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Evaluation of LMP1 of Epstein-Barr virus as a therapeutic target by its inhibition.

ABSTRACT:

Background

The latent membrane protein-1 (LMP1) encoded by Epstein-Barr virus (EBV) is an oncoprotein which acts by constitutive activation of various signalling pathways, including NF-kappaB. In so doing it leads to deregulated cell growth intrinsic to the cancer cell as well as having extrinsic affects upon the tumour microenvironment. These properties and that it is a foreign antigen, lead to the proposition that LMP1 may be a good therapeutic target in the treatment of EBV associated disease. LMP1 is expressed in several EBV-associated malignancies, notably in Hodgkin's lymphoma and nasopharyngeal carcinoma (NPC). However, the viral protein is only detected in approximately 30%-50% of NPC samples, as such its role in carcinogenesis and tumour maintenance can be questioned and thus its relevance as a therapeutic target.

Results

In order to explore if LMP1 has a continuous function in established tumours, its activity was inhibited through expression of a dominant negative LMP1 mutant in tumour cell lines derived from transgenic mice. LMP1 is the tumour predisposing oncogene in two different series of transgenic mice which separately give rise to either B-cell lymphomas or carcinomas. Inhibition of LMP1 activity in the carcinoma cell lines lead to a reduction in clonagenicity and clone viability in all of the cell lines tested, even those with low or below detection levels of LMP1. Inhibition of LMP1 activity in the transgenic B-cell lines was incompatible with growth and survival of the cells and no clones expressing the dominant negative LMP1 mutant could be established.

Conclusions

LMP1 continues to provide a tumour cell growth function in cell lines established from LMP1 transgenic mouse tumours, of both B-cell and epithelial cell origin. LMP1 can perform this function, even when expressed at such low levels as to be undetectable, whereby evidence of its expression can only be inferred by its inhibition being detrimental to the growth of the cell. This raises the possibility that LMP1 still performs a pro-oncogenic function in the 50% to 70% of NPC tumours wherein LMP1 protein expression cannot be detected. This reinforces the basis for pursuing LMP1 as a therapeutic target in EBV associated LMP1-expressing malignancies.

SUBMITTER: Hannigan A

PROVIDER: S-EPMC2913984 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundEpstein-Barr virus (EBV)-encoded LMP1 protein is commonly expressed in nasopharyngeal carcinoma (NPC). LMP1 is a prime candidate for driving tumourigenesis given its ability to activate multiple signalling pathways and to alter the expression and activity of variety of downstream targets. Resistance to TGFbeta-mediated cytostasis is one of the growth transforming effects of LMP1. Of the downstream targets manipulated by LMP1, the induction of Id1 and inactivation of Foxo3a appear particularly relevant to LMP1-mediated effects. Id1, a HLH protein is implicated in cell transformation and plays a role in cell proliferation, whilst Foxo3a, a transcription factor controls cell integrity and homeostasis by regulating apoptosis. The mechanism(s) by which LMP1 induces these effects have not been fully characterised.ResultsIn this study, we demonstrate that the ability of LMP1 to induce the phosphorylation and inactivation of Foxo3a is linked to the upregulation of Id1. Furthermore, we show that the induction of Id1 is essential for the transforming function of LMP1 as over-expression of Id1 increases cell proliferation, attenuates TGFbeta-SMAD-mediated transcription and renders cells refractory to TGFbeta-mediated cytostasis. Id1 silencing in LMP1-expressing epithelial cells abolishes the inhibitory effect of LMP1 on TGFbeta-mediated cell growth arrest and reduces the ability of LMP1 to attenuate SMAD transcriptional activity. In response to TGFbeta stimulation, LMP1 does not abolish SMAD phosphorylation but inhibits p21 protein expression. In addition, we found the induction of Id1 in LMP1-expressing cells upon stimulation by TGFbeta. We provide evidence that LMP1 suppresses the transcriptional repressor ATF3, possibly leading to the TGFbeta-induced Id1 upregulation.ConclusionThe current data provide novel information regarding the mechanisms by which LMP1 suppresses TGFbeta-induced cytostasis, highlighting the importance of Id1 in LMP1 mediated cell transformation.