Paul Changelian - Academia.edu (original) (raw)
Papers by Paul Changelian
Blood, Feb 15, 2008
PF-956980 is a selective inhibitor of JAK3, related in structure to CP-690550, a compound being e... more PF-956980 is a selective inhibitor of JAK3, related in structure to CP-690550, a compound being evaluated in clinical trials for rheumatoid arthritis and prevention of allograft rejection. PF-956980 has been evaluated against a panel of 30 kinases, and found to have nanomolar potency against only JAK3. Cellular and whole blood activity of this compound parallels its potency and selectivity in enzyme assays. It was effective in vivo at inhibiting the delayed type hypersensivity reaction in mice. We compared 2 commercially available JAK3 inhibitors (WHI-P131 and WHI-P154) in the same panel of biochemical and cellular assays and found them to be neither potent nor selective for JAK3. Both were found to be nanomolar inhibi-tors of the EGF receptor family of kinases. As these compounds have been used in numerous publications in the transplant and autoimmune disease literature, their specificity should be considered when interpreting these results.
Journal of Immunology, Mar 1, 1985
De ciency of the C3b/C4b receptor (CR1) of erythrocytes in systemic lupus erythematosus: analysis... more De ciency of the C3b/C4b receptor (CR1) of erythrocytes in systemic lupus erythematosus: analysis of the stability of the defect and of a restriction fragment length polymorphism of the CR1 gene.
Current Opinion in Pharmacology, Aug 1, 2012
Cytokines are critical for normal cell growth and immunoregulation but also contribute to growth ... more Cytokines are critical for normal cell growth and immunoregulation but also contribute to growth of malignant cells and drive immune-mediated disease. A major subset of immunoregulatory cytokines, roughly 60, use the type I and type II cytokine receptors and pharmacological targeting of these cytokines/cytokines receptors has proven to be efficacious in treating immune and inflammatory diseases. These receptors rely on Janus family of kinases (Jaks) for signal transduction and recently the first Jak inhibitor has been approved by the FDA. Many other Jakinibs are likely to follow and in this brief review, we will discuss the state-of-the art of this new class of pharmacological agents.
Trends in Molecular Medicine, Nov 1, 2004
The field of organ transplantation has had tremendous success because of the availability of immu... more The field of organ transplantation has had tremendous success because of the availability of immunosuppressive drugs that efficiently prevent acute organ rejection. Numerous and severe side effects are, however, associated with all current immunosuppressive therapies and justify a search for drugs with better efficacy and safety profiles. Janus kinase (JAK) 3, a tyrosine kinase that is crucial for mediating signals from the common gamma-chain of cytokine receptors, is peculiar in that its expression, contrarily to the targets of most current immunosuppressive drugs, is limited to cells that actively participate to the immune response to allografts. The recent demonstration in stringent preclinical models that JAK3 inhibition results in efficacy for the prevention of allograft rejection with a narrow side-effect profile might lead to a new era in the field of immunosuppression.
American Journal of Transplantation, 2004
Journal of Experimental Medicine, 1986
Complement receptor type 1 (CR1)/the C3b/C4b receptor (1), and CR3, the iC3b receptor (2), of mye... more Complement receptor type 1 (CR1)/the C3b/C4b receptor (1), and CR3, the iC3b receptor (2), of myelomonocytic cells have been shown to be involved in the phagocytosis of particles bearing the C3b and iC3b cleavage fragments of C3. Two states of activity exist for these receptors, a resting state, in which CR1 and CR3 only bind ligand-coated particles, and an activated state, in which these receptors also mediate the phagocytosis of such particles. The activated state was first described (3) when complement receptors on murine peritoneal macrophages were shown to acquire phagocytic capability after treatment of cells with soluble products of activated T cells. Subsequently, PMA was bound to impart phagocytic function on CR1 and CR3 of human monocytes and neutrophils (4, 5), and also to induce ligand-independent internalization of CRI by these cells (6). Finally, the capacity of three proteins of the extracellular matrix, fibronectin (FN) (5, 7, 8), serum amyloid P (5), and laminin (9), and activate CR1 and CR3 suggests that this alteration of receptor function may occur physiologically when cells migrate to the extravascular space. The biochemical reactions accounting for the transition of CR 1 and CR3 from the resting to the activated state are not known, but the ability of PMA to induce this transition suggests that protein phosphorylation by protein kinase C (I0) may be an important event. Therefore, we have assessed the phosphorylation of these proteins by phagocytic and nonphagocytic cells. CR1 of myelomonocytic cells became pbosphorylated following treatment of cells with PMA. In contrast, PMA did not induce phosphorylation of CR1 in tonsilar cells containing B lymphocytes, B lymphoblastoid cells, or erythrocytes, although relatively intense phosphorylation of CR2 (11), the C3d/EBV receptor (12), was observed in both tonsilar B cells and B lymphoblastoid cells. Materials and Methods Chemicals. BSA (fatty acid-free, fraction V; Miles Laboratories, Kankakee, IL), SDS (BioRad Laboratories, Richmond, CA), NP-40 (BDH Chemicals, Ltd., Poole, England),
Nature Reviews Drug Discovery, Jul 1, 2004
Abstract Thousands of organs are transplanted each year and millions of people suffer from autoim... more Abstract Thousands of organs are transplanted each year and millions of people suffer from autoimmune diseases, which creates a need for an armamentarium of immunosuppressive drugs. Unfortunately, immunosuppressants have unwanted side effects owing, in part, to ...
