Cytotoxicity of replication-competent adenoviruses powered by an exogenous regulatory region is not linearly correlated with the viral infectivity/gene expression or with the E1A-activating ability but is associated with the p53 genotypes

Our plain-English summary of Cytotoxicity of replication-competent adenoviruses powered by an exogenous regulatory region is not linearly correlated with the viral infectivity/gene expression or with the E1A-activating ability but is associated with the p53 genotypes, by S. Yamauchi, B. Zhong, K. Kawamura, et al (BMC Cancer, 2017).

The study

• Authors: S. Yamauchi, B. Zhong, K. Kawamura, S. Yang, S. Kubo, M. Shingyoji, I. Sekine, Y. Tada, K. Tatsumi, H. Shimada, K. Hiroshima, M. Tagawa
• Journal: BMC Cancer
• Published: 2017;17(1):622
• Cited by: 6 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: Replication-competent adenoviruses (Ad) produced cytotoxic effects on infected tumors and have been examined for the clinical applicability. A biomarkers to predict the cytotoxicity is valuable in a clinical setting.

Methods: We constructed type 5 Ad (Ad5) of which the expression of E1A gene was activated by a 5' regulatory sequences of survivin, midkine or cyclooxygenase-2, which were highly expressed in human tumors. We also produced the same replication-competent Ad of which the fiber-knob region was replaced by that of Ad35 (AdF35). The cytotoxicity was examined by a colorimetric assay with human tumor cell lines, 4 kinds of pancreatic, 9 esophageal carcinoma and 5 mesothelioma. Ad infectivity and Ad-mediated gene expression were examined with replication-incompetent Ad5 and AdF35 which expressed the green fluorescence protein gene. Expression of cellular receptors for Ad5 and AdF35 was also examined with flow cytometry. A transcriptional activity of the regulatory sequences was investigated with a luciferase assay in the tumor cells. We then investigated a possible correlation between Ad-mediated cytotoxicity and the infectivity/gene expression, the transcriptional activity or the p53 genotype.

Results: We found that the cytotoxicity was greater with AdF35 than with Ad5 vectors, but was not correlated with the Ad infectivity/gene expression irrespective of the fiber-knob region or the E1A-activating transcriptional activity. In contrast, replication-competent Ad produced greater cytotoxicity in p53 mutated than in wild-type esophageal carcinoma cells, suggesting a possible association between the cytotoxicity and the p53 genotype.

Conclusions: Sensitivity to Ad-mediated cytotoxic activity was linked with the p53 genotype but was not lineally correlated with the infectivity/gene expression or the E1A expression.

In plain English

This laboratory study tested replication-competent adenoviruses in human tumour cell lines. The authors built type 5 adenoviruses (Ad5) with E1A driven by 5' regulatory sequences of midkine, survivin or cyclooxygenase-2, and matching vectors whose fiber-knob region came from Ad35 (AdF35). Cytotoxicity (WST assay, IC50 in virus particles per cell) was measured in 4 pancreatic carcinoma, 9 esophageal carcinoma and 5 mesothelioma cell lines, alongside GFP-based infectivity/gene expression, CAR and CD46 expression, and luciferase activity of the regulatory regions as a putative E1 marker. IC50 values were generally lower with AdF35 than Ad5, but cytotoxicity showed no statistically significant linear correlation with infectivity/gene expression or with transcriptional activity, except AdF35/Sur in pancreatic cells. In esophageal carcinoma, IC50 values were higher in p53 wild-type cells (74.4 ± 22.5 x 10^4 virus particles per cell, n = 24 cell line and vector combinations) than in p53-mutated cells (28.0 ± 10.6 x 10^4 virus particles per cell, n = 30 combinations), P = 0.05. The authors concluded that sensitivity was linked with the p53 genotype but was not linearly correlated with the infectivity/gene expression or the E1A expression.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: in vitro laboratory study using human tumour cell lines and engineered adenovirus vectors
• Who was studied: Human tumour cell lines: 4 pancreatic carcinoma, 9 esophageal carcinoma and 5 mesothelioma lines, plus HEK293 cells as a control; work done in Japan
• Exposure: Infection with replication-competent Ad5 or AdF35 whose E1 gene was driven by 5' regulatory regions of midkine (0.6 kb), survivin (0.5 kb) or COX-2 (0.3 kb); replication-incompetent Ad5/GFP and AdF35/GFP at 30 MOI for infectivity; cisplatin 20 microM for p21 experiments
• Main results:
  • IC50 values were in general lower (greater cytotoxicity) with AdF35 than with Ad5 across the regulatory regions; all 4 pancreatic carcinoma lines were more sensitive to AdF35
  • Exceptions where Ad5 was more cytotoxic than the matching AdF35: 5 of 27 esophageal carcinoma pairs and 5 of 15 mesothelioma pairs
  • No significant correlation between receptor expression and infectivity/gene expression: CAR vs Ad5/GFP was -0.13 (P = 0.87) pancreatic, 0.62 (P = 0.07) esophageal, 0.68 (P = 0.21) mesothelioma; CD46 vs AdF35/GFP was -0.20 (P = 0.80), 0.14 (P = 0.71) and 0.72 (P = 0.16)
  • No statistically significant linear correlation between Ad-mediated cytotoxicity and infectivity/gene expression in any of the three tumour types
  • Cytotoxicity was generally independent of the transcriptional activity of the regulatory regions; the single statistically significant correlation was AdF35/Sur cytotoxicity with Sur luciferase activity in pancreatic carcinoma (coefficient 0.99, P = 0.01)
  • In esophageal carcinoma, IC50 values with all replication-competent Ad were higher for p53 wild-type cells (74.4 ± 22.5 x 10^4 vp/cell, n = 24 cell line and vector combinations) than p53-mutated cells (28.0 ± 10.6 x 10^4 vp/cell, n = 30), P = 0.05
  • For AdF35 vectors in esophageal carcinoma, IC50 was 45.7 ± 20.6 (n = 12) in wild-type versus 6.7 ± 1.5 (n = 15) in mutant p53 (P = 0.04); Ad5 vectors showed no statistical difference (103.0 ± 39.2, n = 12 versus 49.4 ± 20.0, n = 15; P = 0.21)
  • IC50 values in the p21-decreased group after cisplatin (26.3 ± 10.5, n = 30) were marginally different from the p21 non-decreased group (74.5 ± 28.0, n = 18), P = 0.06
• The authors' conclusion: The authors concluded that sensitivity to Ad-mediated cytotoxic activity was linked with the p53 genotype but was not linearly correlated with the infectivity/gene expression or the E1A expression, and perhaps was associated with decreased p21 levels
• Limitations the authors state:
  • The percentage of GFP-positive cells was only an indirect estimate of adenovirus infectivity, since fluorescence was affected by promoter activity and GFP protein stability
  • The luciferase activity was only a putative marker of E1 transcription, because adenoviral proteins made during replication influence the E1-activating ability
  • The p53 genotypes were unevenly distributed in the pancreatic carcinoma and mesothelioma lines, so the p53 linkage could not be analysed within those tumour types
  • The authors stated that the p53 genotype as a potential biomarker needs confirmation with clinical specimens
  • The mechanism behind the greater association seen with AdF35 than with Ad5 vectors was unknown to the authors

Topics

Medical terms in this study, explained in our Lexicon: Biomarker, Carcinoma, Mesothelioma.

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• Combination of a p53-activating CP-31398 and an MDM2 or a FAK inhibitor produces growth suppressive effects in mesothelioma with wild-type p53 genotype — B. Zhong, M. Shingyoji, et al. (2020)
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