We investigated the mechanisms involved in KHG26377 regulationof glutamate dehydrogenase (GDH) activity, focusing onthe roles of SIRT4 and SIRT3. Intraperitoneal injection of micewith KHG26377 reduced GDH activity with concomitant repressionof glucose-induced insulin secretion. Consistent withtheir known functions, SIRT4 ribosylated GDH and reduced itsactivity, and SIRT3 deacetylated GDH, increasing its activity.However, KHG26377 did not affect SIRT4-mediated ADP-ribosylation/inhibition or SIRT3-mediated deacetylation/activationof GDH. KHG26377 had no effect on SIRT4 protein levels,and did not alter total GDH, acetylated GDH, or SIRT3 proteinlevels in pancreatic mitochondrial lysates. These results suggestthat the mechanism by which KHG26377 inhibits GDHactivity and insulin secretion does not involve ADP-ribosylationof GDH by SIRT4 or deacetylation of GDH by SIRT3.