IFN-α In Patients Research Articles

Final Analysis of the Daliah Trial: A Randomized Phase III Trial of Interferon-α Versus Hydroxyurea in Patients with MPN

Background: Hydroxyurea (HU) is the most commonly used first-line cytoreductive treatment option for patients with myeloproliferative neoplasms (MPN) worldwide. However, increasing evidence on the efficacy and safety of pegylated interferon-alpha2 (IFNα) is emerging, and optimal first-line treatment is to be established. Aims: To compare the efficacy and safety of HU vs. low-dose IFNα in patients with MPN over five years. Methods: DALIAH (NCT01387763) was a randomized phase III trial of HU vs. IFNα in newly diagnosed or untreated patients with MPN (essential thrombocythemia (ET), polycythemia vera (PV), prefibrotic myelofibrosis (PreMF), and primary myelofibrosis (PMF)). Patients > age 60 were randomized (1:1:1) to HU, IFNα-2a, or IFNα-2b whereas patients ≤ age 60 were randomized to IFNα-2a or IFNα-2b. The primary outcome was the JAK2V617F molecular response (MR) rate at 18, 36, and 60 months per 2009 European LeukemiaNetwork (ELN) (ET, PV, PreMF) or 2005 European Myelofibrosis Network (EUMNET) (PMF) criteria. Secondary outcomes included the complete clinicohematologic response (CHR) rate at 12 months. The JAK2V617F allele burden was measured using quantitative polymerase chain reaction (qPCR) on peripheral blood (assay sensitivity: 0.1%). Primary and secondary outcomes were compared between groups (HU vs. IFNα or HU vs. IFNα > 60 years) using Fisher's exact test (categorical variables) or Wilcoxon rank-sum test (continuous variables). Paired comparisons (within groups) were made using Wilcoxon signed-rank test. Serial JAK2V617F measurements were compared by unadjusted mixed-effects linear regression analysis. Results: We included 203 patients (ET: 73 (36%), PV: 89 (44%), PreMF: 16 (8%), and PMF: 25 (12%)) in the modified intention-to-treat (ITT) population. Baseline characteristics were well balanced except for median age (HU: 68 years vs. IFNα: 59 years, p<0.0001) ( Table 1). The MR rate by ITT analysis was similar between HU and IFNα (18 months: 19% vs. 21%, p=1.00; 36 months: 19% vs. 26%, p=0.64; 60 months: 23% vs. 24%, p=1.00) (Figure 1A). However, the JAK2V617F allele burden was significantly lower in the IFNα group at month 36 and beyond (Figure 1B) and the absolute median (IQR) change in JAK2V617F allele burden (baseline to 60 months) was greater with IFNα (-20% (-9;-49) vs. -7% (3;-15), p=0.0053) (Figure 1C). Two patients (IFNα: n=2) were in complete molecular remission (undetectable JAK2V617F) at 60 months. The CHR rate by ITT analysis was higher with HU at 18 months (58% vs. 38%, p=0.03) but similar at all other time points (12 months: 50% vs. 36%, p=0.21; 60 months: 24% vs. 22%, p=0.83) ( Figure 1D). A post hoc subgroup analysis comparing HU with IFNα > age 60 showed comparable efficacy results. Among patients remaining on treatment (per-protocol analysis), the MR and CHR rates were superior in the IFNα group compared to the HU group at 36 months and beyond. The MR and CHR rates (HU vs. IFNα) by per-protocol analysis were: MR at 36 months: 23% vs. 56%, p=0.01; MR at 48 months: 27% vs. 59%, p=0.02; MR at 60 months: 35% vs. 67%, p=0.03; CHR at 36 months: 33% vs. 67%, p=0.002; CHR at 60 months: 38% vs. 62%, p=0.05. Overall treatment discontinuation at 60 months was 60% (HU: 37% vs. IFNα: 65%, p=0.0019). The most common cause of treatment discontinuation was adverse events (HU: 6/38 (16%); IFNα: 71/165 (43%)). Adverse events ≥ grade 3 occurred in 46% (HU: 58% vs. IFNα: 45%, p=0.15). Nineteen major thrombotic events were reported in 16 patients (HU: 4 events in 4 patients; IFNα>60: 12 events in 10 patients; IFNα≤60: 3 events in 2 patients), corresponding to an incidence rate of 2.6 per 100 person-years for HU and 3.4 per 100 person-years for IFNα (IFNα>60: 6.2; IFNα≤60: 1.2). None of the patients transformed into secondary acute myeloid leukemia. Five patients died during follow-up (HU: 2; IFNα: 3). Bone marrow histologic remission rates at 36 and 60 months will be presented at the meeting. Conclusion: ITT analysis detected no significant difference in the MR or CHR rates between HU and IFNα with long-term treatment (5 years) among patients with MPN, reflecting a higher treatment discontinuation rate in the IFNα group (65%). Thus, using the per-protocol principle, the MR and CHR rates were superior in the IFNα group at 36 months and beyond.

