Impact of Ephrin Receptor A2 Polymorphisms on KSHV Infectivity
Project ended 31 March 2018
Dr Georgia Schäfer
Title of the project
The impact of Ephrin Receptor A2 polymorphisms on KSHV infectivity and Kaposi’s Sarcoma incidence and severity in HIV/AIDS patients in South Africa.
Kaposi’s sarcoma (KS) is an AIDS-defining disease and is the most common AIDS-related malignancy worldwide. Kaposi’s sarcoma-associated herpes virus (KSHV), named also human herpesvirus-8 (HHV8), was discovered in 1994 and is the etiologic agent of KS. Due to the AIDS epidemic, KS is now the most common human tumour found in sub-Saharan African countries with still rising incidence rates [1-7]. KS is an endothelial tumour, and KSHV binding and uptake into endothelial cells involves rather complex virus-host interactions as reviewed by Schäfer et al., 2015 . Crucial for the entry process and downstream signalling, however, is the recently identified ephrin receptor tyrosine kinase A2 (EPHA2) at the host cell membrane [9, 10].
Although KSHV seroprevalence in sub-Saharan Africa is over 30% [11-13] not all HIV/AIDS patients develop KS, suggesting that host genetic factors potentially contribute to susceptibility to KSHV infection and/or KS development. In view of the key role of EPHA2 in KSHV infectious entry and downstream tumorigenesis, it is possible that polymorphisms in the EPHA2 protein coding region and/or its promoter region are a determining factor in KSHV infectivity and/or the incidence and severity of KS. This is particularly relevant in light of genetic variants identified in the coding and 5′ regulatory region of the HIV co-receptor CCR5, several of which have functional consequences for HIV-1 pathogenesis . Similarly, our hypothesis was that mutations in the KSHV host receptor’s coding regions and/or promoter region can result in EPHA2 with altered activity and/or expression patterns that can affect KSHV infectivity and hence affect KS incidence and severity among HIV/AIDS patients. Therefore, the herein proposed research project was aimed at determining if there were sequence polymorphisms in the EPHA2 gene of South African populations and whether these sequence polymorphisms in the promoter and/or the protein sequence had an impact on KSHV infectivity or on KS incidence and severity in HIV/AIDS patients.
How was this project of value in the struggle against cancer:
Research on KS/KSHV is still extremely underrepresented despite the high burden of disease in Southern Africa. In fact, this study for the first time determined the KSHV seroconversion rate in a patient cohort from the Western Cape which was found to be 31.6% which is in agreement with previously reported KSHV seroconversion rates in sub-Saharan Africa to be over 30%. Moreover, this study for the first time describes sequence variations in the KSHV entry receptor gene EPHA2 to affect susceptibility to KSHV infection and KS development in a South African HIV-infected patient cohort. These genetic variations were primarily located in the protein tyrosine kinase domain and the sterile-a-motif. Although the number of patients analysed was limited, this study lays the basis for further investigation into the impact and functional relevance of EPHA2 variants on KSHV infection and subsequent KS development. This will contribute significantly to our understanding of the epidemiology of the KS burden in the South African population additionally burdened by the high HIV/AIDS prevalence. It will open new doors for the development of risk evaluation of KS by using state-of-the-art molecular techniques that will be novel in the South African research landscape.
Future plans for this research project:
We have gratefully received a new CANSA research grant which is based on the findings of this project. We will now test the identified variants in terms of functional involvement for KSHV infection or KS development. We will clone and overexpress them in endothelial cells (which have been silenced for endogenous EPHA2 by CRISP/Cas9 knockdown) and determine their impact on KSHV infection. For this purpose we will establish a KSHV infection system in our laboratory in collaboration with Prof. Thomas Schulz (Medical School Hannover, Germany), as no experimental KSHV infection model is available in South Africa yet. Furthermore, the impact of the identified EPHA2 variants on KS development will be assessed by qRT-PCR to markers associated with KSHV-mediated tumorigenesis. These functional studies will essentially constitute Ms Melissa Blumenthal’s PhD project for which she has already spent 4 months in Prof. Thomas Schulz laboratory in Germany to learn the essential techniques necessary for the successful completion of this project.