# CD19-ReTARGTPR: A Novel Fusion Protein for Physiological Engagement of Anti-CMV Cytotoxic T Cells Against CD19-Expressing Malignancies

## TL;DR

A new therapy called CD19-ReTARGTPR uses the body's virus-fighting T cells to target blood cancers more safely and effectively than current treatments.

## Contribution

CD19-ReTARGTPR is a novel fusion protein that enables physiological T cell activation against CD19-expressing cancers with reduced toxicity.

## Key findings

- CD19-ReTARGTPR effectively redirected anti-CMV CTLs to kill CD19-expressing cancer cells in preclinical tests.
- The therapy retained efficacy even against cancer cells with low CD19 expression.
- Unlike existing therapies, it caused robust cytotoxic activity without excessive cytokine release.

## Abstract

Current treatments for certain blood cancers, like CAR T cells and BiTEs, can overactivate the immune system and cause serious side effects. This study introduces a new therapy called CD19-ReTARGTPR, which works by helping the body’s existing virus-fighting T cells recognize and attack cancer cells in a more natural and controlled way. It achieves this by attaching a small piece of a common virus (CMV) to a targeting molecule that binds to cancer cells, allowing T cells to kill the cancer without excessive immune reactions. In lab tests, this approach effectively killed cancer cells, even those with low levels of the target marker (CD19), and caused fewer harmful side effects compared to current therapies. This new strategy could lead to safer and more precise immunotherapies for patients with CD19-positive blood cancers.

Background/Objectives: The physiological activation of cytotoxic CD8pos T cells (CTLs) relies on the engagement of the TCR/CD3 complex with cognate peptide-HLA class I (pHLA-I) on target cells, triggering cell lysis with appropriate cytokine release and minimized off-target toxicity. In contrast, current immunotherapies for CD19-expressing hematological malignancies, such as chimeric antigen receptor (CAR) T cells and bispecific T cell engagers (BiTEs), bypass TCR/pHLA interactions, resulting in CTL hyperactivation and excessive cytokine release, which frequently cause severe immune-related adverse events (irAEs). Thus, there is a pressing need for T cell-based therapies that preserve physiological activation while maintaining antitumor efficacy. Methods: To address this, we developed CD19-ReTARGTPR, a novel fusion protein consisting of the immunodominant cytomegalovirus (CMV) pp65-derived peptide TPRVTGGAM (TPR) covalently presented by a soluble HLA-B*07:02/β2-microglobulin complex fused to a high-affinity CD19-targeting Fab antibody fragment. The treatment of CD19-expressing cancer cells with CD19-ReTARGTPR makes them recognizable for pre-existing anti-CMVpp65 CTLs via physiological TCR-pHLA engagement. Results: Our preclinical data demonstrate that CD19-ReTARGTPR efficiently redirects anti-CMV CTLs to eliminate CD19-expressing cancer cells, including both established cell lines and primary chronic lymphocytic leukemia (CLL) cells. Unlike CD19-directed CAR T cells or the CD19/CD3 BiTE blinatumomab, CD19-ReTARGTPR mediated robust cytotoxic activity without triggering supraphysiological cytokine release. Importantly, this approach retained efficacy even against cancer cells with low CD19 expression. Conclusions: In summary, we provide a robust proof-of-concept study and show that CD19-ReTARGTPR offers a promising alternative strategy for T cell redirection, enabling the selective and effective killing of CD19-expressing malignancies while minimizing cytokine-driven toxicities through physiological CTL activation pathways.