Abstract
BackgroundSindbis viral vectors are able to efficiently target and kill tumor cells in vivo, as shown using pancreatic and ovarian cancer models. Infection results in apoptosis both in vitro and in vivo. Sindbis vector uptake is mediated by the LAMR, which is upregulated on a number of different tumor types, thus conferring specificity of the vector to a wide range of cancers. In this study we elucidate the mechanism of apoptosis in two tumor cell lines, MOSEC, derived from the ovarian epithelium and Pan02, derived from a pancreatic adenocarcinoma. A comprehensive understanding of the mechanism of apoptosis would facilitate the design of more effective vectors for cancer therapy.ResultsThe initial phase of Sindbis vector induced apoptosis in MOSEC and Pan02 models reconfirms that viral infection is sensed by PKR due to double-stranded RNA intermediates associated with genomic replication. PKR activation results in translation inhibition through eIF2α phosphorylation and initiation of the stress response. Our studies indicate that the roles of two proteins, Mcl-1 and JNK, intimately link Sindbis induced translational arrest and cellular stress. Translational arrest inhibits the synthesis of anti-apoptotic Bcl-2 protein, Mcl-1. JNK activation triggers the release of Bad from 14-3-3, which ultimately results in apoptosis. These signals from translational arrest and cellular stress are propagated to the mitochondria where Bad and Bik bind to Bcl-xl and Mcl-1 respectively. Formation of these heterodimers displaces Bak, which results in caspase 9 cleavage and signaling through the mitochondrial pathway of apoptosis.ConclusionThe host cell response to Sindbis is triggered through PKR activation. Our studies demonstrate that PKR activation and subsequent translational arrest is linked to both cellular stress and apoptosis. We have also found the linkage point between translational arrest and apoptosis to be Mcl-1, a protein whose constant translation is required for inhibition of apoptosis. With this information vectors can be designed, which express or repress proteins implicated in this study, to enhance their therapeutic potential.
Highlights
Sindbis viral vectors are able to efficiently target and kill tumor cells in vivo, as shown using pancreatic and ovarian cancer models
The murine pancreatic adenocarcinoma Pan02 cells have been used to confirm critical results, as an additional cancer model of different tissue origin, verifying that the results shown are not restricted to a single tumor cell line or tissue type
Due to the doublestranded RNA intermediates generated by Sindbis vector replication, we studied the effect of Sindbis infection on PKR
Summary
Sindbis viral vectors are able to efficiently target and kill tumor cells in vivo, as shown using pancreatic and ovarian cancer models. Because of the blood-borne nature of this vector, it is delivered systemically and can treat the primary tumor but sites of metastasis as well [4,5] This vector is able to effectively target and efficiently shrink tumor burden from a number of xenograft models of cancer including pancreatic, colon [6] and ovarian cancers [4,5,6,7]. Sindbis vectors are able to target spontaneous tumors shown in RGR/p15+/- transgenic mice [5] and ovarian xenograft tumors implanted in immune competent mice [6] These vectors have been engineered to deliver genes, such as interleukin 12, which has enhanced the therapeutic potential [4]. The broad range of animal models in which Sindbis vectors have shown therapeutic efficacy, coupled with the ability to tailor therapies through the inclusion of a gene of interest, underscores the benefit of this vector for gene therapy
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