Blood, Nov 15, 2013
Phosphoinositide-3 kinases (PI3Ks) are key cellular signaling proteins that act as a central node... more Phosphoinositide-3 kinases (PI3Ks) are key cellular signaling proteins that act as a central node for relaying signals from cell surface receptors to downstream mediators, such as AKT. The PI3K-δ and PI3K-γ isoforms are preferentially expressed in normal and malignant leukocytes where they play critical roles in cell differentiation, migration, and proliferation. Constitutive activity of the PI3K pathway is common in T-cell acute lymphoblastic leukemia (T-ALL) and frequently involves the deletion of PTEN, the phosphatase that negatively regulates the PI3K pathway. An important role for the PI3K-δ and PI3K-γ isoforms has been demonstrated in the Pten-deleted genetically engineered murine model of T-ALL in conjunction with PI3K-δ and/or PI3K-γ gene knock outs (Subramanian et al, Cancer Cell, 2012). IPI-145 is a potent inhibitor of PI3K-δ and PI3K-γ currently being studied in a Phase 1 trial (IPI-145-02) in patients with advanced hematologic malignancies, including T-ALL (ClinicalTrials.gov NCT01476657). We performed in vitro studies to address the sensitivity of human and murine T-ALL cell lines to IPI-145 and to additional PI3K inhibitors with defined isoform selectivity. The human T-ALL cells examined were from cell lines that lack PTEN protein expression (Loucy, MOLT-4, CCRF-CEM, CEM/C2, p12 Ichikawa, and Karpas-45) and cell lines that express PTEN protein (MOLT13 and MOLT16). In addition, two murine cell lines derived from a Pten-deleted model of T-ALL (LPN049 and LPN236) were studied. The expression levels of class I PI3K isoforms were determined by western blotting and quantitative RT-PCR, which revealed varying levels of protein and RNA expression across the cell lines. In vitro treatment of human T-ALL cells with IPI-145 resulted in variable degrees of growth inhibition, with the PTEN-deficient Loucy cell line demonstrating the greatest sensitivity with an IC50 of 245 nM. In the cell lines tested, growth inhibition to IPI-145 was only seen in PTEN-deficient human cell lines, whereas all PTEN-expressing human T-ALL cell lines were resistant to IPI-145 (IC50 > 10 uM). However, not all PTEN-deficient human T-ALL cells demonstrated sensitivity to IPI-145 (e.g., CEM/C2), indicating that loss of PTEN does not confer sensitivity to PI3K inhibition in all settings. Inhibition of phospho-AKT (pAKT) correlated with growth inhibition, with an IC50 of 286 nM in the Loucy cell line. Studies to evaluate the mechanism of growth inhibition revealed that IPI-145 treatment resulted in apoptosis of sensitive cells as measured by 7-AAD and Annexin V staining. Cell lines derived from the Pten-deleted murine T-ALL model were also sensitive to IPI-145 with IC50s in the 300-600 nM range as measured by MTT assay. In addition, IPI-145 led to apoptosis, as measured by cleaved Caspase 3 and 7-AAD/Annexin V. Interestingly, Pten-deleted murine T-ALL cell lines showed down-regulation of pAKT and c-MYC expression with IPI-145 in a dose responsive manner that corresponded with increasing activated Caspase-3 expression. In NOTCH1-expressing murine T-ALL cell lines, down regulation of NOTCH1 and activated NOTCH1 was also observed in parallel with c-MYC down-regulation. To explore further the individual contributions of the varying PI3K isoforms on T-ALL cell growth, the effect of IPI-145 on tumor cell growth was compared with PI3K-δ, PI3K-δ,γ, and PI3K-β selective compounds in the Loucy PTEN-deficient T-ALL cell line. These experiments support an anti-leukemic effect for both PI3K-δ and PI3K-γ inhibition and indicate that the greatest effect is seen with combined PI3K-δ and PI3K-γ inhibition. A role for PI3K-β in T-ALL cell survival was not observed. Evaluation of the in vivo activity of IPI-145 