Systemic infusion of TLR3-ligand and IFN-α in patients with breast cancer reprograms local tumor microenvironments for selective CTL influx

BackgroundPresence of cytotoxic T lymphocytes (CTL) in the tumor microenvironment (TME) predicts the effectiveness of cancer immunotherapies. The ability of toll-like receptor 3 (TLR3) ligands, interferons (IFNs) and COX2 inhibitors...

Activation of nucleotide-binding oligomerization domain 2 by muramyl dipeptide negatively regulates Toll-like receptor 9-mediated colonic inflammation through the induction of deubiquitinating enzyme A expression.

Mutations in nucleotide-binding oligomerization domain 2 (NOD2) are associated with Crohn's disease (CD). Although NOD2 activation contributes to the maintenance of intestinal homeostasis through the negative regulation of pro-inflammatory cytokine responses mediated by Toll-like receptors (TLRs), the effects of NOD2 activation on interferon (IFN)-α responses induced by TLR9 have been poorly defined. To explore the cross-talk between NOD2 and TLR9, human monocytes or dendritic cells (DCs) were stimulated with NOD2 and/or TLR9 ligands to measure IFN-α production. The severity of dextran sodium sulfate (DSS)-induced colitis was compared in mice treated with NOD2 and/or TLR9 ligands. Expression of IFN-α and IFN-stimulated genes (ISGs) was examined in the colonic mucosa of patients with inflammatory bowel disease (IBD). NOD2 activation reduced TLR9-induced IFN-α production by monocytes and DCs in a deubiquitinating enzyme A (DUBA)-dependent manner. Activation of DUBA induced by the co-stimulation of TLR9 and NOD2 inhibited Lys63-linked polyubiquitination of TRAF3 and suppressed TLR9-mediated IFN-α production. NOD2 activation in hematopoietic cells protected mice from TLR9-induced exacerbation of DSS-induced colitis by down-regulating IFN-α responses and up-regulating DUBA expression. Colonic mucosa of patients with active and remitted IBD phases was characterized by the enhanced and reduced expression of ISGs, respectively. Expression levels of IFN-α and IL-6 positively correlated in the active colonic mucosa of patients with ulcerative colitis and CD, whereas DUBA expression inversely correlated with that of IFN-α in patients with CD. Collectively, these data suggest that DUBA-dependent negative effect of NOD2 on TLR9-mediated IFN-α responses contributes to the maintenance of intestinal homeostasis.

Autoantibodies against IFNα in patients with systemic lupus erythematosus and susceptibility for infection: a retrospective case-control study

IFNα and anti-IFNα autoantibodies have been implicated in susceptibility both for systemic lupus erythematosus (SLE) and viral infection. We aimed to analyze the SLE disease phenotype and risk for infection associated with anti-IFN-α IgG autoantibodies in SLE patients In this multidisciplinary retrospective single referral center study, all consecutive patients with SLE admitted between January 1st and November 30th 2020 were considered. All subjects fulfilled the ACR/EULAR 2019 criteria for SLE. Anti-IFNα IgG autoantibodies were quantified at admission by ELISA. Demographic, medical history, laboratory, treatment, and outcome data were extracted from electronic medical records using a standardized data collection form. 180 patients [female 87.2%, median age of 44.4 (34–54.2) years] were included. The median disease duration was 10 years [4–20] with a median SLEDAI score of 2 [0–4] at study time. Fifty-four (30%) patients had a past-history of lupus nephritis. One hundred and forty-four (80%) had received long-term glucocorticoids and 99 (55%) immunosuppressive drugs. Overall, 127 infections—mostly bacterial and viral—were reported in 95 (52.8%) patients. Twenty SLE patients (11.1%) had positive anti-IFNα IgG autoantibodies with a titer ranging from 10 to 103 UA/mL. Age, sex, SLE phenotype and treatment did not significantly differ between SLE patients with or without anti-IFNα. Infection rate was similar in both groups except for tuberculosis which was more frequent in patients with anti-IFNα (20% vs. 3.1%, p = 0.01). The prevalence of autoantibodies against IFNα is high in SLE and associated with a higher frequency of tuberculosis.

