Seth Corey | Cleveland Clinic (original) (raw)

Papers by Seth Corey

Biomolecules

Inherited bone marrow failure syndromes (IBMFSs) include Fanconi anemia, Diamond–Blackfan anemia,... more Inherited bone marrow failure syndromes (IBMFSs) include Fanconi anemia, Diamond–Blackfan anemia, Shwachman–Diamond syndrome, dyskeratosis congenita, severe congenital neutropenia, and other rare entities such as GATA2 deficiency and SAMD9/9L mutations. The IBMFS monogenic disorders were first recognized by their phenotype. Exome sequencing has validated their classification, with clusters of gene mutations affecting DNA damage response (Fanconi anemia), ribosome structure (Diamond–Blackfan anemia), ribosome assembly (Shwachman–Diamond syndrome), or telomere maintenance/stability (dyskeratosis congenita). The pathogenetic mechanisms of IBMFSs remain to be characterized fully, but an overarching hypothesis states that different stresses elicit TP53-dependent growth arrest and apoptosis of hematopoietic stem, progenitor, and precursor cells. Here, we review the IBMFSs and propose a role for pro-inflammatory cytokines, such as TGF-β, IL-1β, and IFN-α, in mediating the cytopenias. We su...

Purpose: Dasatinib is a dual Src/Abl inhibitor recently approved for Bcr-Abl+ leukemias with resi... more Purpose: Dasatinib is a dual Src/Abl inhibitor recently approved for Bcr-Abl+ leukemias with resistance or intolerance to prior therapy. Because Src kinases contribute to multiple blood cell functions by triggering a variety of signaling pathways, we hypothesized that their molecular targeting might lead to growth inhibition in acute myeloid leukemia (AML).Experimental Design: We studied growth factor–dependent and growth factor–independent leukemic cell lines, including three cell lines expressing mutants of receptor tyrosine kinases (Flt3 or c-Kit) as well as primary AML blasts for responsiveness to dasatinib.Results: Dasatinib resulted in the inhibition of Src family kinases in all cell lines and blast cells at ∼1 × 10−9 mol/L. It also inhibited mutant Flt3 or Kit tyrosine phosphorylation at ∼1 × 10−6 mol/L. Mo7e cells expressing the activating mutation (codon 816) of c-Kit were most sensitive to growth inhibition with a GI50 of 5 × 10−9 mol/L. Primary AML blast cells exhibited a...

Supplementary Data from Dasatinib Inhibits the Growth of Molecularly Heterogeneous Myeloid Leukemias

The Journal of Immunology

We recently reported that Fc mu R on NK cells is a signal transducing protein that stimulates a r... more We recently reported that Fc mu R on NK cells is a signal transducing protein that stimulates a rapid increase in the level of cytoplasmic free calcium upon binding of IgM. This study was designed to examine signal transduction via the Fc mu R on NK cells and to characterize intracellular second messengers activated by IgM. Immunoprecipitation of IgM-bound Fc mu R by IgM-specific Ab coimmunoprecipitated the zeta- and Fc epsilon RI gamma-chains. Furthermore, engagement and clustering of Fc mu R by polyclonal IgM induced tyrosine phosphorylation of the zeta- and Fc epsilon RI gamma-chains, indicating their functional association with the Fc mu R-induced signal transduction cascade. Ligand-induced clustering of the Fc mu R also induced activity of src family kinases, Lck, Fyn, Lyn, and Src, as well as their physical interaction with the receptor. Triggering via Fc mu R also induced the activity of Syk and Zap-70, tyrosine kinases demonstrated to associate with zeta and Lck. Phospholipa...

Blood Advances, 2022

Neutrophils migrate into inflamed tissue, engage in phagocytosis, and clear pathogens or apoptoti... more Neutrophils migrate into inflamed tissue, engage in phagocytosis, and clear pathogens or apoptotic cells. These processes require well-coordinated events involving the actin cytoskeleton. We describe a child with severe neutropenia and episodes of soft tissue infections and pneumonia. Bone marrow examination showed granulocytic hypoplasia with dysplasia. Whole-exome sequencing revealed a de novo heterozygous missense mutation in LCP1, which encodes the F-actin–binding protein Lymphocyte Cytosolic Protein 1. To determine its pathophysiological significance, we stably transduced cells with doxycycline-inducible wild-type LCP1 and LCP1 I232F lentiviral constructs. We observed dysplastic granulocytic 32D cells expressing LCP1 I232F cells. These cells showed decreased proliferation without a block in differentiation. In addition, expression of LCP1 I232F resulted in a cell cycle arrest at the G2/M phase, but it did not lead to increased levels of genes involved in apoptosis or the unfold...