on Loucy xenografts, as well as PTEN-expressing MOLT-13 xenografts, is ongoing. Together, these data provide a strong rationale for combined targeted inhibition of PI3K-δ and PI3K-γ in T-ALL. Disclosures: Huang: Infinity Pharmaceuticals, Inc.: Research Funding. Proctor:Infinity Pharmaceuticals, Inc.: Employment. Yang:Infinity Pharmaceuticals, Inc.: Research Funding. Gao:Infinity Pharmaceuticals, Inc.: Research Funding. Zhang:Infinity Pharmaceuticals, Inc.: Research Funding. Huang:Infinity Pharmaceuticals, Inc.: Research Funding. Changelian:Infinity Pharmaceuticals, Inc.: Employment. Kutok:Infinity Pharmaceuticals, Inc.: Employment. McGovern:Infinity Pharmaceuticals, Inc.: Employment. You:Infinity Pharmaceuticals, Inc.: Research Funding.
iScience, 2021
Summary Inflammatory bowel diseases (IBDs) are genetically complex and exhibit significant inter-... more Summary Inflammatory bowel diseases (IBDs) are genetically complex and exhibit significant inter-patient heterogeneity in disease presentation and therapeutic response. Here, we show that mouse models of IBD exhibit variable responses to inhibition of MK2, a pro-inflammatory serine/threonine kinase, and that MK2 inhibition suppresses inflammation by targeting inflammatory monocytes and neutrophils in murine models. Using a computational approach (TransComp-R) that allows for cross-species comparison of transcriptomic features, we identified an IBD patient subgroup that is predicted to respond to MK2 inhibition, and an independent preclinical model of chronic intestinal inflammation predicted to be non-responsive, which we validated experimentally. Thus, cross-species mouse-human translation approaches can help to identify patient subpopulations in which to deploy new therapies.
The Journal of Immunology, 2020
The discovery of JAKs and STATs and their roles in cytokine and IFN action represented a signific... more The discovery of JAKs and STATs and their roles in cytokine and IFN action represented a significant basic advance and a new paradigm in cell signaling. This was quickly followed by discoveries pointing to their essential functions, including identification of JAK3 mutations as a cause of SCID. This and other findings predicted the use of therapeutically targeting JAKs as a new strategy for treating immune and inflammatory diseases. This now is a reality with seven approved jakinibs being used to treat multiple forms of arthritis, inflammatory bowel disease and myeloproliferative neoplasms, and numerous ongoing clinical trials in other settings. This story provides interesting insights into the process of translating basic discoveries and also reveals the need to return to basic work to fill gaps that now become apparent.
Integrative Biology, 2019
Inflammatory bowel disease (IBD) is a chronic and debilitating disorder that has few treatment op... more Inflammatory bowel disease (IBD) is a chronic and debilitating disorder that has few treatment options due to a lack of comprehensive understanding of its molecular pathogenesis. We used multiplexed mass spectrometry to collect high-content information on protein phosphorylation in two different mouse models of IBD. Because the biological function of the vast majority of phosphorylation sites remains unknown, we developed Substrate-based Kinase Activity Inference (SKAI), a methodology to infer kinase activity from phosphoproteomic data. This approach draws upon prior knowledge of kinase-substrate interactions to construct custom lists of kinases and their respective substrate sites, termed kinase-substrate sets that employ prior knowledge across organisms. This expansion as much as triples the amount of prior knowledge available. We then used these sets within the Gene Set Enrichment Analysis framework to infer kinase activity based on increased or decreased phosphorylation of its s...
