GENERATION OF A HUMAN MELANOCYTE CELL-LINE BY INTRODUCTION OF HPV16 E6 AND E7 GENES

Availability of a standard human melanocyte cell line with unlimited g rowth potential and otherwise normal melanocytic properties will great ly facilitate research in melanocyte biology and in vitro studies on t he etiology of pigmentary disorders and melanoma. Using a retroviral v ector, E6 and E7 open reading frames of human papilloma virus type 16 (HPV 16) have been introduced into cultured normal human melanocytes. Cells selected by increased resistance to geneticin conveyed by the ve ctor and expressing E6E7 mRNA have been cloned to ensure genetic homog eneity. Since their establishment as primary cells, cloned PIG1 cells have undergone more than twice the amount of population doublings of s enescent parental cells. Moreover, in passage numbers when parental ce lls had become senescent, proliferation of clonal cells was retained a t levels exceeding those of normal human melanocytes in third passage by 100%. Further characterization has revealed that the cells remain d ependent on tetradecanoyl phorbol 13-acetate (TPA) for growth and do n ot proliferate in soft agar nor form tumors in nude mice. The antigeni c profile of the cells was slightly altered as compared to parental ce lls, but was incomparable to that of M14 melanoma cells, Importantly, PIG1 cells contain more melanin pigment than parental cells.