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Lars Rönnblom - One of the best experts on this subject based on the ideXlab platform.

  • Interferon-α enhances the IL-12-induced STAT4 activation selectively in carriers of the STAT4 SLE risk allele rs7574865[T].
    Annals of the Rheumatic Diseases, 2018
    Co-Authors: Niklas Hagberg, Lars Rönnblom
    Abstract:

    Intronic single nucleotide polymorphisms (SNP) in STAT4 , tagged by rs7574865[T], are associated with increased susceptibility to develop systemic lupus erythematosus (SLE) and a more severe disease phenotype with nephritis and stroke.1 2 Recently, we demonstrated that T cells from patients with SLE carrying the STAT4 risk allele rs7574865[T] have an enhanced induction of STAT4 Protein following PHA/interleukin (IL)-2 activation, which results in increased IL-12-induced phosphorylation of STAT4 (pSTAT4) and interferon (IFN)-γ production.3 As the majority of STAT4 risk allele carriers do not develop disease, we asked whether the STAT4 risk allele exerts the same effect in healthy individuals. In studies of IL-12-stimulated cells from 72 healthy women (online supplementary materials and methods) we unexpectedly found that STAT4 risk allele carriership was associated with a decreased pSTAT4 in CD8+ and CD4+ T cells from healthy individuals (figure 1A). Notably, in the absence of STAT4 risk alleles healthy individuals and patients with SLE phosphorylated STAT4 to the same extent (figure 1B). However, with increasing numbers of STAT4 risk alleles, patients with SLE responded stronger to IL-12 stimulation. The decreased IL-12 response in healthy STAT4 risk allele carriers resulted in a decreased production of IFN-γ in CD8+ T cells and a slight, but non-significant, decrease in CD4+ T cells (figure 1C). Demonstrating the specificity for IL-12, phorbol 12-myristate 13-acetate-induced IFN-γ …

  • FRI0257 The STAT4 sle risk allele rs7574865[t] is associated with increased il-12-induced ifn-gamma production in t cells from sle patients
    FRIDAY 15 JUNE 2018, 2018
    Co-Authors: Niklas Hagberg, M. Joelsson, Dag Leonard, Sarah Reid, Maija-leena Eloranta, Magnus K Nilsson, A.-c. Syvänen, Yenan T. Bryceson, Lars Rönnblom
    Abstract:

    Background Genetic variants in the transcription factor STAT4 are associated with increased susceptibility to systemic lupus erythematosus (SLE) and a more severe disease phenotype. Objectives This study aimed to clarify how the SLE-associated intronic STAT4 risk variant rs7574865[T] affects the function of immune cells in SLE. Methods Peripheral blood mononuclear cells (PBMCs) were isolated from 52 SLE patients in remission (SLEDAI-2K≤4). STAT4 and STAT1 Protein levels and phosphorylation status in response to interferon (IFN)-α, IFN-γ, or interleukin (IL)−12 were determined, before and after pre-activation of cells with phytohaemagglutinin (PHA) and IL-2, in CD56 dim NK cells, CD56 bright NK cells, B cells, CD4 + T cells, CD8 + T cells and monocytes by flow cytometry. The frequency of IFN-γ + cells upon IL-12 or PMA (phorbol 12-myristate-13-acetate) stimulation and the frequency of T-bet + cells was determined in PHA/IL-2 pre-activated cells. Cellular responses and phenotypes were correlated with STAT4 risk allele carriership (rs7574865[T], 21 homozygous protective, 22 heterozygous and 9 homozygous risk) using an additive linear regression model. Janus kinase inhibitors (JAKi) selective for TYK2 (TYK2i, Compound 35) or JAK2 (JAK2i, BMS-911543) were evaluated for inhibition of IL-12 or IFN-γ-induced activation of SLE PBMCs. Results In resting PBMCs, the STAT4 risk allele was neither associated with Protein levels of STAT4 or STAT1, nor cytokine-induced phosphorylation of STAT4 (pSTAT4) or STAT1 (pSTAT1). However, following PHA/IL-2-activation, CD8 + T cells from STAT4 risk allele carriers displayed increased levels of STAT4 (p=0.04), resulting in increased pSTAT4 in response to IL-12 (p=0.003) and IFN-α (p=0.04). Analysis of T cell subsets revealed that the effect was seen in CD45RA + CD57 – naive and CD45RA – CD57 – memory CD8 + T cells, but not in CD45RA – CD57 + memory or CD45RA + CD57 + effector CD8 + T cells. A slight increase in STAT4 Protein levels and IL-12-induced pSTAT4 was also observed in CD4 + T cells from STAT4 risk allele carriers (p=0.08 and p=0.09, respectively). STAT4 risk allele carriers displayed an augmented IL-12-induced IFN-γ production in CD8 + and CD4 + T cells (p=0.03 for both), whereas PMA-induced IL-12 production was normal (p=0.31 and p=0.10, respectively). T-bet expression was not correlated to the STAT4 genotype. The TYK2i and the JAK2i efficiently blocked IL-12 and IFN-γ-induced activation of PBMCs from STAT4 risk patients, respectively. Conclusions T cells from SLE patients carrying the STAT4 risk allele rs7574865[T] display an augmented response to IL-12 and IFN-α. This subset of patients may benefit from JAKi treatment. Disclosure of Interest N. Hagberg: None declared, M. Joelsson: None declared, D. Leonard: None declared, S. Reid: None declared, M.-L. Eloranta: None declared, J. Mo Employee of: AstraZeneca, M. Nilsson Employee of: AstraZeneca, A.-C. Syvanen: None declared, Y. Bryceson: None declared, L. Ronnblom: None declared

