Development of liposomes conjugated to anti-EGFR monoclonal antibody for selective delivery of oxaliplatin to colorectal cancer cells

Introduction: Oxaliplatin (OxPt) is a chemotherapeutic drug used in the treatment of colorectal cancer, but presents a series of side effects.The epidermal growth factor receptor (EGFR) is a receptor overexpressed in the membrane of cancer cells, which triggers cell proliferation and angiogenesis.Cetuximab is an anti-EGFR monoclonal antibody (Mab) clinically approved to treat colorectal cancer.The immunoliposome technology aims to control drug release, in addition to increasing selective drug delivery, based on the conjugation of the Mab on the liposomal surface.Objectives: This study aimed to develop and evaluate in colorectal cancer cells OxPt immunoliposomes formed by DSPC (1,2-dioctadecanoyl-sn-glycero-3-phosphocholine), CHOL (cholesterol) and DSPE-PEG-Maleimide (distearoylphosphatidylethanolamine associated with polyethylene glycol and maleimide) functionalized with the monoclonal antibody cetuximab. Methodology:The immunoliposomes were functionalized under different temperature and agitation conditions and characterized by particle size, zeta potential, polydispersity index (PDI), encapsulation efficiency (EE%) and conjugation efficiency (EC%).The structural integrity of the antibody was assessed by thermophoresis and electrophoresis.Cellular internalization into the HCT-116 cell line was qualitatively evaluated by confocal microscopy, using the fluorophore 3,3'-Dioctadecyloxacarbocyanine perchlorate (Dio) as a nanoparticle. Results:The selected formulation was the functionalized immunoliposome, incubated for 2 hours at 37C, followed by a period at room temperature without agitation.The immunoliposome results showed mean particle size of 123.96 nm 1.10; PDI of 0.269 0.006; zeta potential of -26.43 mV 0.70; EE% of 67.97 4.71 and EC% of 52,21.In thermophoresis, the cetuximab inflection points were maintained in the immunoliposome samples, which may demonstrate the conjugation of the antibody to the nanoparticle and the maintenance of the structure and stability of the antibody.In electrophoresis, the light and heavy chain bands were maintained in the immunoliposome samples, confirming the presence of the antibody.In the cellular internalization assay by confocal microscopy, the immunoliposomes showed increased internalization over 24 hours in cells that overexpress EGFR. Conclusion:Thus, based on these results, it can be concluded that a formulation with adequate physicochemical characteristics was obtained and that the immunoliposome containing OxPt presented, as expected, high percentage of internalization in colorectal cancer cells with EGFR overexpression.