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570 Background: Over 8900 Canadians are diagnosed with bladder cancer every year, ranking it the fifth most frequent cancer. It can manifest as either non-muscle invasive bladder cancer (NMIBC) or muscle invasive bladder cancer (MIBC). The majority of patients initially receive a diagnosis of NMIBC, although high-grade NMIBC has a 50–70% recurrence rate and 10–30% chance at progressing to MIBC. The transition from high-grade NMIBC to MIBC is poorly understood, and there are currently no accurate biomarkers that predict disease progression. We propose a comprehensive molecular characterization to pinpoint specific copy number alterations (CNAs) related to either MIBC or NMIBC, and hence define the molecular development. Methods: This study analyzed a public dataset from MSKCC and 30 bladder cancer patient samples from Windsor Regional Hospital, both containing NMIBC and MIBC samples. Comprehensive gene sequencing was performed, and CNAs were obtained in over 500 common tumour gene panels. Results: Preliminary data from this study found MIBC may be predicted with 91% accuracy and 95% precision using CNA values of TP53, DDR2 and MLL2. In particular, MIBC correlates with gain of DDR2 or MLL2. Importantly, it has been demonstrated that high expression of DDR2 is associated with a worse prognosis. A panel of bladder carcinoma cell lines were used to validate the DDR2 findings. DDR2 values were quantified across the panels and corresponded with proliferation and invasiveness of cell lines. DDR2 was examined as a possible therapeutic target. Conclusions: Taken together, these findings provide insight to the pathogenesis of muscle invasion in bladder cancer. The potential to identify "genomic triggers" for the transition was facilitated by creating a genetic profile at these two stages.

RNA sequence analysis of emerging SARS-CoV-2 infection is valuable for tracking viral evolution and developing novel diagnostic tools. Furthermore, SARS-CoV-2 sequence analysis can provide insight into potential antigenic drift events that lead to strain speciation and changing clinical outcomes. In this work, we aim to develop a pipeline using next-generation sequencing (NGS) technology in addition to machine learning/bioinformatics to track the accumulation of mutations and viral evolution.

(1) Background:One of the most common cancers that affect North American men and men worldwide is prostate cancer. The Gleason score is a pathological grading system to examine the potential aggressiveness of the disease in the prostate tissue. Advancements in computing and next-generation sequencing technology now allow us to study the genomic profiles of patients in association with their different Gleason scores more accurately and effectively. (2) Methods: In this study, we used a novel machine learning method to analyse gene expression of prostate tumours with different Gleason scores, and identify potential genetic biomarkers for each Gleason group. We obtained a publicly-available RNA-Seq dataset of a cohort of 104 prostate cancer patients from the National Center for Biotechnology Information&rsquo, s (NCBI) Gene Expression Omnibus (GEO) repository, and categorised patients based on their Gleason scores to create a hierarchy of disease progression. A hierarchical model with standard classifiers in different Gleason groups, also known as nodes, was developed to identify and predict nodes based on their mRNA or gene expression. In each node, patient samples were analysed via class imbalance and hybrid feature selection techniques to build the prediction model. The outcome from analysis of each node was a set of genes that could differentiate each Gleason group from the remaining groups. To validate the proposed method, the set of identified genes were used to classify a second dataset of 499 prostate cancer patients collected from cBioportal. (3) Results: The overall accuracy of applying this novel method to the first dataset was 93.3%, the method was further validated to have 87% accuracy using the second dataset. This method also identified genes that were not previously reported as potential biomarkers for specific Gleason groups. In particular, PIAS3 was identified as a potential biomarker for Gleason score 4 + 3 = 7, and UBE2V2 for Gleason score 6. (4) Insight: Previous reports show that the genes predicted by this newly proposed method strongly correlate with prostate cancer development and progression. Furthermore, pathway analysis shows that both PIAS3 and UBE2V2 share similar protein interaction pathways, the JAK/STAT signaling process.