Kazuo Kobayashi - One of the best experts on this subject based on the ideXlab platform.

  • Mycobacterial trehalose 6,6′-dimycolate preferentially induces type 1 helper T cell responses through signal transducer and activator of transcription 4 Protein
    Microbial Pathogenesis, 2005
    Co-Authors: Ryuta Oiso, Nagatoshi Fujiwara, Hirokazu Yamagami, Shinji Maeda, Sohkichi Matsumoto, Shiro Nakamura, Nobuhide Oshitani, Takayuki Matsumoto, Tetsuo Arakawa, Kazuo Kobayashi
    Abstract:

    Abstract Mycobacterium tuberculosis is an intracellular pathogen of tuberculosis and its pathogenicity is related to the ability to escape killing by ingested macrophages and induce delayed-type hypersensitivity (DTH). A major component of the cell wall of M. tuberculosis is trehalose 6,6′-dimycolate (TDM), which has been implicated as a pathogenetic factor. The expression of DTH and cell-mediated immunity is dependent on the macrophage-cytokine-type 1 helper T (Th1) lymphocyte axis. Cytokines, interleukin-12 (IL-12) and interferon-γ (IFN-γ), play a critical role in the process and IL-12-activated signal transducer and activator of transcription (STAT) 4 is required for the development of fully functional Th1 cells. To clarify host responses to mycobacterial TDM, we have analyzed footpad reaction, histopathology and cytokine profile of experimental granulomatous lesions using STAT4-deficient mice. In the present study, we have demonstrated that mycobacterial TDM selectively induces the Th1 response through the STAT4 signaling pathway, because mice lacking STAT4 Protein significantly reduced to develop DTH, hypersensitivity granulomas, and Th1 cytokine responses, when compared to BALB/c mice. These results shed light on the molecular pathogenesis of mycobacterial disease. Taken together with previous studies, TDM is a pleiotropic molecule against the host and participates in the pathogenesis.

  • Mycobacterial trehalose 6,6'-dimycolate preferentially induces type 1 helper T cell responses through signal transducer and activator of transcription 4 Protein.
    Microbial Pathogenesis, 2005
    Co-Authors: Ryuta Oiso, Nagatoshi Fujiwara, Hirokazu Yamagami, Shinji Maeda, Sohkichi Matsumoto, Shiro Nakamura, Nobuhide Oshitani, Takayuki Matsumoto, Tetsuo Arakawa, Kazuo Kobayashi
    Abstract:

