Immunotherapy-resistant acute lymphoblastic leukemia cells exhibit reduced CD19 and CD22 expression and BTK pathway dependency
Sarah Aminov, Orsi Giricz, David T. Melnekoff, R. Alejandro Sica, Veronika Polishchuk, Cristian Papazoglu, Bonnie Yates, Hao-Wei Wang, Srabani Sahu, Yanhua Wang, Shanisha Gordon-Mitchell, Violetta V. Leshchenko, Carolina Schinke, Kith Pradhan, Srinivas Aluri, Moah Sohn, Stefan K. Barta, Beamon Agarwal, Mendel Goldfinger, Ioannis Mantzaris, Aditi Shastri, William Matsui, Ulrich Steidl, Joshua D. Brody, Nirali N. Shah, Samir Parekh, Amit Verma
Sarah Aminov, Orsi Giricz, David T. Melnekoff, R. Alejandro Sica, Veronika Polishchuk, Cristian Papazoglu, Bonnie Yates, Hao-Wei Wang, Srabani Sahu, Yanhua Wang, Shanisha Gordon-Mitchell, Violetta V. Leshchenko, Carolina Schinke, Kith Pradhan, Srinivas Aluri, Moah Sohn, Stefan K. Barta, Beamon Agarwal, Mendel Goldfinger, Ioannis Mantzaris, Aditi Shastri, William Matsui, Ulrich Steidl, Joshua D. Brody, Nirali N. Shah, Samir Parekh, Amit Verma
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Research Article Oncology

Immunotherapy-resistant acute lymphoblastic leukemia cells exhibit reduced CD19 and CD22 expression and BTK pathway dependency

Abstract

While therapies targeting CD19 by antibodies, chimeric antigen receptor T cells (CAR-T), and T cell engagers have improved the response rates in B cell malignancies, the emergence of resistant cell populations with low CD19 expression can lead to relapsed disease. We developed an in vitro model of adaptive resistance facilitated by chronic exposure of leukemia cells to a CD19 immunotoxin. Single-cell RNA-Seq (scRNA-Seq) showed an increase in transcriptionally distinct CD19lo populations among resistant cells. Mass cytometry demonstrated that CD22 was also decreased in these CD19lo-resistant cells. An assay for transposase-accessible chromatin with sequencing (ATAC-Seq) showed decreased chromatin accessibility at promoters of both CD19 and CD22 in the resistant cell populations. Combined loss of both CD19 and CD22 antigens was validated in samples from pediatric and young adult patients with B cell acute lymphoblastic leukemia (B-ALL) that relapsed after CD19 CAR-T–targeted therapy. Functionally, resistant cells were characterized by slower growth and lower basal levels of MEK activation. CD19lo resistant cells exhibited preserved B cell receptor signaling and were more sensitive to both Bruton’s tyrosine kinase (BTK) and MEK inhibition. These data demonstrate that resistance to CD19 immunotherapies can result in decreased expression of both CD19 and CD22 and can result in dependency on BTK pathways.

Authors

Sarah Aminov, Orsi Giricz, David T. Melnekoff, R. Alejandro Sica, Veronika Polishchuk, Cristian Papazoglu, Bonnie Yates, Hao-Wei Wang, Srabani Sahu, Yanhua Wang, Shanisha Gordon-Mitchell, Violetta V. Leshchenko, Carolina Schinke, Kith Pradhan, Srinivas Aluri, Moah Sohn, Stefan K. Barta, Beamon Agarwal, Mendel Goldfinger, Ioannis Mantzaris, Aditi Shastri, William Matsui, Ulrich Steidl, Joshua D. Brody, Nirali N. Shah, Samir Parekh, Amit Verma

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Figure 5

Cells resistant to CD19 immunotherapy exhibit dependency on BCR and MEK signaling.

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Cells resistant to CD19 immunotherapy exhibit dependency on BCR and MEK ...
(A and B) Immunoblotting shows that ibrutinib could inhibit p-BTK in resistant cells. Resistant cells had preserved ERK activation. (C) p-ERK/total ERK ratio in Raji cells shows that p-ERK was maintained in Raji CD19-KO cells. (D) p-BTK/total BTK ratio in NALM6 cells shows that p-BTK was maintained in NALM6-R cells and that ibrutinib effectively inhibited phosphorylation. (E and F) NALM6-R cells were more sensitive to MEK and BTK inhibition (n = 4). (G) A patient with ALL resistant to CD19 immunotoxin (Resistant ALL patient 1) was sensitive to the MEK inhibitor but resistant to methotrexate (n = 3). (H) A patient with ALL resistant to blinatumomab (Resistant ALL patient 2) was sensitive to the MEK inhibitor but resistant to methotrexate (n = 3). (I) Proposed model showing that resistant ALL cells lose CD19 and CD22 expression and maintain BTK expression, leading to slower leukemic proliferation and increased sensitivity to BTK and ERK inhibition. The model was created with BioRender.com.