Mechanisms of action of 3’-deoxyadenosine in treating clear cell renal cell carcinoma

Abstract

Although treatment strategies for advanced and metastatic clear cell renal cell carcinoma (ccRCC) have markedly evolved with the recent use of immune-checkpoint inhibitor (ICI)-based combinations, most patients eventually develop resistance to these therapies. Therefore, there is still an urgent need for the development of effective treatment options. NUC-7738, a novel ProTide transformation of the nucleoside analogue 3ﹶ-deoxyadenosine, releases 3ﹶ-deoxyadenosine monophosphate (3’-dAMP) in cells which is then phosphorylated to the di- (3’-dADP) and tri-phosphate forms (3’-dATP). 3’-dAMP might have the ability to activate AMP-activated protein kinase (AMPK), a key cellular energy sensor, and thus disrupt metabolic homeostasis in cancer cells. Furthermore, 3’-dATP might interfere with RNA synthesis affecting protein expression and survival of cancer cells. The ability of NUC-7738 to activate AMPK through phosphorylation of Th172 was tested in ccRCC cell lines and ex vivo tissue slices of ccRCC from patients. The effect of NUC-7738 on mRNA synthesis and polyadenylation was investigated in two ccRCC cell lines, 786-O and 769-P. AMPK activation by NUC-7738 showed inter-replicate variability and inter-patient variability in ccRCC cell lines and ex vivo tissue slices, respectively, indicating the complexity of the regulation of AMPK phosphorylation. Moreover, mass spectrometry analysis showed that 3’-dATP is the main active metabolite of NUC-7738. Transcriptome data analysis showed mitochondrial gene transcripts of electron transport chain (ETC) complexes were the most significantly altered in both 786-O and 769-P cell lines, with lower expression levels in response to NUC-7738 treatment. This was accompanied by downregulation of the protein expression of ETC complexes subunits. NUC-7738 induced the intrinsic pathway of apoptosis in these cells through the release of cytochrome c from mitochondria and the subsequent activation of caspases -9 and -7. These data suggest that NUC-7738 might inhibit tumour cells growth and proliferation through the inhibition of mitochondrial respiration and the subsequent induction of apoptosis.