Blood, 2020

Background. While childhood leukemia and bone marrow failure disorders always raise suspicions of... more Background. While childhood leukemia and bone marrow failure disorders always raise suspicions of a familial trait, recognition of germ line (GL) contribution to the pathogenesis of adult presentation of these disorders may be difficult because of the absent and unreliable family history, long disease anticipation and incomplete penetrance due to various factors, including competing mortality and interaction with environmental factors. The search for such GL alterations may involve unbiased whole exome sequencing or genome wide analysis studies approaches or, as we conducted here, rely on identification of GL variants in a rationally selected panel of genes previously involved in leukemia and bone marrow failure (BMF). Methods. We hypothesize that, in analogy to pediatric disease, pathogenic GL alterations may exist also in a subset of adult patients with BMF and myeloid neoplasia (MN). Such lesions may contribute to the clinical features and the preferential occurrence of somatic h...

Blood, 2020

Up to 15% of AA patients (pts) treated conservatively with immunosuppression will evolve to myelo... more Up to 15% of AA patients (pts) treated conservatively with immunosuppression will evolve to myeloid neoplasia (MN), either MDS or AML, over a median time of 10 years regardless of response (0-18 years; n=238). The pathogenesis of MN secondary to AA is diverse and will often include antecedent clonal facilitating events that herald progression. Minor clones have been described in AA, some of which are not contributory to later evolution while other may result in subsequent progression. MDS evolution in inherited bone marrow failure (BMF) syndromes suggests that germ line (GL) alterations can be predisposing. In addition, progression to MN may reflect immune escape due to selection pressure e.g., through acquisition of HLA mutations. Here, we studied the molecular landscape of MN arising from AA, to better understand its pathogenesis and ultimately to develop measures of early detection, prevention, and therapeutic strategies. Among 350 pts diagnosed with AA and PNH, 38 (11%) develope...

Frontiers in Physiology, 2020

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, 2018

Blood, 2004

AP23464 is a potent Src and Abl inhibitor (in vitro IC50 < 1 nM), which has been shown to effe... more AP23464 is a potent Src and Abl inhibitor (in vitro IC50 < 1 nM), which has been shown to effectively inhibit growth of imatinib-resistant Bcr-Abl+ cells, and thus, is a promising compound for treating patients with imatinib-resistant leukemia. We conducted a study to evaluate growth inhibition and inhibition of Src versus Abl protein tyrosine kinases in human myeloid cell lines: MV4-11 expressing an internal tandem duplication of Flt3 (Flt3-ITD), the murine pro-B cell line Ba/F3 that expresses the Flt3-ITD, the GM-CSF dependent Mo7e, and the G-CSF-responsive BaF3-GR (Ba/F3 cells expressing the human G-CSF receptor). We compared AP23464 with the PP1, a previously described Src kinase inhibitor (IC50 < 1 uM). We sought to correlate growth inhibition with Src or Abl inhibition. Methods: Growth inhibition was assessed by Trypan blue exclusion and MTT assay using drug concentrations 0.1 uM – 10 uM. Drugs were added daily to the cell suspension during the 3-day experiment. After a ...

Blood, 2008

Recent results have demonstrated that multiple signal transduction pathways are activated in acut... more Recent results have demonstrated that multiple signal transduction pathways are activated in acute myeloid leukemia (AML) cells, however, the tyrosine kinase(s) that phosphorylates these signaling proteins is not identified. We have analyzed AML cells using a phosphoproteomics screen and demonstrate that the Src family kinases, Lyn, Lck and Fgr, are phosphorylated on their activation sites in AML samples. Expression and activation of Lyn has been previously confirmed. Evaluation of Lck demonstrated that Lck is expressed to a variable degree but consistently in AML samples (n=20). Lck kinase assays show activation of Lck in 17/20 samples tested at levels above the level of activation detectable in normal CD34+ progenitor cells. Lyn and Lck both contribute to AML cell growth as siRNA depletion of either kinase leads to decreased leukemia colony forming activity. Interestingly, both Lyn and Lck contribute to phosphorylation of STAT5 as STAT5 phosphorylation is decreased but not abrogat...

