Multi-Omic Single-Cell Dissection of Leukemic T-Cell Lymphoma Following CAR T-Cell Therapy [Spatial Transcriptomics]

A 63-year-old male who received ciltacabtagene autoleucel (cilta-cel) CAR-T cells and the GPRC5DxCD3 bispecific talquetamab for early relapse of his multiple myeloma (MM) developed a leukemic peripheral T-cell lymphoma (PTCL) with cutaneous and intestinal involvement. Longitudinal single-cell RNA and T-cell receptor sequencing of peripheral blood and bone marrow revealed two hyperexpanded CAR-carrying, exhausted effector-memory T-cell clones with marked immunophenotypic as well as transcriptional alterations and different susceptibilities towards treatment with dexamethasone. Spatial transcriptomes of skin lesions confirmed the aberrant CAR-expressing T cells. Whole genome sequencing revealed three distinct integration sites, within the introns of ZGPAT, KPNA4, and polycomb-associated non-coding RNAs. Pre/post-CAR-T whole-genome analyses implicated clonal outgrowth of a TET2-mutated precursor propelled by additional sub-clone specific LOH and other secondary mechanisms Two tissue sections of a formalin-fixed paraffin-embedded (FFPE) skin sample, from a patient treated with CAR-T cells, of the left dorsal region. Three custom probe sets were developed to detect CAR-positive (cilta-cel) spots. To demonstrate the specificity of the cilta-cel custom probes, one slide with a skin sample was subjected to the Visium Spatial Protocol WITHOUT custom probes, another slide with a skin sample was subjected to the Visium Spatial Protocol WITH custom probes and an extramedullary disease sample (sample excision from the lumbar vertebral body) from a patient not treated with CAR-T cells was subjected to the Visium Spatial Protocol WITH custom probes.