    Mycobacterium tuberculosis is an intracellular pathogen of tuberculosis and its pathogenicity is related to the ability to escape killing by ingested macrophages and induce delayed-type hypersensitivity (DTH). A major component of the cell wall of M. tuberculosis is trehalose 6,6'-dimycolate (TDM), which has been implicated as a pathogenetic factor. The expression of DTH and cell-mediated immunity is dependent on the macrophage-cytokine-type 1 helper T (Th1) lymphocyte axis. Cytokines, interleukin-12 (IL-12) and interferon-gamma (IFN-gamma), play a critical role in the process and IL-12-activated signal transducer and activator of transcription (STAT) 4 is required for the development of fully functional Th1 cells. To clarify host responses to mycobacterial TDM, we have analyzed footpad reaction, histopathology and cytokine profile of experimental granulomatous lesions using STAT4-deficient mice. In the present study, we have demonstrated that mycobacterial TDM selectively induces the Th1 response through the STAT4 signaling pathway, because mice lacking STAT4 Protein significantly reduced to develop DTH, hypersensitivity granulomas, and Th1 cytokine responses, when compared to BALB/c mice. These results shed light on the molecular pathogenesis of mycobacterial disease. Taken together with previous studies, TDM is a pleiotropic molecule against the host and participates in the pathogenesis.

Niklas Hagberg - One of the best experts on this subject based on the ideXlab platform.

  • Interferon-α enhances the IL-12-induced STAT4 activation selectively in carriers of the STAT4 SLE risk allele rs7574865[T].
    Annals of the Rheumatic Diseases, 2018
    Co-Authors: Niklas Hagberg, Lars Rönnblom
    Abstract:

    Intronic single nucleotide polymorphisms (SNP) in STAT4 , tagged by rs7574865[T], are associated with increased susceptibility to develop systemic lupus erythematosus (SLE) and a more severe disease phenotype with nephritis and stroke.1 2 Recently, we demonstrated that T cells from patients with SLE carrying the STAT4 risk allele rs7574865[T] have an enhanced induction of STAT4 Protein following PHA/interleukin (IL)-2 activation, which results in increased IL-12-induced phosphorylation of STAT4 (pSTAT4) and interferon (IFN)-γ production.3 As the majority of STAT4 risk allele carriers do not develop disease, we asked whether the STAT4 risk allele exerts the same effect in healthy individuals. In studies of IL-12-stimulated cells from 72 healthy women (online supplementary materials and methods) we unexpectedly found that STAT4 risk allele carriership was associated with a decreased pSTAT4 in CD8+ and CD4+ T cells from healthy individuals (figure 1A). Notably, in the absence of STAT4 risk alleles healthy individuals and patients with SLE phosphorylated STAT4 to the same extent (figure 1B). However, with increasing numbers of STAT4 risk alleles, patients with SLE responded stronger to IL-12 stimulation. The decreased IL-12 response in healthy STAT4 risk allele carriers resulted in a decreased production of IFN-γ in CD8+ T cells and a slight, but non-significant, decrease in CD4+ T cells (figure 1C). Demonstrating the specificity for IL-12, phorbol 12-myristate 13-acetate-induced IFN-γ …

  • FRI0257 The STAT4 sle risk allele rs7574865[t] is associated with increased il-12-induced ifn-gamma production in t cells from sle patients
    FRIDAY 15 JUNE 2018, 2018
    Co-Authors: Niklas Hagberg, M. Joelsson, Dag Leonard, Sarah Reid, Maija-leena Eloranta, Magnus K Nilsson, A.-c. Syvänen, Yenan T. Bryceson, Lars Rönnblom
    Abstract:

