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Homozygous BCMA gene deletion in response to anti-BCMA CAR T cells in a patient with multiple myeloma

Abstract

B cell maturation antigen (BCMA) is a target for various immunotherapies and a biomarker for tumor load in multiple myeloma (MM). We report a case of irreversible BCMA loss in a patient with MM who was enrolled in the KarMMa trial (NCT03361748) and progressed after anti-BCMA CAR T cell therapy. We identified selection of a clone with homozygous deletion of TNFRSF17 (BCMA) as the underlying mechanism of immune escape. Furthermore, we found heterozygous TNFRSF17 loss or monosomy 16 in 37 out of 168 patients with MM, including 28 out of 33 patients with hyperhaploid MM who had not been previously treated with BCMA-targeting therapies, suggesting that heterozygous TNFRSF17 deletion at baseline could theoretically be a risk factor for BCMA loss after immunotherapy.

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Fig. 1: Clinical characteristics and BCMA expression from baseline to relapse.
Fig. 2: WGS and read coverage of CD138+ purified plasma cells at progression from CAR T cell therapy.

Data availability

All requests for raw and analyzed data and materials will be promptly reviewed by University Hospital Würzburg and Münchner Leukämielabor to verify if the request is subject to any confidentiality and data protection obligations. Any data and materials that can be shared will be released via a material transfer agreement. All raw and analyzed scRNA-seq data can be found in the National Center for Biotechnology Information Gene Expression Omnibus (accession no. GSE143317). Adherence to the Datenschutz-Grundverordnung (https://dsgvo-gesetz.de/ and https://data.consilium.europa.eu/doc/document/ST-5419-2016-REV-1/en/pdf) is mandatory for sharing WGS data. Thus, the complete whole-genome raw dataset of a single patient cannot be shared according to European law. Processed data from which the identification of a patient is not possible can be made available. WGS data derived from public datasets are deposited in the European Genome-phenome Archive with the references EGAS00001002111 (ref. 7) and EGAS00001001810 (ref. 9). Source data are provided with this paper.

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Acknowledgements

We thank S. Roth for her excellent technical support. We acknowledge the work and contribution of T. Mack from Bristol Myers Squibb. L.R. was supported by the German Cancer Aid via the MSNZ program and IZKF Würzburg. H.E and M.H. were supported by the DFG via SFB/TR 221 project A3. A.-E.S. and O.D. were supported by the German Research Society graduate college 2157. Bristol Myers Squibb provided clinical trial data related to the presented case but no financial support for the study.

Author information

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Authors

Contributions

L.R., M.C.D.V., K.H., A.-E.S. and H.E. contributed to study design and manuscript preparation. K.M.K., M.G., M.C., J.D., S. Kraus, A.H., X.Z., S.D., M.S.T., M.H. and S.P. contributed to the collection of clinical specimens. A.R. and V.F. performed the immunohistochemistry analysis. M.C.D.V., O.D., P.A., V.F. and A.-E.S. performed the scRNA-seq. O.D. performed the scRNA-seq data analysis. M.T., S.M., S.T., M.M. and C.H. performed whole-genome and bulk RNA sequencing. F.E. contributed to the bioinformatics analysis. N.W. analyzed the additional patent cohorts. S. Kaiser and K.H. provided clinical trial data. All authors approved the manuscript.

Corresponding author

Correspondence to Leo Rasche.

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Competing interests

K.H. and S.K. are employees of Bristol Myers Squibb. M.H. is listed as an inventor on patent applications and granted patents related to CAR T cell therapy and CAR T cell technologies; he has received speaker honoraria from Celgene/Bristol Myers Squibb, Janssen and Kite/Gilead. H.E. received research funding from Janssen, Bristol Myers Squibb/Celgene, Amgen, Novartis, Sanofi, GSK and Janssen and has received speaker honoraria from Bristol Myers Squibb/Celgene, Amgen, Novartis, Takeda, Sanofi and GSK. L.R. has received speaker honoraria from Bristol Myers Squibb/Celgene, Sanofi, GSK, Oncopeptides and Janssen. The remaining authors declare no competing interests.

Additional information

Peer review information Nature Medicine thanks Eric Smith, Premal Lulla and Alfred Garfall for their contribution to the peer review of this work. Saheli Sadanand was the primary editor on this article and managed its editorial process and peer review in collaboration with the rest of the editorial team.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Extended Data Fig. 1 Baseline characteristics and previous therapies.

Summary of patient baseline characteristics and previous therapies before inclusion on the KarMMa trial. Abbreviations: Rev, Lenalidomide; Dara, Daratumumab; SC mob, stem cell mobilization; auto SCT, autologous stem cell transplantation; Pom, Polmalidomide; Carf, Carfilzomib; Cyclo, Cyclophosphamide; Dexa, Dexamethasone; Thal, Thalidomide.

