Estudo pré-clínico do novo fármaco antitumoral LASSBio-1920: toxicidade aguda, farmacocinética e nanoformulação lipossomal

Abstract

Cancer is an important public health problem, with an estimated twenty million new cases by 2025, according to the World Health Organization (WHO). Treatment management depends on the characteristics of the tumor and the patient, which suggests the need for developing effective therapeutic alternatives. LASSBio-1920 is a combretastatin (CA-4) analogous, is an antimitotic compound, and was developed from reactions with N-acylhydrazones. Its cytotoxic activity was confirmed in several tumor cell lines, with emphasis on IC50 corresponding to 0.06 µM in the colorectal cancer line (HCT-116), demonstrating a promising prototype with cytotoxic potency in the nanomolar order. In early in vitro pharmacokinetic studies, it was demonstrated that it is highly absorbed by the gastrointestinal tract and is also able to cross the blood-brain barrier. Furthermore, in the microsomal stability assay, four possible cofactor-dependent metabolites became apparent. Thus, based on the suggestion of continuity in studies to advance the stages of pre-clinical analyses and development of a suitable formulation for in vivo trials, the objective of this work was to evaluate LASSBio-1920 regarding the profile of acute oral and intravenous toxicity in Wistar rats, determine pharmacokinetic parameters in the non compartmental model, develop and characterize a liposomal nanoformulation, followed by evaluation of physicochemical properties and determination of cytotoxic activity. From acute toxicity tests in Wistar rats, using the up and down method, there was no toxicity by the oral route up to the maximum dose of 175 mg/kg. Regarding the evaluation of toxicity by the intravenous route, it was possible to determine the maximum tolerated dose of 27.5 mg/kg, with the appearance of reversible histopathological changes, characteristics of nonspecific reactive hepatitis and mild nephrosis. Then, the determination of the pharmacokinetic data was performed in silico, with simulation in the SimCyp Certara® program, and in vivo, with quantification by the LC/MS technique, with data from up to 6h after IV administration. The simulation indicated first-order pharmacokinetics and increased biodistribution to the kidney in the animal module. Regarding the in vivo parameters, AUC0-inf of 0.32 mg.h/L, clearance of 6.24 L/h.kg, volume of distribution of 53.26 L/kg and t1/2 of 5.91 h were determined. Furthermore, in relation to tissue biodistribution, in a maximum time of 5 min, LASSBio-1920 had high quantification identified in the kidney, lung and adipose tissue. Additionally, in the development of liposomal nanoformulation the parameters indicated particle size (TM) of 113.4 nm, polydispersity index (PDI) of 0.372; and zeta potential (PZ) of -18.1. The most stable formulation showed an encapsulation efficiency (EE) of 85%, preserving the characteristics for longer when stored after freeze-drying and in the absence of humidity. Regarding cytotoxicity, no significant changes were observed in the IC50 of the liposomes, when compared to the free molecule, that is, nanoencapsulation did not interfere with the cytotoxic activity. Therefore, based on our results, it is possible to infer that the drug LASSBio-1920 is well tolerated orally when compared to intravenously and has rapid tissue biodistribution when administered intravenously. The liposomal nanoformulation presented acceptable characteristics in all analyses. Therefore, the new nanoencapsulated LASSBio-1920 prototype could be mandatory for the stages of determining intravenous antitumor efficacy.