CD33 splicing SNP regulates expression levels of CD33 in normal regenerating monocytes in AML patients

Lamba, Jatinder K.; Voigt, Andrew P.; Chauhan, Lata; Shin, Miyoung; Aplenc, Richard; Brodersen, Lisa Eidenschink; Gamis, Alan S.; Meshinchi, Soheil; Loken, Michael R.

doi:10.1080/10428194.2017.1421756

Cited by 8 publications

(5 citation statements)

References 11 publications

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“…The function of CD33 in AML is poorly understood, however CD33 ligation by a monoclonal antibody has been shown to inhibit AML proliferation ( 21 ). The expression of CD33 molecule in AML is regulated, in part, by alternative splicing ( 22 ). At least two distinct CD33 isoforms have been identified to date.…”

Section: Introductionmentioning

confidence: 99%

A Unique Human Immunoglobulin Heavy Chain Variable Domain-Only CD33 CAR for the Treatment of Acute Myeloid Leukemia

Schneider

Xiong

et al. 2018

Front. Oncol.

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Acute myeloid leukemia (AML) remains a challenging pediatric and adult disease. Given the elevated expression of the CD33 antigen on leukemic blasts, therapeutic approaches to AML now feature the approved antibody drug conjugate (Mylotarg, GO) and investigational CART cell approaches incorporating CD33-binding domains derived from humanized scFvs. We designed a functional chimeric antigen receptor utilizing a human targeting sequence, derived from a heavy chain variable domain, termed CAR33VH. Lentiviral-based expression vectors which encoded CAR constructs incorporating the novel binding domain (CAR33VH), or the My96 scFv control binder (My96CAR) in frame with a CD8 hinge and transmembrane domain, a 4-1BB costimulatory domain and a CD3 zeta activation domain, were transduced into primary human CD4+ and CD8+ T cells, and CAR expression was confirmed by flow cytometry. CAR33VH, similar to My96CAR, demonstrated robust and specific cytotoxicity in short-term and long-term co-incubation killing assays against CD33+ AML lines. In overnight cytokine release assays in which CAR T cells were challenged with the CD33+ tumor cells HL-60, MOLM-14 and KG-1a, CAR33VH elicited IFN-gamma, TNF-alpha and IL-2. This was seen with CD33+ cell lines, but not when CAR T were cultured alone. Studies with a CD33− cell line engineered to stably express the full length CD33 variant 1, or the naturally occurring CD33 splice variant 2, revealed that both CAR33VH and My96CAR, target the V domain of CD33, suggesting a similar therapeutic profile. Colony-formation assays utilizing peripheral blood CD34+ hematopoietic stem cells treated with CAR33VH, My96CAR, or with an untransduced T cell control, yielded similar numbers of BFU-E erythroid and CFU-GM myeloid colonies, suggesting a lack of CAR-related overt toxicity. In an in vivo AML model, NSG mice engrafted with MOLM-14 cells stably expressing firefly luciferase, both CAR33VH and CARMy96 efficiently eliminated tumors. In conclusion, we demonstrate for the first time the feasibility and efficacy of employing human variable domain-only binder derived from a phage display library in an anti-AML CAR design. CAR33VH, comprised of a human heavy-chain variable fragment-only antigen binding domain, was efficient in tumor killing in vitro and in vivo, and showed comparable functionality to the scFv-based My96CAR.

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Section: Introductionmentioning

confidence: 99%

A Unique Human Immunoglobulin Heavy Chain Variable Domain-Only CD33 CAR for the Treatment of Acute Myeloid Leukemia

Schneider

Xiong

et al. 2018

Front. Oncol.

