(490e) Dendrimer-Based Nanodevices for Targeted Cancer Therapy Based on Mechanistic Understanding of Device Performance

Innovations in chemistry are resulting in a large variety of drugs, yet delivering them with maximal therapeutic benefit and minimal side effects remains a scientific and medical challenge. Dendritic polymers (size ~ 5 nm) are emerging as promising candidates for novel targeted drug delivery vehicles, because they present a large density of tailorable functional groups at the periphery. Their nanoscale branching architecture and significantly smaller size provides several advantages over current delivery vehicles, such as linear polymers, nanoparticles and liposomes. Even though folate-dendrimers have been shown to be effective in targeting tumors with drugs, this has not translated to significant improvements in therapeutic effectiveness. The dendrimer surface charge can play a key role in the cellular uptake and release of conjugated drugs. There is a knowledge gap relating to how the dendrimer surface charge and arrangement of the targeting ligand affect the cellular/tumor uptake and intracellular drug release. Such mechanistic understanding will lead to the development of drug delivery vehicles with significantly improved efficacy. We have investigated the effect of dendrimer end-functionality on the activity of polyamido amine (PAMAM) dendrimer-methotrexate (MTX) conjugates in MTX-sensitive and MTX-resistant human acute lymphoblastoid leukemia (CCRF-CEM) and Chinese hamster ovary (CHO) cell lines. Two amide-bonded PAMAM dendrimer-MTX conjugates were prepared using a dicyclohexylcarbodiimide (DCC) coupling reaction; one between a carboxylic acid terminated G2.5 dendrimer and the amine groups of the MTX (Conjugate A) and another between an amine-terminated G3 dendrimer and the carboxylic acid group of the MTX (Conjugate B). Our studies suggest that Conjugate A showed an increased drug activity compared to equimolar amount of free MTX toward both sensitive and resistant cell lines, whereas Conjugate B did not show significant activity on any of the cell lines. In spite of substantially impaired MTX transport by MTX-resistant CEM/MTX and RII cells, Conjugate A showed sensitivity increases of approximately 8- and 24-fold (based on IC50 values), respectively, compared to free MTX. These are the first polymer-MTX conjugates that show increased sensitivity, even without any targeting moiety. Co-incubation of the cells with adenosine and thymidine along with either Conjugate A or MTX resulted in almost complete protection, suggesting that the conjugate achieves its effect on dihyrofolate reductase (DHFR) enzyme, through the same mechanism as that of MTX. The differences in cytotoxicity of these amide-bonded conjugates may be indicative of differences in the intracellular drug release from the cationic dendrimer (Conjugate B) versus the anionic dendrimer (Conjugate A), perhaps due to the differences in lysosomal residence times dictated by the surface functionality. These findings demonstrate the feasibility of using dendrimers as drug delivery vehicles for achieving higher therapeutic effects in chemotherapy, especially in drug resistant cells. To investigate the effect of the linker on the intracellular drug release, we have synthesized ester-linked and peptide-linked dendrimer-MTX conjugates. Once the linker is established, dendrimer-MTX-folic acid conjugates (targeting nanodevices) will be synthesized, and will be tested in already established leukemic tumor models that over-express the folate receptor. References:

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