University of Pittsburgh scientists have developed a novel approach to reactivate latent Epstein-Barr virus (EBV) in cancer cells. Reactivation of EBV treatment with ganciclovir (GCV), a known antiviral agent, can induce cancer cell death and this approach could revolutionize cancer treatments for EBV-positive cancer cells.
Description
EBV infection is linked to various cancers. As a ubiquitous herpesvirus, EBV can establish lifelong persistent infection in B cells and epithelial cells. A novel approach called CRISPR/dCas9-Mediated EBV Reactivation (CMER) has been developed to reactivate EBV in EBV-positive cancer cells. This approach uses single guide RNAs (sgRNAs) specifically
targeting EBV genome and leads to EBV reactivation in EBV-positive cancer cells with subsequent GCV-induced cell death. This cell death was selective to only EBV-positive cells and provides high potential for a targeted therapy for EBV-linked cancers.
Applications
• Lymphoma
• Gastric cancer
• Nasopharyngeal carcinoma
• Other EBV-related cancers
Advantages
Given the EBV links to some cancers, previous efforts have been made to treat cancer by overexpressing EBV Immediate-Early (IE) genes. Other approaches including -irradiation and chemotherapeutic agents (e.g., Bortezomib, cis-platinum, and fluorouracil) can also induce EBV reactivation. However, none of these are selective to only EBV-positive cells. This novel strategy of CMER with sgRNA can effectively reactivate EBV directly without lytic inducing agents. As a result, only EBV-positive cells can be targeted. Once EBV is reactivated, encoded protein kinase BGLF4 is expressed which phosphorylates GCV. Phosphorylated GCV inhibits both viral and cellular DNA polymerases resulting in cell death of only EBV-positive cells, regardless of cell origin. This selective targeting of EBV-positive cells should lead to less adverse effects compared to other therapies.
Invention Readiness
Using the power of CRISPR/dCas9-VP64 gene activation system, the promoter of EBV ZTA was targeted. A series of 10 unique sgRNAs were identified and cloned into a lentiviral system, and Akata (EBV+) Burkitt lymphoma cell lines were then transduced with the lentiviruses. Of these cell lines, 9 out of 10 CMER sgRNAs reactivated EBV, with CMER sg-5 triggering the highest viral reactivation. Similar results were observed in other EBV-positive lymphoma and epithelial cancers. CMER with sg-5 and subsequent treatment of cells with GCV led to cell death of EBV-positive cells only, leaving EBV-negative cells unaffected, regardless of cell type. Further work is required to optimize the CRISPR/dCas9-VP64 and sgRNA delivery system to treat EBV-positive cancers.
IP Status
https://patents.google.com/patent/WO2025072640A1