    Background Genetic variants in the transcription factor STAT4 are associated with increased susceptibility to systemic lupus erythematosus (SLE) and a more severe disease phenotype. Objectives This study aimed to clarify how the SLE-associated intronic STAT4 risk variant rs7574865[T] affects the function of immune cells in SLE. Methods Peripheral blood mononuclear cells (PBMCs) were isolated from 52 SLE patients in remission (SLEDAI-2K≤4). STAT4 and STAT1 Protein levels and phosphorylation status in response to interferon (IFN)-α, IFN-γ, or interleukin (IL)−12 were determined, before and after pre-activation of cells with phytohaemagglutinin (PHA) and IL-2, in CD56 dim NK cells, CD56 bright NK cells, B cells, CD4 + T cells, CD8 + T cells and monocytes by flow cytometry. The frequency of IFN-γ + cells upon IL-12 or PMA (phorbol 12-myristate-13-acetate) stimulation and the frequency of T-bet + cells was determined in PHA/IL-2 pre-activated cells. Cellular responses and phenotypes were correlated with STAT4 risk allele carriership (rs7574865[T], 21 homozygous protective, 22 heterozygous and 9 homozygous risk) using an additive linear regression model. Janus kinase inhibitors (JAKi) selective for TYK2 (TYK2i, Compound 35) or JAK2 (JAK2i, BMS-911543) were evaluated for inhibition of IL-12 or IFN-γ-induced activation of SLE PBMCs. Results In resting PBMCs, the STAT4 risk allele was neither associated with Protein levels of STAT4 or STAT1, nor cytokine-induced phosphorylation of STAT4 (pSTAT4) or STAT1 (pSTAT1). However, following PHA/IL-2-activation, CD8 + T cells from STAT4 risk allele carriers displayed increased levels of STAT4 (p=0.04), resulting in increased pSTAT4 in response to IL-12 (p=0.003) and IFN-α (p=0.04). Analysis of T cell subsets revealed that the effect was seen in CD45RA + CD57 – naive and CD45RA – CD57 – memory CD8 + T cells, but not in CD45RA – CD57 + memory or CD45RA + CD57 + effector CD8 + T cells. A slight increase in STAT4 Protein levels and IL-12-induced pSTAT4 was also observed in CD4 + T cells from STAT4 risk allele carriers (p=0.08 and p=0.09, respectively). STAT4 risk allele carriers displayed an augmented IL-12-induced IFN-γ production in CD8 + and CD4 + T cells (p=0.03 for both), whereas PMA-induced IL-12 production was normal (p=0.31 and p=0.10, respectively). T-bet expression was not correlated to the STAT4 genotype. The TYK2i and the JAK2i efficiently blocked IL-12 and IFN-γ-induced activation of PBMCs from STAT4 risk patients, respectively. Conclusions T cells from SLE patients carrying the STAT4 risk allele rs7574865[T] display an augmented response to IL-12 and IFN-α. This subset of patients may benefit from JAKi treatment. Disclosure of Interest N. Hagberg: None declared, M. Joelsson: None declared, D. Leonard: None declared, S. Reid: None declared, M.-L. Eloranta: None declared, J. Mo Employee of: AstraZeneca, M. Nilsson Employee of: AstraZeneca, A.-C. Syvanen: None declared, Y. Bryceson: None declared, L. Ronnblom: None declared

Michael J. Robertson - One of the best experts on this subject based on the ideXlab platform.

  • ACQUIRED STAT4 DEFICIENCY AS A CONSEQUENCE OF CANCER CHEMOTHERAPY
    Blood, 2011
    Co-Authors: Ivan P. Lupov, Larry Voiles, Ling Han, Allysia Schwartz, Manuel De La Rosa, Kinnari Oza, David Pelloso, Ravi P. Sahu, Jeffrey B. Travers, Michael J. Robertson
    Abstract:

    Signal Transducer and Activator of Transcription 4 (STAT4) is a transcription factor that is activated by IL-12 signaling and promotes Th1-cell differentiation and IFN-γ production. Defective IFN-γ production because of STAT4 mRNA and Protein deficiency occurs after autologous stem cell transplantation for lymphoma. In the present study, we investigated the mechanisms of STAT4 deficiency in lymphoma patients. The tumor-bearing state is not responsible, because STAT4 levels were not significantly different in PBMCs obtained from healthy control subjects compared with those from lymphoma patients before treatment. STAT4 Protein levels were significantly decreased in PBMCs and T cells obtained from lymphoma patients after standard-dose chemotherapy. Furthermore, treatment of control PBMC cultures or a natural killer cell line with chemotherapy drugs in vitro also resulted in reduced STAT4 Protein and diminished, IL-12–induced IFN-γ production. Translation of STAT4 Protein was not impaired in chemotherapy-treated cells, whereas the STAT4 Protein half-life was significantly reduced. Chemotherapy drugs promoted the ubiquitination and proteasomal degradation of STAT4. Treatment with the proteasome inhibitor bortezomib reversed chemotherapy-induced STAT4 deficiency and defective IFN-γ production. We conclude that acquired STAT4 deficiency in lymphoma patients is a consequence of treatment with chemotherapy, results that have important implications for the design of optimal immunotherapy for lymphoma.

