Publications by authors named "Svetlana Gaidarova"

13 Publications

  • Page 1 of 1

Harnessing features of adaptive NK cells to generate iPSC-derived NK cells for enhanced immunotherapy.

Cell Stem Cell 2021 Sep 9. Epub 2021 Sep 9.

University of Minnesota, Department of Medicine, Minneapolis, MN 55455, USA. Electronic address:

Select subsets of immune effector cells have the greatest propensity to mediate antitumor responses. However, procuring these subsets is challenging, and cell-based immunotherapy is hampered by limited effector-cell persistence and lack of on-demand availability. To address these limitations, we generated a triple-gene-edited induced pluripotent stem cell (iPSC). The clonal iPSC line was engineered to express a high affinity, non-cleavable version of the Fc receptor CD16a and a membrane-bound interleukin (IL)-15/IL-15R fusion protein. The third edit was a knockout of the ecto-enzyme CD38, which hydrolyzes NAD. Natural killer (NK) cells derived from these uniformly engineered iPSCs, termed iADAPT, displayed metabolic features and gene expression profiles mirroring those of cytomegalovirus-induced adaptive NK cells. iADAPT NK cells persisted in vivo in the absence of exogenous cytokine and elicited superior antitumor activity. Our findings suggest that unique subsets of the immune system can be modeled through iPSC technology for effective treatment of patients with advanced cancer.
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http://dx.doi.org/10.1016/j.stem.2021.08.013DOI Listing
September 2021

iPSC-derived NK cells maintain high cytotoxicity and enhance in vivo tumor control in concert with T cells and anti-PD-1 therapy.

Sci Transl Med 2020 11;12(568)

Department of Medicine, University of Minnesota, Minneapolis, MN 55455, USA.

The development of immunotherapeutic monoclonal antibodies targeting checkpoint inhibitory receptors, such as programmed cell death 1 (PD-1), or their ligands, such as PD-L1, has transformed the oncology landscape. However, durable tumor regression is limited to a minority of patients. Therefore, combining immunotherapies with those targeting checkpoint inhibitory receptors is a promising strategy to bolster antitumor responses and improve response rates. Natural killer (NK) cells have the potential to augment checkpoint inhibition therapies, such as PD-L1/PD-1 blockade, because NK cells mediate both direct tumor lysis and T cell activation and recruitment. However, sourcing donor-derived NK cells for adoptive cell therapy has been limited by both cell number and quality. Thus, we developed a robust and efficient manufacturing system for the differentiation and expansion of high-quality NK cells derived from induced pluripotent stem cells (iPSCs). iPSC-derived NK (iNK) cells produced inflammatory cytokines and exerted strong cytotoxicity against an array of hematologic and solid tumors. Furthermore, we showed that iNK cells recruit T cells and cooperate with T cells and anti-PD-1 antibody, further enhancing inflammatory cytokine production and tumor lysis. Because the iNK cell derivation process uses a renewable starting material and enables the manufacturing of large numbers of doses from a single manufacture, iNK cells represent an "off-the-shelf" source of cells for immunotherapy with the capacity to target tumors and engage the adaptive arm of the immune system to make a "cold" tumor "hot" by promoting the influx of activated T cells to augment checkpoint inhibitor therapies.
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http://dx.doi.org/10.1126/scitranslmed.aaz5618DOI Listing
November 2020

Pluripotent stem cell-derived NK cells with high-affinity noncleavable CD16a mediate improved antitumor activity.

Blood 2020 02;135(6):399-410

Department of Medicine, University of California, San Diego, La Jolla, CA.

Antibody-dependent cellular cytotoxicity (ADCC) is a key effector mechanism of natural killer (NK) cells that is mediated by therapeutic monoclonal antibodies (mAbs). This process is facilitated by the Fc receptor CD16a on human NK cells. CD16a appears to be the only activating receptor on NK cells that is cleaved by the metalloprotease a disintegrin and metalloproteinase-17 upon stimulation. We previously demonstrated that a point mutation of CD16a prevents this activation-induced surface cleavage. This noncleavable CD16a variant is now further modified to include the high-affinity noncleavable variant of CD16a (hnCD16) and was engineered into human induced pluripotent stem cells (iPSCs) to create a renewable source for human induced pluripotent stem cell-derived NK (hnCD16-iNK) cells. Compared with unmodified iNK cells and peripheral blood-derived NK (PB-NK) cells, hnCD16-iNK cells proved to be highly resistant to activation-induced cleavage of CD16a. We found that hnCD16-iNK cells were functionally mature and exhibited enhanced ADCC against multiple tumor targets. In vivo xenograft studies using a human B-cell lymphoma demonstrated that treatment with hnCD16-iNK cells and anti-CD20 mAb led to significantly improved regression of B-cell lymphoma compared with treatment utilizing anti-CD20 mAb with PB-NK cells or unmodified iNK cells. hnCD16-iNK cells, combined with anti-HER2 mAb, also mediated improved survival in an ovarian cancer xenograft model. Together, these findings show that hnCD16-iNK cells combined with mAbs are highly effective against hematologic malignancies and solid tumors that are typically resistant to NK cell-mediated killing, demonstrating the feasibility of producing a standardized off-the-shelf engineered NK cell therapy with improved ADCC properties to treat malignancies that are otherwise refractory.
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http://dx.doi.org/10.1182/blood.2019000621DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7005364PMC
February 2020