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“…Because alternative splicing impacts CD33 expression ( 8 , 12 , 13 ), we included cDNAs encoding both full-length CD33M and the major alternative shorter transcript CD33m, which lacks the membrane-distal Ig-like IgV2 sialic acid–binding domain encoded by exon 2 ( 14 – 16 ), in our screening library. Expression of CD33M resulted in a strong fluorescent signal following staining with all three V H H-Fc fusion proteins, while expression of CD33m resulted in a strong fluorescent signal only following staining with V H H1-Fc ( Supplemental Figure 1A and Supplemental Figure 3A ).…”

Section: Resultsmentioning

confidence: 99%

Drug-regulated CD33-targeted CAR T cells control AML using clinically optimized rapamycin dosing

Appelbaum,

Price,

Oda

et al. 2024

Journal of Clinical Investigation

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Chimeric antigen receptor (CAR) designs that incorporate pharmacologic control are desirable; however, designs suitable for clinical translation are needed. We designed a fully human, rapamycin-regulated drug product for targeting CD33 + tumors called dimerizaing agent–regulated immunoreceptor complex (DARIC33). T cell products demonstrated target-specific and rapamycin-dependent cytokine release, transcriptional responses, cytotoxicity, and in vivo antileukemic activity in the presence of as little as 1 nM rapamycin. Rapamycin withdrawal paused DARIC33-stimulated T cell effector functions, which were restored following reexposure to rapamycin, demonstrating reversible effector function control. While rapamycin-regulated DARIC33 T cells were highly sensitive to target antigen, CD34 + stem cell colony-forming capacity was not impacted. We benchmarked DARIC33 potency relative to CD19 CAR T cells to estimate a T cell dose for clinical testing. In addition, we integrated in vitro and preclinical in vivo drug concentration thresholds for off-on state transitions, as well as murine and human rapamycin pharmacokinetics, to estimate a clinically applicable rapamycin dosing schedule. A phase I DARIC33 trial has been initiated (PLAT-08, NCT05105152), with initial evidence of rapamycin-regulated T cell activation and antitumor impact. Our findings provide evidence that the DARIC platform exhibits sensitive regulation and potency needed for clinical application to other important immunotherapy targets.

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“…CD33 is transmembrane receptor expressed on cells of myeloid lineage and is usually considered to be myeloid specific . FCM analysis showed that CD33 expression was reduced in the CRT siRNA‐transfected HL‐60 cells compared to controls (51.21% vs 93.47%, respectively; Figure B).…”

Section: Resultsmentioning

confidence: 99%

Diallyl disulfide down‐regulates calreticulin and promotes C/EBPα expression in differentiation of human leukaemia cells

Sun

et al. 2018

J Cellular Molecular Medi

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Diallyl disulfide (DADS), the main active component of the cancer fighting allyl sulfides found in garlic, has shown potential as a therapeutic agent in various cancers. Previous studies showed DADS induction of HL‐60 cell differentiation involves down‐regulation of calreticulin (CRT). Here, we investigated the mechanism of DADS‐induced differentiation of human leukaemia cells and the potential involvement of CRT and CCAAT enhancer binding protein‐α (C/EBPα). We explored the expression of CRT and C/EBPα in clinical samples (20 healthy people and 19 acute myeloid leukaemia patients) and found that CRT and C/EBPα expressions were inversely correlated. DADS induction of differentiation of HL‐60 cells resulted in down‐regulated CRT expression and elevated C/EBPα expression. In severe combined immunodeficiency mice injected with HL‐60 cells, DADS inhibited the growth of tumour tissue and decreased CRT levels and increased C/EBPα in vivo. We also found that DADS‐mediated down‐regulation of CRT and up‐regulation of C/EBPα involved enhancement of reactive oxidative species. RNA immunoprecipitation revealed that CRT bound C/EBPα mRNA, indicating its regulation of C/EBPα mRNA degradation by binding the UG‐rich element in the 3′ untranslated region of C/EBPα. In conclusion, the present study demonstrates the C/EBPα expression was correlated with CRT expression in vitro and in vivo and the molecular mechanism of DADS‐induced leukaemic cell differentiation.

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CD33 splicing SNP regulates expression levels of CD33 in normal regenerating monocytes in AML patients

Cited by 8 publications

References 11 publications

A Unique Human Immunoglobulin Heavy Chain Variable Domain-Only CD33 CAR for the Treatment of Acute Myeloid Leukemia

A Unique Human Immunoglobulin Heavy Chain Variable Domain-Only CD33 CAR for the Treatment of Acute Myeloid Leukemia

Drug-regulated CD33-targeted CAR T cells control AML using clinically optimized rapamycin dosing

Diallyl disulfide down‐regulates calreticulin and promotes C/EBPα expression in differentiation of human leukaemia cells

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