  • Reconstitution of STAT4 Restores Defective Interferon-gamma Production and Allows Normal IFN-gamma-Dependent Responses after Autologous Stem Cell Transplantation.
    Blood, 2006
    Co-Authors: Michael J. Robertson, David Pelloso, Hua-chen Chang, Mark H. Kaplan
    Abstract:

    Production of IFN-gamma is critical for optimal antitumor immune responses in several preclinical animal models. Patients with lymphoma who have undergone autologous hematopoietic stem cell transplantation exhibit profoundly diminished IFN-gamma production in vivo during IL-12 therapy (Clin Cancer Res.2002; 8: 3383). Furthermore, post-transplant patient peripheral blood mononuclear cells (PBMCs) stimulated directly in vitro with IL-12 secrete little if any IFN-gamma. To determine if restoration of IFN-gamma production would be sufficient to reconstitute IFN-gamma-dependent responses after transplantation, the integrity of IFN-gamma signaling was assessed in post-transplant patient PBMCs. Compared to control subject PBMCs, post-transplant patient PBMCs expressed equivalent levels of CD119 (IFN-gamma receptor alpha) and STAT1. Moreover, tyrosine phosphorylation of STAT1 in response to IFN-gamma did not differ in post-transplant patient as compared to control PBMCs. Thus, IFN-gamma signaling is intact after transplantation. Our prior studies have shown that STAT4 Protein levels are decreased by ~97% in post-transplant patient PBMCs, whereas levels of Jak2, Tyk2, and STAT3 are similar to control PBMCs (Blood2005; 106: 963). We wished to demonstrate directly that STAT4 deficiency is the mechanism of defective IFN-gamma production after autologous transplantation. Enriched CD4+ T cells were isolated from post-transplant patient and control PBMCs and cultured under conditions that promote Th1 differentiation. STAT4 deficiency persisted in post-transplant patient CD4+ T cells cultured under Th1 conditions, whereas these cells expressed normal levels of T-bet. Compared to control Th1 cells, post-transplant patient CD4+ T cells cultured under Th1 conditions exhibited markedly reduced IFN-gamma production after restimulation with CD3 mAb. These results are consistent with expected impairment of Th1 differentiation due to profound STAT4 deficiency. STAT4 cDNA was transiently transfected into Th1-cultured CD4+ T cells from post-transplant patients. After stimulation with CD3 mAb plus IL-12, the amounts of IFN-gamma secreted by STAT4-transfected patient cells were equivalent to or higher than those secreted by control subject cells. These data indicate that reconstitution of STAT4 expression is sufficient to restore IFN-gamma production by post-transplant patient PBMCs. As IFN-gamma signaling is normal post-transplant, circumventing STAT4 deficiency should therefore restore IFN-gamma-dependent antitumor immune responses after autologous transplantation. Future studies will determine the molecular mechanisms of STAT4 deficiency after transplantation and develop clinically feasible methods to circumvent this deficiency.

  • Impaired interferon-γ production as a consequence of STAT4 deficiency after autologous hematopoietic stem cell transplantation for lymphoma
    Blood, 2005
    Co-Authors: Michael J. Robertson, David Pelloso, Hua-chen Chang, Mark H. Kaplan
    Abstract:

    Production of interferon γ (IFN-γ) is critical for optimal antitumor immunotherapy in several preclinical animal models. Interleukin-12 (IL-12)–induced IFN-γ production is markedly defective after autologous stem cell transplantation. Quantitative deficiency in CD4 T cells, relative increase in CD25+CD4+ T cells, and bias toward T helper 2 (Th2) differentiation are not the primary mechanisms of defective IFN-γ production. IL-12 receptor β1 (IL-12Rβ1) and IL-12Rβ2 are expressed at equivalent or higher levels on posttransplantation patient peripheral blood mononuclear cells (PBMCs) as compared with control PBMCs. IL-12–induced tyrosine phosphorylation of signal transducer and activator of transcription 4 (STAT4) was undetectable or barely detectable in posttransplantation patient PBMCs, whereas IL-4–induced tyrosine phosphorylation of STAT6 did not differ in posttransplantation patient and control PBMCs. Levels of STAT4 Protein were decreased by 97% in posttransplantation patient PBMCs. Levels of STAT4 mRNA were also significantly decreased in posttransplantation patient PBMCs. Incubation with IL-12 and IL-18 in combination partially reversed the defective IFN-γ production by posttransplantation patient PBMCs. IFN-γ production in response to IL-12 plus IL-18 did not require increased expression of STAT4 but was dependent on the activity of p38 mitogen-activated Protein kinase (MAPK). These results indicate that defective IFN-γ production is due to an intrinsic deficiency in STAT4 expression by posttransplantation patient lymphocytes and suggest strategies for circumventing this deficiency in cancer immunotherapy.

