Endocrine Abstracts

Class 2a HDACs (i.e. HDAC 4, 5, 7 and 9) are involved in the regulation of gluconeogenesis and accordingly, their inhibition has been shown to result in lower blood glucose and improved pyruvate tolerance in diabetic mice. However, pan-inhibition of HDACs is not a viable approach for chronic treatment of type 2 diabetes (T2D) due to induction of severe side effects in various tissues. We now have investigated in vitro and in vivo efficacy and safety of a liver-selective HDAC knock-down via single or combinatorial siRNAs for HDAC 4, 5 or 7. In mouse and human hepatocytes, siRNAs directed against HDAC4, 5 or 7, as well as the combination of all three siRNAS, led to a selective knock-down of the respective HDAC mRNA(s) by about 80-90%. In parallel, genes involved in the regulation of gluconeogenesis, namely G6PC and PCK1, were significantly down-regulated by 70–80% in primary hepatocytes. Those siRNAs were used either alone or in dual or triple combinations (0.75 mg/kg) to treat healthy 9-week old C57BL/6J mice with five intravenous injections for 25 days, as control served either PBS, or a non-silencing control siRNA. After the 4th injection, data from an intraperitoneal pyruvate tolerance test (PTT) in 16 h fasted mice showed no significant differences in glucose excursion. Furthermore, no significant effects on fasting blood glucose and plasma insulin were observed between treatment groups. Quantitative real-time PCR demonstrated a significant hepatic downregulation of the respective HDACs mRNA by 60–80%. However, no reduction of the gluconeogenic genes PCK1 and G6PC was detected in livers of mice. During microscopic examination of key tissues, adverse findings were increased hematopoiesis in spleens of 15/49 mice and chronic purulent pyelonephritis in individual animals (5/49) treated with the different targeting siRNAs. In contrast, spleens and kidneys of mice treated with the non-silencing control siRNA displayed no adverse findings. While the pyelonephritis may indicate an ascending bacterial infection after immunosuppression, the pathomechanism of increased hematopoiesis remains unclear. In summary, although active in cell culture, liver-targeted siRNA knockdown of class 2a HDACs did not translate in an inhibition of genes regulating gluconeogenesis in vivo. Lack of glucose lowering properties in vivo as well as the detection of increased hematopoiesis hamper a clear path forward for development of this approach for treatment of T2D.