Discovery of small molecule: Regulates the cryptochrome stability and controls blood glucose levels in diabetic mice

The circadian rhythm controls the behavioral, biochemical, and physiological variables in most living organisms, from bacteria to mammals. The circadian rhythm is an endogenous, the most well-known biological rhythm, and refers to a 24-hour cycle that regulates various physiological processes like hormone secretion, food intake, sleep-wake cycles, body temperature regulation, and metabolism. The mammalian circadian clock mechanism is endogenous, although it is synchronized with the environmental cues through the suprachiasmatic nuclei (SCN) located in the anterior part of the hypothalamus. Circadian clock disruption influences metabolic health and is closely associated with many diseases, including sleep disorders, neurological conditions, immune dysregulation, obesity, diabetes, and cancer. Cryptochromes (CRYs), one of the core clock proteins, are transcriptional repressors of the circadian clock in mammals. The stability of CRYs is important because of the ability to alter the period and amplitude of the circadian rhythm. The gluconeogenesis pathway is regulated by a variety of factors, including hormones such as glucagon and insulin and the circadian clock mechanisms. On the molecular level, CRY interacts directly with the glucagon-mediated G-protein coupled receptors, in turn, inhibits the gluconeogenesis pathway that controls glucose metabolism. Therefore, we aimed to discover small organic molecules which increase the stability of CRYs and inhibit gluconeogenesis. Such molecules could be used as anti-diabetic drugs, which control blood glucose levels. To this end, we used a structure-based drug design approach against the primary pocket of CRY, which is responsible for their degradation. Around 2 million small molecules with non-characterized functions were screened through molecular docking in silico. The candidate small molecules were tested for cellular toxicity and their effects on CRYs stability, circadian clock, and gluconeogenesis. We identified a novel molecule, TW68, which increases the stability of both CRY1 and CRY2, lengthens the period of circadian rhythm, and represses gluconeogenic genes in further characterization of pre-clinical studies. In transgenic and fat-induced diabetic animal models, TW68 regulated the fasting blood glucose levels in biochemical and physiological studies. These results signify the therapeutic potential of TW68 on the circadian clock – gluconeogenesis-related metabolic diseases, including type 2 diabetes mellitus.