Feasibility and preclinical efficacy of CD7-unedited CD7 CAR T cells for T cell malignancies

Watanabe, Norihiro; Mo, Feiyan; Zheng, Rong; Ma, Royce; Bray, Vanesa C.; Leeuwen, Dayenne G. van; Sritabal-Ramirez, Juntima; Hu, Hongxiang; Wang, Sha; Mehta, Birju; Srinivasan, Madhuwanti; Scherer, Lauren; Zhang, Huimin; Thakkar, Sachin G.; Hill, LaQuisa; Heslop, Helen E.; Brenner, Malcolm K.; Mamonkin, Maksim

doi:10.1016/j.ymthe.2022.09.003

Cited by 34 publications

(35 citation statements)

References 39 publications

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“…The expression of CAR45 and CD45 was verified on days 7 and 14 post CD45 knockout ( Figure 2e) . Inspired by previous studies showing that fratricide of CAR T cells can be fully prevented using dasatinib, the pharmacologic inhibitor of key CAR/CD3ζ signaling kinases 46 , we supplemented the T cell culture with 50 nM dasatinib, which rescued the expansion and viability of CAR45 and CD45 Δ CAR45 T cells ( Figure 2d right and f). Interestingly, in the culture of CD45 Δ CAR45 T cells, despite the inclusion of dasatinib, we observed the elimination of residual CD45-positive cells and the growth of the CD45 Δ CAR45 population on day 14 ( Figure 2g).…”

Section: Resultsmentioning

confidence: 99%

“…Only recent studies that developed gene-editing approaches to generate CD5- and CD7-specific CAR-T cells demonstrated the possibility of obviating fratricide. Of note, naturally CD5-negative and CD7-negative T cells can be selected to produce CAR T cells of the relevant specificity, a phenomenon not described for CD45 31,36,46 . (2) Considering the severe combined immune deficiency phenotype generated by human constitutional CD45 deficiency 61 , the idea that the removal of such an important molecule as CD45 will retain T-cell functionality seems counterintuitive.…”

Section: Discussionmentioning

confidence: 99%

Section: Generation Of Fratricide-resistant Cd45-specific Car T Cellsmentioning

confidence: 99%

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Targeting CD45 by gene-edited CAR-T cells for leukemia eradication and hematopoietic stem cell transplantation preconditioning

Stepanova,

Volkov,

Osipova

et al. 2023

Preprint

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Hematopoietic stem cell transplantation (HSCT) is widely used to treat patients with life-threatening hematologic and immune system disorders. The currently used nontargeted chemo- /radiotherapy conditioning regimens cause tissue injury and induce an array of immediate and delayed adverse effects, which limits the use of this potentially curative treatment. The growing demand to replace canonical conditioning regimens has led to the development of alternative approaches based on antibody‒drug conjugates, naked antibodies and CAR T cells. Here, we propose a preconditioning strategy based on targeting CD45 on hematopoietic cells with CAR45 T cells. To avoid fratricide of CD45 CAR T cells, targeted genomic disruption of the CD45 gene was performed in human CD45 CAR T cells in combination with dasatinib treatment. CD45DCAR45 T cells showed impressive activity in terms of target cell elimination in vitro and depletion of tumor cells in vivo or human hematopoietic cells in humanized immunodeficient mice engrafted with human blood-derived HSCs. CD45DCAR45 NK cells also exhibited potent killing activity against tumor cell lines and human hematopoietic cells. Therefore, fratricide-resistant CAR45 T and NK cells have the potential to provide the benefits of full myeloablative conditioning and therapy for hematologic malignancies. Thus, we provide the proof of concept for the generation and preclinical efficacy of CAR T cells directed against CD45-expressing cells.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Generation Of Fratricide-resistant Cd45-specific Car T Cellsmentioning

confidence: 99%

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Targeting CD45 by gene-edited CAR-T cells for leukemia eradication and hematopoietic stem cell transplantation preconditioning

Stepanova,

Volkov,

Osipova

et al. 2023

Preprint

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“…The above study demonstrated the persistence of naturally selected CD7 CAR T cells in the in vitro phase of the study. However, further validation of its durability in comparison with other reported CD7 CAR-T cells and in clinical trials of R/R-AML needs to be performed [15,[51][52][53][54].…”

Section: Discussionmentioning

confidence: 99%

“…In addition, CD7 CAR-Ts may be less costly and exhibit lower risks associated with gene editing. In addition, in recent years, the construction of CAR-T cells by isolating CD7-negative cell populations or the use of ibrutinib and dasatinib to inhibit fratricide has demonstrated good potential value [52,54]. Hai-Ping Dai et al demonstrated the safety and efficacy of CD7 CAR-T cells using protein expression blockers to block CD7 expression at the CAR-T-cell membrane for the treatment of R/R early T-cell precursor lymphoblastic leukaemia/lymphoma (ETP-ALL/LBL) in patients with TP53 mutations [51].…”