Extended Data Fig. 2 Cell type annotation in bone marrow biopsies.

a, Schematic of the experimental workflow. b, UMAP embedding of 5,592 single-cell transcriptomes from bone marrow biopsies before treatment (Baseline) and after relapse (Relapse). c, as b showing the annotated cell types. d, Dotplot showing the mean (of scaled log-normalized counts) expression (color) and the proportion in percentage of positive (non-zero expression) cells per cluster (size). MM, Multiple Myeloma. e, Log-normalized expression of marker genes specific for the identified cell types (MM: IGHG1+; B/DC: HLA-DRA+, CD79B+, LILRA4+; T/NK: CD3D+, NKG7+; Neutrophil: HLA-DRA-, FCGR3B+; Monocyte: HLA-DRA+, CD68+; Ery: ALAS2+) color coded on the UMAP embedding as panels b and c. Source data

Extended Data Fig. 3 TNFRSF17 (BCMA) expression from baseline to relapse.

a, UMAP embedding of 2,471 single-cell transcriptomes of Multiple Myeloma cells (MM cells from Extended Data Fig. 2 extracted and re-analyzed) from bone marrow biopsies before treatment (45 MM single cells, Baseline, blue) and after relapse (2,426 MM single cells, Relapse, orange). b, As panel a colored by the log-normalized expression of TNFRSF17 (BCMA). c, As panel a colored by cluster. d, Dotplot of scaled, log-normalized expression (color) of marker genes identified for the different clusters shown in panel d and the proportion in percentage of positive (non-zero expression) cells per cluster (size). e, Violin plot showing the expression (log-normalized) of target antigens of immunotherapy between baseline and relapse. Horizontal line shows median of expression value. f, Expression of indicated genes shown in e color coded on top of the UMAP embedding as in panels a and c. Source data

Extended Data Fig. 4 T cell state characterization at baseline and relapse.

a, UMAP embedding of 1,044 T cells from bone marrow biopsies before (635, blue dots) and at relapse (409, orange dots). b, UMAP embedding (as panel a) showing T cell subsets. c, Dotplot showing the scaled, log-normalized expression (color) of canonical marker genes for each cell subset and the proportion in percentage of positive (non-zero expression) cells per cluster (size). d, Violin-plots showing log-normalized expression of T cell exhaustion (LAG3, TIGIT, PDCD1, HAVCR2) and activation markers (IFNG, IFIT1, IFIT3, IFI16, HLA-DRA, HLA-DRB1, HLA-DRB5) between baseline (orange) and relapse (blue) across different cell subsets. P-values were calculated using the two-sided wilcoxon rank sum test, no adjustment for multiple comparisons was performed. Horizontal line shows median of expression value. Abbreviations: Th: Helper T cells; Treg: Regulatory T cells; NK: Natural killer cells; Prolif: Proliferating T cells. Source data

Extended Data Fig. 5 Genomic architecture of baseline sample.

Circos plot of CD138+ selected cells from the baseline sample including copy number variations, structural variants, and single nucleotide variations based on WGS data. Outer track runs clockwise from chromosome 1 to Y. Inner track shows gains > 1 Mb in blue and losses > 1 Mb in red. Red lines inside the circle represent interchromosomal reciprocal translocations with vaf > 0.1. Genes with mutations (NRAS, ERBB4 and TP53) are depicted in red and genes with detected variants of unknown significance (TTN) are shown in grey.

Extended Data Fig. 6 Copy number variant analysis.

a, Normalized Log2-ratio of gene expression within the cytogenetic band 16p13.13 between baseline and progression (n = number MM cells=45 MM cells at baseline and n = 2,426 MM cells at relapse). Error bars represent 90 % credible intervals. b, Total UMI count in non-malignant control cells at baseline vs. progression. We focused on the changes of expression levels from baseline to progression for genes located in the heterozygous deletion region. If the patient acquired the heterozygous deletion after treatment, systematic changes of gene expression are expected in this region. In contrast, this is not expected with the heterozygous deletion already present at baseline. In our patient, there were strong expression changes for genes in this region in malignant plasma cells (median > 3.7-fold difference) but not in monocytes (median < 1.3-fold difference) that we analyzed as negative control.

Extended Data Fig. 7 Interphase FISH of the 16p heterozygous deletion.

Interphase Fluorescent-in-situ-hybridization (FISH) using the CBFB/MYH11 probe set in a patient with heterozygous del(16p) and concomitant del(16q). Two aberrant cells show only 1 signal instead of 2 for the respective probes in line with two deletions encompassing the MYH11 (located on 16p13.1) and CBFB (located on 16q22) locus.

Extended Data Fig. 8 TNFRSF17 RNA-seq expression data from 28 relapsed/refractory multiple myeloma (R/R MM) patients.

TNFRSF17 (BCMA) expression levels in patients with and w/o BCMA heterozygous deletions. Bulk RNAseq data for 28 R/R MM patients with availability of WGS data. Two patients showed a heterozygous TNFRSF17 deletion (red dots), whereas the remaining patients did not show BCMA aberrations (grey dots). Expression levels did not differ between the two groups.

Supplementary information

Supplementary Information

Supplementary Tables 1–3.

Reporting Summary

Source data

Source Data Extended Data Fig. 2

Statistical source data.

Source Data Extended Data Fig. 3

Statistical source data.

Source Data Extended Data Fig. 4

Statistical source data.

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Da Vià, M.C., Dietrich, O., Truger, M. et al. Homozygous BCMA gene deletion in response to anti-BCMA CAR T cells in a patient with multiple myeloma. Nat Med (2021). https://doi.org/10.1038/s41591-021-01245-5

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