Conjugation of methotrexate to poly (L-lysine) as a potential way to overcome drug resistance

Methotrexate (MTX) was conjugated through a carbodiimide-catalyzed reaction to poly (L-lysines) of various molecular sizes at a ratio of approximately one molecule per 27 lysyl residues. These conjugates were tested on cultured, Chinese hamster, ovary cells known to be drug resistant because of a deficient methotrexate transport. The cellular uptake of conjugated drug far exceeded the uptake of free drug in both drug sensitive and resistant lines. The conjugated drug inhibited the growth of the transport-deficient cells at concentrations at which free drug had no effect. The conjugate failed to inhibit dihydrofolate reductase in vitro. This and other evidence indicate that the strong pharmacologic effect of the MTX-poly(Lys) conjugate is due to the intracellular--presumably intralysosomal--hydrolysis of its polymeric backbone followed by the release inside the cell of an active form of MTX. This conclusion is supported by data obtained with conjugates using poly(D-lysine) as a carrier. This optical isomer is not susceptible to common proteolytic enzymes and MTX-poly(D-Lys) has not growth inhibitory effect whatever on either transport proficient or deficient CHO-cells. MTX-poly(L-lys) can thus be seen as a lysosome-activated drug.