568 Background: Bladder cancer is the fifth most common cancer and eighth leading cause of cancer related-death in North America. It can present as non-muscle invasive bladder cancer (NMIBC) and/or muscle invasive bladder (MIBC). Although genomic profiling studies have established that low-grade NMIBC and MIBC are genetically distinct, high-grade NMIBC can recur and progress to MIBC [ Knowles, M.A. and C.D. Hurst, 2015]. Low grade, non-invasive bladder cancers are characterized by activating mutations in fibroblast growth factor receptor 3 (FGFR3), HRAS or other pathways of receptor kinase activation. High-grade disease, which is often becomes invasive, is characterized by inactivation of TP53 and Rb pathways [Kim, J., et al.]. Finding a subtype of invasive carcinoma with FGFR3 mutation may suggest an alternate pathway by which low grade, non-invasive pathology could transform into invasive disease [Knowles, M.A. and C.D. Hurst, 2015]. Methods: In this study, using a total of 30 bladder cancer (NMIBC and MIBC) patient samples from Windsor Regional Hospital Cancer Program, we performed comprehensive targeted gene sequencing to identify single nucleotide variants, small insertions / deletions, copy number variants and splice variants in over 500 common tumor genes panel. Results: Preliminary data from our study correlates with previously published mutation landscape for NMIBC and MIBC, and includes mutations in EGFR, FGFR3, FGFR4, PIK3CA, CDK6, ALK, JAK, as well as RET. While mutations in AKT1, BRCA1, CCND1, ERBB2, FGFR1, FGFR2, HRAS, and MET appear to be prevalent in NMIBC, mutations in IDH1 and MAP2K2 appear to be more common in MIBC. Three of the samples used in the study are from patients who progressed from high-grade NMIBC to MIBC. Conclusions: Therefore, have the genomic profiling performed at these two stages, which provides a unique ability to identify the potential “genomic triggers” for the transition.

Cyclin J (CycJ) is a poorly characterized member of the Cyclin superfamily of cyclin-dependent kinase regulators, many of which regulate the cell cycle or transcription. Although CycJ is conserved in metazoans its cellular function has not been identified and no mutant defects have been described. In Drosophila, CycJ transcript is present primarily in ovaries and very early embryos, suggesting a role in one or both of these tissues. The CycJ gene (CycJ) lies immediately downstream of armitage (armi), a gene involved in the Piwi-associated RNA (piRNA) pathways that are required for silencing transposons in the germline and adjacent somatic cells. Mutations in armi result in oogenesis defects but a role for CycJ in oogenesis has not been defined. Here we assessed oogenesis in CycJ mutants in the presence or absence of mutations in armi or other piRNA pathway genes. CycJ null ovaries appeared normal, indicating that CycJ is not essential for oogenesis under normal conditions. In contrast, armi null ovaries produced only two egg chambers per ovariole and the eggs had severe axis specification defects, as observed previously for armi and other piRNA pathway mutants. Surprisingly, the CycJ armi double mutant failed to produce any mature eggs. The double null ovaries generally had only one egg chamber per ovariole and the egg chambers frequently contained an overabundance of differentiated germline cells. Production of these compound egg chambers could be suppressed with CycJ transgenes but not with mutations in the checkpoint gene mnk, which suppress oogenesis defects in armi mutants. The CycJ null showed similar genetic interactions with the germline and somatic piRNA pathway gene piwi, and to a lesser extent with aubergine (aub), a member of the germline-specific piRNA pathway. The strong genetic interactions between CycJ and piRNA pathway genes reveal a role for CycJ in early oogenesis. Our results suggest that CycJ is required to regulate egg chamber production or maturation when piRNA pathways are compromised.

ObjectiveTo describe the pattern and frequency of oncogene mutations in white and African American women with endometrial cancer and to determine if racial differences in oncogene mutations exist among women with pathologically similar tumors.MethodsPatients with endometrial cancer from a large urban hospital were identified through medical records, and representative formalin-fixed paraffin-embedded tumor blocks were retrieved. The study sample included 150 patients (84 African Americans) who underwent total abdominal hysterectomy for endometrial cancer. The Sequenom MassARRAY system and the OncoCarta Assay version 1.0 (Sequenom) were used to test for 238 mutations in 19 common oncogenes. The χ2test and the Fisher exact test were used to assess differences in distribution of variables by race and oncogene mutation status.ResultsThere were 20 mutations identified in 2 oncogenes (PIK3CAandKRAS) in tumors from 19 women (12.7%). Most of the mutations were found inPIK3CA(16/20). Thirteen percent of endometrioid tumors harbored mutations (11PIK3CAand 2KRAS) as did 29% of the malignant mixed Mullerian tumors (3PIK3CAand 1KRAS). There were no observed mutations in serous, clear cell, or mucinous tumor types. Among low-grade endometrioid cancers, tumors from African American patients were significantly associated with harboring either aKRASorPIK3CAmutation (P= 0.04), with 7PIK3CAmutations and all 4KRASmutations identified in African American women.ConclusionsThis study provides preliminary evidence that oncogene mutation frequency of some subtypes of histologically similar endometrial carcinoma differ by race. Additional studies are needed to further explore this phenomenon in patients with endometrial carcinoma.

