Molecular Subtype Profiling of Urothelial Carcinoma Using a Subtype-Specific Immunohistochemistry Panel

BACKGROUND: Improved therapies are needed for patients with invasive urothelial carcinoma (InvUC). Tailoring treatment to molecular subtypes holds promise, but requires further study, including studies in pre-clinical animal models. Naturally-occurring canine InvUC harbors luminal and basal subtypes, mimicking those observed in humans, and could offer a relevant model for the disease in people. OBJECTIVE: To further validate the canine InvUC model, clinical and tumor characteristics associated with luminal and basal subtypes in dogs were determined, with comparison to findings from humans. METHODS: RNA sequencing (RNA-seq) analyses were performed on 56 canine InvUC tissues and bladder mucosa from four normal dogs. Data were aligned to CanFam 3.1, and differentially expressed genes identified. Data were interrogated with panels of genes defining luminal and basal subtypes, immune signatures, and other tumor features. Subject and tumor characteristics, and outcome data were obtained from medical records. RESULTS: Twenty-nine tumors were classified as luminal and 27 tumors as basal subtype. Basal tumors were strongly associated with immune infiltration (OR 52.22, 95%CI 4.68–582.38, P = 0.001) and cancer progression signatures in RNA-seq analyses, more advanced clinical stage, and earlier onset of distant metastases in exploratory analyses (P = 0.0113). Luminal tumors were strongly associated with breeds at high risk for InvUC (OR 0.06, 95%CI 0.01 –0.37, P = 0.002), non-immune infiltrative signatures, and less advanced clinical stage. CONCLUSIONS: Dogs with InvUC could provide a valuable model for testing new treatment strategies in the context of molecular subtype and immune status, and the search for germline variants impacting InvUC onset and subtype.

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Epigenetic regulation of mammalian Hedgehog signaling to the stroma determines the molecular subtype of bladder cancer

eLife ◽

10.7554/elife.43024 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 5

Author(s):

SungEun Kim ◽

Yubin Kim ◽

JungHo Kong ◽

Eunjee Kim ◽

Jae Hyeok Choi ◽

...

Keyword(s):

Bladder Cancer ◽

Urothelial Carcinoma ◽

Sonic Hedgehog ◽

Hedgehog Signaling ◽

Molecular Mechanisms ◽

Molecular Subtype ◽

Early Stage ◽

Human Bladder Cancer ◽

Human Bladder ◽

Stromal Response

In bladder, loss of mammalian Sonic Hedgehog (Shh) accompanies progression to invasive urothelial carcinoma, but the molecular mechanisms underlying this cancer-initiating event are poorly defined. Here, we show that loss of Shh results from hypermethylation of the CpG shore of the Shh gene, and that inhibition of DNA methylation increases Shh expression to halt the initiation of murine urothelial carcinoma at the early stage of progression. In full-fledged tumors, pharmacologic augmentation of Hedgehog (Hh) pathway activity impedes tumor growth, and this cancer-restraining effect of Hh signaling is mediated by the stromal response to Shh signals, which stimulates subtype conversion of basal to luminal-like urothelial carcinoma. Our findings thus provide a basis to develop subtype-specific strategies for the management of human bladder cancer.

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Molecular Subtype-Specific Immunocompetent Models of High-Grade Urothelial Carcinoma Reveal Differential Neoantigen Expression and Response to Immunotherapy

Cancer Research ◽

10.1158/0008-5472.can-18-0173 ◽

2018 ◽

Vol 78 (14) ◽

pp. 3954-3968 ◽

Cited By ~ 25

Author(s):

Ryoichi Saito ◽

Christof C. Smith ◽

Takanobu Utsumi ◽

Lisa M. Bixby ◽

Jordan Kardos ◽

...

Keyword(s):

Urothelial Carcinoma ◽

Molecular Subtype ◽

High Grade

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Hypomethylated RRBP1 Potentiates Tumor Malignancy and Chemoresistance in Upper Tract Urothelial Carcinoma

International Journal of Molecular Sciences ◽

10.3390/ijms22168761 ◽

2021 ◽

Vol 22 (16) ◽

pp. 8761

Author(s):

Hao-Lun Luo ◽

Hui-Ying Liu ◽

Yin-Lun Chang ◽

Ming-Tse Sung ◽

Po-Yen Chen ◽

...

