SBIR-STTR Award

Breast cancer remains one of the top three cancers to affect and cause mortality in women. Major shortcomings with current treatment are the high level of side effects and insufficient efficacy, particularly for patients with metastatic disease. Targeted cancer therapy is designed to treat only the cancer cells and minimize damage to healthy cells. These targets are critical to the tumor's malignant phenotype but not to the host's normal tissues, improving outcomes while minimizing systemic side effects. Anti-HER2, Herceptin, leads the way for targeted therapy in breast cancer with a definite efficacy in a specific but small subset of patients. There is a clear need for additional therapeutic options including other novel targeted therapies. The 88 kDa autocrine growth factor PC-Cell Derived Growth Factor (PCDGF), also known as granulin precursor (GP88), is a novel and preclinically validated candidate of choice as it plays a critical role in breast cancer cell biology, exemplified by the following: 1) GP88 is an autocrine growth/survival factor for breast cancer cells, 2) increased GP88 expression in breast cancer cells is associated with increased tumorigenicity, 3) GP88 mediates tumor cell angiogenesis and invasiveness, 4) breast cancer cells overexpressing GP88 are resistant to current therapies - Tamoxifen, Faslodex, doxorubicin and Herceptin, 5) increased GP88 expression in breast cancer tissue correlates with clinical parameters of poor prognosis while normal and benign breast tissue are negative, 6) patients with poor prognosis have elevated GP88 serum levels. These results highlight GP88 as a novel targeted therapy of breast cancer via the development of anti-GP88 therapeutic neutralizing monoclonal antibodies (Mabs). We have generated a large library of mouse monoclonal antibodies specific to GP88 using a variety of GP88-specific immunogens. Initial preliminary results indicate that an anti-GP88 antibody can abrogate GP88 functional activity. This phase I SBIR application is focused on fully characterizing this Mabs library in order to identify the Mabs with the optimal characteristics to serve as development candidates. The Specific Aims are: 1. Characterize the binding kinetics and specificity of anti-GP88 Mabs by Biacore analysis. 2. Identify Mab's with maximal efficacy and potency in relevant in vitro functional cellular models. 3. Determine efficacy and potency in nude mouse xenograft tumorigenicity models. At the conclusion of this Phase I, we will have identified a mouse Mab (or Mabs) with the optimal in vitro and in vivo pre-clinical efficacy and potency for targeting breast cancer. These Mabs will require further development activities as potential therapeutic candidates in order to be considered for clinical trials in breast cancer during phase II. The first step will be to generate a mouse-human chimeric Mab from the mouse Mab candidate selected during phase I. This chimeric Mab will then be taken further into pre-clinical development. This and other development activities will serve as the basis for a Phase II SBIR application as a basis for the development of novel biological therapy for breast cancer. Breast cancer remains one of the top three cancers to affect and cause mortality in women. Major shortcomings with current treatment are the high level of side effects induced in patients and insufficient efficacy, particularly for patients with advanced disease. The innovative breast cancer therapy to be developed in this proposal will target a mechanism inherent in breast cancer but avoid the side effects associated with many current breast cancer therapies

Breast cancer remains one of the top three cancers to affect and cause mortality in women. Major shortcomings with current treatment are the high level of side effects induced in patients and insufficient efficacy, particularly for patients with metastatic disease. Targeted cancer therapy is designed to treat only the cancer cells and minimize damage to healthy cells. These targets are critical to the tumor's malignant phenotype but not to the host's normal tissues, improving outcomes while minimizing systemic side effects. Anti-HER2, Herceptin, leads the way for targeted therapy in breast cancer with a much lower incidence of side effects and definite efficacy in a specific but small subset of patients. However, there is a clear