Title |
Alternative splicing of its 5′-UTR limits CD20 mRNA translation and enables resistance to CD20-directed immunotherapies
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Published in |
Blood, September 2023
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DOI | 10.1182/blood.2023020400 |
Pubmed ID | |
Authors |
Zhiwei Ang, Luca Paruzzo, Katharina E. Hayer, Carolin Schmidt, Manuel Torres Diz, Feng Xu, Urvi Zankharia, Yunlin Zhang, Samantha Soldan, Sisi Zheng, Catherine D. Falkenstein, Joseph P. Loftus, Scarlett Y. Yang, Mukta Asnani, Patricia King Sainos, Vinodh Pillai, Emeline Chong, Marilyn M. Li, Sarah K. Tasian, Yoseph Barash, Paul M. Lieberman, Marco Ruella, Stephen J. Schuster, Andrei Thomas-Tikhonenko |
Abstract |
Aberrant skipping of coding exons in CD19 and CD22 compromises responses to immunotherapy for B-cell malignancies. Here, we show that the MS4A1 gene encoding human CD20 also produces several mRNA isoforms with distinct 5' untranslated regions (5'-UTR). Four variants (V1-4) were detectable by RNA-seq in distinct stages of normal B-cell differentiation and B-lymphoid malignancies, with V1 and V3 being the most abundant by far. During B-cell activation and Epstein-Barr virus infection, redirection of splicing from V1 to V3 coincided with increased CD20 positivity. Similarly, in diffuse large B-cell lymphoma only V3, but not V1, correlated with CD20 protein levels, suggesting that V1 might be translation-deficient. Indeed, the longer V1 isoform was found to contain upstream open reading frames (uORFs) and a stem-loop structure, which cooperatively inhibited polysome recruitment. By modulating CD20 isoforms with splice-switching Morpholino oligomers, we enhanced CD20 expression and anti-CD20 antibody rituximab-mediated cytotoxicity in a panel of B-cell lines. Furthermore, reconstitution of CD20-knockout cells with V3 mRNA led to the recovery of CD20 positivity, while V1-reconstituted cells had undetectable levels of CD20 protein. Surprisingly, in vitro CD20-directed CAR T cells were able to kill both V3- and V1-expressing cells, but the bispecific T cell engager mosunetuzumab was only effective against V3-expressing cells. To determine whether CD20 splicing is involved in immunotherapy resistance, we performed RNA-seq on four post-mosunetuzumab follicular lymphoma relapses and discovered that in two of them downregulation of CD20 was accompanied by the V3-to-V1 shift. Thus, splicing-mediated mechanisms of epitope loss extend to CD20-directed immunotherapies. |
X Demographics
Geographical breakdown
Country | Count | As % |
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United States | 11 | 35% |
France | 2 | 6% |
Italy | 1 | 3% |
Canada | 1 | 3% |
Comoros | 1 | 3% |
Unknown | 15 | 48% |
Demographic breakdown
Type | Count | As % |
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Members of the public | 20 | 65% |
Scientists | 10 | 32% |
Science communicators (journalists, bloggers, editors) | 1 | 3% |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
Unknown | 24 | 100% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Researcher | 6 | 25% |
Unspecified | 2 | 8% |
Student > Ph. D. Student | 2 | 8% |
Student > Bachelor | 1 | 4% |
Student > Doctoral Student | 1 | 4% |
Other | 1 | 4% |
Unknown | 11 | 46% |
Readers by discipline | Count | As % |
---|---|---|
Medicine and Dentistry | 4 | 17% |
Biochemistry, Genetics and Molecular Biology | 3 | 13% |
Unspecified | 2 | 8% |
Immunology and Microbiology | 2 | 8% |
Agricultural and Biological Sciences | 1 | 4% |
Other | 0 | 0% |
Unknown | 12 | 50% |