GSK3 Inhibition Drives Maturation of NK Cells and Enhances Their Antitumor Activity.

Cancer Res 2017 10 8;77(20):5664-5675. Epub 2017 Aug 8.

Division of Hematology, Oncology and Transplantation, Department of Medicine, University of Minnesota Cancer Center, Minneapolis, Minnesota.

Maturation of human natural killer (NK) cells as defined by accumulation of cell-surface expression of CD57 is associated with increased cytotoxic character and TNF and IFNγ production upon target-cell recognition. Notably, multiple studies point to a unique role for CD57 NK cells in cancer immunosurveillance, yet there is scant information about how they mature. In this study, we show that pharmacologic inhibition of GSK3 kinase in peripheral blood NK cells expanded with IL15 greatly enhances CD57 upregulation and late-stage maturation. GSK3 inhibition elevated the expression of several transcription factors associated with late-stage NK-cell maturation including T-BET, ZEB2, and BLIMP-1 without affecting viability or proliferation. When exposed to human cancer cells, NK cell expanded in the presence of a GSK3 inhibitor exhibited significantly higher production of TNF and IFNγ, elevated natural cytotoxicity, and increased antibody-dependent cellular cytotoxicity. In an established mouse xenograft model of ovarian cancer, adoptive transfer of NK cells conditioned in the same way also displayed more robust and durable tumor control. Our findings show how GSK3 kinase inhibition can greatly enhance the mature character of NK cells most desired for effective cancer immunotherapy. .
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http://dx.doi.org/10.1158/0008-5472.CAN-17-0799DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5645243PMC
October 2017

A novel cereblon modulator recruits GSPT1 to the CRL4(CRBN) ubiquitin ligase.

Nature 2016 07 22;535(7611):252-7. Epub 2016 Jun 22.

Celgene Corporation, 10300 Campus Point Drive, Suite 100, San Diego, California 92121, USA.

Immunomodulatory drugs bind to cereblon (CRBN) to confer differentiated substrate specificity on the CRL4(CRBN) E3 ubiquitin ligase. Here we report the identification of a new cereblon modulator, CC-885, with potent anti-tumour activity. The anti-tumour activity of CC-885 is mediated through the cereblon-dependent ubiquitination and degradation of the translation termination factor GSPT1. Patient-derived acute myeloid leukaemia tumour cells exhibit high sensitivity to CC-885, indicating the clinical potential of this mechanism. Crystallographic studies of the CRBN-DDB1-CC-885-GSPT1 complex reveal that GSPT1 binds to cereblon through a surface turn containing a glycine residue at a key position, interacting with both CC-885 and a 'hotspot' on the cereblon surface. Although GSPT1 possesses no obvious structural, sequence or functional homology to previously known cereblon substrates, mutational analysis and modelling indicate that the cereblon substrate Ikaros uses a similar structural feature to bind cereblon, suggesting a common motif for substrate recruitment. These findings define a structural degron underlying cereblon 'neosubstrate' selectivity, and identify an anti-tumour target rendered druggable by cereblon modulation.
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http://dx.doi.org/10.1038/nature18611DOI Listing
July 2016

Lenalidomide inhibits the proliferation of CLL cells via a cereblon/p21(WAF1/Cip1)-dependent mechanism independent of functional p53.

Blood 2014 Sep 2;124(10):1637-44. Epub 2014 Jul 2.

Moores Cancer Center, University of California, San Diego, La Jolla, CA; and.

