Poly(lactic- co-glycolic acid)-loaded nanoparticles of betulinic acid for improved treatment of hepatic cancer: characterization, in vitro and in vivo evaluations

Int J Nanomedicine. 2018 Feb 16:13:975-990. doi: 10.2147/IJN.S157391. eCollection 2018.

Abstract

Purpose: The application of betulinic acid (B), a potent antineoplastic agent, is limited due to poor bioavailability, short plasma half-life and inappropriate tissue distribution. Thus, we aimed to prepare novel 50:50 poly(lactic-co-glycolic acid) (PLGA)-loaded B nanoparticles (BNP) and to compare its anti-hepatocellular carcinoma (HCC) activity with parent B.

Methods: BNP were synthesized and characterized using different methods such as scanning electron microscopy (SEM), fourier-transform infrared (FTIR) spectrometry and particle size analyses. Particle size of BNP was optimized through the application of the stabilizer, polyvinyl alcohol (PVA). The anti-HCC response was evaluated through in vitro cell line study using Hep-G2 cells, confocal microscopy, in vivo oral pharmacokinetics and animal studies. Further, quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis was conducted to observe the changes in the expression of specific genes.

Results: Particle size of BNP was optimized through the application of the stabilizer, polyvinyl alcohol. Physicochemical characterization exhibited particle size of 257.1 nm with zeta potential -0.170 mV (optimized batch B, BNP). SEM and FTIR analyses of BNP showed that cylindrical particles of B converted to spherical particles in BNP and there were no interaction between B and used polymers. The release study of optimized BNP was highest (≥80%) than any other formulation. Later, in vitro cell culture analysis using Hep-G2 cells and confocal microscopy studies revealed that BNP had the highest inhibition and penetration properties than parent B. Oral pharmacokinetics studies using albino Wistar rats at single 100 mg dose again exhibited BNP had the higher 50% of plasma concentration (t1/2), a higher maximum plasma concentration (Cmax) and took longer to reach the maximum plasma concentration (Tmax) than parent B. Next, our in vivo study using nitrosodiethyl amine (NDEA)-induced HCC model documented BNP decreased in number of nodules, restored body weight, oxidative stress parameters, liver marker enzymes and histological architecture than parent B. Lastly, qRT-PCR studies further demonstrated that anti-HCC properties of BNP may be due to over expression of antiapoptotic caspases i.e., caspase 3 and 8.

Conclusion: The prepared BNP showed a better therapeutic response against HCC and could be attributed as future candidate molecule for HCC treatment.

Keywords: HepG2 cells; PLGA-loaded nanoparticles; betulinic acid; caspase-3 and -8; hepatocellular carcinoma.

MeSH terms

  • Administration, Oral
  • Animals
  • Antineoplastic Agents / blood
  • Antineoplastic Agents / chemistry
  • Antineoplastic Agents / pharmacokinetics
  • Antineoplastic Agents / therapeutic use
  • Apoptosis / drug effects
  • Betulinic Acid
  • Carcinoma, Hepatocellular / drug therapy
  • Cell Proliferation / drug effects
  • Chromatography, High Pressure Liquid
  • Drug Carriers / chemistry
  • Drug Liberation
  • Half-Life
  • Hep G2 Cells
  • Humans
  • Inflammation Mediators / metabolism
  • Kinetics
  • Lactic Acid / chemistry*
  • Liver / drug effects
  • Liver / pathology
  • Liver / physiopathology
  • Liver Neoplasms / drug therapy*
  • Liver Neoplasms / pathology
  • Liver Neoplasms / ultrastructure
  • Male
  • Nanoparticles / chemistry*
  • Nanoparticles / ultrastructure
  • Oxidative Stress / drug effects
  • Particle Size
  • Pentacyclic Triterpenes
  • Polyglycolic Acid / chemistry*
  • Polylactic Acid-Polyglycolic Acid Copolymer
  • Rats, Wistar
  • Spectroscopy, Fourier Transform Infrared
  • Static Electricity
  • Triterpenes / blood
  • Triterpenes / chemistry
  • Triterpenes / pharmacokinetics
  • Triterpenes / therapeutic use*

Substances

  • Antineoplastic Agents
  • Drug Carriers
  • Inflammation Mediators
  • Pentacyclic Triterpenes
  • Triterpenes
  • Polylactic Acid-Polyglycolic Acid Copolymer
  • Polyglycolic Acid
  • Lactic Acid
  • Betulinic Acid