The long-term objective of this project is to understand the nature and significance of the vasopressin (VP) gene-related products present in small-cell carcinoma (SCCL), and to use this understanding to develop more rational approaches to treatment. Our data show VP gene-related products are a constant features of all small-cell tumors and result from the expression of both normal and abnormal VP genes. This expression gives rise not only to VP, a mitogen, but also to Neurophysin-Related cell-Surface Antigen (NRSA) that can be targeted in patients with antibodies. The hypotheses being tested are that NRSA and/or VP receptors will provide sensitive and reliable targets for the detection, diagnosis, and effective treatment of SCCL, and that expression of VP gene(s) is essential for tumor growth and survival.
Specific Aims are focused on either NRSA proteins or VP receptors, and are directed towards (i) using known targets on NRSA proteins in a new screening method for SCCL; (ii) identifying new targets through completing our characterization of NRSA proteins and the mRNAs giving rise to them; (iii) evaluating antibodies against known and newly discovered targets on NRSA proteins for their ability to provide sensitive and reliable immunotargeting of SCCL in vivo; (iv) ascertaining the involvement in VP-induced mitogenesis of the three, and perhaps four, VP receptor sub-types expressed by SCCL; (v) determining the potential value of VP receptor subtypes as diagnostic/prognostic markers and targets for treatment from their distribution and abundance in SCCL; (vi) determining how VP gene over-expression, and under-expression, influences cell growth in vitro and in vivo, and; (vii) assessing the ability of VP antagonists to target SCCL in vivo and influence growth. The methods employed in our approach to achieving these Specific Aims will include ABC immunohistochemistry, RT-PCR, cloning, DNA sequencing, Edman degradation, antibody generation, Northern and Western analysis with densiometric quantitation, radioimmunoassay, tumor-directed targeting with antibodies and receptor antagonists in mice, whole-body scintigraphy for 131Iodine and 99mTechnetium, cytofluorographic and radiometric quantitation, radioligand binding, pulse-chase of radiolabel, flow cytometry, vector-mediated stable transfections, and cell and tumor growth assessments. They are intended to provide both qualitative and quantitative evaluations. Short term goals are focused on developing and testing methods that can then be translated to patient care within the time-frame of the requested funding. The information obtained is expected to result in effective chemotherapeutic and immunotherapeutic interventions for all patients with limited and extensive SCCL.
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