Customer case study: Rapid identification of effective dual-targeting CAR T-cell strategies for multiple myeloma
Prof. Carlos Fernández de Larrea
Associate professor of Medicine University of Barcelona
About Carlos Fernández de Larrea and his research focus
Prof. Carlos Fernández de Larrea and his research group are dedicated to developing innovative strategies redirecting the patient’s immune system against hematological diseases, such as multiple myeloma.
Multiple myeloma is a cancer derived from malignant plasma cells. Over 100.000 people die globally every year because of this disease which remains incurable. The goal of Prof. Carlos Fernández de Larrea is to induce long-term remission by enhancing the immune response through novel immunotherapeutic approaches such as CAR T-cell therapy. His development of effective CAR T-cell therapy aims to suppress tumor growth, enhance a durable remission, and to improve the life quality of patients suffering from multiple myeloma.
The recent work of Prof. Carlos Fernández de Larrea in collaboration with the team led by Dr. Eric Smith from Dana Farber Cancer Institute focuses on developing novel dual-targeting CAR T-cell therapy against multiple myeloma. Conventional mono-targeting CAR T-cell therapies target a single antigen expressed on tumor cells. B cell maturation antigen (BCMA) is a notable treatment target for multiple myeloma because of its highly selective expression in malignant cells. Looking at mono-targeting BCMA CAR T cells, they have yielded remission in treated patients, however, a large proportion relapses due to BCMA antigen escape as a result of low antigen density. Simultaneous targeting of two tumor antigens using dual-targeting CAR T-cell approaches can yield a more robust and specific killing. By developing dual-targeting approaches, Prof. Carlos Fernández de Larrea aims to overcome BCMA antigen escape and prevent relapse observed in patients treated with BCMA mono-targeting CAR T cells. As an early adopter of the z-Movi® Cell Avidity Analyzer, he and his research group pioneer cell avidity measurements in CAR T-cell therapy development.
Cell avidity
A new parameter to rapidly evaluate different CAR T-cell strategies
In this application note, Prof. Carlos Fernández de Larrea and his team demonstrated in preclinical models of multiple myeloma that targeting BCMA and GPRC5D at the same time using CAR T cells may prevent tumor escape. For this, they created 4 different targeting approaches and compared them using cell avidity measurements and in vivo assays (Figure 1).
1. Representation of the CAR designs used in the experiments.
Cell avidity analysis provides the means to distinguish between low and high efficacy CAR T cells (Figure 2). Unlike conventional in vitro methods, avidity measurements allow to determine the enhancement in antigen targeting due to the second CAR. This detailed evaluation of cell-cell interactions enabled Prof. Carlos Fernández de Larrea and his colleagues to assess the functionality and outcome of their CAR T-cell approaches in a fast, predictive and reproducible manner.
2. Schematic figure of the z-Movi workflow depicting measurements of two different immune-cell populations with different avidities to the monolayer of target cells and the resulting avidity curves.
Cell avidity measurements correspond with in vivo outcomes
During this study, Prof. Carlos Fernández de Larrea and his team evaluated how dual-targeting CAR T cells performed against multiple myeloma, and compared them against populations of mono-targeting CAR T cells targeting BCMA or GPRC5D. For this, they tested the overall survival of mono-targeting or dual-targeting CAR-treated xenograft mouse models injected with myeloma cells expressing both antigens, and compared their results with cell avidity measurements using the z-Movi. Their results showed that:
- Dual-targeting CARs exhibit superior binding interactions to target cells expressing both antigens compared with mono-targeting CARs (Figure 3).
- Dual-targeting CARs display stronger intercellular binding strength in BCMA-deficient target cells expressing GPRC5D, validating the ability of dual-targeting approaches to overcome BCMA antigen escape (Figure 4).
- Measurements with the z-Movi correlate with treatment outcomes in mouse models, suggesting that cell avidity measurements are reliable predictors of CAR T-cell efficacy (de Larrea et al., 2020).
The cell avidity data align with the mouse survival outcome in in vivo study (de Larrea et al., 2020). This means by implementing cell avidity measurements at an early stage, researchers can expedite their research by being able to predict in vivo and clinical outcomes in the preliminary phase.
3. Avidity curve showing the average proportion of CAR T cells and nontransduced (NTD) T cells bound to BCMA- and GPRC5D-expressing target cells upon increasing acoustic forces. The dashed line is set at 200 pN and indicates the plateau of the NTD curve (left). Bar graph representing fold increases of bound CAR T cells at 200 pN compared with antigen-negative target cells; gated from the avidity curve (left). rForce represents forces calibrated on 10 μm polystyrene beads (right).
4. Avidity curves showing the percentage of target-bound CAR T cells and non-transduced (NTD) T cells upon increasing acoustic forces. Cell avidity measurements of CAR T cells were performed on target cell without BCMA and GPRC5D expression (left) or BCMA-deficient target cells expressing GPRC5D (right). Dashed line indicates the required force to detach all the non-specific T cells. rForce indicates relative force.
3. Avidity curve showing the average proportion of CAR T cells and nontransduced (NTD) T cells bound to BCMA- and GPRC5D-expressing target cells upon increasing acoustic forces. The dashed line is set at 200 pN and indicates the plateau of the NTD curve (left). Bar graph representing fold increases of bound CAR T cells at 200 pN compared with antigen-negative target cells; gated from the avidity curve (left). rForce represents forces calibrated on 10 μm polystyrene beads (right).
4. Avidity curves showing the percentage of target-bound CAR T cells and non-transduced (NTD) T cells upon increasing acoustic forces. Cell avidity measurements of CAR T cells were performed on target cell without BCMA and GPRC5D expression (left) or BCMA-deficient target cells expressing GPRC5D (right). Dashed line indicates the required force to detach all the non-specific T cells. rForce indicates relative force.
Rapid and easy workflow
Cell avidity analysis using the z-Movi enables a quick quantification of CAR T-cell interaction. Compared to cell-killing assays and in vivo mouse studies that require tedious and time-consuming validations, cell avidity measurements represent a rapid and time-saving solution to identify effective lead candidates. The entire workflow to retrieve data about T cell functionality takes less than 10 minutes.
Get the app note!
Download the application note to learn how cell avidity analysis enables Prof. Carlos Fernández de Larrea and his team to explore and validate CAR T-cell functionality and obtain predictive, reproducible and rapid results at the single-cell level.
Watch the webinar!
In this webinar, Prof. Carlos Fernández de Larrea explains how expressing two CARs on a single cell enhances the strength of CAR T cell/target cell interactions, prevents BCMA escape-driven relapse, and how cell avidity measurements contributed to their findings.
