In-vitro characterization and evaluation of apoptotic potential of bicistronic plasmid encoding HN gene of Newcastle disease virus and human TNF-α.
The viral gene oncotherapy in combination with cytokines emerges as an exciting strategy for cancer therapy due to its minimal side effects and tumor specificity. HN is the surface protein of NDV which is involved in virus infectivity and is known to kill many cancerous cell types. TNF-α, a multifactorial cytokine has direct anti-tumor activity by activating the extrinsic pathways of apoptosis. In the present study, HN gene of NDV and TNF-α of human were cloned at multiple cloning sites (MCS) 1 and 2 of bicistronic expression vector pVIVO2. Expression pattern of recombinant clone was checked on transcriptional and translational level by RT-PCR, Immunofluorescence assay and flow cytometry. On flow cytometric analysis HN gene expression was found to be 28.30 ± 1.21; 5.22 ± 0.60%, and TNF-α gene expression was found to be 15.44 ± 0.42; 6.51 ± 0.757%, in HeLa cells transfected with pVIVO.nd.hn.hu.tnf and pVIVO2 empty vector control, respectively. These assays confirm that HN and TNF-α act synergistically in the induction of apoptosis in HeLa cells.
a Molecular Biology Lab, Division of Veterinary Biotechnology , Indian Veterinary Research Institute , Bareilly , Uttar Pradesh , India.
R S
RS
Rajmani
Prafull Kumar
PK
Singh
G
G
Ravi Kumar
Shikha
S
Saxena
Lakshya Veer
LV
Singh
Rajiv
R
Kumar
Aditya P
AP
Sahoo
Shishir Kumar
SK
Gupta
Uttara
U
Chaturvedi
Ashok K
AK
Tiwari
D017209:Apoptosis / Q000235:genetics*; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics*; D015707:HN Protein / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D009522:Newcastle disease virus / Q000235:genetics*; D010957:Plasmids / Q000235:genetics*; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics*
D015707:HN Protein; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha
Apoptosis; Cancer; Gene therapy; HN gene; NDV; TNF-α