Potential of Herpesvirus Saimiri-Based Vectors To Reprogram a Somatic Ewing's Sarcoma Family Tumor Cell Line

Abstract

cHerpesvirus saimiri (HVS) infects a range of human cell types with high efficiency. Upon infection, the viral genome can persist as high-copy-number, circular, nonintegrated episomes that segregate to progeny cells upon division. This allows HVS-based vectors to stably transduce a dividing cell population and provide sustained transgene expression in vitro and in vivo. Moreover, the HVS episome is able to persist and provide prolonged transgene expression during in vitro differentiation of mouse and human hemopoietic progenitor cells. Together, these properties are advantageous for induced pluripotent stem cell (iPSC) technology, whereby stem cell-like cells are generated from adult somatic cells by exogenous expression of specific reprogramming factors. Here we assess the potential of HVS-based vectors for the generation of induced pluripotent cancer stem-like cells (iPCs). We demonstrate that HVS-based exogenous delivery of Oct4, Nanog, and Lin28 can reprogram the Ewing's sarcoma family tumor cell line A673 to produce stem cell-like colonies that can grow under feeder-free stem cell culture conditions. Further analysis of the HVS-derived putative iPCs showed some degree of reprogramming into a stem cell-like state. Specifically, the putative iPCs had a number of embryonic stem cell characteristics, staining positive for alkaline phosphatase and SSEA4, in addition to expressing elevated levels of pluripotent marker genes involved in proliferation and self-renewal. However, differentiation trials suggest that although the HVS-derived putative iPCs are capable of differentiation toward the ectodermal lineage, they do not exhibit pluripotency. Therefore, they are hereby termed induced multipotent cancer cells.

Epigenetic reprogramming and re-differentiation of a Ewing sarcoma cell line

“…While hierarchical cluster analysis of our EWS-iPS cells demonstrated similar patterns of global methylation to other ES and iPS cell lines, the degree of promoter hypomethylation of several stem cell-specific DMRs and some endogenous pluripotency genes was not as pronounced in our EWS-iPS cells, thus suggesting that the EWS-derived iPS cells may have been only partially reprogrammed. This appeared to be the case in a recent study which attempted to reprogram a EWS cell line via employing a herpesvirus saimiri-based (HSV) 3 factor (OCT4, NANOG, and LIN28) vector; these cells, termed “induced multipotent cancer cells (iPCs),” yielded only a partially reprogrammed phenotype capable of differentiation toward the ectodermal lineage (Brown et al, 2013 ). Contrary to this report, our EWS-derived iPS cell clones (specifically B and C) exhibited capacity to differentiate into a number of embryonic tissues.…”

Herpesvirüs Saimiri’nin Alternatif Gen Terapi Vektörü Olarak Kullanımı

“…Bu nedenlerden dolayı alternatif gen terapi vektörlerine ihtiyaç duyulmaktadır. Herpesvirus saimiri vektörleri insan T lenfesitleri dışında birçok hücre türünü enfekte ederek, yüksek kapasitede heterolog transgen ekspresyonu sağlayabildiği için farklı enfeksiyon, kanser hastalıklarında, romatoid artrit ve akut karaciğer hastalıklarının adoptif immünoterapisinde kullanılabilme potansiyeli göstermiştir [27][28][29][30][31][32][33] .…”