Metabolic dysfunction and impairments in the DNA Damage Response: dissecting a pathomechanistic link between Microcephalic Primordial Dwarfisms and cancer cachexia

ATR (ataxia telangiectasia and Rad3-related) encodes master regulator of the DNA damage response ATR. Hypomorphic mutations in ATR result in microcephalic primordial dwarfism disorder Seckel Syndrome (SS). ATR-SS also presents with an apparent lack of subcutaneous fat. This potentially suggests ATR deficiency impairs lipogenic function. This is concerning considering the proposed use of small molecule kinase inhibitors of ATR (ATRis) as cancer chemotherapeutics. ATRi is a highly selective anti-cancer agent due to synthetic lethality in ATMor p53-deficient cells. An invariant feature of cancer is metabolic dysregulation, aggressive cancers can enforce systemic metabolic reprogramming resulting in drastic weight loss. Several screens have identified putative substrates for ATR in insulin signalling and metabolic pathways, indicating uncharacterised roles for ATR may exist here. Using several clinically relevant ATRis, I dissected the metabolic consequences of impaired ATR functionality on adipogenesis and lipogenic function of the 3T3-L1 cell line - widely utilised to investigate adipogenic differentiation. Acute ATRi treatment attenuated transcription of key adipogenic factors in differentiating preadipocytes, resulting in a failure to complete adipogenesis. I treated mature adipocytes chronically with ATRis, observing a striking transdifferentiation process known as browning – fat-storing white adipocytes underwent transcriptional reprogramming towards a thermogenic, brown adipocyte-like status. ATR deficiency generated this phenomenon by impinging on multiple pathways associated with metabolic regulation. I also observed striking cytoplasmic vacuolation and a disrupted autophagy response in every cell line treated with ATRis. I discovered ATR at the ER membrane, where ATRi-induced vacuolisation was derived from swollen endoplasmic reticulum (ER) concomitant with ER stress. I have characterised novel effects of ATR deficiency in adipocyte differentiation and metabolism, and ER and autophagic functionality. Autophagy and lipid metabolism are consistently deregulated in cancers, suggesting these results could lead to the generation of novel synthetic lethality approaches utilising ATRis and compounds targeting metabolic or autophagic pathways.