  • Impaired Interferon-Gamma Production as a Consequence of Profound STAT4 Deficiency in Lymphocytes of Lymphoma Patients after Autologous Hematopoietic Stem Cell Transplantation.
    Blood, 2004
    Co-Authors: Michael J. Robertson, David Pelloso, Hua-chen Chang, Mark H. Kaplan
    Abstract:

    Production of IFN-gamma has been found to be critical for optimal antitumor immunotherapy in several preclinical animal models. IL-12-induced IFN-gamma production is markedly defective in patients with lymphoma who have undergone autologous hematopoietic stem cell transplantation (AHSCT). We have investigated the mechanism of defective IFN-gamma production after AHSCT. Increasing the number of CD4 T cells in IFN-gamma assays did not substantially improve IFN-gamma production by post-transplant patient peripheral blood mononuclear cells (PBMCs). Thus, the quantitative deficiency in CD4 T cells observed for up to a year after AHSCT was not the primary mechanism of defective IFN-gamma production. Post-transplant patient T cells and NK cells expressed the IL-12 receptor beta1 subunit at significantly higher levels than did control PBMCs. Moreover, IL-12 receptor beta2 was expressed by a higher proportion of NK cells from post-transplant patients as compared to control subjects. Thus, down-regulation of IL-12 receptor subunits did not account for defective IFN-gamma production by post-transplant patient PBMCs. Levels of Jak2 and Tyk2 were comparable in control and post-transplant patient PBMCs. In contrast, IL-12-induced tyrosine phosphorylation of STAT4 was undetectable or barely detectable in post-transplant patient PBMCs, whereas it was readily detectable in control PBMCs. The total levels of STAT4 Protein were also markedly decreased (P

Mark H. Kaplan - One of the best experts on this subject based on the ideXlab platform.

  • Reconstitution of STAT4 Restores Defective Interferon-gamma Production and Allows Normal IFN-gamma-Dependent Responses after Autologous Stem Cell Transplantation.
    Blood, 2006
    Co-Authors: Michael J. Robertson, David Pelloso, Hua-chen Chang, Mark H. Kaplan
    Abstract:

    Production of IFN-gamma is critical for optimal antitumor immune responses in several preclinical animal models. Patients with lymphoma who have undergone autologous hematopoietic stem cell transplantation exhibit profoundly diminished IFN-gamma production in vivo during IL-12 therapy (Clin Cancer Res.2002; 8: 3383). Furthermore, post-transplant patient peripheral blood mononuclear cells (PBMCs) stimulated directly in vitro with IL-12 secrete little if any IFN-gamma. To determine if restoration of IFN-gamma production would be sufficient to reconstitute IFN-gamma-dependent responses after transplantation, the integrity of IFN-gamma signaling was assessed in post-transplant patient PBMCs. Compared to control subject PBMCs, post-transplant patient PBMCs expressed equivalent levels of CD119 (IFN-gamma receptor alpha) and STAT1. Moreover, tyrosine phosphorylation of STAT1 in response to IFN-gamma did not differ in post-transplant patient as compared to control PBMCs. Thus, IFN-gamma signaling is intact after transplantation. Our prior studies have shown that STAT4 Protein levels are decreased by ~97% in post-transplant patient PBMCs, whereas levels of Jak2, Tyk2, and STAT3 are similar to control PBMCs (Blood2005; 106: 963). We wished to demonstrate directly that STAT4 deficiency is the mechanism of defective IFN-gamma production after autologous transplantation. Enriched CD4+ T cells were isolated from post-transplant patient and control PBMCs and cultured under conditions that promote Th1 differentiation. STAT4 deficiency persisted in post-transplant patient CD4+ T cells cultured under Th1 conditions, whereas these cells expressed normal levels of T-bet. Compared to control Th1 cells, post-transplant patient CD4+ T cells cultured under Th1 conditions exhibited markedly reduced IFN-gamma production after restimulation with CD3 mAb. These results are consistent with expected impairment of Th1 differentiation due to profound STAT4 deficiency. STAT4 cDNA was transiently transfected into Th1-cultured CD4+ T cells from post-transplant patients. After stimulation with CD3 mAb plus IL-12, the amounts of IFN-gamma secreted by STAT4-transfected patient cells were equivalent to or higher than those secreted by control subject cells. These data indicate that reconstitution of STAT4 expression is sufficient to restore IFN-gamma production by post-transplant patient PBMCs. As IFN-gamma signaling is normal post-transplant, circumventing STAT4 deficiency should therefore restore IFN-gamma-dependent antitumor immune responses after autologous transplantation. Future studies will determine the molecular mechanisms of STAT4 deficiency after transplantation and develop clinically feasible methods to circumvent this deficiency.