Lenalidomide has demonstrated clinical activity in patients with chronic lymphocytic leukemia (CLL), even though it is not cytotoxic for primary CLL cells in vitro. We examined the direct effect of lenalidomide on CLL-cell proliferation induced by CD154-expressing accessory cells in media containing interleukin-4 and -10. Treatment with lenalidomide significantly inhibited CLL-cell proliferation, an effect that was associated with the p53-independent upregulation of the cyclin-dependent kinase inhibitor, p21(WAF1/Cip1) (p21). Silencing p21 with small interfering RNA impaired the capacity of lenalidomide to inhibit CLL-cell proliferation. Silencing cereblon, a known molecular target of lenalidomide, impaired the capacity of lenalidomide to induce expression of p21, inhibit CD154-induced CLL-cell proliferation, or enhance the degradation of Ikaros family zinc finger proteins 1 and 3. We isolated CLL cells from the blood of patients before and after short-term treatment with low-dose lenalidomide (5 mg per day) and found the leukemia cells were also induced to express p21 in vivo. These results indicate that lenalidomide can directly inhibit proliferation of CLL cells in a cereblon/p21-dependent but p53-independent manner, at concentrations achievable in vivo, potentially contributing to the capacity of this drug to inhibit disease-progression in patients with CLL.
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http://dx.doi.org/10.1182/blood-2014-03-559591DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4155272PMC
September 2014

Measuring cereblon as a biomarker of response or resistance to lenalidomide and pomalidomide requires use of standardized reagents and understanding of gene complexity.

Br J Haematol 2014 Jan 28;164(2):233-44. Epub 2013 Oct 28.

Celgene Corporation, Summit, NJ, USA.

Cereblon, a member of the cullin 4 ring ligase complex (CRL4), is the molecular target of the immunomodulatory drugs (IMiDs) lenalidomide and pomalidomide and is required for the antiproliferative activity of these agents in multiple myeloma (MM) and immunomodulatory activity in T cells. Cereblon's central role as a target of lenalidomide and pomalidomide suggests potential utility as a predictive biomarker of response or resistance to IMiD therapy. Our studies characterized a cereblon monoclonal antibody CRBN65, with high sensitivity and specificity in Western analysis and immunohistochemistry that is superior to commercially available antibodies. We identified multiple cereblon splice variants in both MM cell lines and primary cells, highlighting challenges with conventional gene expression assays given this gene complexity. Using CRBN65 antibody and TaqMan quantitative reverse transcription polymerase chain reaction assays, we showed lack of correlation between cereblon protein and mRNA levels. Furthermore, lack of correlation between cereblon expression in MM cell lines and sensitivity to lenalidomide was shown. In cell lines made resistant to lenalidomide and pomalidomide, cereblon protein is greatly reduced. These studies show limitations to the current approaches of cereblon measurement that rely on commercial reagents and assays. Standardized reagents and validated assays are needed to accurately assess the role of cereblon as a predictive biomarker.
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http://dx.doi.org/10.1111/bjh.12622DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4253085PMC
January 2014

Lenalidomide downregulates the cell survival factor, interferon regulatory factor-4, providing a potential mechanistic link for predicting response.

Br J Haematol 2011 Aug 24;154(3):325-36. Epub 2011 Jun 24.

Translational Development Department, Celgene, San Diego, CA 92122, USA.

Overexpression of the transcription factor interferon regulatory factor-4 (IRF4), which is common in multiple myeloma (MM), is associated with poor prognosis. Patients with higher IRF4 expression have significantly poorer overall survival than those with low IRF4 expression. Lenalidomide is an IMiD immunomodulatory compound that has both tumouricidal and immunomodulatory activity in MM. This study showed that lenalidomide downregulated IRF4 levels in MM cell lines and bone marrow samples within 8 h of drug exposure. This was associated with a decrease in MYC levels, as well as an initial G1 cell cycle arrest, decreased cell proliferation, and cell death by day 5 of treatment. In eight MM cell lines, high IRF4 levels correlated with increased lenalidomide sensitivity. The clinical significance of this observation was investigated in 154 patients with MM. Among MM patients with high levels of IRF4 expression, treatment with lenalidomide led to a significantly longer overall survival than other therapies in a retrospective analysis. These data confirm the central role of IRF4 in MM pathogenesis; indicate that this is an important mechanism by which lenalidomide exerts its antitumour effects; and may provide a mechanistic biomarker to predict response to lenalidomide.
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http://dx.doi.org/10.1111/j.1365-2141.2011.08689.xDOI Listing
August 2011

IL-1beta induces a MyD88-dependent and ceramide-mediated activation of Src in anterior hypothalamic neurons.

J Neurochem 2006 Sep 12;98(5):1379-89. Epub 2006 Jun 12.