Transplantation, 2004
Introduction: The long-term success of ABO incompatible and positive crossmatch kidney transplant... more Introduction: The long-term success of ABO incompatible and positive crossmatch kidney transplantation is still unclear. We compared protocol kidney biopsies 1 year after positive crossmatch, ABO incompatible and negative crossmatch/ABO compatible ("conventional") kidney transplants with regard chronic interstitial fibrosis and glomerulopathy. Methods: Patients with at least one year of follow-up were included in this analysis. From 9/2000-3/2003, 25 AHG-CDC positive crossmatch transplants and 23 ABO incompatible transplants were performed. Desensitization consisted primarily of pre-transplant plasmapheresis with splenectomy in the earlier patients. During an overlaping time period, 404 conventional transplants were performed. Immunosuppression consisted of anti-thymocyte globulin induction, tacrolimus, MMF and prednisone. Protocol allograft biopsies were performed at time zero and at 4 and 12 months after transplant and were scored according to Banff '97 criteria. Statistical analyses were performed using Student's t-test and Fisher's exact test. Results: No significant differences were seen in serum creatinine levels between the groups at 4 and 12 months. The results are summarized in Table 1. No significant difference was seen in the rates of patients having a "ci" or "cg" score of zero at any time point when comparing the conventional recipients to either the positive crossmatch or ABO incompatible recipients. During the first year post-transplant, the respective cellular and humoral rejection rates were 12% and 16% in the positive crossmatch transplants, 9% and 17 % in the ABO incompatible transplants, and 4% and 1% in the conventional recipients. The incidence of glomerulopathy was low (Ͻ1%) in all groups. While not statistically significant, there was a trend toward an increase in ci2 or greater fibrosis in recipients of positive crossmatch transplants (33% vs 23% vs 18%). Importantly, more than 1/3 of patients in all groups had no histologic abnormalities at 1 year. Conclusions: Renal allograft histology of recipients of ABOi and ϩXM recipients at one year after transplantation is comparable to that of conventional transplants. Despite a humoral rejection rate of 17% in the antibody groups, the renal function was comparable at 4 and 12 months. While longer follow-up is needed, these intermediate-term results suggest that the ABOi incompatible and positive crossmatch recipients may yet have long-term outcomes similar to conventional transplants.
Journal of Medicinal Chemistry, Nov 24, 2010
There is a critical need for safer and more convenient treatments for organ transplant rejection ... more There is a critical need for safer and more convenient treatments for organ transplant rejection and autoimmune disorders such as rheumatoid arthritis. Janus tyrosine kinases (JAK1, JAK3) are expressed in lymphoid cells and are involved in the signaling of multiple cytokines important for various T cell functions. Blockade of the JAK1/JAK3-STAT pathway with a small molecule was anticipated to provide therapeutic immunosuppression/immunomodulation. The Pfizer compound library was screened against the catalytic domain of JAK3 resulting in the identification of a pyrrolopyrimidine-based series of inhibitors represented by CP-352,664 (2a). Synthetic analogues of 2a were screened against the JAK enzymes and evaluated in an IL-2 induced T cell blast proliferation assay. Select compounds were evaluated in rodent efficacy models of allograft rejection and destructive inflammatory arthritis. Optimization within this chemical series led to identification of CP-690,550 1, a potential first-in-class JAK inhibitor for treatment of autoimmune diseases and organ transplant rejection.
Proceedings of the National Academy of Sciences, 1997
JAK3 is a protein tyrosine kinase that specifically associates with the common γ chain (γ c ), a ... more JAK3 is a protein tyrosine kinase that specifically associates with the common γ chain (γ c ), a shared subunit of receptors for interleukin (IL) 2, 4, 7, 9, and 15. Patients deficient in either JAK3 or γ c presented with virtually identical forms of severe combined immunodeficiency (SCID), underscoring the importance of the JAK3–γ c interaction. Despite the key roles of JAK3 and γ c in lymphocytic development and function, the molecular basis of this interaction remains poorly understood. In this study, we have characterized the regions of JAK3 involved in γ c association. By developing a number of chimeric JAK3–JAK2 constructs, we show that the binding specificity to γ c can be conferred to JAK2 by transferring the N-terminal domains of JAK3. Moreover, those JAK3–JAK2 chimeras capable of binding γ c were also capable of reconstituting IL-2 signaling as measured by inducible phosphorylation of the chimeric JAK3–JAK2 protein, JAK1, the IL-2 receptor β chain, and signal transducer an...