Blood, 2011

61 To identify new therapeutic strategies for AML, we compiled and screened an in-house library o... more 61 To identify new therapeutic strategies for AML, we compiled and screened an in-house library of on-patent and off-patent drugs to identify agents cytotoxic to leukemia cells. From this screen, we identified mefloquine, an off-patent drug indicated for the treatment and prophylaxis of malaria. In secondary assays, mefloquine decreased the viability of 9/10 human and murine leukemia cell lines (EC50 3.25–8.0 μM). Moreover, it reduced the viability of 4/5 primary AML samples, but was not cytotoxic to normal hematopoietic cells (EC50>31 μM). Importantly, mefloquine reduced the clonogenic growth of primary AML samples, but not normal hematopoietic cells, and completely inhibited engraftment of primary AML cells into immune deficient mice. Finally, systemic treatment with oral mefloquine (50 mg/kg/day) decreased leukemic burden without evidence of toxicity in 4 mouse models of leukemia, including mice engrafted with primary AML cells. Thus, mefloquine effectively targets leukemic ce...

Blood, 2011

705 Using the Src kinase Lyn as bait in yeast two-hybrid screen, we isolated Cdc42-interacting pr... more 705 Using the Src kinase Lyn as bait in yeast two-hybrid screen, we isolated Cdc42-interacting protein 4 (CIP4). CIP4 is an F-BAR protein implicated in membrane curvature, membrane tubulation, and vesicle formation. CIP4 contains an SH3 domain that interacts with Wiskott-Aldrich Syndrome protein (WASP). We have reported thrombocytopenia in CIP4 knockout (KO) male mice, which was of similar severity as observed in WASP knockout male mice. Because a cardinal feature of Wiskott-Aldrich Syndrome is thrombocytopenia, we hypothesized that the CIP4-dependent thrombocytopenia occurs through the same mechanism as that in WASP-dependent thrombocytopenia. Interestingly, we have just generated a CIP4, WASP double knockout that displays a more severe thrombocytopenia. Neither CFU-MK progenitor studies nor ploidy studies showed a difference between CIP4 KO, WASP KO, double KO, and Wild Type (WT) mice. Thus, we reasoned that the defect lies in the structural production of platelets. We first studi...

Blood, 2015

Shwachman-Diamond syndrome (SDS) is a multisystem disorder characterized by neutropenia, exocrine... more Shwachman-Diamond syndrome (SDS) is a multisystem disorder characterized by neutropenia, exocrine pancreatic insufficiency, and skeletal abnormalities. Individuals with SDS are also at increased risk for developing myelodysplastic syndrome (MDS) and/or acute myeloid leukemia (AML). SDS results from mutations in the SBDS gene, which encodes a protein involved in 80S ribosome maturation. Genetic ablation of Sbds results in early embryonic lethality (ED 7.5) in mice. Zebrafish provide an attractive model organism with demonstrated relevance to mammalian hematopoiesis. Genome analysis of zebrafish (Danio rerio) revealed the presence of a single sbds gene, encoding a protein 90% identical to the human orthologue. qPCR demonstrated presence of sbds transcript in in all stages of the development including unfertilized egg. We used CRISPR/Cas9 genome editing technique to generate indel mutations in sbds. We established multiple zebrafish lines with mutations in the sbds gene. We have now ch...

Blood, 2012

3792 Myelodysplastic Syndromes (MDS) are a group of bone marrow disorders closely related to acut... more 3792 Myelodysplastic Syndromes (MDS) are a group of bone marrow disorders closely related to acute myeloid leukemia (AML). Even though a number of genetic mutations have been recently identified in patients with MDS, their contributions to MDS pathogenesis remains poorly understood. Some of these genetic mutations involve transcription factors, but they are also found in AML: TET2, EZH2, and ASXL1. One group of mutations distinct to MDS are those encoding proteins involved in RNA splicing (e.g. U2AF1/U2AF35, ZRSR2, SRSF2, SF3B1). Based on RNA-Seq of MDS/AML patients, we report exon skipping in the 3′ end of the CSF3R transcript, which encodes the granulocyte colony-stimulating factor receptor (GCSFR), in a patient carrying the S34F mutation in the U2AF1 gene. U2AF1 is one of the more recurrent genes affected by mutation in MDS, and it is associated with progression to secondary AML. The S34F mutation in U2AF1 is a gain of function mutation that promotes excess splicing and exon skip...

