Targeting T-cells to human cancer associated fibroblasts using an oncolytic virus expressing a FAP-specific T-cell engager Joshua Freedman1, Rebecca Ashfield2, Margaret R. Duffy1, Alice C.N. Brown3, Leonard W. Seymour1, Kerry D. Fisher and Brian R. Champion3 1Department of Oncology, University of Oxford, OX3 7DQ; 2Jenner Institute, University of Oxford, OX3 7DQ; 3PsiOxus Therapeutics Ltd, Abingdon, Oxfordshire, OX14 3YS Enadenotucirev (EnAd) is a chimeric oncolytic group B adenovirus with potent and selective anti-tumor activity against a range of epithelial cancer cells, with a blood stability profile that enables systemic dosing and has been administered intravenously to over 100 cancer patients. As an approach to immunogene therapy targeting stromal rich tumors, we have created transgene-modified variants of EnAd expressing a bi-specific T-cell engager (BiTE) molecule recognizing human fibroblast activating protein (FAP) on cancer associated fibroblasts (CAFs) and CD3 on T-cells. We hypothesize that production of BiTE proteins by virus infected tumor cells could be an effective way of modifying the stromal microenvironment to drive effective anti-tumor immunity and would bypass delivery and safety issues related to systemic dosing of BiTE proteins.
Bi-specific T-cell engager constructs comprising linked ScFv antibodies specific for human FAP and CD3 were designed and used to generate EnAd viruses expressing the BiTE such that transgene expression was under the control the virus major late promoter to allow tumor-selective expression.
We have shown that the viruses selectively express and secrete functional FAP-specific BiTEs following infection of tumor cell lines. In co-culture with FAP+ fibroblasts and PBMC-derived T cells this leads to T cell-activation and cytotoxicity towards the fibroblasts. Unlike activation of T-cells with anti-CD3/CD28, this FAP-BiTE activation was also shown to be effective even in the presence of immunosuppressive (cell free) ascitic fluid samples added to the cultures. Similarly, infection of primary malignant cancer ascites (unseparated cells replete with autologous fluid) with FAP-BiTE-encoding viruses led to polyclonal activation of endogenous T-cells, depletion of FAP+ cells from the cultures and reduction in the expression of immunosuppressive molecules such as TGFβ. Systemic dosing of an EnAd-FAP BiTE virus to patients with stromal rich tumors may be effective at driving therapeutic anti-tumor immunity.
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