Section: Discussionmentioning

confidence: 99%

Naturally selected CD7 CAR-T therapy without genetic editing demonstrates significant antitumour efficacy against relapsed and refractory acute myeloid leukaemia (R/R-AML)

Liu

Wen

et al. 2022

J Transl Med

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Background The survival rate for patients with relapsed and refractory acute myeloid leukaemia (R/R-AML) remains poor, and treatment is challenging. Chimeric antigen receptor T cells (CAR-T cells) have been widely used for haematologic malignancies. Current CAR-T therapies for acute myeloid leukaemia mostly target myeloid-lineage antigens, such as CD123 and CD33, which may be associated with potential haematopoietic toxicity. As a lineage-specific receptor, CD7 is expressed in acute myeloid leukaemia cells and T cells but is not expressed in myeloid cells. Therefore, the use of CD7 CAR-T cells for R/R-AML needs to be further explored. Methods In this report, immunohistochemistry and flow cytometry were used to analyse CD7 expression in clinical samples from R/R-AML patients and healthy donors (HDs). We designed naturally selected CD7 CAR-T cells to analyse various functions and in vitro antileukaemic efficacy based on flow cytometry, and xenograft models were used to validate in vivo tumour dynamics. Results We calculated the percentage of cells with CD7 expression in R/R-AML patients with minimal residual disease (MRD) (5/16, 31.25%) from our institution and assessed CD7 expression in myeloid and lymphoid lineage cells of R/R-AML patients, concluding that CD7 is expressed in T cells but not in myeloid cells. Subsequently, we designed and constructed naturally selected CD7 CAR-T cells (CD7 CAR). We did not perform CD7 antigen knockdown on CD7 CAR-T cells because CD7 molecule expression is naturally eliminated at Day 12 post transduction. We then evaluated the ability to target and kill CD7+ acute myeloid leukaemia cells in vitro and in vivo. Naturally selected CD7 CAR-T cells efficiently killed CD7+ acute myeloid leukaemia cells and CD7+ primary blasts of R/R-AML patients in vitro and significantly inhibited leukaemia cell growth in a xenograft mouse model. Conclusion Naturally selected CD7 CAR-T cells represent an effective treatment strategy for relapsed and refractory acute myeloid leukaemia patients in preclinical studies.

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CD38‐Specific CAR Integrated into CD38 Locus Driven by Different Promoters Causes Distinct Antitumor Activities of T and NK Cells

et al. 2023

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The robust and stable expression of CD38 in T‐cell acute lymphoblastic leukemia (T‐ALL) blasts makes CD38 chimeric antigen receptor (CAR)‐T/natural killer (NK) a potential therapy for T‐ALL. However, CD38 expression in normal T/NK cells causes fratricide of CD38 CAR‐T/NK cells. Here a “2‐in‐1” gene editing strategy is developed to generate fratricide‐resistant locus‐specific CAR‐T/NK cells. CD38‐specific CAR is integrated into the disrupted CD38 locus by CRISPR/Cas9, and CAR is placed under the control of either endogenous CD38 promoter (CD38KO/KI) or exogenous EF1α promoter (CD38KO/KIEF1α). CD38 knockout reduces fratricide and allows the expansion of CAR‐T cells. Meanwhile, CD38KO/KIEF1α results in higher CAR expression than CD38KO/KI in both CAR‐T and CAR‐NK cells. In a mouse T‐ALL model, CD38KO/KIEF1α CAR‐T cells eradicate tumors better than CD38KO/KI CAR‐T cells. Surprisingly, CD38KO/KI CAR‐NK cells show superior tumor control than CD38KO/KIEF1α CAR‐NK cells. Further investigation reveals that endogenous regulatory elements in NK cells lead to higher expression of CD38 CAR than in T cells, and the expression levels of CAR affect the therapeutic outcome of CAR‐T and CAR‐NK cells differently. Therefore, these results support the efficacy of CD38 CAR‐T/NK against T‐ALL and demonstrate that the “2‐in‐1” strategy can resolve fratricide and enhance tumor eradication, paving the way for clinical translation.

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Feasibility and preclinical efficacy of CD7-unedited CD7 CAR T cells for T cell malignancies

Cited by 34 publications

References 39 publications

Targeting CD45 by gene-edited CAR-T cells for leukemia eradication and hematopoietic stem cell transplantation preconditioning

Targeting CD45 by gene-edited CAR-T cells for leukemia eradication and hematopoietic stem cell transplantation preconditioning

Naturally selected CD7 CAR-T therapy without genetic editing demonstrates significant antitumour efficacy against relapsed and refractory acute myeloid leukaemia (R/R-AML)

CD38‐Specific CAR Integrated into CD38 Locus Driven by Different Promoters Causes Distinct Antitumor Activities of T and NK Cells

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