Abstract 6128: Evaluation of therapeutic efficacy of combination of 4HPR and SAHA for inhibition of growth of human glioblastoma T98G xenografts in zebrafish
Authors: Khathayer Firas , Kathrein Katie L. , Nagarkatti Mitzi , Ray Swapan K. 
Published: September 2020

Abstract
Glioblastoma is the most aggressive and deadly primary brain cancer in humans. Despite the current treatments using surgery, radiotherapy, and chemotherapy, glioblastoma reappears due to drug resistance. New approaches for treatment of glioblastoma suggest that combination of two drugs may overcome the drug resistance. In this study, we have evaluated the efficacy of the combination of N-(4-hydroxphenyl) retinamide (4HPR) and suberoylanilide hydroxamic acid (SAHA) for inhibition of human glioblastoma T98G cells in a zebrafish xenograft model in our laboratory. Both 4HPR and SAHA have been approved by the United States Food and Drug Administration for treatment of cancers in the clinics. The synthetic retinoid 4HPR works via retinoic acid receptor signaling pathways to inhibit cancer cell proliferation and induce apoptosis, while SAHA is a histone deacetylase (HDAC) inhibitor that targets class I, II, and IV HDACs for controlling growth of cancer cells. Although rodent models have provided valuable insights into the biology and pathology of glioblastoma, these models are frankly unsuitable and increasingly unaffordable for high-throughput screening and profiling of novel therapeutic agents for rapid translation to the clinics. The optical transparency of zebrafish embryos and larvae provides the unique opportunity to track tumors in a way that is impossible in rodent models. Small size of zebrafish embryos allows for study design in small plates, making them a more useful laboratory system than other cancer models. Zebrafish lack a fully functioning adaptive immune response until 4 days post-fertilization, allowing rejection-free xenografts of human tumors during this period. Recently, zebrafish xenograft models have emerged as a new powerful tool for studying different types of human tumors including glioblastoma and for evaluating therapeutic efficacy of different drugs alone and in combination. Organ systems and cell types are common in human and zebrafish. Tumors that occur in the same organs in human and zebrafish look and behave alike, often share common genetic makeups, and employ basic mechanisms for tumor formation, expansion, and spread. Zebrafish xenograft model of human glioblastoma offers us a major opportunity for discovering key pathways of this tumor growth and for evaluating efficacy of a novel combination therapy. We loaded exponentially grown T98G cells labeled with CellTrace™ Red into the glass needle (0.75 mm internal diameter without filament, World Precision Instruments, FL) at a density of about 1 × 106 cells/200 µl. Using a micro-injector, each anesthetized zebrafish larva was microinjected with about 100 T98G cells into the duct of Cuvier at 1.5 - 3.0 psi. After injection, larvae were transferred to fresh egg water for recovery during 1 h at 28 ± 0.5°C and then incubated at 33°C for the rest of the experiment. After 2 days of tumor growth, we treated zebrafish larvae with 4 µM 4HPR and 8 µM SAHA alone and in combination and incubated for 3 days. Results showed that 4HPR and SAHA alone inhibited growth of T98G xenografts in zebrafish, but the combination of the two drugs most significantly reduced T98G cell proliferation inside of the zebrafish. Our work suggested that the zebrafish xenograft model could be useful to identify the therapeutic efficacy of combination of drugs in the treatment of glioblastoma.
Citation Format: Firas Khathayer, Katie L. Kathrein, Mitzi Nagarkatti, Swapan K. Ray. Evaluation of therapeutic efficacy of combination of 4HPR and SAHA for inhibition of growth of human glioblastoma T98G xenografts in zebrafish [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6128.