Background: Epidermal growth factor receptor (EGFR) mutations in non-small cell lung cancer (NSCLC) predict response to tyrosine kinase inhibitors. Mutations occur more commonly in never smokers and East Asians, but there are conflicting reports on the frequency of EGFR mutations in tumors from African Americans. Methods: Tumors from 67 African American and 77 white participants in previous case-control studies of lung cancer were selected to determine EGFR mutational status. Mutation analysis was performed using the Sequenom mass array analyzer (Sequenom, San Diego, CA). Results: Overall, 13.9% of the study population carried an EGFR mutation. EGFR mutations occurred in 11.9% of tumors from African Americans compared with 15.6% in whites ( p = 0.53). All mutations found in African Americans were deletions in exon 19. The majority of mutations were found in nonsmokers among both African Americans (7/8) and whites (8/12). Conclusion: These results indicate that African Americans with NSCLC harbor somatic EGFR mutations at a frequency similar to whites with NSCLC. Thus, clinicians should not use race as a clinical decision parameter for the use of EGFR-tyrosine kinase inhibitors.

Activated oncogenes are the genetic drivers of human cancer progression. Oncogenes become activated by structural DNA alterations including gene amplification and over expression, point mutations, and gene translocations/rearrangements. Data from existing targeted cancer treatments clearly show that the best response is observed when the activated oncogene is targeted. On the other hand, among the patients with activated oncogene who respond to a targeted treatment, the level and duration of response is highly variable. This could be explained by the genomic context of an activated oncogene that is unique to any given patient. Therefore, the ability to detect complex oncogene signatures in human cancer specimens is vital for better predictions of the efficacy of targeted agents, and to design therapeutic strategies with multiple targeted agents. In our laboratory, we have been working to identify candidate amplified and point mutated oncogenes in multiple human cancer specimens and cell lines. To obtain individual oncogene signatures, we have used high resolution aCGH coupled with a novel gene expression algorithm that allows for the identification of genes that are over expressed within amplicons in individual specimens. We have also used a high-throughput, mass spectrometry based method for detection of point mutations in oncogenes using the same specimens. These approaches have allowed us to identify tens to hundreds of candidate oncogenes in each specimen which were then refined by on-line computational tools such as Gene Tracker. Using this approach, we have elucidated complex oncogene signatures for over 50 cancer cell lines and specimens. Analysis of these data sets points to highly variable oncogene combinations in individual specimens. In other studies using a subset of breast cancers selected for the presence of the 8p11 amplicon, we found that even in these highly selected specimens, the oncogene signatures were unique to each patient. These signatures make clear predictions about combinations of targeted agents that could be considered. For example, we have observed activation of oncogenes in the 8p11 amplicon to be associated with other oncogenes such the EGFR, cdk4, MYST3, CRKL, and others. Additionally, the presence of PIK3CA mutations occurs variably in these specimens as well. Most of the breast cancer specimens and cell lines in our study have a well-defined signature of oncogenes, and understanding these signatures is important not only for the design of strategies using multiple targeted agents, but also for the proper interpretation of clinical trial results using single targeted agents. Thus, we have developed a high-throughput, cost effective molecular diagnostic platform that can accurately and rapidly identify the causal driving oncogenes in an individual cancer biopsy specimens and therefore can be adapted to analysis of large numbers of patient's specimens. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5099. doi:1538-7445.AM2012-5099

