Poly[N-(2-hydroxypropyl)methacrylamide] conjugates of bovine seminal ribonuclease. Synthesis, physicochemical, and preliminary biological evaluation

The synthesis of three conjugates of poly(HPMA) with bovine seminal ribonuc lease (BS-RNase) differing in their structure is described. Two conjugates contained BS-RNase conjugated with the polymer via functional group situate d at the end of the polymer chain (star-shaped conjugate I) or attached to the poly(HPMA) carrier via biodegradable oligopeptide spacers randomly dist ributed along the polymer chain ("classic" conjugate II). These two conjuga tes differ in structure, molecular weight, and molecular weight distributio n. In addition, a conjugate combining the activity of two compounds, BS-RNa se and doxorubicin, both attached to the same polymer chain via biodegradab le spacers was synthesized ("classic" conjugate III). Biological activity o f all BS-RNase conjugates was compared with that of free BS-RNase and to th e polymer-bound anticancer drug doxorubicin (conjugate IV). Unlike the bovi ne pancreatic ribonuclease (RNase A), BS-RNase displays a potent antitumor activity when tested in vitro and, if administered intratumorally, also in vivo. BS-RNase in its polymer-conjugated forms (conjugates I, II and III) t ested on various human tumor cell lines has lost at least part of its antit umor activity. In in vivo experiments (nude mice bearing human melanoma), i ntratumoral (i.t.) therapy with BS-RNase or with its conjugates II and III showed a significant antitumor effect. Intravenous (i.v.) application of fr ee BS-RNase was totally ineffective, while both BS-RNase conjugates II and III caused significant inhibition of tumor growth. BS-RNase bound to a star -shaped polymer (conjugate I) and administered i.t. or i.v. at the same con centration showed very high toxicity. Our results demonstrate that modifica tion of BS-RNase with poly(HPMA) can prevent it from degrading or inactivat ing events occurring in blood vessels after intravenous application, signif icantly enhancing its potential for therapeutical application.