need for additional therapeutic options including other novel targeted therapies. The 88 kDa autocrine growth factor called PC-Cell Derived Growth Factor (PCDGF), also known as granulin precursor (GP88), has been shown to play a critical role in breast cancer cell biology, exemplified by the following: 1) GP88 is an autocrine growth/survival factor for breast cancer cells, 2) increased GP88 expression in breast cancer cells is associated with increased tumorigenicity, 3) GP88 mediates tumor cell angiogenesis and invasiveness, 4) breast cancer cells overexpressing GP88 are resistant to current therapies - Tamoxifen, Faslodex, doxorubicin, 5) increased GP88 expression in breast cancer tissue correlates with clinical parameters of poor prognosis while normal and benign breast tissue are negative, 6) patients with poor prognosis have elevated GP88 serum levels. These experiments show the essential role that GP88 plays in breast cancer cell tumorigenesis and that the inhibition of GP88 expression or function leads to inhibition of tumor growth. These results highlight the importance of GP88 for targeted therapy of breast cancer via the development of anti-GP88 therapeutic neutralizing monoclonal antibodies (Mabs). Our Phase I research proposed to evaluate a library of murine anti-GP88 for antigen binding properties and for biological activity in GP88-dependent in vitro and in vivo functional tumor cell models. These Phase I specific aims were accomplished and resulted in the identification of anti-GP88 neutralizing Mabs with the in vitro and in vivo potency and efficacy to be candidates for targeted therapy in breast cancer to address a vitally important unmet clinical need. We have selected one Mab, AG1, for further pre-clinical development activities, specifically the generation and characterization of a recombinant Mab (chimerized and humanized Mabs). The following Specific Aims are proposed for this Phase II SBIR application: 1. Cloning of the cDNA encoding the variable sequences of the light chain and heavy chain of the antibody expressed by the AG1 hybridoma cells and generation of expression vectors for both a humanized (HuAG1) and a mouse-human chimeric antibody (ChAG1). 2. Transient expression of the HuAG1 and ChAG1 chimeric antibodies and validation of their binding properties. The antigen binding characteristics and biological activities of the cloned, expressed HuAG1 and ChAG1 will be evaluated and compared with the murine AG1 Mab in order to select candidate recombinant AG1 that fit acceptance criteria for further development. 3. Generate stable HuAG1- or ChAG1-producing cell clones. HuAG1 or ChAG1 selected above will be stably expressed in CHO cells. The HuAG1- or ChAG1-producing clones will be compared for cell growth performance, antibody productivity, and metabolic profile in order to select a candidate clone for scale-up production. 4. Express, purify and characterize the biological properties of the HuAG1 or ChAG1 Mab. The Mab will be evaluated in relevant in vitro and in vivo functional tumor cell models and compared directly with the murine AG1 Mab. GP88 represents a novel, pre-clinically validated target for breast cancer. At the conclusion of this research in this Phase II project, we will have generated a mouse/human chimeric Mab with the in vitro and in vivo pre-clinical efficacy and potency for targeted therapy of breast cancer and we will have developed an optimized clonal antibody production cell line suitable for transfer to a CMO for contract manufacturing. However, once this chimeric anti-GP88 Mab has been generated and characterized it will require further pre-clinical development activities (e.g. formulation, pharmacokinetics and safety) before IND filing for clinical trials in breast cancer. These additional development activities will serve as the basis for a subsequent continuation/renewal Phase II SBIR application.

Public Health Relevance:
Breast cancer remains one of the top three cancers to affect and cause mortality in women. Major shortcomings with current treatment are the high level of side effects induced in patients and insufficient efficacy, particularly for patients with advanced disease. The innovative breast cancer therapy to be developed in this proposal will target a mechanism inherent in breast cancer but avoid the side effects associated with many current breast cancer therapies.

