Leila Munaretto

CAR T cell therapy has revolutionized the treatment of a spectrum of blood-related malignancies. However, treatment responses vary among cancer types and patients. Accurate monitoring of CAR T cell dynamics is crucial for understanding and evaluating treatment efficacy. Positron emission tomography (PET) offers a comprehensive view of CAR T cell homing, especially in critical organs such as lymphoid structures and bone marrow. This information will help assess treatment response and predict relapse risk. Current PET imaging methods for CAR T require genetic modifications, limiting clinical use. To overcome this, we developed an antigen-based imaging approach enabling whole-body CAR T cell imaging. The probe detects CAR T cells in vivo without affecting their function. In an immunocompetent B cell leukemia model, CAR-PET signal in the spleen predicted early mortality risk. The antigen-based CAR-PET approach allows assessment of CAR T therapy responses without altering established clinical protocols. It seamlessly integrates with FDA-approved and future CAR T cell generations, facilitating broader clinical application.

Pemphigus vulgaris is a B cell-mediated autoimmune disease characterized by autoantibodies targeting desmoglein-3 (Dsg3), a critical adhesion molecule in epithelial tissues. Current treatments rely on broad immunosuppression, highlighting the need for more targeted therapeutic approaches in pemphigus vulgaris and other autoantibody-driven disorders. We engineered a therapeutic fusion protein consisting of the pathogenic domains of Dsg3 linked to either human immunoglobulin G1 (IgG1) or mouse IgG2a (Dsg3-Fc). In vitro, Dsg3-Fc selectively eliminated Dsg3-autoreactive B cells. In vivo, Dsg3-Fc effectively depleted human B cells expressing patient-derived anti-Dsg3 B cell receptors, even in the presence of circulating autoantibodies. Moreover, Dsg3-Fc inhibited both disease initiation and progression in a polyclonal, active pemphigus vulgaris model in immunocompetent mice. In addition, Dsg3-Fc rapidly neutralized pathogenic autoantibodies without inducing systemic toxicity. These findings demonstrate that targeting pathogenic B cells and neutralizing autoantibodies through autoantigen-Fc fusion proteins may represent a promising therapeutic strategy for pemphigus vulgaris and potentially other autoantibody-mediated diseases without the need for global immunosuppression.

BACKGROUND: Limited durability of clinical responses remains a major challenge in chimeric antigen receptor (CAR)-T therapy. CAR-enhancers (CAR-Es), which fuse tumor antigens to interleukin (IL)-2 muteins, provide a targeted strategy to enhance CAR-T persistence and function. It remained unclear whether CAR-Es are effective across distinct tumor contexts, when using patient-derived T cells, or in preventing exhaustion and sustaining persistence. It was also unknown whether CAR-Es can selectively expand CAR-Ts in humanized mice with pre-existing T cells, and to what extent their efficacy depends on IL-2Rβγ vs IL-2Rα engagement. While IL-2Rα (CD25) has been classically linked to potent antitumor responses and memory formation, it also drives IL-2-associated toxicities, including vascular leak and preferential regulatory T cell expansion. METHODS: We systematically dissected CAR-E signaling requirements by engineering IL-2 variants with selective receptor affinities. Multiple CAR-E constructs were developed and tested across a range of in vitro and in vivo models. RESULTS: We demonstrate that CAR-E activity is entirely independent of IL-2Rα and critically dependent on IL-2Rβγ signaling. A next-generation IL-2Rα-sparing CAR-E maintained full potency, driving robust CAR-T expansion, persistence, and tumor clearance, even at low doses and when using CAR-T cells derived from previously treated multiple myeloma patients. These CAR-T cells not only resisted exhaustion but also re-expanded months later to eradicate tumor rechallenges. In humanized mice with pre-established T cells, CAR-Es selectively expanded CAR-Ts to dominate the circulating T-cell pool. CAR-E exerted a dominant influence on CAR-T fate, overriding tumor-derived cues and enforcing consistent phenotypes across diverse preclinical models. CONCLUSIONS: These findings nominate a lead B-cell maturation antigen (BCMA)-IL-2 CAR-E candidate with strong translational potential for clinical development and establish IL-2Rβγ as a key driver of CAR-E activity. The results also identify IL-2Rα as dispensable and provide a mechanistic framework for designing safer, IL-2Rα-sparing CAR-Es.