Journal of Biological Chemistry, 1996
Inflammation Research, 1995
Recent developments in our understanding of lymphocyte receptor-associated signalling events have... more Recent developments in our understanding of lymphocyte receptor-associated signalling events have offered many new potential targets for modifying antigen and cytokine receptor signalling events in immune-related diseases such as allergy, autoimmunity and transplant rejection. As discussed below, these targets are largely tissue-restricted and are functionally confined to a limited set of receptors. Therefore, it is anticipated that selective inhibitors of
Blood, Feb 15, 2008
PF-956980 is a selective inhibitor of JAK3, related in structure to CP-690550, a compound being e... more PF-956980 is a selective inhibitor of JAK3, related in structure to CP-690550, a compound being evaluated in clinical trials for rheumatoid arthritis and prevention of allograft rejection. PF-956980 has been evaluated against a panel of 30 kinases, and found to have nanomolar potency against only JAK3. Cellular and whole blood activity of this compound parallels its potency and selectivity in enzyme assays. It was effective in vivo at inhibiting the delayed type hypersensivity reaction in mice. We compared 2 commercially available JAK3 inhibitors (WHI-P131 and WHI-P154) in the same panel of biochemical and cellular assays and found them to be neither potent nor selective for JAK3. Both were found to be nanomolar inhibi-tors of the EGF receptor family of kinases. As these compounds have been used in numerous publications in the transplant and autoimmune disease literature, their specificity should be considered when interpreting these results.
Journal of Immunology, Mar 1, 1985
De ciency of the C3b/C4b receptor (CR1) of erythrocytes in systemic lupus erythematosus: analysis... more De ciency of the C3b/C4b receptor (CR1) of erythrocytes in systemic lupus erythematosus: analysis of the stability of the defect and of a restriction fragment length polymorphism of the CR1 gene.
Current Opinion in Pharmacology, Aug 1, 2012
Cytokines are critical for normal cell growth and immunoregulation but also contribute to growth ... more Cytokines are critical for normal cell growth and immunoregulation but also contribute to growth of malignant cells and drive immune-mediated disease. A major subset of immunoregulatory cytokines, roughly 60, use the type I and type II cytokine receptors and pharmacological targeting of these cytokines/cytokines receptors has proven to be efficacious in treating immune and inflammatory diseases. These receptors rely on Janus family of kinases (Jaks) for signal transduction and recently the first Jak inhibitor has been approved by the FDA. Many other Jakinibs are likely to follow and in this brief review, we will discuss the state-of-the art of this new class of pharmacological agents.
Trends in Molecular Medicine, Nov 1, 2004
The field of organ transplantation has had tremendous success because of the availability of immu... more The field of organ transplantation has had tremendous success because of the availability of immunosuppressive drugs that efficiently prevent acute organ rejection. Numerous and severe side effects are, however, associated with all current immunosuppressive therapies and justify a search for drugs with better efficacy and safety profiles. Janus kinase (JAK) 3, a tyrosine kinase that is crucial for mediating signals from the common gamma-chain of cytokine receptors, is peculiar in that its expression, contrarily to the targets of most current immunosuppressive drugs, is limited to cells that actively participate to the immune response to allografts. The recent demonstration in stringent preclinical models that JAK3 inhibition results in efficacy for the prevention of allograft rejection with a narrow side-effect profile might lead to a new era in the field of immunosuppression.
American Journal of Transplantation, 2004
Journal of Experimental Medicine, 1986
Complement receptor type 1 (CR1)/the C3b/C4b receptor (1), and CR3, the iC3b receptor (2), of mye... more Complement receptor type 1 (CR1)/the C3b/C4b receptor (1), and CR3, the iC3b receptor (2), of myelomonocytic cells have been shown to be involved in the phagocytosis of particles bearing the C3b and iC3b cleavage fragments of C3. Two states of activity exist for these receptors, a resting state, in which CR1 and CR3 only bind ligand-coated particles, and an activated state, in which these receptors also mediate the phagocytosis of such particles. The activated state was first described (3) when complement receptors on murine peritoneal macrophages were shown to acquire phagocytic capability after treatment of cells with soluble products of activated T cells. Subsequently, PMA was bound to impart phagocytic function on CR1 and CR3 of human monocytes and neutrophils (4, 5), and also to induce ligand-independent internalization of CRI by these cells (6). Finally, the capacity of three proteins of the extracellular matrix, fibronectin (FN) (5, 7, 8), serum amyloid P (5), and laminin (9), and activate CR1 and CR3 suggests that this alteration of receptor function may occur physiologically when cells migrate to the extravascular space. The biochemical reactions accounting for the transition of CR 1 and CR3 from the resting to the activated state are not known, but the ability of PMA to induce this transition suggests that protein phosphorylation by protein kinase C (I0) may be an important event. Therefore, we have assessed the phosphorylation of these proteins by phagocytic and nonphagocytic cells. CR1 of myelomonocytic cells became pbosphorylated following treatment of cells with PMA. In contrast, PMA did not induce phosphorylation of CR1 in tonsilar cells containing B lymphocytes, B lymphoblastoid cells, or erythrocytes, although relatively intense phosphorylation of CR2 (11), the C3d/EBV receptor (12), was observed in both tonsilar B cells and B lymphoblastoid cells. Materials and Methods Chemicals. BSA (fatty acid-free, fraction V; Miles Laboratories, Kankakee, IL), SDS (BioRad Laboratories, Richmond, CA), NP-40 (BDH Chemicals, Ltd., Poole, England),
Nature Reviews Drug Discovery, Jul 1, 2004
Abstract Thousands of organs are transplanted each year and millions of people suffer from autoim... more Abstract Thousands of organs are transplanted each year and millions of people suffer from autoimmune diseases, which creates a need for an armamentarium of immunosuppressive drugs. Unfortunately, immunosuppressants have unwanted side effects owing, in part, to ...