729P Adoptive T-cell therapy during chemotherapy with or without peginterferon-α (IFNα) in patients with platinum sensitive recurrent epithelial ovarian cancer (EOC)

The correlation between T-cell infiltration and EOC survival suggests that EOC may be sensitive to adoptive cell therapy with autologous tumor-infiltrating lymphocytes (TIL). Carboplatin-paclitaxel chemotherapy (CPC) lowers the number of tumor induced immune suppressive myeloid cells, creating a window-of-opportunity for TILs to exert their effective anti-tumor response. IFNα supports TIL. A phase I/II trial (NCT04072263) was initiated to study the feasibility and safety of TIL during CPC +/- IFNα in patients with recurrent platinum sensitive EOC.

Inferring the dynamics of mutated hematopoietic stem and progenitor cells induced by IFNα in myeloproliferative neoplasms

AbstractClassical BCR-ABL–negative myeloproliferative neoplasms (MPNs) are clonal disorders of hematopoietic stem cells (HSCs) caused mainly by recurrent mutations in genes encoding JAK2 (JAK2), calreticulin (CALR), or the thrombopoietin receptor (MPL). Interferon α (IFNα) has demonstrated some efficacy in inducing molecular remission in MPNs. To determine factors that influence molecular response rate, we evaluated the long-term molecular efficacy of IFNα in patients with MPN by monitoring the fate of cells carrying driver mutations in a prospective observational and longitudinal study of 48 patients over more than 5 years. We measured the clonal architecture of early and late hematopoietic progenitors (84 845 measurements) and the global variant allele frequency in mature cells (409 measurements) several times per year. Using mathematical modeling and hierarchical Bayesian inference, we further inferred the dynamics of IFNα-targeted mutated HSCs. Our data support the hypothesis that IFNα targets JAK2V617F HSCs by inducing their exit from quiescence and differentiation into progenitors. Our observations indicate that treatment efficacy is higher in homozygous than heterozygous JAK2V617F HSCs and increases with high IFNα dose in heterozygous JAK2V617F HSCs. We also found that the molecular responses of CALRm HSCs to IFNα were heterogeneous, varying between type 1 and type 2 CALRm, and a high dose of IFNα correlates with worse outcomes. Our work indicates that the long-term molecular efficacy of IFNα implies an HSC exhaustion mechanism and depends on both the driver mutation type and IFNα dose.

A Translational Investigation of IFN-α and STAT1 Signaling in Endothelial Cells during Septic Shock Provides Therapeutic Perspectives.

Septic shock and disseminated intravascular coagulation (DIC) are known to be characterized by an endothelial cell dysfunction. The molecular mechanisms underlying this relationship are, however, poorly understood. In this work, we aimed to investigate human circulating IFN-α in patients with septic shock-induced DIC and tested the potential role of endothelial Stat1 (signal transducer and activator of transcription 1) as a therapeutic target in a mouse model of sepsis. For this, circulating type I, type II, and type III IFNs and procoagulant microvesicles were quantified in a prospective cohort of patients with septic shock. Next, we used a septic shock model induced by cecal ligation and puncture in wild-type mice, in Ifnar1 (type I IFN receptor subunit 1)-knockout mice, and in Stat1 conditional knockout mice. In human samples, we observed higher concentrations of circulating IFN-α and IFN-α1 in patients with DIC compared with patients without DIC, whereas concentrations of IFN-β, IFN-γ, IFN-λ1, IFN-λ2, and IFN-λ3 were not different. IFN-α concentration was positively correlated with CD105 microvesicle concentrations, reflecting endothelial injury. In Ifnar1-/- mice, cecal ligation and puncture did not induce septic shock and was characterized by lesser endothelial cell injury, with lower aortic inflammatory cytokine expression, endothelial inflammatory-related gene expression, and fibrinolysis. In mice in which Stat1 was specifically ablated in endothelial cells, a marked protection against sepsis was also observed, suggesting the relevance of an endothelium-targeted strategy. Our work highlights the key roles of type I IFNs as pathogenic players in septic shock-induced DIC and the potential pertinence of endothelial STAT1 as a therapeutic target.

