SBIR-STTR Award

Phase I and II clinical trials with BR96-doxorubicin, an immunoconjugate that recognizes receptors on human carcinomas, have demonstrated that the monoclonal antibody component, BR96, is capable of safely delivering active doxorubicin to tumor masses, albeit at concentrations that are sub-optimal. We propose to construct and test significantly improved conjugates consisting of highly potent drugs attached to BR96 through a new generation of optimized peptide-based linkers. The resulting conjugates should be stable in serum, but labile inside tumor cell lysosomes, leading to the release of active drug at the target site. The drugs will consist of two classes. Minor groove binders containing a distamycin unit will be attached to the DNA alkylator cyclopropylpyrroloindole, forming a construct that will covalently modify the DNA of target cancer cells. The second drug will be combretastatin A4, a potent antimitotic agent that acts both on tumor cells and tumor vasculature. It is expected that conditionally stable conjugates prepared with these agents will be potent and capable of effecting antitumor activities at biologically relevant doses. The aims of the proposed study are to synthesize potent drug derivatives, link them to BR96 and to a ,monoclonal antibody against the CD40 antigen, and evaluate their stability characteristics in vitro cytotoxic activities, and in vivo toxicities and activities in nude mice with human tumor xenografts. PROPOSED COMMERCIAL APPLICATIONS: There is a very large unmet clinical need for treating carcinomas of the breast, lung, colon, and prostate. The BR96 antibody recognizes the Lewis-Y antigen, which is widely expressed on these tumors. Using the BR96 antibody for the delivery of potent drugs to tumors may lead to pronounced anticancer activity with acceptable levels of systemic toxicity. This would constitute a major advancement in the clinical treatment of cancer.

In light of the efficacy of several new clinically approved monoclonal antibodies (mAbs) for cancer therapy, there has been a resurgence of interest in their use for the delivery of cytotoxic drugs to tumor cells. Recent studies have identified critical parameters that must be addressed for therapeutic efficacy, including intracellular uptake of the mAb within target cells, use of potent and stable drugs for delivery, and incorporation of conditionally stable linkage systems that allow for the selective release of active drug. In our Phase I SBIR study, we addressed these issues, and the work led to the development of cAC1O-valine-citrulline- monomethylauristatin E (cAC1O-Val-Cit-MMAE), a conjugate composed of three elements: the cAC1O mAb that recognizes the CD30 antigen on lymphomas, the Val-Cit peptide linker that is cleaved from the drug by lysosomal enzymes, and the highly potent antimitotic agent, MMAE. The conjugate, which contained 8 drugs/mAb, was stable in plasma, released drug in the presence of cathepsin B, and induced cures and regressions of established tumors with unprecedented therapeutic windows. We subsequently found that these conjugates were prone to aggregation and had accelerated in vivo clearance, in contrast to equally efficacious conjugates containing only 4 drugs/mAb (cAC10-Val-Cit-MMAE4). However, the current procedure for producing cAC10-Val-Cit-MMAE4 leads to a complex mixture of conjugates with varying degrees of drug substitution at different positions on the mAb. The objective of the Phase II SBIR research is to establish an optimized form of cAC10-Val-Cit-MMAE4 in terms of in vitro potency and specificity, in vivo safety and efficacy, pharmacokinetics and biodistribution, product uniformity, and ease of production. This will be accomplished by exploring a variety of chemical conjugation strategies for the preparation of cAClOVal- Cit-MMAE4. In parallel, we will engineer cACIO with cysteine to serine replacements, thereby enabling the construction of conjugates with precisely defined sites and stoichiometries of drug attachment. These studies will lead to the identification of a highly optimized form of cAC10-Val-Cit-MMAE4 for clinical trials in humans with Hodgkin's disease, anaplastic large cell lymphoma, and cutaneous T-cell lymphoma