methods (55 ECs, 19 CCs, 112 HGSCs). Association of gene expression with progression-free survival (PFS) was completed with Cox proportional hazards models. Eight additional multi-histotype expression array datasets (N = 852 patients) were used for replication. Results: In the discovery set, tumors generally clustered together by histotype. Thirty-two protein-coding genes were differentially expressed across histotype (P &lt; 1 × 10−10) and showed similar associations in replication datasets, including MAP2K6, KIAA1324, CDH1, ENTPD5, LAMB1, and DRAM1. Nine genes associated with PFS (P &lt; 0.0001) showed similar associations in replication datasets. In particular, we observed shorter PFS time for CC and EC patients with high gene expression for CCNB2, CORO2A, CSNK1G1, FRMD8, LIN54, LINC00664, PDK1, and PEX6, whereas, the converse was observed for HGSC patients. Conclusions: The results suggest important histotype differences that may aid in the development of treatment options, particularly those for patients with EC or CC. Impact: We present replicated findings on transcriptomic differences and how they relate to clinical outcome for two of the rarer ovarian cancer histotypes of EC and CC, along with comparison with the common histotype of HGSC. Cancer Epidemiol Biomarkers Prev; 27(9); 1101–9. ©2018 AACR.