Anti-Neoplastic Cytotoxicity of Gemcitabine-(C(4)-amide)-[anti-HER2/neu] in Combination with Griseofulvin against Chemotherapeutic-Resistant Mammary Adenocarcinoma (SKBr-3)

INTRODUCTION: Gemcitabine is a pyrimidine nucleoside analog that becomes triphosphorylated and in this form it competitively inhibits cytidine incorporation into DNA strands. Diphosphorylated gemcitabine irreversibly inhibits ribonucleotide reductase thereby preventing deoxyribonucleotide synthesis. Functioning as a potent chemotherapeutic, gemcitabine decreases neoplastic cell proliferation and induces apoptosis which accounts for its effectiveness in the clinical treatment of several leukemia and carcinoma cell types. A brief plasma half-life due to rapid deamination, chemotherapeuticresistance and sequelae restricts gemcitabine utility in clinical oncology. Selective “targeted” gemcitabine delivery represents a molecular strategy for prolonging its plasma half-life and minimizing innocent tissue/organ exposure. METHODS: A previously described organic chemistry scheme was applied to synthesize a UV-photoactivated gemcitabine intermediate for production of gemcitabine-(C(4)-amide)-[anti-HER2/neu]. Immunodetection analysis (Western-blot) was applied to detect the presence of any degradative fragmentation or polymerization. Detection of retained binding-avidity for gemcitabine-(C(4)-amide)-[anti-HER2/neu] was determined by cell-ELISA using populations of chemotherapeutic-resistant mammary adenocarcinoma (SKBr-3) that highly over-express the HER2/neu trophic membrane receptor. Anti-neoplastic cytotoxicity of gemcitabine-(C(4)-amide)-[anti-HER2/neu] and the tubulin/microtubule inhibitor, griseofulvin was established against chemotherapeutic-resistant mammary adenocarcinoma (SKBr-3). Related investigations evaluated the potential for gemcitabine-(C(4)-amide)-[anti-HER2/neu] in dual combination with griseofulvin to evoke increased levels of anti-neoplastic cytotoxicity compared to gemcitabine-(C(4)-amide)-[anti-HER2/neu]. RESULTS: Covalent gemcitabine-(C(4)-amide)-[anti-HER2/neu] immunochemotherapeutic and griseofulvin exerted anti-neoplastic cytotoxicity against chemotherapeutic-resistant mammary adenocarcinoma (SKBr-3). Covalent gemcitabine-(C(4)-amide)-[anti-HER2/neu] immunochemotherapeutic or gemcitabine in dual combination with griseofulvin created increased levels of anti-neoplastic cytotoxicity that were greater than was attainable with gemcitabine-(C(4)-amide)-[anti-HER2/neu] or gemcitabine alone. CONCLUSION: Gemcitabine-(C(4)-amide)-[anti-HER2/neu] in dual combination with griseofulvin can produce enhanced levels of anti-neoplastic cytotoxicity and potentially provide a basis for treatment regimens with a wider margin-of-safety. Such benefits would be possible through the collective properties of; [i] selective “targeted” gemcitabine delivery; [ii] relatively lower toxicity of griseofulvin compared to many if not most conventional chemotherapeutics; [iii] reduced total dosage requirements faciliated by additive or synergistic anti-cancer properties; and [iv] differences in sequelae for gemcitabine-(C(4)-amide)-[anti-HER2/neu] compared to griseofulvin functioning as a tubulin/microtubule inhibitor.