Abstracts from the New England Section of the AUA 2020: A Virtual Experience

© The Canadian Journal of Urology TM : International Supplement, August 2020 Scientific Session VI: Stones II/ Basic Science 64 Sendai Virus as a Novel Oncolytic Virus for Urothelial Carcinoma AndrewJ.Charap,BS 1 ,JohnHeard,MS 2 ,MatthewLin,BS 1 ,JorgeDaza,MD 1 ,JohnSfakianos, MD 1 , Amir Horowitz, PhD 1 1 Icahn School of Medicine at Mount Sinai, New York, NY; 2 SUNY Downstate College of Medicine, New York, NY Introduction: The use of oncolytic viruses is an active area of investigation in the field of urologic oncology. Several clinical trials have been completed investigating recombinant adenovirus as a second-line treatment for non-muscle invasive bladder cancer after BCG- failure. Sendai virus is a murine paramyxovirus with no innate virulence in humans but with documented oncolytic activity and tumor cell specificity. Sendai virus treatment has been investigated in several murine solid tumor models, but to date has not been studied in bladder cancer. Materials & Methods: We modified the genome of the Sendai virus to express green- fluorescent protein (GFP) as an indicator of successful viral infection. We then infected the urothelial carcinoma cell line 639V with Sendai virus at various titrations. Three days post-infection we performed a flow cytometric analysis to quantify oncolytic activity and changes in cell-surface protein expression. Finally, we infected primary urothelial carcinoma cells derived from pathologic T1, high grade urothelial tumor in vitro . Results: At three-days post-infection, we observed GFP fluorescence in 639V cells exposed to the Sendai virus at titers greater than 1000 infectious units (IU). Successful viral infection, defined as a positive GFP-signal during cell acquisition via flow cytometry, directly correlated with increasing viral titer (Figure 1). Higher viral titers associated with increased 639V cell death. We observed upregulation of the immune checkpoint PD-L1, in addition to increased expression of the MHC class I proteins HLA-A, B, and C (Figure 2). We observed gross fluorescence of primary tumor cells 72-hours post-infection (Figure 3) using fluorescence microscopy. Conclusions: Functionally wild-type Sendai virus was capable of both infecting and killing 639V urothelial carcinoma cells in vitro . Flow cytometric analysis revealed dynamic changes in expression of important surface proteins after infection, including PD-L1. These proof-of-concept experiments demonstrate the feasibility of Sendai virus as an alternative to adenovirus for the treatment of non-muscle invasive bladder cancer. Figure 1: 639V cells exposed to GFP-expressing Sendai virus were analyzed 72 hours post-infection to determine successful infection and cell death. A : Values indicate the median fluorescence intensity (MFI) of 10000 639V cells at a given quantity of infectious units (IU). B : Cell death was determined using an amine viability dye. C: Histograms displayingMFI of HLA-A/B/C and PD-L1 of 639V cells 72-hours post exposure to Sendai virus, gated on live cells. miRNA Expression Profile Predicts Local Invasion in cT1 Renal Cancer James Trussler, MD, MS 1 , Kareem Alazem, MD 1 , Thomas Kalantzakos, BS 1 , Travis Sullivan, MS 1 , Eric Burks, MD 2 , Carmen Sarita-Reyes, MD 2 , Kimberly Rieger-Christ, PhD 1 , David Canes, MD 1 1 Lahey Hospital and Medical Center, Burlington, MA; 2 BostonMedical Center, Boston, MA, USA Introduction: Clinical staging of incidentally discovered renal masses largely relies on radiographic size criteria and gross invasion of venous structures or perirenal fat. However, early involvement of segmental veins or microscopic fat invasion in clear cell renal carcinoma (ccRCC) is not adequately detected on imaging leading to upstaging at the time of surgery. This has been estimated to occur in as many as 14% of patients with an associated 40% increase in mortality risk. Additionally, the incidence of pre-operative renal mass biopsy is increasing as active surveillance becomes more common. MicroRNA (miRNA) has been implicated as an early aberration identifiable in cancerous tissues. We therefore sought to identify alterations in miRNA expression profiles of clinical T1 (cT1) ccRCC which could predict pathologic upstaging to pT3 at the time of surgery. Materials &Methods: Seventy eight age and Fuhrman grade matched patients with cT1 renal masses were identifiedwho had undergone partial or radical nephrectomy for ccRCC at our institution, 40 pT1 and 38 pT3. Two patients were excluded due to synchronous tumours, and 2 patients were excluded due to the pre-operative presence of metastases. miRNA was isolated from the remaining patients using the Qiagen FFPE AllPrep RNA isolation kit. An array of 751 candidate miRNAs were screened via qRT-PCR using a balanced subset of 20 patient samples. Results: miRNA screening identified 24 candidate miRNA with differential expression profiles between patients with pT1 and pT3 disease (Figure 1). 20 of these miRNA were previously identified as potential mediators of ccRCC progression while the remaining 4 have been implicated in other cancers including prostate, breast, colorectal, and lung. miR-26a-5p, -26b-5p, -24-3p, and -25-3p were the best predictors of pT1 versus pT3 disease (all p<.001, q<.05) with AUC >0.9. The ROC for miR-26a-5p is shown in Figure 2. Conclusions: The differentiation of renal masses into subsets by probability of progression may aid in more appropriate and informed selection of patients for both surgery and active surveillance. As renal mass biopsy becomes more common, the development of novel techniques to examine factors influencing invasion may improve this selection process. We identified 24 candidate miRNAwhich could form a profile predictive of local invasion in ccRCC. Phase 2 of this study seeks to identify and validate a smaller subset of these miRNA as a predictive profile of pathologic upstaging, therefore creating a tool to aid in surgical decision making. 63 28 Figure 2: Gross fluorescence of Sendai-infected primary tumor cells derived from a pathologic T1, high-grade tumor after cystectomy, visualized using fluorescence microscopy.