PSMA-Targeted Stably Linked “Dendrimer-Glutamate Urea-Methotrexate” as a Prostate Cancer Therapeutic

Abstract

One of the important criteria for achieving efficient nanoparticle-based targeted drug delivery is that the drug is not prematurely released at off-target sites. Here we report the preclinical evaluation of a serum-stable dendrimer-based drug conjugate capable of actively targeting into prostate cancer (PC) cells, delivered through the prostate-specific membrane antigen (PSMA). Multiple molecules of PSMA-binding small molecule glutamate urea (GLA; targeting agent) and the drug methotrexate (MTX) were conjugated to generation 5 PAMAM dendrimer (G5) through Cu-free "click" chemistry. The GLA was conjugated through a stable amide bond, and the MTX was conjugated either through ester (Es)- or amide (Am)-coupling, to generate G5-GLA(m)-(Es)MTX(n) and G5-GLA(m)-(Am)MTX(n), respectively. In serum-containing medium, free MTX was slowly released from "G5-GLA(m)-(Es)MTX(n)", with ~8% MTX released from the dendrimer in 72 h, whereas the MTX on G5-GLA(m)-(Am)MTX(n) was completely stable. The G5-GLA(m)-(Am)MTX(n) bound and internalized into PSMA-expressing LNCaP cells, but not into PSMA-negative PC3 cells. The conjugate-inhibited recombinant dihydrofolate reductase and induced potent cytotoxicity in the LNCaP cells, but not in the PC3 cells. Similar to the action of free GLA, stable amide-linked dendrimer-GLA was capable of inhibiting the enzyme N-acetylated α-linked acidic dipeptidase (NAALADase) activity of PSMA. The G5-GLA(m)-MTX(n) may serve as a serum-stable nanoparticle conjugate to specifically and effectively target and treat PSMA-overexpressing prostate tumors.

Applications of Nanoparticles for Anticancer Drug Delivery: A Review

“…In another example, methotrexate was conjugated either through ester-or amide-coupling PAMAM dendrimer through "click" chemistry. 80 PAMAM dendrimers can also be conjugated to cisplatin. The dendrimer-cisplatin nanoparticles were highly water soluble and released cisplatin slowly in vitro.…”

Dendrimer antibody conjugate to target and image HER-2 overexpressing cancer cells

“…Therefore, dendrimers are attractive macromolecules for drug delivery and molecular imaging [5, 6]. In particular, polyamidoamine (PAMAM) dendrimers have a narrow polydispersity and contain terminal amines that can be conjugated with targeting agents, therapeutics, and imaging agents for drug delivery and in vitro imaging [7–9]. …”

Poly(amidoamine) Dendrimer–Methotrexate Conjugates: The Mechanism of Interaction with Folate Binding Protein

“…1−7 In particular, conjugation of methotrexate (MTX) to poly(amidoamine) (PAMAM) dendrimer has been extensively studied, with over 100 related publications since 2002. 8−19 Acetylated, neutral G5 PAMAM has been of particular interest as a drug delivery vector because of its narrow polydispersity index (PDI), low toxicity and immunogenicity, and molecular weight (MW) of about 30 kDa that allows it to solubilize multiple hydrophobic ligands while still being small enough to diffuse through tissue for cell-level targeting. 20 MTX is a structural derivative and competitive inhibitor of folic acid (FA); 21 therefore, it was proposed that MTX conjugates may also be able to provide increased binding and uptake when multivalently displayed on a polymer scaffold.…”