Blood, 2010

1555 Hematopoietic cytokine receptors, such as the G-CSFR, use Janus and Src kinases to transduce... more 1555 Hematopoietic cytokine receptors, such as the G-CSFR, use Janus and Src kinases to transduce their signal. Less well known is how the receptors activate these cytosolic protein tyrosine kinases. The phosphorylation/dephosphorylation of inhibitory and stimulatory tyrosine residues of the Src kinases are critical regulatory steps. For Lyn, phosphorylation of Y507 inhibits its activity, whereas phosphorylation of Y396 promotes it. We hypothesized that the tyrosine phosphatase SHP2 is activated by G-CSFR signaling, resulting in dephosphorylation at the negative regulatory site Lyn Y507, and that the adaptor protein Gab2 directs SHP2 effects on phospho-Lyn Y507. To address this hypothesis, we established mouse IL-3-dependent Ba/F3 cells which express the G-CSFR (Ba/F3GR cells). (1) After G-CSF stimulation, phosphorylation status of Lyn (Y507, Y396) was determined by immunoblotting, and protein-protein binding between Gab2-SHP2 and Gab2-Lyn was assessed by the immunoprecipitation and...

Event-free and overall survival remains poor for acute myeloid leukemia (AML). Chemo-resistant cl... more Event-free and overall survival remains poor for acute myeloid leukemia (AML). Chemo-resistant clones contributing to relapse of the disease arise from minimal residual disease (MRD) rather than resulting from newly acquired mutations during or after chemotherapy. MRD is the presence of measurable leukemic cells using non-morphologic assays. It is considered a strong predictor of relapse. The dynamics of clones comprising MRD is poorly understood and is considered influenced by a form of Darwinian selection. We propose a stochastic model based on a multitype (multi-clone) age-dependent Markov branching process to study how random events in MRD contribute to the heterogeneity in response to treatment in a cohort of six patients from The Cancer Genome Atlas database with whole genome sequencing data at two time points. Our model offers a more accurate understanding of how relapse arises and which properties allow a leukemic clone to thrive in the Darwinian competition among leukemic a...

Biomolecules

Inherited bone marrow failure syndromes (IBMFSs) include Fanconi anemia, Diamond–Blackfan anemia,... more Inherited bone marrow failure syndromes (IBMFSs) include Fanconi anemia, Diamond–Blackfan anemia, Shwachman–Diamond syndrome, dyskeratosis congenita, severe congenital neutropenia, and other rare entities such as GATA2 deficiency and SAMD9/9L mutations. The IBMFS monogenic disorders were first recognized by their phenotype. Exome sequencing has validated their classification, with clusters of gene mutations affecting DNA damage response (Fanconi anemia), ribosome structure (Diamond–Blackfan anemia), ribosome assembly (Shwachman–Diamond syndrome), or telomere maintenance/stability (dyskeratosis congenita). The pathogenetic mechanisms of IBMFSs remain to be characterized fully, but an overarching hypothesis states that different stresses elicit TP53-dependent growth arrest and apoptosis of hematopoietic stem, progenitor, and precursor cells. Here, we review the IBMFSs and propose a role for pro-inflammatory cytokines, such as TGF-β, IL-1β, and IFN-α, in mediating the cytopenias. We su...

Purpose: Dasatinib is a dual Src/Abl inhibitor recently approved for Bcr-Abl+ leukemias with resi... more Purpose: Dasatinib is a dual Src/Abl inhibitor recently approved for Bcr-Abl+ leukemias with resistance or intolerance to prior therapy. Because Src kinases contribute to multiple blood cell functions by triggering a variety of signaling pathways, we hypothesized that their molecular targeting might lead to growth inhibition in acute myeloid leukemia (AML).Experimental Design: We studied growth factor–dependent and growth factor–independent leukemic cell lines, including three cell lines expressing mutants of receptor tyrosine kinases (Flt3 or c-Kit) as well as primary AML blasts for responsiveness to dasatinib.Results: Dasatinib resulted in the inhibition of Src family kinases in all cell lines and blast cells at ∼1 × 10−9 mol/L. It also inhibited mutant Flt3 or Kit tyrosine phosphorylation at ∼1 × 10−6 mol/L. Mo7e cells expressing the activating mutation (codon 816) of c-Kit were most sensitive to growth inhibition with a GI50 of 5 × 10−9 mol/L. Primary AML blast cells exhibited a...

