Preclinical and Clinical Trial Results Using Talazoparib and Low-Dose Chemotherapy.

PURPOSE: On the basis of preclinical data, we hypothesized that low doses of chemotherapy (10% of therapeutic doses) with full dose of a PARP inhibitor could have improved efficacy and tolerability. PATIENTS AND METHODS: In this phase I dose-escalation study, patients with BRCA-normal advanced malignancies were assigned to either talazoparib/temozolomide or talazoparib/irinotecan. Talazoparib was dose-escalated from 500 mcg to 1 mg daily before dose escalation of temozolomide/irinotecan. The starting dose of temozolomide was 25 mg/m2/day orally on days 1 to 5 and irinotecan was 25 mg/m2/day intravenously on days 1 and 15. The primary objectives of this trial were safety and tolerability, dose-limiting toxicities (DLT), and maximum tolerated dose (MTD). RESULTS: Of 40 patients enrolled, 18 (mean: 7 prior therapies) were enrolled in talazoparib + temozolomide and 22 in talazoparib + irinotecan. DLTs were hematologic in both arms, but all hematologic adverse events resolved with either treatment interruption and/or dose reductions of talazoparib. The MTDs were talazoparib 1 mg + temozolomide 37.5 mg/m2 and talazoparib 1 mg + irinotecan 37.5 mg/m2. There were four partial responses in the talazoparib + temozolomide arm and five in the talazoparib + irinotecan arm for a response rate of 23% (9/40). The pharmacokinetic profiles of talazoparib + temozolomide/irinotecan were similar to that of talazoparib monotherapy. Responses were seen independent of homologous recombination (HR) status and HR deficiency score. CONCLUSIONS: These results show that talazoparib with low-dose temozolomide or irinotecan is reasonably well tolerated and demonstrates clinical activity in a wide range of cancers. Randomized trials of talazoparib with or without low-dose chemotherapy are ongoing in small cell lung cancer and ovarian cancer.

Keeping immune checkpoint inhibitor myocarditis in check: advanced circulatory mechanical support as a bridge to recovery.

Immune checkpoint inhibitor (ICI)-associated myocarditis is a rare, potentially life-threatening complication of immunotherapy. We report a case of a 60-year-old female with a history of colorectal cancer treated with nivolumab immunotherapy who presented with new cardiomyopathy complicated by cardiogenic shock and ventricular arrhythmias. Treatment of ICI-associated myocarditis requires aggressive immunosuppression and supportive therapy. In this case, the patient required advanced mechanical circulatory support as a bridge to recovery. This case highlights the complexity of diagnosis, haemodynamic management, and treatment of fulminant ICI myocarditis.

The sphingosine-1-phosphate/sphingosine-1-phosphate receptor 2 axis regulates early airway T-cell infiltration in murine mast cell-dependent acute allergic responses.

BACKGROUND: Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid produced by mast cells (MCs) on cross-linking of their high-affinity receptors for IgE by antigen that can amplify MC responses by binding to its S1P receptors. An acute MC-dependent allergic reaction can lead to systemic shock, but the early events of its development in lung tissues have not been investigated, and S1P functions in the onset of allergic processes remain to be examined. OBJECTIVE: We used a highly specific neutralizing anti-S1P antibody (mAb) and the sphingosine-1-phosphate receptor 2 (S1PR2) antagonist JTE-013 to study the signaling contributions of S1P and S1PR2 to MC- and IgE-dependent airway allergic responses in mice within minutes after antigen challenge. METHODS: Allergic reaction was triggered by a single intraperitoneal dose of antigen in sensitized mice pretreated intraperitoneally with anti-S1P, isotype control mAb, JTE-013, or vehicle before antigen challenge. RESULTS: Kinetics experiments revealed early pulmonary infiltration of mostly T cells around blood vessels of sensitized mice 20 minutes after antigen exposure. Pretreatment with anti-S1P mAb inhibited in vitro MC activation, as well as in vivo development of airway infiltration and MC activation, reducing serum levels of histamine, cytokines, and the chemokines monocyte chemoattractant protein 1/CCL2, macrophage inflammatory protein 1α/CCL3, and RANTES/CCL5. S1PR2 antagonism or deficiency or MC deficiency recapitulated these results. Both in vitro and in vivo experiments demonstrated MC S1PR2 dependency for chemokine release and the necessity for signal transducer and activator of transcription 3 activation. CONCLUSION: Activation of S1PR2 by S1P and downstream signal transducer and activator of transcription 3 signaling in MCs regulate early T-cell recruitment to antigen-challenged lungs through chemokine production.

K63-linked polyubiquitination of transcription factor IRF1 is essential for IL-1-induced production of chemokines CXCL10 and CCL5.