Keyword(s):

Urothelial Carcinoma ◽

Epithelial Mesenchymal Transition ◽

Molecular Subtype ◽

Nodal Metastasis ◽

Upper Tract Urothelial Carcinoma ◽

Body Region ◽

Gene Body ◽

Mesenchymal Transition ◽

Upper Tract ◽

Specific Pcr

Ribosome-binding protein 1 (RRBP1) is a potential oncogene in several cancer types. However, the correlation between RRBP1 expression and the prognosis of patients with upper tract urothelial carcinoma (UTUC) remains unclear. In this study, we identified that RRBP1 is associated with carcinogenesis and metastasis in UTUC using a methylation profiling microarray. High correlations between RRBP1 and cancer stages, nodal metastasis status, molecular subtypes, and prognosis in bladder urothelial cancer (BLCA) were found. Aberrant DNA methylation in the gene body region of RRBP1 was determined in UTUC tissues by methylation-specific PCR. RRBP1 expression was significantly increased in UTUC tissues and cell lines, as determined by real-time PCR and immunohistochemistry. RRBP1 depletion significantly reduced BFTC909 cell growth induced by specific shRNA. On the other hand, molecular subtype analysis showed that the expression of RRBP1 was associated with genes related to cell proliferation, epithelial–mesenchymal transition, and basal markers. A patient-derived organoid model was established to analyze patients’ responses to different drugs. The expression of RRBP1 was related to chemoresistance. Taken together, these results provide the first evidence that RRBP1 gene body hypomethylation predicts RRBP1 high expression in UTUC. The data highlight the importance of RRBP1 in UTUC malignancy and chemotherapeutic tolerance.

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APOBEC-mediated mutagenesis in urothelial carcinoma is associated with improved survival, mutations in DNA damage response genes, and immune response

10.1101/123802 ◽

2017 ◽

Cited By ~ 1

Author(s):

Alexander P. Glaser ◽

Damiano Fantini ◽

Kalen J. Rimar ◽

Joshua J. Meeks

Keyword(s):

Gene Expression ◽

Immune Response ◽

Dna Damage ◽

Bladder Cancer ◽

Urothelial Carcinoma ◽

Dna Damage Response ◽

Molecular Subtype ◽

Regulatory Genes ◽

Enrichment Score ◽

Damage Response

AbstractBackgroundThe APOBEC family of enzymes is responsible for a mutation signature characterized by a TCW>T/G mutation. APOBEC-mediated mutagenesis is implicated in a wide variety of tumors, including bladder cancer. In this study, we explore the APOBEC mutational signature in bladder cancer and the relationship with specific mutations, molecular subtype, gene expression, and survival. We hypothesized that tumors with high levels of APOBEC-mediated mutagenesis would be enriched for mutations in DNA damage response genes and associated with higher expression of genes related to activation of the immune system.MethodsGene expression (n=408) and mutational (n=395) data from the Cancer Genome Atlas (TCGA) bladder urothelial carcinoma provisional dataset was utilized for analysis. Tumors were split into “APOBEC-high” and “APOBEC-low” tumors based on APOBEC enrichment score. Analysis was performed with R.FindingsPatients with APOBEC-high tumors have better overall survival compared to those with APOBEC-low tumors (38.2 vs 18.5 months, p=0.005). Tumors enriched for APOBEC mutagenesis are more likely to have mutations in DNA damage response genes (TP53, ATR, BRCA2), and chromatin regulatory genes (MLL, MLL3), while APOBEC-low tumors are more likely to have mutations inFGFR3andKRAS. APOBEC3AandAPOBEC3Bexpression correlates with total mutational burden, regardless of bladder tumor molecular subtype. APOBEC mutagenesis and enrichment is associated with increased expression of immune-related genes, including interferon signaling.InterpretationTumors enriched for APOBEC mutagenesis are more likely to have mutations in DNA damage response genes and chromatin regulatory genes, potentially providing more single-strand DNA substrate forAPOBEC3AandAPOBEC3B, leading to a hypermutational phenotype and the subsequent immune response.HighlightsABPOEC enzymes, particularlyAPOBEC3AandAPOBEC3B, are responsible for the predominant pattern of mutagenesis in bladder cancerTumors enriched for APOBEC-mediated mutagenesis are more likely to have mutations in DNA damage response genes and chromatin regulatory genes, while tumors not enriched for APOBEC-mediated mutagenesis are more likely to have mutations inKRASandFGFR3APOBEC enrichment is associated with upregulation of genes involved in the immune response

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Re: Molecular Subtype-Specific Immunocompetent Models of High-Grade Urothelial Carcinoma Reveal Differential Neoantigen Expression and Response to Immunotherapy

The Journal of Urology ◽

10.1097/ju.0000000000000999.01 ◽

2020 ◽

Vol 203 (6) ◽

pp. 1061-1061

Author(s):

Sam S. Chang

Keyword(s):

Urothelial Carcinoma ◽

Molecular Subtype ◽

High Grade

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Evaluation of basal and luminal subtypes of urothelial carcinoma in African American and non-African American patients.

Journal of Clinical Oncology ◽

10.1200/jco.2015.33.7_suppl.305 ◽

2015 ◽

Vol 33 (7_suppl) ◽

pp. 305-305 ◽

Cited By ~ 3

Author(s):

Jordan Kardos ◽

Jonathan J Melquist ◽

David D. Chism ◽

Woonyoung Choi ◽

Katherine Cockerill ◽

...

