Adenovirus‐mediated Transfer of p53‐related Genes Induces Apoptosis of Human Cancer Cells

Abstract

Two p53‐related genes, p73 and p51, were recently identified as structural homologues of the p53 tumor suppressor gene, suggesting that the roles of these two genes may be similar to those of p53, including growth suppression and induction of apoptosis. Here we show that introduction of p73 or p51 cDNAs into cultured human cancer cells suppressed colony formation in the presence of G418. We then examined the ability of various isoforms of p73 and p51 to activate transcription of a reporter gene. This assay showed that p73 β and p51A activated transcription through a consensus p53 binding sequence, while p73 α and p51B isoforms minimally transactivated the p53 reporter gene. To characterize further the biological functions of the p53‐related genes, we constructed recombinant adenoviruses containing the p73 and p51 cDNAs. Ad‐p73 β and Ad‐p51A induced endogenous p21 gene expression more effectively than Ad‐p73 β and Ad‐p51B, respectively. To evaluate the mode of cell death induced by p53‐related genes, Ad‐p73 and Ad‐p51 were used to infect human cancer cells. Infection of Ad‐p73 β, Ad‐p51A or Ad‐p51B resulted in DNA fragmentation in a subset of cancer cell lines more efficiently than did infection of Ad‐p53. We then examined the combined effect of each p53‐related gene and the E1A oncogene in the induction of apoptosis. The E1A oncogene cooperated with p51 as well as p53 to induce apoptosis, while p73 resulted in a weak induction of apoptosis by E1A. Overall, apoptosis induction by p51B and p73 α isoforms may be due to mechanisms other than transcriptional activation of p53‐target genes. Our results suggest that p53‐related genes are both similar to and different from p53 in their pathways leading to growth suppression.

Adenovirus-mediated transfer of the p53 family genes, p73 and p51/p63 induces cell cycle arrest and apoptosis in colorectal cancer cell lines: potential application to gene therapy of colorectal cancer

“…Similar results were previously reported in different cell types. 14,20 These differences in response were not simply attributable to differences in the expression levels, as determined by immunoblot analysis. 20 Therefore, we analyzed p73␤ and p51A/p63␥ isoforms in further detail.…”

“…14,20 These differences in response were not simply attributable to differences in the expression levels, as determined by immunoblot analysis. 20 Therefore, we analyzed p73␤ and p51A/p63␥ isoforms in further detail. Cell lines that underwent cell cycle arrest after infection were accompanied by a significant increase in the proportion of cells in the G0-G1 phase.…”

“…20,39 Essentially, we used a replicationdeficient E1-deleted Ad5 vector pJM17 (Microbix Biosystem, Toronto, Ontario, Canada) as a backbone and the corresponding genes were regulated under the control of a human cytomegalovirus promoter/enhancer and a bovine growth hormone polyadenylation signal. Adenovirus titer in plaque forming units (p.f.u.)…”

“…[13][14][15][16] p73 and p51/p63 can bind to the p53 responsive elements and upregulate some p53 target genes, which suggest that the p53 family member genes have a potential for functional overlap with p53 itself. [17][18][19][20] However, in contrast to p53, p73 and p51/p63 are rarely mutated in human cancers. 21 Li-Fraumeni syndrome patients have inherited mutations of the p53 gene and develop normally, but are predisposed to cancer, 22 while heterozygous germline mutations in the p51/p63 gene are the cause of ectrodactyly, ectodermal dysplasia and facial clefts (EEC) syndrome.…”

“…However, the studies of p53 and p53 family members have used different cell types and have used different methods to induce gene expression. [17][18][19][20] It has therefore been difficult to make direct comparisons between the apoptotic activity observed in independent studies. In the present study, to evaluate the anti-cancer activity of p73 and p51/p63 in comparison with that of p53, we sought to analyze growth arrest and apoptosis induced by the p53 family genes using adenoviral vectors in a panel of human colorectal cancer cell lines.…”