Treatment with connexin 46 siRNA suppresses the growth of human Y79 retinoblastoma cell xenografts in vivo.

ABSTRACT

Tumors with a hypoxic component, including human Y79 retinoblastoma cells, express a specific gap junction protein, Connexin 46 (Cx46), which is usually only found in naturally hypoxic tissues such as the differentiated lens. The aim of this study was to investigate if Cx46 downregulation would suppress Y79 tumor formation in vivo. Five-week old nude mice were subcutaneously implanted with human Y79 retinoblastoma cells and treated with intratumor siRNA injections of 30 µg Cx46 siRNA (n = 6), 30 µg non-silencing siRNA (n = 6), or no siRNA treatment (n = 6) every 2 days for a maximum of 10 treatments. Tumor volume (TV) was calculated from the recorded caliper measurements of length and width. Excised tumors were measured and weighed. Western blot analyses were performed to evaluate Cx46 and Cx43 expression in tumors which received Cx46 siRNA, non-silencing siRNA, or no siRNA treatment. Tumor histopathology was used to assess tumor features. Cx46 siRNA treated Y79 tumors had a reduced TV (287 mm(3) ± 77 mm(3)) when compared to the tumors of mice receiving the negative control siRNA (894 mm(3) ± 218 mm(3); P ≤ 0.03) or no siRNA (1068 mm(3) ± 192 mm(3); P ≤ 0.002). A 6-fold knockdown of Cx46 and a 3-fold rise in Cx43 protein expression was observed from western blots of tumors treated with Cx46 siRNA compared to mice treated with non-silencing siRNA. Knockdown of Cx46 with siRNA had an antitumor effect on human Y79 retinoblastoma tumors in the nude mouse model. The results suggest that anti-Cx46 therapy may be a potential target in the future treatment of retinoblastoma.