The Harold L. Dorris Neurological Research Institute and Molecular and Integrative Neurosciences Department, The Scripps Research Institute, La Jolla, California 92037, USA.

The proinflammatory cytokine interleukin 1beta (IL-1beta), acting at IL-1R1 receptors, affects neuronal signaling under both physiological and pathophysiological conditions. The molecular mechanism of the rapid synaptic actions of IL-1beta in neurons is not known. We show here that within minutes of IL-1beta exposure, the firing rate of anterior hypothalamic (AH) neurons in culture was inhibited. This effect was prevented by pre-exposure of the cells to the Src family inhibitor, PP2, suggesting the involvement of Src in the hyperpolarizing effects of IL-1beta. The IL-1beta stimulation of neurons induced a rapid increase in the phosphorylation of the tyrosine kinase Src and kinase suppressor of Ras (ceramide activated protein kinase (CAPK)/KSR) in neurons grown on glia from IL-1RI(-/-) mice. These effects of IL-1beta were dependent on the association of the cytosolic adaptor protein, MyD88, to the IL-1 receptor, and on the activation of the neutral sphingomyelinase, leading to production of ceramide. A cell-permeable analog of ceramide mimicked the effects of IL-1beta on the cultured AH neurons. These results suggest that ceramide may be the second messenger of the fast IL-1beta actions in AH neurons, and that this IL-1beta/ceramide pathway may underlie the fast non-transcription-dependent, electrophysiological effects of IL-1beta observed in AH neurons in vivo.
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http://dx.doi.org/10.1111/j.1471-4159.2006.03951.xDOI Listing
September 2006

MyD88-dependent and -independent signaling by IL-1 in neurons probed by bifunctional Toll/IL-1 receptor domain/BB-loop mimetics.

Proc Natl Acad Sci U S A 2006 Feb 13;103(8):2953-8. Epub 2006 Feb 13.

The Harold L. Dorris Neurological Institute, La Jolla, CA 92037, USA.

Interleukin (IL)-1beta is a pluripotent proinflammatory cytokine that signals through the type-I IL-1 receptor (IL-1RI), a member of the Toll-like receptor family. In hypothalamic neurons, binding of IL-1beta to IL-1RI mediates transcription-dependent changes that depend on the recruitment of the cytosolic adaptor protein myeloid differentiation primary-response protein 88 (MyD88) to the IL-1RI/IL-1 receptor accessory protein (IL-1RAcP) complex through homomeric Toll/IL-1 receptor (TIR)-TIR interactions. Through design and synthesis of bifunctional TIR mimetics that disrupt the interaction of MyD88 with the IL-1RI/IL-1RAcP complex, we analyzed the involvement of MyD88 in the signaling of IL-1beta in anterior hypothalamic neurons. We show here that IL-1beta-mediated activation of the protein tyrosine kinase Src depended on a MyD88 interaction with the IL-1RI/IL-1RAcP complex. The activation of the protein kinase Akt/PKB depended on the recruitment of the p85 subunit of PI3K to IL-1RI and independent of MyD88 association with the IL-1RI/IL-1RAcP complex. These bifunctional TIR-TIR mimetics represent a class of low-molecular-weight compounds with both an antiinflammatory and neuroprotective potential. These compounds have the potential to inhibit the MyD88-dependent proinflammatory actions of IL-1beta, while permitting the potential neuronal survival supporting actions mediated by the MyD88-independent activation of the protein kinase Akt.
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http://dx.doi.org/10.1073/pnas.0510802103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1413805PMC
February 2006

Uncoupling protein 2 protects dopaminergic neurons from acute 1,2,3,6-methyl-phenyl-tetrahydropyridine toxicity.

J Neurochem 2005 Apr;93(2):493-501

Harold L. Dorris Neurological Research Center, Scripps Research Institute, La Jolla, California 92037, USA.

Oxidative stress is implicated in the death of dopaminergic neurons in sporadic forms of Parkinson's disease. Because oxidative stress can be modulated endogenously by uncoupling proteins (UCPs), we hypothesized that specific neuronal expression of UCP2, one member of the UCP family that is rapidly induced in the CNS following insults, could confer neuroprotection in a mouse model of Parkinson's disease. We generated transgenic mice overexpressing UCP2 in catecholaminergic neurons under the control of the tyrosine hydroxylase promoter (TH-UCP2). In these mice, dopaminergic neurons of the substantia nigra showed a twofold elevation in UCP2 expression, elevated uncoupling of their mitochondria, and a marked reduction in indicators of oxidative stress, an effect also observed in the striatum. Upon acute exposure to 1,2,3,6-methyl-phenyl-tetrahydropyridine, TH-UCP2 mice showed neuroprotection and retention of locomotor functions. Our data suggest that UCP2 may represent a drug target for slowing the progression of Parkinson's disease.
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http://dx.doi.org/10.1111/j.1471-4159.2005.03052.xDOI Listing
April 2005

A low molecular weight mimic of the Toll/IL-1 receptor/resistance domain inhibits IL-1 receptor-mediated responses.