Supplementary Data from Dasatinib Inhibits the Growth of Molecularly Heterogeneous Myeloid Leukemias

The Journal of Immunology

We recently reported that Fc mu R on NK cells is a signal transducing protein that stimulates a r... more We recently reported that Fc mu R on NK cells is a signal transducing protein that stimulates a rapid increase in the level of cytoplasmic free calcium upon binding of IgM. This study was designed to examine signal transduction via the Fc mu R on NK cells and to characterize intracellular second messengers activated by IgM. Immunoprecipitation of IgM-bound Fc mu R by IgM-specific Ab coimmunoprecipitated the zeta- and Fc epsilon RI gamma-chains. Furthermore, engagement and clustering of Fc mu R by polyclonal IgM induced tyrosine phosphorylation of the zeta- and Fc epsilon RI gamma-chains, indicating their functional association with the Fc mu R-induced signal transduction cascade. Ligand-induced clustering of the Fc mu R also induced activity of src family kinases, Lck, Fyn, Lyn, and Src, as well as their physical interaction with the receptor. Triggering via Fc mu R also induced the activity of Syk and Zap-70, tyrosine kinases demonstrated to associate with zeta and Lck. Phospholipa...

Blood Advances, 2022

Neutrophils migrate into inflamed tissue, engage in phagocytosis, and clear pathogens or apoptoti... more Neutrophils migrate into inflamed tissue, engage in phagocytosis, and clear pathogens or apoptotic cells. These processes require well-coordinated events involving the actin cytoskeleton. We describe a child with severe neutropenia and episodes of soft tissue infections and pneumonia. Bone marrow examination showed granulocytic hypoplasia with dysplasia. Whole-exome sequencing revealed a de novo heterozygous missense mutation in LCP1, which encodes the F-actin–binding protein Lymphocyte Cytosolic Protein 1. To determine its pathophysiological significance, we stably transduced cells with doxycycline-inducible wild-type LCP1 and LCP1 I232F lentiviral constructs. We observed dysplastic granulocytic 32D cells expressing LCP1 I232F cells. These cells showed decreased proliferation without a block in differentiation. In addition, expression of LCP1 I232F resulted in a cell cycle arrest at the G2/M phase, but it did not lead to increased levels of genes involved in apoptosis or the unfold...

Blood, 2020

Background. While childhood leukemia and bone marrow failure disorders always raise suspicions of... more Background. While childhood leukemia and bone marrow failure disorders always raise suspicions of a familial trait, recognition of germ line (GL) contribution to the pathogenesis of adult presentation of these disorders may be difficult because of the absent and unreliable family history, long disease anticipation and incomplete penetrance due to various factors, including competing mortality and interaction with environmental factors. The search for such GL alterations may involve unbiased whole exome sequencing or genome wide analysis studies approaches or, as we conducted here, rely on identification of GL variants in a rationally selected panel of genes previously involved in leukemia and bone marrow failure (BMF). Methods. We hypothesize that, in analogy to pediatric disease, pathogenic GL alterations may exist also in a subset of adult patients with BMF and myeloid neoplasia (MN). Such lesions may contribute to the clinical features and the preferential occurrence of somatic h...

Blood, 2020

Up to 15% of AA patients (pts) treated conservatively with immunosuppression will evolve to myelo... more Up to 15% of AA patients (pts) treated conservatively with immunosuppression will evolve to myeloid neoplasia (MN), either MDS or AML, over a median time of 10 years regardless of response (0-18 years; n=238). The pathogenesis of MN secondary to AA is diverse and will often include antecedent clonal facilitating events that herald progression. Minor clones have been described in AA, some of which are not contributory to later evolution while other may result in subsequent progression. MDS evolution in inherited bone marrow failure (BMF) syndromes suggests that germ line (GL) alterations can be predisposing. In addition, progression to MN may reflect immune escape due to selection pressure e.g., through acquisition of HLA mutations. Here, we studied the molecular landscape of MN arising from AA, to better understand its pathogenesis and ultimately to develop measures of early detection, prevention, and therapeutic strategies. Among 350 pts diagnosed with AA and PNH, 38 (11%) develope...