Blood, Nov 15, 2013
Phosphoinositide-3 kinases (PI3Ks) are key cellular signaling proteins that act as a central node... more Phosphoinositide-3 kinases (PI3Ks) are key cellular signaling proteins that act as a central node for relaying signals from cell surface receptors to downstream mediators, such as AKT. The PI3K-δ and PI3K-γ isoforms are preferentially expressed in normal and malignant leukocytes where they play critical roles in cell differentiation, migration, and proliferation. Constitutive activity of the PI3K pathway is common in T-cell acute lymphoblastic leukemia (T-ALL) and frequently involves the deletion of PTEN, the phosphatase that negatively regulates the PI3K pathway. An important role for the PI3K-δ and PI3K-γ isoforms has been demonstrated in the Pten-deleted genetically engineered murine model of T-ALL in conjunction with PI3K-δ and/or PI3K-γ gene knock outs (Subramanian et al, Cancer Cell, 2012). IPI-145 is a potent inhibitor of PI3K-δ and PI3K-γ currently being studied in a Phase 1 trial (IPI-145-02) in patients with advanced hematologic malignancies, including T-ALL (ClinicalTrials.gov NCT01476657). We performed in vitro studies to address the sensitivity of human and murine T-ALL cell lines to IPI-145 and to additional PI3K inhibitors with defined isoform selectivity. The human T-ALL cells examined were from cell lines that lack PTEN protein expression (Loucy, MOLT-4, CCRF-CEM, CEM/C2, p12 Ichikawa, and Karpas-45) and cell lines that express PTEN protein (MOLT13 and MOLT16). In addition, two murine cell lines derived from a Pten-deleted model of T-ALL (LPN049 and LPN236) were studied. The expression levels of class I PI3K isoforms were determined by western blotting and quantitative RT-PCR, which revealed varying levels of protein and RNA expression across the cell lines. In vitro treatment of human T-ALL cells with IPI-145 resulted in variable degrees of growth inhibition, with the PTEN-deficient Loucy cell line demonstrating the greatest sensitivity with an IC50 of 245 nM. In the cell lines tested, growth inhibition to IPI-145 was only seen in PTEN-deficient human cell lines, whereas all PTEN-expressing human T-ALL cell lines were resistant to IPI-145 (IC50 > 10 uM). However, not all PTEN-deficient human T-ALL cells demonstrated sensitivity to IPI-145 (e.g., CEM/C2), indicating that loss of PTEN does not confer sensitivity to PI3K inhibition in all settings. Inhibition of phospho-AKT (pAKT) correlated with growth inhibition, with an IC50 of 286 nM in the Loucy cell line. Studies to evaluate the mechanism of growth inhibition revealed that IPI-145 treatment resulted in apoptosis of sensitive cells as measured by 7-AAD and Annexin V staining. Cell lines derived from the Pten-deleted murine T-ALL model were also sensitive to IPI-145 with IC50s in the 300-600 nM range as measured by MTT assay. In addition, IPI-145 led to apoptosis, as measured by cleaved Caspase 3 and 7-AAD/Annexin V. Interestingly, Pten-deleted murine T-ALL cell lines showed down-regulation of pAKT and c-MYC expression with IPI-145 in a dose responsive manner that corresponded with increasing activated Caspase-3 expression. In NOTCH1-expressing murine T-ALL cell lines, down regulation of NOTCH1 and activated NOTCH1 was also observed in parallel with c-MYC down-regulation. To explore further the individual contributions of the varying PI3K isoforms on T-ALL cell growth, the effect of IPI-145 on tumor cell growth was compared with PI3K-δ, PI3K-δ,γ, and PI3K-β selective compounds in the Loucy PTEN-deficient T-ALL cell line. These experiments support an anti-leukemic effect for both PI3K-δ and PI3K-γ inhibition and indicate that the greatest effect is seen with combined PI3K-δ and PI3K-γ inhibition. A role for PI3K-β in T-ALL cell survival was not observed. Evaluation of the in vivo activity of IPI-145 