Distinct effects of ruxolitinib and interferon-alpha on murine JAK2V617F myeloproliferative neoplasm hematopoietic stem cell populations.

JAK2V617F is the most common mutation in patients with BCR-ABL negative myeloproliferative neoplasms (MPNs). The eradication of JAK2V617F hematopoietic stem cells (HSCs) is critical for achieving molecular remissions and cure. We investigate the distinct effects of two therapies, ruxolitinib (JAK1/2 inhibitor) and interferon-alpha (IFN-α), on the disease-initiating HSC population. Whereas ruxolitinib inhibits Stat5 activation in erythroid progenitor populations, it fails to inhibit this same pathway in HSCs. In contrast, IFN-α has direct effects on HSCs. Furthermore, STAT1 phosphorylation and pathway activation is greater after IFN-α stimulation in Jak2V617F murine HSCs with increased induction of reactive oxygen species, DNA damage and reduction in quiescence after chronic IFN-α treatment. Interestingly, ruxolitinib does not block IFN-α induced reactive oxygen species and DNA damage in Jak2V617F murine HSCs in vivo. This work provides a mechanistic rationale informing how pegylated IFN-α reduces JAK2V617F allelic burden in the clinical setting and may inform future clinical efforts to combine ruxolitinib with pegylated IFN-α in patients with MPN.

Angiotensin-converting enzyme inhibitor suppresses Type I interferon responses, decreases neuroinflammation and improves depression-like behavior in MRL/lpr lupusprone mice

Abstract Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by multi-organ damage. Interferon- α (IFN-α) is a central mediator in disease pathogenesis. Neuropsychiatric lupus (NPSLE) is one of the most common manifestations of human SLE often causing depression. Administration of IFN-α in patients with chronic viral infections or cancers causes depressive symptoms. Angiotensin-converting enzyme (ACE) is part of the kallikrein-kinin/renin-angiotensin (KKS/RAS) system that regulates many physiological processes, including inflammation and brain functions. It is known that ACE degrades bradykinin into inactive peptides. We have previously shown in an in vitro model of mouse bone-marrow-derived dendritic cells that the effects of captopril (an ACE inhibitor) on IFN responsive gene inhibition were reversed when bradykinin receptors were blocked, suggesting that ACE, in part, acts via kinin receptors to bring about the suppressive effects. We used the MRL/lpr lupus-prone mouse model, an established model to study NPSLE, to check the in vivo effects of captopril on IFN-induced immune responses, neuroinflammation, and behavior. Exposing MRL/lpr mice to IFN-α increased depressive-like behavior and enhanced ACE expression in the brain. Administering captopril in MRL/lpr mice decreased expression of IFN responsive genes in brain and kidney, decreased microglial activation (IBA1 expression) and reduced depressive-like behavior as assessed by the forced swim test. Thus, ACE inhibitors may have potential for therapeutic use for systemic and neurolupus.

Hypomethylation of STAT1 and HLA-DRB1 is associated with type-I interferon-dependent HLA-DRB1 expression in lupus CD8+ T cells

ObjectiveWe examined genome-wide DNA methylation changes in CD8+ T cells from patients with lupus and controls and investigated the functional relevance of some of these changes in lupus.MethodsGenome-wide DNA methylation...

Hepatitis B virus sensitivity to interferon-α in hepatocytes is more associated with cellular interferon response than with viral genotype.