Frontiers in Physiology, 2020

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature, 2018

Blood, 2004

AP23464 is a potent Src and Abl inhibitor (in vitro IC50 < 1 nM), which has been shown to effe... more AP23464 is a potent Src and Abl inhibitor (in vitro IC50 < 1 nM), which has been shown to effectively inhibit growth of imatinib-resistant Bcr-Abl+ cells, and thus, is a promising compound for treating patients with imatinib-resistant leukemia. We conducted a study to evaluate growth inhibition and inhibition of Src versus Abl protein tyrosine kinases in human myeloid cell lines: MV4-11 expressing an internal tandem duplication of Flt3 (Flt3-ITD), the murine pro-B cell line Ba/F3 that expresses the Flt3-ITD, the GM-CSF dependent Mo7e, and the G-CSF-responsive BaF3-GR (Ba/F3 cells expressing the human G-CSF receptor). We compared AP23464 with the PP1, a previously described Src kinase inhibitor (IC50 < 1 uM). We sought to correlate growth inhibition with Src or Abl inhibition. Methods: Growth inhibition was assessed by Trypan blue exclusion and MTT assay using drug concentrations 0.1 uM – 10 uM. Drugs were added daily to the cell suspension during the 3-day experiment. After a ...

Blood, 2008

Recent results have demonstrated that multiple signal transduction pathways are activated in acut... more Recent results have demonstrated that multiple signal transduction pathways are activated in acute myeloid leukemia (AML) cells, however, the tyrosine kinase(s) that phosphorylates these signaling proteins is not identified. We have analyzed AML cells using a phosphoproteomics screen and demonstrate that the Src family kinases, Lyn, Lck and Fgr, are phosphorylated on their activation sites in AML samples. Expression and activation of Lyn has been previously confirmed. Evaluation of Lck demonstrated that Lck is expressed to a variable degree but consistently in AML samples (n=20). Lck kinase assays show activation of Lck in 17/20 samples tested at levels above the level of activation detectable in normal CD34+ progenitor cells. Lyn and Lck both contribute to AML cell growth as siRNA depletion of either kinase leads to decreased leukemia colony forming activity. Interestingly, both Lyn and Lck contribute to phosphorylation of STAT5 as STAT5 phosphorylation is decreased but not abrogat...

Blood, 2011

61 To identify new therapeutic strategies for AML, we compiled and screened an in-house library o... more 61 To identify new therapeutic strategies for AML, we compiled and screened an in-house library of on-patent and off-patent drugs to identify agents cytotoxic to leukemia cells. From this screen, we identified mefloquine, an off-patent drug indicated for the treatment and prophylaxis of malaria. In secondary assays, mefloquine decreased the viability of 9/10 human and murine leukemia cell lines (EC50 3.25–8.0 μM). Moreover, it reduced the viability of 4/5 primary AML samples, but was not cytotoxic to normal hematopoietic cells (EC50>31 μM). Importantly, mefloquine reduced the clonogenic growth of primary AML samples, but not normal hematopoietic cells, and completely inhibited engraftment of primary AML cells into immune deficient mice. Finally, systemic treatment with oral mefloquine (50 mg/kg/day) decreased leukemic burden without evidence of toxicity in 4 mouse models of leukemia, including mice engrafted with primary AML cells. Thus, mefloquine effectively targets leukemic ce...

Blood, 2011

705 Using the Src kinase Lyn as bait in yeast two-hybrid screen, we isolated Cdc42-interacting pr... more 705 Using the Src kinase Lyn as bait in yeast two-hybrid screen, we isolated Cdc42-interacting protein 4 (CIP4). CIP4 is an F-BAR protein implicated in membrane curvature, membrane tubulation, and vesicle formation. CIP4 contains an SH3 domain that interacts with Wiskott-Aldrich Syndrome protein (WASP). We have reported thrombocytopenia in CIP4 knockout (KO) male mice, which was of similar severity as observed in WASP knockout male mice. Because a cardinal feature of Wiskott-Aldrich Syndrome is thrombocytopenia, we hypothesized that the CIP4-dependent thrombocytopenia occurs through the same mechanism as that in WASP-dependent thrombocytopenia. Interestingly, we have just generated a CIP4, WASP double knockout that displays a more severe thrombocytopenia. Neither CFU-MK progenitor studies nor ploidy studies showed a difference between CIP4 KO, WASP KO, double KO, and Wild Type (WT) mice. Thus, we reasoned that the defect lies in the structural production of platelets. We first studi...

