Parvoviral Virothereapy for Melanoma

P/ f# 57 -“细小病毒治疗黑色素瘤”

P/F #57 - "Parvoviral Virotherapy for Melanoma" Peter Tattersall, PhD (Laboratory Medicine), Ruth Halaban, PhD (Dermatology) Eligibility Category: Established investigator with no previous work in skin diseases We have identified the orphan parvovirus Lulll as an effective oncolytic virus from a screen of five parvoviral species infecting step-wise transformed melanocytes in cell culture. We have demonstrated that Lulll efficiently infects these cells, expressing abundant viral structural and non-structural proteins. Furthermore, Lulll generates significant amounts of progeny virions that rapidly spread to, and kill, other transformed cells in the culture. The further aims of this research are to generate and characterize replication competent derivatives of Lulll for their potential as virotherapeutic agents against malignant melanoma. We will seek to confirm the applicability of Lulll by examining its ability to grow productively in human melanocyte cell lines transformed by different stepwise procedures. Lulll will also be assessed for its ability to productively infect normal melanocytes versus a panel of primary and established melanoma cell cultures, in order to explore its therapeutic index and the generality of its melanotropism. We will further enhance the oncoselectivity of the Lulll derivatives in neoplastically transformed human melanocyte cultures by genetic manipulation. For this, we will construct Lulll P4 promoter variants predicted to be responsive to activation of the Notch signaling pathway, a frequent event in the evolution of melanoma. In parallel, we will use a Lulll P4 promoter derivative to generate an infectious clone library of degenerate oligonucleotide sequences replacing the Ets1 site, a major determinant of oncoselectivity in transformed fibroblasts. This library will be used to attempt selection of mutants with P4 proximal transcription site(s) further optimized for growth in fully transformed melanocytes. Selected mutants will be screened for their ability to expand in and kill a panel of melanoma tumor cells, while sparing those from a parallel panel of normal, primary melanocyte cultures. Viruses are often very specific for the host cell types they will infect and kill. This project will explore whether Lulll, a relatively melanoma-specific member of the parvovirus family, can be further modified so as to preferentially destroy malignant melanoma cells, compared to their normal counterparts.