on Loucy xenografts, as well as PTEN-expressing MOLT-13 xenografts, is ongoing. Together, these data provide a strong rationale for combined targeted inhibition of PI3K-δ and PI3K-γ in T-ALL. Disclosures: Huang: Infinity Pharmaceuticals, Inc.: Research Funding. Proctor:Infinity Pharmaceuticals, Inc.: Employment. Yang:Infinity Pharmaceuticals, Inc.: Research Funding. Gao:Infinity Pharmaceuticals, Inc.: Research Funding. Zhang:Infinity Pharmaceuticals, Inc.: Research Funding. Huang:Infinity Pharmaceuticals, Inc.: Research Funding. Changelian:Infinity Pharmaceuticals, Inc.: Employment. Kutok:Infinity Pharmaceuticals, Inc.: Employment. McGovern:Infinity Pharmaceuticals, Inc.: Employment. You:Infinity Pharmaceuticals, Inc.: Research Funding.
iScience, 2021
Summary Inflammatory bowel diseases (IBDs) are genetically complex and exhibit significant inter-... more Summary Inflammatory bowel diseases (IBDs) are genetically complex and exhibit significant inter-patient heterogeneity in disease presentation and therapeutic response. Here, we show that mouse models of IBD exhibit variable responses to inhibition of MK2, a pro-inflammatory serine/threonine kinase, and that MK2 inhibition suppresses inflammation by targeting inflammatory monocytes and neutrophils in murine models. Using a computational approach (TransComp-R) that allows for cross-species comparison of transcriptomic features, we identified an IBD patient subgroup that is predicted to respond to MK2 inhibition, and an independent preclinical model of chronic intestinal inflammation predicted to be non-responsive, which we validated experimentally. Thus, cross-species mouse-human translation approaches can help to identify patient subpopulations in which to deploy new therapies.
The Journal of Immunology, 2020
The discovery of JAKs and STATs and their roles in cytokine and IFN action represented a signific... more The discovery of JAKs and STATs and their roles in cytokine and IFN action represented a significant basic advance and a new paradigm in cell signaling. This was quickly followed by discoveries pointing to their essential functions, including identification of JAK3 mutations as a cause of SCID. This and other findings predicted the use of therapeutically targeting JAKs as a new strategy for treating immune and inflammatory diseases. This now is a reality with seven approved jakinibs being used to treat multiple forms of arthritis, inflammatory bowel disease and myeloproliferative neoplasms, and numerous ongoing clinical trials in other settings. This story provides interesting insights into the process of translating basic discoveries and also reveals the need to return to basic work to fill gaps that now become apparent.
Integrative Biology, 2019
Inflammatory bowel disease (IBD) is a chronic and debilitating disorder that has few treatment op... more Inflammatory bowel disease (IBD) is a chronic and debilitating disorder that has few treatment options due to a lack of comprehensive understanding of its molecular pathogenesis. We used multiplexed mass spectrometry to collect high-content information on protein phosphorylation in two different mouse models of IBD. Because the biological function of the vast majority of phosphorylation sites remains unknown, we developed Substrate-based Kinase Activity Inference (SKAI), a methodology to infer kinase activity from phosphoproteomic data. This approach draws upon prior knowledge of kinase-substrate interactions to construct custom lists of kinases and their respective substrate sites, termed kinase-substrate sets that employ prior knowledge across organisms. This expansion as much as triples the amount of prior knowledge available. We then used these sets within the Gene Set Enrichment Analysis framework to infer kinase activity based on increased or decreased phosphorylation of its s...