Although interleukin 1 (IL-1) induces expression of the transcription factor IRF1 (interferon-regulatory factor 1), the roles of IRF1 in immune and inflammatory responses and mechanisms of its activation remain elusive. Here we found that IRF1 was essential for IL-1-induced expression of the chemokines CXCL10 and CCL5, which recruit mononuclear cells into sites of sterile inflammation. Newly synthesized IRF1 acquired Lys63 (K63)-linked polyubiquitination mediated by the apoptosis inhibitor cIAP2 that was enhanced by the bioactive lipid S1P. In response to IL-1, cIAP2 and the sphingosine kinase SphK1 (the enzyme that generates S1P) formed a complex with IRF1, which led to its activation. Thus, IL-1 triggered a hitherto unknown signaling cascade that controlled the induction of IRF1-dependent genes that encode molecules important for sterile inflammation.

Sphingosine-1-phosphate links persistent STAT3 activation, chronic intestinal inflammation, and development of colitis-associated cancer.

Inflammatory bowel disease is an important risk factor for colorectal cancer. We show that sphingosine-1-phosphate (S1P) produced by upregulation of sphingosine kinase 1 (SphK1) links chronic intestinal inflammation to colitis-associated cancer (CAC) and both are exacerbated by deletion of Sphk2. S1P is essential for production of the multifunctional NF-κB-regulated cytokine IL-6, persistent activation of the transcription factor STAT3, and consequent upregulation of the S1P receptor, S1PR1. The prodrug FTY720 decreased SphK1 and S1PR1 expression and eliminated the NF-κB/IL-6/STAT3 amplification cascade and development of CAC, even in Sphk2(-/-) mice, and may be useful in treating colon cancer in individuals with ulcerative colitis. Thus, the SphK1/S1P/S1PR1 axis is at the nexus between NF-κB and STAT3 and connects chronic inflammation and CAC.

A specific sphingosine kinase 1 inhibitor attenuates airway hyperresponsiveness and inflammation in a mast cell-dependent murine model of allergic asthma.

BACKGROUND: Sphingosine-1-phosphate (S1P), which is produced by 2 sphingosine kinase (SphK) isoenzymes, SphK1 and SphK2, has been implicated in IgE-mediated mast cell responses. However, studies of allergic inflammation in isotype-specific SphK knockout mice have not clarified their contribution, and the role that S1P plays in vivo in a mast cell- and IgE-dependent murine model of allergic asthma has not yet been examined. OBJECTIVE: We used an isoenzyme-specific SphK1 inhibitor, SK1-I, to investigate the contributions of S1P and SphK1 to mast cell-dependent airway hyperresponsiveness (AHR) and airway inflammation in mice. METHODS: Allergic airway inflammation and AHR were examined in a mast cell-dependent murine model of ovalbumin (OVA)-induced asthma. C57BL/6 mice received intranasal delivery of SK1-I before sensitization and challenge with OVA or only before challenge. RESULTS: SK1-I inhibited antigen-dependent activation of human and murine mast cells and suppressed activation of nuclear factor κB (NF-κB), a master transcription factor that regulates the expression of proinflammatory cytokines. SK1-I treatment of mice sensitized to OVA in the absence of adjuvant, in which mast cell-dependent allergic inflammation develops, significantly reduced OVA-induced AHR to methacholine; decreased numbers of eosinophils and levels of the cytokines IL-4, IL-5, IL-6, IL-13, IFN-γ, and TNF-α and the chemokines eotaxin and CCL2 in bronchoalveolar lavage fluid; and decreased pulmonary inflammation, as well as activation of NF-κB in the lungs. CONCLUSION: S1P and SphK1 play important roles in mast cell-dependent, OVA-induced allergic inflammation and AHR, in part by regulating the NF-κB pathway.

Sphingosine-1-phosphate produced by sphingosine kinase 2 in mitochondria interacts with prohibitin 2 to regulate complex IV assembly and respiration.

The potent lipid mediator sphingosine-1-phosphate (S1P) regulates diverse physiological processes by binding to 5 specific GPCRs, although it also has intracellular targets. Here, we demonstrate that S1P, produced in the mitochondria mainly by sphingosine kinase 2 (SphK2), binds with high affinity and specificity to prohibitin 2 (PHB2), a highly conserved protein that regulates mitochondrial assembly and function. In contrast, S1P did not bind to the closely related protein PHB1, which forms large, multimeric complexes with PHB2. In mitochondria from SphK2-null mice, a new aberrant band of cytochrome-c oxidase was detected by blue native PAGE, and interaction between subunit IV of cytochrome-c oxidase and PHB2 was greatly reduced. Moreover, depletion of SphK2 or PHB2 led to a dysfunction in mitochondrial respiration through cytochrome-c oxidase. Our data point to a new action of S1P in mitochondria and suggest that interaction of S1P with homomeric PHB2 is important for cytochrome-c oxidase assembly and mitochondrial respiration.