Thesaurus Terms:
"(8s-Cis)-10-[(3-Amino-2,3,6-Trideoxy-Alpha-L-Lyxo-Hexopyranosyl)Oxy]-7,8,9,10-Tetrahydro-6,8,11-Trihydroxy-8-(Hydroacetyl)-1-Methoxy-5,12-Naphthacenedione; (Z)-2-[4(1,2-Diphenyl-1-Butenyl)-Phenoxyl]-N,N-Dimethylethanamine; 1-P-Beta-Dimethylamino-Ethoxyphenyl-Trans-1,2-Diphenylbut-1-Ene; 14-Hydroxydaunomycin; 5,12-Naphthacenedione, 10-((3-Amino-2,3,6-Trideoxy-Alpha-L-Lyxo-Hexopyranosyl)Oxy)-7,8,9,10-Tetrahydro-6,8,11-Trihydroxy-8-(Hydroxyacetyl)-1-Methoxy-, (8s-Cis)-; Address; Adriamycine; Adverse Effects; Affect; Antibodies; Antibody Formation; Antibody Production; Antibody Response; Autocrine Systems; Benign; Binding; Binding (Molecular Function); Biologic Characteristic; Biological; Biological Characteristics; Blood Serum; Body Tissues; Breast Cancer Cell; Breast Tissue; Cho Cells; Cancer Treatment; Cancer Of Breast; Cancers; Cell Line; Cell Lines, Strains; Cell Model; Cellline; Cells; Cellular Expansion; Cellular Growth; Cellular Biology; Cellular Model; Characteristic, Biologic; Chinese Hamster Ovary Cell; Clinical; Clinical Trials; Clinical Trials, Unspecified; Clone Cells; Cloning; Complementary Dna; Contracting Opportunities; Contracts; Dna, Complementary; Dox; Development; Disease; Disorder; Doxorubicin; Doxorubicina; Drug Formulations; Drug Kinetics; Ethanamine, 2-(4-(1,2-Diphenyl-1-Butenyl)Phenoxy)-N,N-Dimethyl-, (Z)-; Faslodex; Faslodex (Fulvestrant, Ici182780); Faslodex(Ici 182,780); Forecast Of Outcome; Formulation; Formulations, Drug; Gfac; Genentech Brand Of Trastuzumab; Generalized Growth; Generations; Growth; Growth Agents; Growth Factor; Growth Factors, Proteins; Growth Substances; Herceptin; Hoffman-La Roche Brand Of Trastuzumab; Human; Human Breast Cancer Cell; Human, General; Hybridomas; Hydroxyl Daunorubicin; Hydroxyldaunorubicin; In Vitro; Incidence; Libraries; Light; Malignant Cell; Malignant Neoplasm Therapy; Malignant Neoplasm Treatment; Malignant Neoplasms; Malignant Tumor; Malignant Tumor Of The Breast; Malignant Neoplasm Of Breast; Mammals, Mice; Mammary Gland Parenchyma; Mammary Gland Tissue; Man (Taxonomy); Man, Modern; Mediating; Metabolic; Mice; Molecular Interaction; Mortality; Mortality Vital Statistics; Murine; Mus; Normal Tissue; Normal Tissue Morphology; Oncogenesis; Outcome; Pc Cell-Derived Growth Factor; Pcdgf; Patients; Performance; Pharmacokinetics; Phase; Photoradiation; Play; Principal Investigator; Production; Productivity; Prognosis; Programs (Pt); Programs [publication Type]; Property; Property, Loinc Axis 2; Recombinants; Research; Resistance; Roche Brand Of Trastuzumab; Role; Sbir; Sbirs (R43/44); Safety; Serum; Small Business Innovation Research; Small Business Innovation Research Grant; Tam; Tamoxifen; Therapeutic; Tissue Growth; Tissues; Treatment Side Effects; Tumor Cell; Tumorigenicity; Validation; Woman; Advanced Disease; Angiogenesis; Antibody Biosynthesis; Anticancer Therapy; Antigen Binding; Autocrine; Base; Cdna; Cancer Cell; Cancer Therapy; Cell Biology; Cell Growth; Chimeric Antibody; Clinical Efficacy; Clinical Investigation; Cultured Cell Line; Design; Designing; Disease/Disorder; Experiment; Experimental Research; Experimental Study; Expression Vector; Granulin Precursor; Immunoglobulin Biosynthesis; Improved; In Vivo; Innovate; Innovation; Innovative; Malignancy; Malignant Breast Neoplasm; Malignant Phenotype; Neoplasm/Cancer; Neoplastic Cell; Neutralizing Mab; Neutralizing Monoclonal Antibodies; Novel; Ontogeny; Outcome Forecast; Overexpression; Pre-Clinical; Preclinical; Programs; Public Health Relevance; Research Study; Resistant; Scale Up; Side Effect; Social Role; Therapy Adverse Effect; Treatment Adverse Effect; Tumor; Tumor Growth; Tumorigenesis"