Monoclonal Antibody Conjugates of Doxorubicin Prepared with Branched Linkers:  A Novel Method for Increasing the Potency of Doxorubicin Immunoconjugates

Abstract

Immunoconjugates of monoclonal antibody BR96 and Doxorubicin have been prepared using a novel series of branched hydrazone linkers. Since each linker bound to the mAb carries two DOX molecules, the DOX/mAb molar ratios of these conjugates were approximately 16, twice that of those previously prepared with single-chain hydrazone linkers. The conjugates were stable at a physiological pH of 7, but released DOX rapidly at lysosomal pH 5. The branched series of BR96 conjugates demonstrated antigen-specific cytotoxicity, and were more potent in vitro than the single-chain conjugate on both a DOX (4-14-fold) and mAb (7-23-fold) basis. The results suggest that, by using the branched linker methodology, it is possible to significantly reduce the amount of mAb required to achieve antigen-specific cytotoxic activity. In this paper, the synthesis and in vitro biology of branched chain immunoconjugates are described.

Synthesis of a Covalent Epirubicin-(C3-amide)-Anti-HER2/neuImmunochemotherapeutic Utilizing a UV-Photoactivated Anthracycline Intermediate

“…. Such findings are also analogous to the greater 27,29,35,42,43,101,102,127 or relatively high (effective) 39 levels of cytotoxic anti-neoplastic potency noted for similar covalent anthracycline-immunochemotherapeutics synthesized under optimal conditions when evaluated against other neoplastic cell types applying standardized anthracyclineequivalent concentrations.…”

Effects of Drug Loading on the Antitumor Activity of a Monoclonal Antibody Drug Conjugate

“…For example, doxorubicin-antibody-drug conjugates displayed a correlation between drug loading and cell cytotoxicity in vitro (20,26). The use of branched linkers increased molar ratios as high as 16 doxorubicin molecules per mAb, which additionally improved the in vitro potency without affecting antigen binding (20). The activities of TA.1-maytansinoid conjugates with an average ranging from one to six drugs per mAb were compared in vitro, and optimal cytotoxicity was achieved with an average of four drugs per mAb (16).…”

“…These include the use of highly potent drugs that are attenuated and stable while attached to the mAbs (12)(13)(14)16) and the use of drug-mAb linkers that allow for the release of active drug only when the mAb has reached the target site (12,(17)(18)(19)(20). Another key factor is the choice of target antigen.…”

“…Historically, augmenting the potency of an antibody-drug conjugate involved increasing the number of drugs on an antibody (20) or conjugating more potent drugs (12). Although previous reports describe how drug loading influences in vitro characteristics (16,20,26), the effects of drug multiplicity on in vivo parameters of antibody-drug conjugates has not been described. In the present work, we have evaluated the effect of drug loading on the in vitro and in vivo properties of cAC10 antibody-drug conjugates.…”

Prodrug chemotherapeutics bypass p-glycoprotein resistance and kill tumors in vivo with high efficacy and target-dependent selectivity

“…Several approaches have been developed in order to deliver specifically Doxorubicin to tumor cells using monoclonal antibodies (mAbs) or small peptides as carrier molecules (Arap et al, 1998;Nagy et al, 1998;King et al, 1999;Stan et al, 1999;Garsky et al, 2001;Langer et al, 2001). Targeted chemotherapy couples a drug to a carrier molecule (ligand), and depends on the ability of the ligand to bind selectively to the target tumor cells in such a way that it may increase the drug concentration near the tumor while decreasing systemic toxicity.…”