New prodrugs of metformin do not influence the overall haemostasis potential and integrity of the erythrocyte membrane

Abstract

Although metformin, an oral anti-diabetic drug, has been found to have multidirectional effects over the past decade, it is characterised by unfavourable pharmacokinetic properties. This study discusses the effects of metformin, phenformin and three prodrugs of metformin on the haemostasis and integrity of Red Blood Cells (RBCs). The influence of examined biguanide derivatives on haemostasis was evaluated spectrophotometrically by clot formation and lysis test (CL-test) at 405nm. The extrinsic and intrinsic coagulation pathway were examined by measuring the PT (Prothrombin Time) and aPTT (Activated Partial Tromboplastin Time). Haemolysis assay, microscopy and flow cytometry studies were used to assess the effect of the tested compounds on RBCs. Although none of the tested biguanide derivatives significantly influenced the overall potential of clot formation and fibrinolysis (CL constants), statistically significant changes were seen in the values of the kinetic parameters of fibrinolysis. Furthermore, only prodrug 2, with an 8-carbon alkyl chain, unfavourably affected RBCs by interaction with the erythrocyte membrane leading to significant haemolysis. Our results provide a further insight into the effects of metformin and its prodrugs on haemostasis and RBCs and underscore the necessity for further research.

Novel Sulfonamide-Based Analogs of Metformin Exert Promising Anti-Coagulant Effects without Compromising Glucose-Lowering Activity

“…All other synthesized compounds contributed to the significant increase in APTT and TT. Results presented in this study as well as those obtained in a previous one [ 22 ], in which metformin did not appear to affect APTT and PT, revealed that benzenesulfonamides with methyl substituents in the aromatic ring significantly affected the intrinsic coagulation pathway and the process of fibrin polymerization, and thus, may exert beneficial effects on plasma hemostasis. Importantly, also other sulfonamide-based metformin analogs, such as those with an o - or p -nitro substituent in the aromatic ring exert highly beneficial anti-coagulation properties, manifested by prolonged PT and APTT [ 41 ].…”

“…Only a derivative with a propyl chain in the aromatic ring (compound 5 ) at a concentration of 1.5 µmol/mL contributed to hemolysis reaching the value of 9.99 ± 2.95%. In our previous study, metformin did not exert any effect on the integrity of erythrocyte membrane over the entire concentration range (0.006–1.5 µmol/mL) [ 22 ]. For instance, one-hour incubation of human RBC with metformin at 1.5 µmol/mL resulted in 1.81 ± 0.78% hemolysis [ 22 ].…”

“…Based on the results of basic coagulation tests, and the effects on integrity of RBCs membrane, compounds 2 , 3 , 4 , 6 and 10 were examined for further in-depth insight into anti-coagulation potential using the CL-test. This assay allows to estimate in vitro effects of the compounds on the overall potential of clot formation and fibrinolysis (CL AUC ) as well as several kinetic parameters of the clot formation phase, clot stabilization and fibrinolysis [ 22 ].…”

Pleiotropic Activity of Metformin and Its Sulfonamide Derivatives on Vascular and Platelet Haemostasis

“…Metformin, and its derivatives 1-4 were tested at the concentration range 0.006-1.5 µmol/mL, based on therapeutic plasma concentration of metformin (0.129 to 90 mg/L [38], which is 0.8 nmol/mL-0.6 µmol/mL) as well as our previous studies [32,34]. The exception was cell viability studies, where higher concentrations were used to determine IC 50 values.…”

“…Sulfonamide-based compounds ( Figure S1, Supplementary Materials) were found to exhibit beneficial effects on plasma, vascular and platelet haemostasis, which is frequently impaired in T2DM patients [32][33][34]. For instance, o-nitrobenzenesulfonamide was found to demonstrate more prominent effects on certain plasma and vascular haemostasis parameters than the parent drug metformin [32][33][34]. The beneficial effects of sulfenamide and sulfonamide derivatives of metformin on plasma and vascular haemostasis have been summarized in Figure S1 (Supplementary Materials).…”