In the cell-culture-based HBV infection models, the sensitivity of HBV to IFN-α in hepatocytes is determined more by the cell-intrinsic IFN response than by viral genotype, and improvement of the IFN response in HepG2-NTCP cells promotes the efficacy of IFN-α against HBV. (Hepatology 2018;67:1237-1252).

Benefit of adjuvant immunotherapy in renal cell carcinoma: A myth or a reality?

BackgroundThe benefit of adjuvant immunotherapy after nephrectomy in renal cell carcinoma (RCC) is controversial. The present study aimed to examine the possible benefit of adjuvant immunotherapy in various clinical settings.MethodsWe retrospectively reviewed 436 patients with pT1-3N0-2M0 RCC who underwent radical or partial nephrectomy with curative intent at our institution between 1981 and 2009. Of them, 98 (22.5%) patients received adjuvant interferon-α (IFN-α) after surgery (adjuvant IFN-α group), while 338 (77.5%) did not (control group). The primary endpoint was cancer-specific survival (CSS). Univariate and multivariate analyses were conducted using log-rank tests and Cox proportional hazards models, respectively.ResultsFifty-two (11.9%) patients died from RCC with a median follow-up period of 96 months. Preliminary univariate analyses comparing CSS among treatment groups in each TNM setting revealed that CSS in the control group was equal or superior to that in the adjuvant IFN-α group in earlier stages, while the opposite trend was observed in more advanced stages. We evaluated the TNM cutoffs and demonstrated maximized benefit of adjuvant IFN-α in patients with pT2b-3cN0 (P = 0.0240). In multivariate analysis, ≥pT3 and pN1-2 were independent predictors for poor CSS in all patients. In the subgroups with ≥pT2 disease (n = 123), pN1-2 and no adjuvant treatment were significant poor prognostic factors.ConclusionsAdjuvant immunotherapy after nephrectomy may be beneficial in pT2b-3cN0 RCC. Careful consideration is, however, required for interpretation of this observational study because of its selection bias and adverse effects of IFN-α.

Interferon-induced pulmonary hypertension: an update.

Pulmonary adverse effects of interferon (IFN) therapies are rare but can be life threatening. This article proposes to review clinical and experimental data suggesting a causal link between interferon exposure and pulmonary arterial hypertension (PAH). Interferon has recently been added to the list of possible risk factors for PAH. This was justified by the reporting of many cases of pulmonary hypertension potentially associated with IFN-α or IFN-β exposure. Some of them were reversible after cessation of interferon exposure, especially in patients without concomitant risk factors for pulmonary hypertension. In contrast, it remains a challenge to definitively confirm the causal role of IFN-α in patients treated for hepatitis C viral infection because of frequent concomitant PAH risk factors such as portal hypertension and/or HIV infection. In these patients, temporal and clinical arguments suggest that interferon may potentially act as an additional trigger for PAH. Moreover, the information obtained from clinical experience with interferon therapy has been enriched by basic science research on this topic suggesting that interferon is involved in both human and experimental pulmonary hypertension. Many clinical and experimental data corroborate the link between interferon exposure and the risk to develop PAH.

Pathogenesis of antimicrobial peptides LL-37 and CpG-ODN in ANCA associated vasculitis.