Blood, 2015

Shwachman-Diamond syndrome (SDS) is a multisystem disorder characterized by neutropenia, exocrine... more Shwachman-Diamond syndrome (SDS) is a multisystem disorder characterized by neutropenia, exocrine pancreatic insufficiency, and skeletal abnormalities. Individuals with SDS are also at increased risk for developing myelodysplastic syndrome (MDS) and/or acute myeloid leukemia (AML). SDS results from mutations in the SBDS gene, which encodes a protein involved in 80S ribosome maturation. Genetic ablation of Sbds results in early embryonic lethality (ED 7.5) in mice. Zebrafish provide an attractive model organism with demonstrated relevance to mammalian hematopoiesis. Genome analysis of zebrafish (Danio rerio) revealed the presence of a single sbds gene, encoding a protein 90% identical to the human orthologue. qPCR demonstrated presence of sbds transcript in in all stages of the development including unfertilized egg. We used CRISPR/Cas9 genome editing technique to generate indel mutations in sbds. We established multiple zebrafish lines with mutations in the sbds gene. We have now ch...

Blood, 2012

3792 Myelodysplastic Syndromes (MDS) are a group of bone marrow disorders closely related to acut... more 3792 Myelodysplastic Syndromes (MDS) are a group of bone marrow disorders closely related to acute myeloid leukemia (AML). Even though a number of genetic mutations have been recently identified in patients with MDS, their contributions to MDS pathogenesis remains poorly understood. Some of these genetic mutations involve transcription factors, but they are also found in AML: TET2, EZH2, and ASXL1. One group of mutations distinct to MDS are those encoding proteins involved in RNA splicing (e.g. U2AF1/U2AF35, ZRSR2, SRSF2, SF3B1). Based on RNA-Seq of MDS/AML patients, we report exon skipping in the 3′ end of the CSF3R transcript, which encodes the granulocyte colony-stimulating factor receptor (GCSFR), in a patient carrying the S34F mutation in the U2AF1 gene. U2AF1 is one of the more recurrent genes affected by mutation in MDS, and it is associated with progression to secondary AML. The S34F mutation in U2AF1 is a gain of function mutation that promotes excess splicing and exon skip...

Blood, 2010

1555 Hematopoietic cytokine receptors, such as the G-CSFR, use Janus and Src kinases to transduce... more 1555 Hematopoietic cytokine receptors, such as the G-CSFR, use Janus and Src kinases to transduce their signal. Less well known is how the receptors activate these cytosolic protein tyrosine kinases. The phosphorylation/dephosphorylation of inhibitory and stimulatory tyrosine residues of the Src kinases are critical regulatory steps. For Lyn, phosphorylation of Y507 inhibits its activity, whereas phosphorylation of Y396 promotes it. We hypothesized that the tyrosine phosphatase SHP2 is activated by G-CSFR signaling, resulting in dephosphorylation at the negative regulatory site Lyn Y507, and that the adaptor protein Gab2 directs SHP2 effects on phospho-Lyn Y507. To address this hypothesis, we established mouse IL-3-dependent Ba/F3 cells which express the G-CSFR (Ba/F3GR cells). (1) After G-CSF stimulation, phosphorylation status of Lyn (Y507, Y396) was determined by immunoblotting, and protein-protein binding between Gab2-SHP2 and Gab2-Lyn was assessed by the immunoprecipitation and...

Event-free and overall survival remains poor for acute myeloid leukemia (AML). Chemo-resistant cl... more Event-free and overall survival remains poor for acute myeloid leukemia (AML). Chemo-resistant clones contributing to relapse of the disease arise from minimal residual disease (MRD) rather than resulting from newly acquired mutations during or after chemotherapy. MRD is the presence of measurable leukemic cells using non-morphologic assays. It is considered a strong predictor of relapse. The dynamics of clones comprising MRD is poorly understood and is considered influenced by a form of Darwinian selection. We propose a stochastic model based on a multitype (multi-clone) age-dependent Markov branching process to study how random events in MRD contribute to the heterogeneity in response to treatment in a cohort of six patients from The Cancer Genome Atlas database with whole genome sequencing data at two time points. Our model offers a more accurate understanding of how relapse arises and which properties allow a leukemic clone to thrive in the Darwinian competition among leukemic a...