DGK-α: A Checkpoint in Cancer-Mediated Immuno-Inhibition and Target for Immunotherapy

Noeßner, Elfriede

doi:10.3389/fcell.2017.00016

Cited by 29 publications

(33 citation statements)

References 48 publications

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“…In particular, our screen showed that knockdown of diacylglycerol kinase (DGK) family of genes resulted in increased sensitivity to CAR-T cells. Previous groups have shown that inhibition of DGK-α activity in T cells using pharmacological inhibitors induced T cell activation, thus improving its cytotoxic activity on cancer cells (33,38). Our study indicates that knockdown of DGK kinases in myeloma cells increases sensitivity to T cells.…”

Section: Bcma Car-t Cell Co-culture Screen Reveals Myeloma Cell-intrisupporting

confidence: 58%

CRISPR-based screens uncover determinants of immunotherapy response in Multiple Myeloma

Ramkumar

Abarientos

Tian

et al. 2019

Preprint

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Cancer cells commonly develop resistance to immunotherapy by loss of antigen expression. Combinatorial treatments that increase levels of the target antigen on the surface of cancer cells have the potential to restore efficacy to immunotherapy. Here, we use our CRISPR interference and CRISPR activation-based functional genomics platform to systematically identify pathways controlling cell-surface expression of the multiple myeloma immunotherapy antigen - B cell maturation antigen, BCMA. We discovered that pharmacological inhibition of HDAC7 and the Sec61 complex increased cell-surface BCMA, including in primary patient cells. Importantly, pharmacological Sec61 inhibition enhanced the anti-myeloma efficacy of a BCMA-targeted antibody-drug conjugate. A CRISPR interference CAR-T coculture screen enabled us to identify both antigen-dependent and -independent mechanisms controlling response of myeloma cells to BCMA-targeted CAR-T cells. Thus, our study demonstrates the potential of CRISPR screens to uncover mechanisms controlling response of cancer cells to immunotherapy and to suggest potential combination therapies.Key PointsUsing CRISPR screens, we systematically identify mechanisms increasing expression of the immunotherapy target BCMA and ADC efficacy.We also identify antigen-independent mechanisms regulating response of cancer cells to BCMA-CAR-T cells.

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Section: Bcma Car-t Cell Co-culture Screen Reveals Myeloma Cell-intrisupporting

confidence: 58%

CRISPR-based screens uncover determinants of immunotherapy response in Multiple Myeloma

Ramkumar

Abarientos

Tian

et al. 2019

Preprint

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“…In addition to these issues, immunologic differences between human and mouse T cells exist. For instance, unlike mouse T cells, human CD8 T cells lose CD28 expression during the developmental process (27). This is critical because CD28 inhibits DGKa activity and DGKa-mediated anergy.…”

Section: Introductionmentioning

confidence: 99%

CRISPR/Cas9-Mediated Knockout of DGK Improves Antitumor Activities of Human T Cells

et al. 2018

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The efficacy of T-cell therapy is inhibited by various tumor-associated immunosuppressive ligands and soluble factors. Such inhibitory signals turn specific T-cell signaling pathways on or off, impeding the anticancer functions of T cells. Many studies have focused on PD-1 or CTLA-4 blockade to invigorate T-cell functions through CD28/B7 signaling, but obtaining robust clinical outcomes remains challenging. In this study, we use CRISPR/Cas9 to potentiate T-cell function by increasing CD3 signaling via knockout of diacylglycerol kinase (DGK), an enzyme that metabolizes diacylglycerol to phosphatidic acid. Knockout of DGK augmented the effector functions of CAR-T cells via increased TCR signaling. DGK knockout from CAR-T cells rendered them resistant to soluble immunosuppressive factors such as TGFβ and prostaglandin E2 and sustained effector functions under conditions of repeated tumor stimulation. Moreover, DGK knockout caused significant regression of U87MGvIII glioblastoma tumors through enhanced effector functions in a xenograft mouse model. Collectively, our study shows that knockout of DGK effectively enhances the effector functions of CAR-T cells, suggesting that CRISPR/Cas9-mediated knockout of DGK could be applicable as part of a multifaceted clinical strategy to treat solid cancers. This novel study demonstrates efficient ablation of diacylglycerol kinase in human CAR-T cells that leads to improved antitumor immunity and may have significant impact in human cancer immunotherapy. .

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“…1A ). Recently, increasing attention has been paid to DGKα as a potential target for anti-cancer treatments including cancer immunotherapy ( Dominguez et al, 2013 ; Purow, 2015 ; Sakane, Mizuno & Komenoi, 2016 ; Liu et al, 2016 ; Noessner, 2017 ). Expression of DGKα has been reported to be upregulated in melanoma cells (but not in noncancerous melanocytes) ( Yanagisawa et al, 2007 ), lymphoma ( Bacchiocchi et al, 2005 ), hepatocellular carcinoma ( Takeishi et al, 2012 ), breast cancer cells ( Torres-Ayuso et al, 2014 ), and glioblastoma cells ( Dominguez et al, 2013 ) where DGKα promotes cancer cell survival, proliferation, migration, and invasion ( Merida et al, 2017 ).…”

Section: Introductionmentioning

confidence: 99%

Expression and purification of human diacylglycerol kinase α from baculovirus-infected insect cells for structural studies

Takahashi¹,

Sakane²

2018

PeerJ

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Diacylglycerol kinases (DGKs) are lipid kinases that modulate the levels of lipid second messengers, diacylglycerol and phosphatidic acid. Recently, increasing attention has been paid to its α isozyme (DGKα) as a potential target for cancer immunotherapy. DGKα consists of the N-terminal regulatory domains including EF-hand motifs and C1 domains, and the C-terminal catalytic domain (DGKα-CD). To date, however, no structures of mammalian DGKs including their CDs have yet been reported, impeding our understanding on the catalytic mechanism of DGKs and the rational structure-based drug design. Here we attempted to produce DGKα-CD or a full-length DGKα using bacterial and baculovirus-insect cell expression system for structural studies. While several DGKα-CD constructs produced using both bacterial and insect cells formed insoluble or soluble aggregates, the full-length DGKα expressed in insect cells remained soluble and was purified to near homogeneity as a monomer with yields (1.3 mg/mL per one L cell culture) feasible for protein crystallization. Following enzymatic characterization showed that the purified DGKα is in fully functional state. We further demonstrated that the purified enzyme could be concentrated without any significant aggregation, and characterized its secondary structure by circular dichroism. Taken together, these results suggest that the presence of N-terminal regulatory domains suppress protein aggregation likely via their intramolecular interactions with DGKα-CD, and demonstrate that the baculovirus-insect cell expression of the full-length form of DGKα, not DGKα-CD alone, represents a promising approach to produce protein sample for structural studies of DGKα. Thus, our study will encourage future efforts to determine the crystal structure of DGK, which has not been determined since it was first identified in 1959.

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DGK-α: A Checkpoint in Cancer-Mediated Immuno-Inhibition and Target for Immunotherapy

Cited by 29 publications

References 48 publications

CRISPR-based screens uncover determinants of immunotherapy response in Multiple Myeloma

CRISPR-based screens uncover determinants of immunotherapy response in Multiple Myeloma

CRISPR/Cas9-Mediated Knockout of DGK Improves Antitumor Activities of Human T Cells

Expression and purification of human diacylglycerol kinase α from baculovirus-infected insect cells for structural studies

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