Anti-neutrophil cytoplasmic antibodies (ANCA)-associated vasculitides (AAV) are a group of systemic autoimmune disorders characterized by necrotizing inflammation of small- to medium-sized blood vessels. The pathogenesis of patients with AAV are still in investigation. In this study, we explored the involvement of LL-37 and nucleic acids in AAV. 15 patients with AAV diagnosed according to the Chapel Hill definition between October 2014 and July 2015 in the department of Nephrology of Huangdao, affiliated Hospital of Qingdao University were enrolled. 16 patients with chronic bronchitis (CB) were selected as disease control group. 15 cases of healthy people from Medical Healthy Center were as healthy control group. Peripheral blood mononuclear cells (PBMCs) were collected from these groups and stimulated by LL-37and (or) two types ofCpG-ODN for 7days. The IFN-α and ANCA in vitro were measured by ELISA. The serum IFN-α, LL-37 and ANCA were measured also. The serum level of IFN-α in AAV group was much higher than that in CB group (692.13±407.28 vs 397.07±211.62pg/ml, p=0.019), and that in healthy control group (692.13±407.28 vs 251.54±190.46pg/ml, p<0.001). The serum level of LL-37 in AAV group was much higher than that in CB group (101.18±66.59 vs 40.23±13.51ng/ml, p<0.001, and that in healthy control group (101.18±66.59 vs 27.80±16.86ng/ml, p<0.001). Also the level of IFN-α showed a significant positive relationship with ANCA in AAV group both in serum and in supernatant of cultured PBMCs stimulated by LL-37 and (or) CpG-ODN (r=0.783, p=0.001; r=0.575, p=0.064; r=0.649, p=0.031; r=0.806, p=0.003). In patients with AAV, the supernatant levels of IFN-α in cultured PBMCs stimulated by LL-37 and (or) CpG-ODN were higher than that without stimulating factor (p<0.05). The supernatant level of IFN-α in cultured PBMC stimulated by LL-37 alone was lower than that stimulated by CpGA alone (699.57±476.26 vs 2342.63±2025.11pg/ml, p=0.001). But the supernatant level of IFN-α in cultured PBMCs stimulated by LL-37 alone was higher than in that stimulated by CpGB alone (699.57±476.26 vs 153.35±78.08pg/ml, p<0.001). The supernatant level of IFN-α in cultured PBMCs stimulated by both LL-37 and CpG-ODN was higher than that stimulated by LL-37 or CpG-ODN alone (2550.57±2217.41 vs 699.57±476.26pg/ml, p=0.003; 2550.57±2217.41 vs 153.35±78.08pg/ml, p=0.001; 2660.95±391.31 vs 699.57±476.26pg/ml, p<0.001; 2660.95±391.31 vs 153.35±78.08pg/ml, p<0.001). Either it is stimulated by LL-37 or CpG-ODN or both, the supernatant level of IFN-α in cultured PBMCs in AAV patients wasthe highest, that in healthy controls was the lowest. Either stimulated by LL-37 or CPG-ODN, or both, the levels of ANCA production in vitro in AAV groups were statistically significantly higher than those in CB group and healthy control group. There were higherserum levels of LL-37and IFN-α in patients with AAV. IFN-α could reach a higher level stimulated by LL-37 and nucleic acidsboth of which are related to infection. Patients with AAV have ANCA-producing B lymphocytes in the circulation even in remission stage. Infections could induce the relapse of AAV.

Altered molecular expression of TLR-signaling pathways affects the steady-state release of IL-12p70 and IFN-α in patients with relapsing-remitting multiple sclerosis.

Recent evidence suggests a key role of dendritic cells (DC) in the immunopathogenesis of multiple sclerosis (MS). Whereas dysfunction of DC was reported in MS patients, the underlying cause for this is not fully elucidated yet. The aim of the present study was to compare the gene expression profile of molecules involved in TLR4 and TLR7 signaling in DC from patients with MS and healthy controls. For this, circulating DC subsets were purified from patients with relapsing-remitting MS (RRMS) and from healthy controls for quantitative real-time PCR analysis. Additionally, TLR responsiveness in peripheral blood was investigated. We observed an aberrant steady-state release of IL-12p70 and IFN-α in patients with RRMS compared with healthy controls. Expression of IRF1 and JUN was reduced in conventional DC from patients with RRMS. In plasmacytoid DC from patients with RRMS, expression of IRF7 and IFNGR1 was reduced, while higher expression levels of TLR4 and LY86 were found compared with DC from healthy controls. The observed alterations in the gene expression of molecules involved in the TLR4 and TLR7 signaling pathways in circulating DC subsets may underlie the impaired IL-12p70 and IFN-α secretion in patients with RRMS, thereby potentially contributing to the disease pathogenesis of MS.