Keyword(s):

Breast Cancer ◽

Gene Expression ◽

African American ◽

Urothelial Carcinoma ◽

Molecular Subtypes ◽

Molecular Subtype ◽

Tumor Stage ◽

Intrinsic Subtype ◽

Breast Cancer Patients ◽

Muscle Invasive

305 Background: African American (AA) patients with urothelial carcinoma (UC) have been known to have a worse prognosis even when corrected for variables such as tumor stage and grade. Analysis of gene expression of several malignancies has resulted in the discovery of molecular subtypes with well-defined intrinsic biology. Recent studies in high grade (HG), muscle-invasive UC have led to the identification of two intrinsic, molecular subsets termed “luminal” and “basal” with characteristics of stages of urothelial differentiation, and that remarkably reflect the luminal and basal-like molecular subtypes of breast cancer. Patients with basal-like UC have a significantly worse overall survival. Methods: A total of 215 HG muscle-invasive UC tumors from the MDACC (n=75) and TCGA (n=140) were used to make intrinsic subtype calls using gene expression profiling (MDACC: DASL [cDNA-mediated Annealing, Selection, extension, and Ligation] and TCGA: RNA seq). Basal and luminal subtype calls were derived using previously published subtype classifiers (Damrauer et. al. PNAS, 2014 and Choi et. al. Cancer Cell, 2014). Patients were classified into AA and non-AA (white, Hispanic, or Asian) based upon self-reported race. Results: In total there were 16 and 199 tumors from AA and non-AA patients respectively. In non-AA patients, the proportion of tumors that were classified as basal and luminal were approximately equal (93 and 106 respectively), while in AA patients, there was enrichment of basal tumors (12 basal and 4 luminal) (p=0.03735, Fisher’s exact test). Conclusions: AA patients are enriched in the basal molecular subtype of UC. Similar findings have been previously documented in AA women with breast cancer. The enrichment of basal UC in AAs suggests that a biological explanation may in part underlie the poor outcomes seen in AA patients. Future studies will explore the prognostic and predictive implications of basal subtype in AA patients with UC.

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Genomic analysis of same-patient metachronous upper-tract and bladder urothelial carcinoma.

Journal of Clinical Oncology ◽

10.1200/jco.2018.36.6_suppl.449 ◽

2018 ◽

Vol 36 (6_suppl) ◽

pp. 449-449

Author(s):

Firas G. Petros ◽

Yuan Qi ◽

Woonyoung Choi ◽

Roger Li ◽

Xiaoping Su ◽

...

Keyword(s):

Urothelial Carcinoma ◽

Molecular Subtype ◽

Genomic Analysis ◽

Endoscopic Biopsy ◽

Upper Tract ◽

Formalin Fixed Paraffin ◽

Bladder Urothelial Carcinoma ◽

Formalin Fixed Paraffin Embedded ◽

Group 2 ◽

Next Generation Sequencing Ngs

449 Background: Despite similarities between upper-tract (UTUC) and bladder urothelial carcinoma (BUC), distinctive clinicopathologic and genomic differences are being described. We further investigate the genomic landscape of these two interrelated malignancies in same-patient metachronous (m) UTUC and BUC using next generation sequencing (NGS). Methods: Following institutional board approval, UTUC and BUC samples were obtained from patients via surgical resection or endoscopic biopsy. Tumors were macrodissected from unstained formalin-fixed, paraffin-embedded slides. Study inclusion was untreated patient samples of UTUC and/or BUC divided into 4 groups: 1) UTUC with mBUC, 2) BUC with mUTUC, 3) Synchronous BUC and UTUC, 4) UTUC with no bladder history. Exclusions were for inadequate clinical data or histological tumor purity < 30%. Whole transcriptome RNA sequencing was performed and analyzed using BASE47 panel (includes basal, luminal, p53-like and cell cycle genes). Results: A total of 95 (UTUC = 61, BUC = 34) samples from 40 patients were analyzed. UTUC samples were 33 primary ureter and 28 renal pelvis cancer. Median age was 72 years, 68% male, 76% Caucasian, 60% former smokers. Groups samples were: 1) UTUC (n = 19), mBUC (n = 12); 2) BUC (n = 12), mUTUC (n = 9); 3) Synchronous UTUC/BUC (n = 10); and 4) UTUC (n = 23). Unsupervised hierarchical clustering segregated tumors into basal-like and luminal subtypes, with 87.5% of metachronous tumors displaying conserved subtype membership. For the groups with UTUC and BUC, only 3/24 (12.5%) clusters (2 patients in Group 2, and 1 patient in Group 3) had unmatched basal/luminal subtypes. Conclusions: NGS analysis of same-patient metachronous UTUC and BUC shows that the majority stay within the same molecular subtype regardless of chronologic development or anatomic origin. Additional studies are necessary to explore differences that may occur within the subtypes, the role of methylation, and clinical correlates.

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