Dual potency anti-HER2/neu and anti-EGFR anthracycline immunoconjugates in chemotherapeutic-resistant mammary carcinoma combined with cyclosporin A and verapamil P-glycoprotein inhibition

Abstract

Immunoconjugates of epirubicin were synthesized with monoclonal antibodies against the epidermal growth factor receptors, HER2/neu and EGFR, by creating a sulfhydryl-reactive epirubicin intermediate applying heterobifunctional succinimidyl-4-(N-maleimidomethyl)cyclohexane-1-carboxylate (SMCC), which was introduced at α-monoamide groups of the epirubicin carbohydrate moiety. In parallel, N-succinimidyl-S-acetylthioacetate (SATA) was used to incorporate a sulfhydryl group into immunoglobulin at the terminal amine position of ϵ-lysine amino acid residues. Eprirubicin–SMCC–SATA–IgG immunoconjugates were produced by reacting epirubicin–SMCC and SATA–IgG at appropriate molar ratios. Epirubicin–(anti-HER2/neu) and epirubicin–(anti-EGFR) had greater potency against chemotherapeutic-resistant SKBr-3 mammary carcinoma than did epirubicin at epirubicin-equivalent concentrations. Epirubicin–(anti-HER2/neu) was more potent than epirubicin–(anti-EGFR), and a synergistic level of antineoplastic activity was detected with an epirubicin immunoconjugate 50/50 combination. Competitive P-glycoprotein inhibition with cyclosporin A or verapamil enhanced the potency of the epirubicin immunoconjugate 50/50 combination. Minor levels of antineoplastic activity were detected only with an immunoglobulin 50/50 combination of anti-HER2/neu and anti-EGFR. The investigations represent a potential strategy for enhancing the selective internalization, intracellular deposition, and antineoplastic potency of chemotherapeutics in multidrug-resistant neoplasias.

Keywords::

• Epirubicin
• immunoconjugate
• HER2/neu
• EGFR
• P-glycoprotein
• mammary carcinoma

Acknowledgments

The authors would like to acknowledge Mrs. Toni Jones for laboratory technical assistance and Mr. Tom Thompson in the Office of Agriculture Communications for assistance with the photographic illustration. Research investigations were supported by indirect funds generated from an extramural grant devoted to the development of a vaccine preparation.

Declaration of interest: The authors report no conflict of interest. The authors alone are responsible for the content and writing of the manuscript. The authors do not have and financial or personal relationships with other people organizations that would inappropriately influence the conduction of this research investigation or the integrity of laboratory results or their interpretation.