Endometrial cancer is the most common gynecologic cancer, with an estimated 42,000 new cases in 2010. Histologic types of endometrial cancer and stage at diagnosis differ substantially between African Americans (AA) and whites, but AA women experience poorer survival even after accounting for these factors. Additionally, other research suggests this disparity persists after controlling for co-morbid conditions, access to care, and treatment decisions. We hypothesize that there are differences in the molecular biology of tumors from white and AA women, beyond histologic type and grade, which may lead to the increased risk of death seen in AA women after an endometrial cancer diagnosis. After IRB approval, we performed a case-only retrospective analysis of patients with endometrial cancer, for whom we had tumor blocks, diagnosed from 1990-2005 at a single institution. Demographic and clinical variables were obtained from medical record review. Tumor histologic type, stage, and grade were obtained from the Metropolitan Detroit Cancer Surveillance System, part of the SEER program. Tumor histologic type and grade were confirmed by a gynecologic pathologist by slide review. We selected 150 archived tumors, including all high grade cases of any race (35 white, 40 AA), all low grade, endometroid tumors from AA women (n=44), and 31 low grade tumors from white women. Tumor DNA was extracted from formalin fixed paraffin embedded tissue using standard methods. We examined 238 mutations in 19 common oncogenes using the OncoCarta Assay v1.0 (Sequenom). Chi-square tests and Fisher's exact tests were used to assess differences in distribution of clinical and demographic variables by race or oncogene mutation status. We identified 20 mutations in 2 oncogenes in tumors from 19 women (12.7%). The most common mutation was in PIK3CA, the gene coding for the catalytic subunit p110α of PI3K, representing 15/20 (75%) of detected mutations. In PIK3CA, the following mutations were identified: R88Q in the p85-binding domain, E542K, M1043I and P539R in the helical domain, and H1047R in the kinase domain. Previous data suggest these mutations induce a gain of PI3K function and prognosis may vary by location. An additional 4 samples had activating mutations in KRAS (1 G12D, 1 G12V and 2 Q61H). No other mutations were identified. 13% of endometroid tumors harbored mutations, (11 PIK3CA and 2 KRAS). Among tumors from women with low grade endometroid cancers, race was significantly associated with harboring a mutation (p=0.04), with all 7 PIK3CA mutations were identified in AA women. All 4 KRAS mutations were found in AA women (p=0.13). This study provides preliminary evidence that the underlying molecular biology of some subtypes of histologically similar endometrial cancers differ by race. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1137. doi:10.1158/1538-7445.AM2011-1137

Oncogenomic mutations found in NSCLC are potential therapeutic targets, as seen in the dramatic responses individuals with EGFR mutations have to tyrosine kinase inhibitors. We selected 149 patients with NSCLC from previous population-based studies of lung cancer which included 80 (54%) Caucasian samples and 69 African American (46%) samples. Tumor DNA was extracted from formalin fixed paraffin embedded tissue. We examined 238 mutations in 19 oncogenes using the OncoCarta Assay v1.0 (Sequenom). We identified a total of 52 mutations in tumors from 47 individuals; the remaining 102 tumors did not have mutations in the oncogenes we assayed. Mutations were identified in EGFR (n=20), KRAS (n=19), PIK3CA (n=6), PDGFRA (n=2), AKT2 (n=1), FLT3 (n=1) and CDK1 (n=1). Consistent with previous studies, EGFR mutation was more likely to occur in non-smokers (p=3.3E-08 Fisher's test) and the significance of this effect was more pronounced when analyzed against the number of pack years smoked (p=7.3E-08, Kruskal-Wallis test). Previously published reports have suggested that African American patients are less likely to exhibit EGFR mutations compared to Caucasian NSCLC patients and are therefore less likely to respond to tyrosine kinase inhibitors. We found no difference in the proportion of EGFR mutations in African American patients (n=8, 11.6%) compared to Caucasian (n=12, 15.0%, p-value=0.54). Furthermore, EGFR mutations in African American patients were exclusively in exon 19 (n=8, 100%), whereas Caucasian patients had mutations in exon 21 (n=3, 25%) and exon 18 (n=1, 8.3%) in addition to those in exon 19 (n=8, 66.7%). Interestingly, several reports have suggested a higher rate of response of NSCLC to erlotinib, an EGFR tyrosine kinase inhibitor, associated with exon 19 deletions relative to the exon 21 L858R mutation. Thus, our oncogenomic data demonstrates that African American NSCLC patients not only harbor a similar frequency of EGFR mutation as Caucasians, but that the specific nature of the mutation may result in a higher frequency of response to EGFR targeted therapeutics.