The medicinal properties of guanidine derivatives

The medicinal properties of guanidine derivatives are also of great interest due to their diverse anti-microbial, anti-inflammatory, anti-viral, and anti-cancer, activities (C and D). Guanidine-containing drugs, such as -iodobenzylguanidine and methylglyoxal bis(guanylhydrazone), were shown several decades ago to have anti-tumor properties and have since been subjected to intense preclinical and clinical evaluation. In our previous work, we reported the design and synthesis of a series of UA derivatives containing heterocyclic moieties at the C-28 position of UA, and one peficitinib (E) was shown to potently inhibit HIF-1α transcriptional activity under hypoxic conditions. Building on this, our ongoing research seeks to identify HIF-1α inhibitors with the potential to be developed as anti-cancer agents. Here, we report on the activities of three series of UA derivatives in which we changed the hydroxyl moiety of UA to an aminoguanidine group at C-3 and simultaneously (i) introduced smaller moieties, such as benzene and alkyl groups, at the C-28 position; (ii) retained the carboxyl group at C-28; or (iii) introduced different substituted benzene rings or heterocyclic groups at the C-2 position. These novel series, totaling 28 compounds, were evaluated for their ability to inhibit hypoxia-induced HIF-1α transcriptional activity. The biological mechanism of action of one selected compound, , was investigated in detail. The synthetic pathway of the target compounds –, –, and – is presented in . The intermediate was synthesized by reacting UA with Jones reagent in acetone at 0 °C. Compounds , , and were prepared by reacting intermediate with aminoguanidine bicarbonate, semicarbazide hydrochloride, and thiosemicarbazide, respectively, in refluxing ethanol. Compounds , , and were prepared by reacting intermediate with methyl iodide, ethyl bromide, and amyl bromide, respectively. Intermediate was reacted with different substituents of benzyl chloride to provide compounds –. Compounds – were prepared by Claisen–Schmidt condensation of intermediate with different aldehydes. Compounds – and – were reacted with aminoguanidine bicarbonate in refluxing ethanol to yield the target compounds – and –. The structures of the desired compounds were characterized by H and C NMR spectroscopy and high-resolution mass spectrometry. The activities of the UA derivatives are summarized in . To examine the effects on hypoxia-induced HIF-1α transcriptional activity, Hep3B cells were transfected with a luciferase reporter construct driven by six hypoxia-response elements and then exposed to hypoxia (1% O). After 24 h, luciferase activity in the supernatants was measured. UA was used as a positive control. We found that most of the tested compounds inhibited HIF-1α transcriptional activity. For the substituted guanidine derivatives 3a (IC > 100 µM), 3b (IC 88.8 µM), and 3c (IC 13.3 µM), the inhibitory activity was in the order O > S > NH. It indicated that the semicarbazide derivatives have good HIF-1α inhibitory activities and merit further investigation as potential anti-cancer agents. For the compounds in series , alkyl groups of different carbon chain lengths were introduced at C-28 of UA. Of these, only compound showed potent anti-cancer activity (IC 6.7 µM), indicating that a methyl group at C-28 improves the anti-cancer activity UA, whereas extending the carbon chain may have the reverse effect. Compounds – which contained different substituents on the phenyl ring, exhibited significant HIF-1α inhibition, with IC values between 10.3 and 40.5 µM. For the chlorinated compounds (IC 30.3 µM), (IC 21.1 µM), (IC 40.5 µM), and (IC 13.3 µM), the inhibitory activity was in the order 2,6-Cl > 4-Cl > 3-Cl > 2,4-Cl. For compounds (IC 21.1 µM), (IC 13.2 µM), and (IC 10.3 µM), electron-donating groups showed beneficial effects compared with electron-withdrawing groups (4-CH > 4-F > 4-Cl). No clear structure–activity relationship could be found in series . Compounds (H), (4-Cl), (3-Br), and (2,4-Cl) exhibited potent anti-cancer activities when a benzene ring was introduced at the C-2 position of UA. Among these compounds, was the most potent inhibitor of HIF-1α transcriptional activity (IC 4.0 µM). Compound also potently inhibited HIF-1α, suggesting that the thiophene moiety is more favorable than furan for inhibition.