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 diseasesWe have identified the orphan parvovirus Lulll as an effective oncolytic virus from a screen of five parvoviralspecies infecting step-wise transformed melanocytes in cell culture. We have demonstrated that Lulllefficiently 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 inthe culture. The further aims of this research are to generate and characterize replication competentderivatives of Lulll for their potential as virotherapeutic agents against malignant melanoma. We will seek toconfirm the applicability of Lulll by examining its ability to grow productively in human melanocyte cell linestransformed by different stepwise procedures. Lulll will also be assessed for its ability to productively infectnormal melanocytes versus a panel of primary and established melanoma cell cultures, in order to explore itstherapeutic index and the generality of its melanotropism. We will further enhance the oncoselectivity of theLulll 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 signalingpathway, a frequent event in the evolution of melanoma. In parallel, we will use a Lulll P4 promoter derivativeto generate an infectious clone library of degenerate oligonucleotide sequences replacing the Ets1 site, amajor determinant of oncoselectivity in transformed fibroblasts. This library will be used to attempt selection ofmutants 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, whilesparing 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 whetherLulll, a relatively melanoma-specific member of the parvovirus family, can be further modified so as topreferentially destroy malignant melanoma cells, compared to their normal counterparts.
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