Transplantation, 2004
Introduction: The long-term success of ABO incompatible and positive crossmatch kidney transplant... more Introduction: The long-term success of ABO incompatible and positive crossmatch kidney transplantation is still unclear. We compared protocol kidney biopsies 1 year after positive crossmatch, ABO incompatible and negative crossmatch/ABO compatible ("conventional") kidney transplants with regard chronic interstitial fibrosis and glomerulopathy. Methods: Patients with at least one year of follow-up were included in this analysis. From 9/2000-3/2003, 25 AHG-CDC positive crossmatch transplants and 23 ABO incompatible transplants were performed. Desensitization consisted primarily of pre-transplant plasmapheresis with splenectomy in the earlier patients. During an overlaping time period, 404 conventional transplants were performed. Immunosuppression consisted of anti-thymocyte globulin induction, tacrolimus, MMF and prednisone. Protocol allograft biopsies were performed at time zero and at 4 and 12 months after transplant and were scored according to Banff '97 criteria. Statistical analyses were performed using Student's t-test and Fisher's exact test. Results: No significant differences were seen in serum creatinine levels between the groups at 4 and 12 months. The results are summarized in Table 1. No significant difference was seen in the rates of patients having a "ci" or "cg" score of zero at any time point when comparing the conventional recipients to either the positive crossmatch or ABO incompatible recipients. During the first year post-transplant, the respective cellular and humoral rejection rates were 12% and 16% in the positive crossmatch transplants, 9% and 17 % in the ABO incompatible transplants, and 4% and 1% in the conventional recipients. The incidence of glomerulopathy was low (Ͻ1%) in all groups. While not statistically significant, there was a trend toward an increase in ci2 or greater fibrosis in recipients of positive crossmatch transplants (33% vs 23% vs 18%). Importantly, more than 1/3 of patients in all groups had no histologic abnormalities at 1 year. Conclusions: Renal allograft histology of recipients of ABOi and ϩXM recipients at one year after transplantation is comparable to that of conventional transplants. Despite a humoral rejection rate of 17% in the antibody groups, the renal function was comparable at 4 and 12 months. While longer follow-up is needed, these intermediate-term results suggest that the ABOi incompatible and positive crossmatch recipients may yet have long-term outcomes similar to conventional transplants.
Journal of Medicinal Chemistry, Nov 24, 2010
There is a critical need for safer and more convenient treatments for organ transplant rejection ... more There is a critical need for safer and more convenient treatments for organ transplant rejection and autoimmune disorders such as rheumatoid arthritis. Janus tyrosine kinases (JAK1, JAK3) are expressed in lymphoid cells and are involved in the signaling of multiple cytokines important for various T cell functions. Blockade of the JAK1/JAK3-STAT pathway with a small molecule was anticipated to provide therapeutic immunosuppression/immunomodulation. The Pfizer compound library was screened against the catalytic domain of JAK3 resulting in the identification of a pyrrolopyrimidine-based series of inhibitors represented by CP-352,664 (2a). Synthetic analogues of 2a were screened against the JAK enzymes and evaluated in an IL-2 induced T cell blast proliferation assay. Select compounds were evaluated in rodent efficacy models of allograft rejection and destructive inflammatory arthritis. Optimization within this chemical series led to identification of CP-690,550 1, a potential first-in-class JAK inhibitor for treatment of autoimmune diseases and organ transplant rejection.
Proceedings of the National Academy of Sciences, 1997
JAK3 is a protein tyrosine kinase that specifically associates with the common γ chain (γ c ), a ... more JAK3 is a protein tyrosine kinase that specifically associates with the common γ chain (γ c ), a shared subunit of receptors for interleukin (IL) 2, 4, 7, 9, and 15. Patients deficient in either JAK3 or γ c presented with virtually identical forms of severe combined immunodeficiency (SCID), underscoring the importance of the JAK3–γ c interaction. Despite the key roles of JAK3 and γ c in lymphocytic development and function, the molecular basis of this interaction remains poorly understood. In this study, we have characterized the regions of JAK3 involved in γ c association. By developing a number of chimeric JAK3–JAK2 constructs, we show that the binding specificity to γ c can be conferred to JAK2 by transferring the N-terminal domains of JAK3. Moreover, those JAK3–JAK2 chimeras capable of binding γ c were also capable of reconstituting IL-2 signaling as measured by inducible phosphorylation of the chimeric JAK3–JAK2 protein, JAK1, the IL-2 receptor β chain, and signal transducer an...
Journal of Biological Chemistry, 1996
Inflammation Research, 1995
Recent developments in our understanding of lymphocyte receptor-associated signalling events have... more Recent developments in our understanding of lymphocyte receptor-associated signalling events have offered many new potential targets for modifying antigen and cytokine receptor signalling events in immune-related diseases such as allergy, autoimmunity and transplant rejection. As discussed below, these targets are largely tissue-restricted and are functionally confined to a limited set of receptors. Therefore, it is anticipated that selective inhibitors of