  • Impaired interferon-γ production as a consequence of STAT4 deficiency after autologous hematopoietic stem cell transplantation for lymphoma
    Blood, 2005
    Co-Authors: Michael J. Robertson, David Pelloso, Hua-chen Chang, Mark H. Kaplan
    Abstract:

    Production of interferon γ (IFN-γ) is critical for optimal antitumor immunotherapy in several preclinical animal models. Interleukin-12 (IL-12)–induced IFN-γ production is markedly defective after autologous stem cell transplantation. Quantitative deficiency in CD4 T cells, relative increase in CD25+CD4+ T cells, and bias toward T helper 2 (Th2) differentiation are not the primary mechanisms of defective IFN-γ production. IL-12 receptor β1 (IL-12Rβ1) and IL-12Rβ2 are expressed at equivalent or higher levels on posttransplantation patient peripheral blood mononuclear cells (PBMCs) as compared with control PBMCs. IL-12–induced tyrosine phosphorylation of signal transducer and activator of transcription 4 (STAT4) was undetectable or barely detectable in posttransplantation patient PBMCs, whereas IL-4–induced tyrosine phosphorylation of STAT6 did not differ in posttransplantation patient and control PBMCs. Levels of STAT4 Protein were decreased by 97% in posttransplantation patient PBMCs. Levels of STAT4 mRNA were also significantly decreased in posttransplantation patient PBMCs. Incubation with IL-12 and IL-18 in combination partially reversed the defective IFN-γ production by posttransplantation patient PBMCs. IFN-γ production in response to IL-12 plus IL-18 did not require increased expression of STAT4 but was dependent on the activity of p38 mitogen-activated Protein kinase (MAPK). These results indicate that defective IFN-γ production is due to an intrinsic deficiency in STAT4 expression by posttransplantation patient lymphocytes and suggest strategies for circumventing this deficiency in cancer immunotherapy.

  • Impaired Interferon-Gamma Production as a Consequence of Profound STAT4 Deficiency in Lymphocytes of Lymphoma Patients after Autologous Hematopoietic Stem Cell Transplantation.
    Blood, 2004
    Co-Authors: Michael J. Robertson, David Pelloso, Hua-chen Chang, Mark H. Kaplan
    Abstract:

    Production of IFN-gamma has been found to be critical for optimal antitumor immunotherapy in several preclinical animal models. IL-12-induced IFN-gamma production is markedly defective in patients with lymphoma who have undergone autologous hematopoietic stem cell transplantation (AHSCT). We have investigated the mechanism of defective IFN-gamma production after AHSCT. Increasing the number of CD4 T cells in IFN-gamma assays did not substantially improve IFN-gamma production by post-transplant patient peripheral blood mononuclear cells (PBMCs). Thus, the quantitative deficiency in CD4 T cells observed for up to a year after AHSCT was not the primary mechanism of defective IFN-gamma production. Post-transplant patient T cells and NK cells expressed the IL-12 receptor beta1 subunit at significantly higher levels than did control PBMCs. Moreover, IL-12 receptor beta2 was expressed by a higher proportion of NK cells from post-transplant patients as compared to control subjects. Thus, down-regulation of IL-12 receptor subunits did not account for defective IFN-gamma production by post-transplant patient PBMCs. Levels of Jak2 and Tyk2 were comparable in control and post-transplant patient PBMCs. In contrast, IL-12-induced tyrosine phosphorylation of STAT4 was undetectable or barely detectable in post-transplant patient PBMCs, whereas it was readily detectable in control PBMCs. The total levels of STAT4 Protein were also markedly decreased (P