Optimization of biguanide derivatives as selective antitumor agents blocking adaptive stress responses in the tumor microenvironment

Abstract

Adaptive cellular responses resulting from multiple microenvironmental stresses, such as hypoxia and nutrient deprivation, are potential novel drug targets for cancer treatment. Accordingly, we focused on developing anticancer agents targeting the tumor microenvironment (TME). In this study, to search for selective antitumor agents blocking adaptive responses in the TME, thirteen new compounds, designed and synthesized on the basis of the arylmethylbiguanide scaffold of phenformin, were used in structure activity relationship studies of inhibition of hypoxia inducible factor (HIF)-1 and unfolded protein response (UPR) activation and of selective cytotoxicity under glucose-deprived stress conditions, using HT29 cells. We conducted luciferase reporter assays using stable cell lines expressing either an HIF-1-responsive reporter gene or a glucose-regulated protein 78 promoter-reporter gene, which were induced by hypoxia and glucose deprivation stress, respectively, to screen for TME-targeting antitumor drugs. The guanidine analog (compound 2), obtained by bioisosteric replacement of the biguanide group, had activities comparable with those of phenformin (compound 1). Introduction of various substituents on the phenyl ring significantly affected the activities. In particular, the o-methylphenyl analog compound 7 and the o-chlorophenyl analog compound 12 showed considerably more potent inhibitory effects on HIF-1 and UPR activation than did phenformin, and excellent selective cytotoxicity under glucose deprivation. These compounds, therefore, represent an improvement over phenformin. They also suppressed HIF-1- and UPR-related protein expression and secretion of vascular endothelial growth factor-A. Moreover, these compounds exhibited significant antiangiogenic effects in the chick chorioallantoic membrane assay. Our structural development studies of biguanide derivatives provided promising candidates for a novel anticancer agent targeting the TME for selective cancer therapy, to be subjected to further in vivo study.

Molecular features of biguanides required for targeting of mitochondrial respiratory complex I and activation of AMP-kinase

“…Only biguanides that enter mitochondria and inhibit complex I activate AMP kinase. Our results can be applied in the design of new complex I-targeted biguanides with improved anti-proliferative activity [ 39 ], and to assist in deconvoluting the targets and mechanisms of action of anti-hyperglycemic biguanides like metformin.…”

Anticancer properties of novel pyrazole‐containing biguanide derivatives with activating the adenosine monophosphate‐activated protein kinase signaling pathway

“…It was observed that HL156A significantly downregulated p‐mTOR and enhanced p‐AMPK expression . Furthermore, the new biguanide derivatives that were designed and synthesized on the basis of the arylmethylbiguanide scaffold of phenformin showed significantly stronger inhibition on cancer cell growth than phenformin and exhibited remarkable antiangiogenic effects . We report here for the first time that compounds 10b and 10d , by the introduction of side chains containing chloro‐substituted phenylpyrazole, are more beneficial for enhancing activity than the previous modified derivatives.…”

“…Pyrazole‐containing biguanide derivatives were prepared by modifying the procedure reported in the literature . All reagents used in the synthesis were obtained commercially and used without further purification, unless otherwise specified.…”

Development of antitumor biguanides targeting energy metabolism and stress responses in the tumor microenvironment

“…Phenformin is also considerably potent than metformin with respect to these biological effects 22 , 23 . Therefore, we have focused on the arylbiguanide scaffold and developed new arylbiguanides with higher activity than phenformin by screening based on selective cytotoxicity during glucose deprivation 24 . These arylbiguanide derivatives led to significant suppression of hypoxia-induced factor-1 (HIF-1)- and UPR-dependent transcriptional activation, the expression of various ER stress-related proteins and c-Myc, as well as angiogenesis 24 – 27 .…”

“…Therefore, we have focused on the arylbiguanide scaffold and developed new arylbiguanides with higher activity than phenformin by screening based on selective cytotoxicity during glucose deprivation 24 . These arylbiguanide derivatives led to significant suppression of hypoxia-induced factor-1 (HIF-1)- and UPR-dependent transcriptional activation, the expression of various ER stress-related proteins and c-Myc, as well as angiogenesis 24 – 27 . Cancer cells have various adaptive response systems, including HIF-1 and UPR signaling pathways, which enable them to survive under hypoxia and low nutrition stresses in the tumor microenvironment (TME) 28 .…”