Essential roles of sphingosine-1-phosphate receptor 2 in human mast cell activation, anaphylaxis, and pulmonary edema.

Systemic exacerbation of allergic responses, in which mast cells play a critical role, results in life-threatening anaphylactic shock. Sphingosine-1-phosphate (S1P), a ligand for a family of G protein-coupled receptors, is a new addition to the repertoire of bioactive lipids secreted by activated mast cells. Yet little is known of its role in human mast cell functions and in anaphylaxis. We show that S1P(2) receptors play a critical role in regulating human mast cell functions, including degranulation and cytokine and chemokine release. Immunoglobulin E-triggered anaphylactic responses, including elevation of circulating histamine and associated pulmonary edema in mice, were significantly attenuated by the S1P(2) antagonist JTE-013 and in S1P(2)-deficient mice, in contrast to anaphylaxis induced by administration of histamine or platelet-activating factor. Hence, S1P and S1P(2) on mast cells are determinants of systemic anaphylaxis and associated pulmonary edema and might be beneficial targets for anaphylaxis attenuation and prophylaxis.

Sphingosine-1-phosphate induces development of functionally mature chymase-expressing human mast cells from hematopoietic progenitors.

Mast cells (MCs) play a critical role in both acute and chronic inflammation and mature in peripheral tissues from bone marrow-derived progenitors that circulate in the blood as immature precursors. MCs developed from cord blood-derived progenitors cultured with stem cell factor (SCF) alone express intragranular tryptase (MC(T)s), the phenotype predominant in the lung. MC progenitors are likely to encounter the serum-borne bioactive sphingolipid metabolite, sphingosine-1-phosphate (S1P), during migration to target tissues. S1P accelerated the development of cord blood-derived MCs (CB-MCs) and strikingly increased the numbers of MC-expressing chymase. These MCs have functional Fc epsilonRIs, and similar to skin MC(TC)s that express both tryptase and chymase, also express CD88 and are activated by anaphylatoxin C5a and the secretagogue compound 48/80. S1P induced release of IL-6, a cytokine known to promote development of functionally mature MC(TC)s, from cord blood cultures containing adherent macrophages, and from highly purified macrophages, but not from macrophage-depleted CB-MCs. In contrast, S1P stimulated secretion of the chemokine, monocyte chemoattractant protein 1 (MCP-1/CCL2), from these macrophage-depleted and purified CB-MCs. These results suggest crucial roles for S1P in regulating development of human MCs and their functions and reveal a complex interplay between macrophages and MC progenitors in the development of mature human MCs.

Distinct roles of sphingosine kinases 1 and 2 in human mast-cell functions.

Sphingosine-1-phosphate (S1P) is now emerging as a potent lipid mediator produced by mast cells that contributes to inflammatory and allergic responses. In contrast to its weak effect on degranulation of murine mast cells, S1P potently induced degranulation of the human LAD2 mast-cell line and cord blood-derived human mast cells (hMCs). S1P also stimulated production and secretion of cytokines, TNF-alpha and IL-6, and markedly enhanced secretion of a chemokine, CCL2/MCP-1, important modulators of inflammation. S1P is produced in mast cells by the 2 sphingosine kinases, SphK1 and SphK2. SphK1 but not SphK2 plays a critical role in IgE/Ag-induced degranulation, migration toward antigen, and CCL2 secretion from hMCs, as determined by specifically down-regulating their expression. However, both isoenzymes were required for efficient TNF-alpha secretion. Taken together, our data suggest that differential formation of S1P by SphK1 and SphK2 has distinct and important actions in hMCs.

Sphingosine-1-phosphate synthesis and functions in mast cells.

Sphingolipids are not only major lipid components of all eukaryotic cell membranes, but they also comprise an important family of bioactive signaling molecules that regulate a diverse array of biological responses. The sphingolipid metabolite sphingosine-1-phosphate (S1P), is a key regulator of immune responses. Cellular levels of S1P are determined by the balance between its synthesis, involving two sphingosine kinases (SphK1 and SphK2), and its degradation, involving S1P lyase and S1P phosphatases. S1P mainly signals through its cell-surface receptors and may also have intracellular functions. S1P has important functions in mast cells - the major effectors of allergic responses. Antigen triggering of IgE receptors on mast cells activates both SphKs resulting in the production of S1P that is released and regulates and amplifies mast cell functions, including degranulation as well as cytokine and chemokine release.