Simultaneous interrogation of genetic evolution and immune evasion of multiple myeloma using blood biopsy in patients treated with CAR-T cells.

• Applicant: Dr. Joanna Blocka
• Subject Area: Hematology, Oncology
• Term: since 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 496758595

Background:Immunotherapies have been an effective treatment in many malignant diseases. However, resistance develops frequently, because tumor cells change their immunological profile to escape killing and engineered T cells become exhausted. In multiple myeloma (MM), intratumoral heterogeneity plays an important role and may contribute to therapy resistance due to somatic mutations or adaptive changes of the subclones. To detect the drug-resistant subclones, frequent assessment of MM genetics and adaptation during treatment is necessary. Single-site bone marrow biopsy is confounded by the patchy distribution of myeloma in the bone marrow and may thus not sufficiently capture all existing subclones. Moreover, in patients treated with CAR-T cells an efficacy loss is observed due to T-cell exhaustion. Interrogation of distinct T-cell differentiation states may be predictive of therapy success.Objective:We will use innovative liquid biopsy approaches for isolation of circulating tumor cells (CTCs), cell-free (cf)DNA and T cells to comprehensively define immediately actionable vulnerabilities of both the tumor and CAR-T cells, develop novel biomarkers, and use these as predictors of clonal evolution, disease burden, therapy response and early relapse.Research plan and methods:We will use peripheral blood and bone marrow samples of patients treated with anti-BCMA CAR-T cells on KarMMa 2- and -3 trials. Through whole genome and exome sequencing, we will examine if clonal and subclonal events found in myeloma cells in the bone marrow are also detectable in cfDNA in the peripheral blood and define the events that are exclusively detectable in cfDNA but not in the bone marrow. We will determine the sensitivity and specificity of interrogating myeloma CTCs to detect established genetic risk factors by single-cell sequencing and compare the single nucleotide/ copy number variants in the CTCs to those in the bone marrow. We will examine the correlation between the tumor burden, remission status and progression-free survival with the decline of myeloma CTCs and cfDNA. By means of single-cell RNA sequencing, we will investigate how the CAR-T cells evolve in MM patient as they lose efficacy and how the CAR-T-cell differentiation states differ between MM patients with good vs. poor response and long vs. short progression-free survival. Conclusion:The expected outcomes of our project are: - Development of cfDNA and CTCs as robust biomarkers of disease burden and clonal evolution, to guide treatment decisions in MM with minimal sampling effort, discomfort, and patient risk.- Assessment of better tools to predict the prognosis and treatment outcome in MM patients through interrogation of the activation and exhaustion status of CAR-T cells that recognize BCMA on myeloma cells as well as comparison of the differentiation states of CAR-T cells in good vs. poor therapy responders.

• DFG Programme: WBP Fellowship
• International Connection: USA
• Host: Professor Jens Lohr, Ph.D.