Design and In vitro Validation of Multivalent Dendrimer Methotrexates as a Folate-targeting Anticancer Therapeutic

Thommey   P.   Thomas; Melvin      Joice; Madhuresh      Sumit; Justin   E.   Silpe; Alina      Kotlyar; Sophia      Bharathi; Jolanta      Kukowska-Latallo; James   R.   Baker; Seok   Ki   Choi

Abstract

Design of cancer-targeting nanotherapeutics relies on a pair of two functionally orthogonal molecules, one serving as a cancer cell-specific targeting ligand, and the other as a therapeutic cytotoxic agent. The present study investigates the validity of an alternative simplified strategy where a dual-acting molecule which bears both targeting and cytotoxic activity is conjugated to the nanoparticle as cancer-targeting nanotherapeutics. Herein, we demonstrate that methotrexate is applicable for this dual-acting strategy due to its reasonable affinity to folic acid receptor (FAR) as a tumor biomarker, and cytotoxic inhibitory activity of cytosolic dihydrofolate reductase. This article describes design of new methotrexate-conjugated poly(amidoamine) (PAMAM) dendrimers, each carrying multiple copies of methotrexate attached through a stable amide linker. We evaluated their dual biological activities by performing surface plasmon resonance spectroscopy, a cell-free enzyme assay and cell-based experiments in FAR-overexpressing cells. This study identifies the combination of an optimal linker framework and multivalency as the two key design elements that contribute to achieving potent dual activity.

Keywords: Folate receptor, methotrexate, dihydrofolate reductase, PAMAM dendrimer, multivalent binding, drug delivery.

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