Conjugation of poly(styrene-co-maleic acid) derivatives to the antitumor protein neocarzinostatin: pronounced improvements in pharmacological properties.

An anticancer agent of intermediate molecular weight and having both a hydrophilic and hydrophobic nature was developed by utilizing the antitumor protein neocarzinostatin (NCS; Mr = 12000) as a prototype drug. The modification was achieved by reacting the two amino groups on NCS with an anhydride group of partially half-esterified (p-E-) or partially hydrolyzed (p-H-) poly(styrene-co-maleic anhydride) (SMA) in 0.8 M NaHCO3. The SMA samples with narrow molecular weights distributions (Mw = ca. 2000) were prepared by copolymerizing styrene and maleic anhydride in cumene followed by fractionation by means of a column-elution method. The derivatives p-E- or p-H-SMA were then formed by using the appropriate monoalcohols or H2O, respectively. These SMA derivatives contain about 2 mol of anhydride residues/mol of SMA. The reaction product, SMA-conjugated NCS (designated as SMANCS), was purified by dialysis followed by gel filtration with Sephadex G-75. The complete reaction yielded essentially a single product, biantennary SMANCS. The molecular weight of the pure SMAMCS was estimated by various methods, including polyacrylamide gel electrophoresis with NaDodSO4, HPLC in the gel permeation mode, fluorescence polarization, and a decrease in both nitrogen and protein contents. These results agree with the apparent molecular weight of about 16000. Characters of SMANCS was considerably altered from that of parental NCS: solubility characteristics in both organic and aqueous solvents were changed, the biological half-life in blood was prolonged 10 times, and antitumor activity became more pronounced, but the toxicity was reduced to one-fourth of the parental NCS. Thus, the present study has provided a method of improving biologically active substances by polymer conjugation.