Proc Natl Acad Sci U S A 2003 Jun 10;100(13):7971-6. Epub 2003 Jun 10.

Department of Neuropharmacology, The Harold L. Dorris Neurological Research Center, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.

Toll-like receptors (TLRs) and the type I IL-1 receptor (IL-1RI) are key components of the innate immune system activated by microbial infections and inflammation. The signaling cascade from agonist-occupied TLRs and IL-1Rs involves recruitment of the small cytosolic adapter protein MyD88 that binds to IL-1RI via homotypic interactions mediated by Toll/IL-1R/resistance (TIR) domains. Dominant negative forms and null mutations of MyD88 have recently been shown to preclude bacterial product or IL-1-mediated activation of NF-kappaB pathways, demonstrating that MyD88 is an essential component of the Toll receptor signaling. Here, we report the synthesis and pharmacological effects of a low molecular weight MyD88 mimic, hydrocinnamoyl-l-valyl pyrrolidine (compound 4a), modeled on a tripeptide sequence of the BB-loop [(F/Y)-(V/L/I)-(P/G)] of the TIR domain. Results are presented showing that compound 4a interferes with the interactions between mouse MyD88 and IL-1RI at the TIR domains. Compound 4a inhibited IL-1beta-induced phosphorylation of the mitogen-activated protein kinase p38 in EL4 thymoma cells and in freshly isolated murine lymphocytes in a concentration-dependent manner. In vivo, compound 4a produced a significant attenuation of the IL-1beta-induced fever response (200 mg/kg, i.p.). Inhibition of the TIR domain-mediated MyD88/IL1-RI interaction by a low molecular weight, cell-penetrating TIR domain mimic suggests an intracellular site for antiinflammatory drug action.
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http://dx.doi.org/10.1073/pnas.0932746100DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC164697PMC
June 2003

Inhibition of basal and transforming growth factor-beta-induced stimulation of COL1A1 transcription by the DNA intercalators, mitoxantrone and WP631, in cultured human dermal fibroblasts.

J Biol Chem 2002 Oct 23;277(41):38737-45. Epub 2002 Jul 23.

Division of Rheumatology, Department of Medicine, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.

The Sp1 transcription factor plays a crucial role in COL1A1 transcriptional regulation under normal and pathologic conditions and under the effects of transforming growth factor-beta (TGF-beta). Sp1 activity is elevated in numerous diseases characterized by tissue fibrosis. Therefore, inhibition of Sp1 binding to COL1A1 regulatory elements may represent an effective treatment for these diseases. Here we examined the effect of two DNA intercalators that prevent Sp1 binding on the expression of COL1A1 in human dermal fibroblasts. Cultured human adult dermal fibroblasts were treated with WP631 (50 pm/ml to 500 nm/ml) or mitoxantrone (5-500 nm/ml). Cytotoxicity, cellular apoptosis, and collagen deposition were examined by fluorescence microscopy. Collagen production was examined by enzyme-linked immunosorbent assay and metabolic labeling, COL1A1 steady-state mRNA levels, and stability were assessed by Northern hybridizations, and COL1A1 transcription by in vitro nuclear transcription assays and transient transfections. Competition of the drugs for Sp1 binding and their effect on TGF-beta-induced stimulation of COL1A1 transcription was also examined. Both drugs caused a dose-related inhibition of COL1A1 production and mRNA levels without cytotoxicity or apoptosis. COL1A1 transcriptional activity showed a profound reduction mediated by a short proximal promoter region containing an Sp1-binding element at -87 to -82 bp. Furthermore, both drugs inhibited Sp1 DNA complex formation and abrogated the stimulation of COL1A1 transcription induced by TGF-beta. WP631 showed 10-fold higher potency than mitoxantrone. These data indicate that mitoxantrone and WP631 are very potent inhibitors of basal and TGF-beta-stimulated COL1A1 expression and suggest that Sp1-DNA intercalators may be an effective and novel approach for the treatment of fibrotic diseases and modulation of profibrogenic effects of TGF-beta.
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http://dx.doi.org/10.1074/jbc.M201742200DOI Listing
October 2002
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