All-Trans Retinoic Acid and Interferon-Alpha Increase CD38 Expression on Adult T-Cell Leukemia Cells and Sensitize Them to T Cells Bearing Anti-CD38 Chimeric Antigen Receptors

The survival of patients with adult T-cell leukemia (ATL) has been improved by the introduction of anti-CCR4 monoclonal antibody and the expanding use of allogeneic hematopoietic stem cell transplantation. However, not all patients benefit from these treatment modalities, warranting development of a novel therapeutic strategy. CD38, a cell surface ectoenzyme that functions as cyclic ADP ribose hydrolase, is an attractive target of chimeric antigen receptor (CAR) therapy for lymphoid neoplasms because it is widely expressed on the cells of B- or T-lymphoid malignancies. We have previously demonstrated the prominent cytotoxicity of T cells engineered to express an anti-CD38-CAR against B-lymphoma cells and myeloma cells expressing CD38. To expand the applicability of anti-CD38-CAR against ATL cells that usually express undetectable or low levels of CD38, notably, we were successfully able to induce cell surface CD38 expression in HTLV-1-infected cell lines with all-trans retinoic acid (ATRA) (Yoshida T, et al. 2013 ASH Meeting). In ATL cells freshly isolated obtained from the patients, we were able to induce CD38 with ATRA in 60-80% of the cells; the remaining cells survived under the anti-CD38-CAR treatment. We hereby report our attempts in improving the efficacy of anti-CD38-CAR T cells against ATL cells from the patients through the expression of CD38 enhanced with the entry of agents, which are clinically used. Firstly, we investigated whether ATL cells from patients could be transduced with anti-CD38-CAR and what is the efficiency of transduction into T cells in our settings. ATL cells (CD4+ CD25+ GFP+) transduced with retroviral vector were little detected. CD4- CD25- GFP+ T cells alone were detected in our transduction methods. Transduction efficiency was over 40%. To increase the expression of CD38 on ATL cells, we took notice of the CD38 gene upstream region that contains binding sites for interferon regulatory factor-1 (IRF-1) and peroxisome proliferator-activated receptor (PPAR). We thus investigated whether IFN-α, IFN-γ or troglitazone, which is a PPAR-α and -γ agonist, could enhance CD38 expression in ATL cell lines (MT-4, Su9T, ED, and S1T cells), which are negative for CD38. IFN-α and IFN-γ efficiently enhanced CD38 expression in MT-4 cells in a dose-dependent manner but not in Su9T, ED, and S1T cells. As little as 2.5U/ml of IFN-α induced CD38 expression in MT-4 cells for 18 hours in vitr o (>95% at positivity of CD38). 10-25% increase in CD38 expression was observed in ED cells with 125-250 pM troglitazone after 18 hours of treatment, but not in MT-4, Su9T, and S1T cells. Prolonged exposure to troglitazone was toxic to cells. Combined treatment with 10nM ATRA and IFN-α, which induced higher expression of CD38 than IFN-γ, synergistically enhanced CD38 expression of ATL cells from the patients (>90%at positivity of CD38). We next co-cultured ATL cells form three patients with T cells transduced with mock or anti-CD38-CAR in the presence of both ATRA and IFN-α at effector (E): target (T) ratio of 1: 2 for 3 days. The treatment eradicated more than 95% of these ATL cells, demonstrating that ATL cells can be eliminated by T cells harboring anti-CD38-CAR in the presence of ATRA and IFN-α, which is actively used for ATL patients. CD38 targeting therapy is a feasible method, because an anti-CD38 antibody, daratuzumab, has been used to treat plasma cell myeloma. The safety regarding the clinical use of T cells bearing anti-CD38-CAR still needs to be established. As CAR therapy reportedly causes cytokine storm and can potentially be lethal, we envision an inducible immunotherapy with CAR to be a preferred modality with increased efficacy and safety. Our results provide a rationale for a novel therapeutic strategy involving T cells carrying anti-CD38-CAR in combination with ATRA and IFN-α for patients with ATL. DisclosuresNo relevant conflicts of interest to declare.

A Novel Score to Predict Interferon-Alpha Therapy Responsiveness in Patients with Essential Thrombocythemia

Introduction.Interferon alpha (IFN-α) is an attractive agent for the treatment of Essential Thrombocythemia (ET) due to its ability to induce clonal complete remission, sometimes lasting beyond treatment discontinuation, and to its recognized non-leukemogenicity.However, despite decades of clinical experience with IFN-α in patients with MPNs, optimal dose schedules, treatment duration and the ultimate molecular basis of the heterogeneous response still remain undefined.Hence, the early identification of IFN-sensitive patients may help limit IFN-α exposure to those who really benefit from treatment.Aim.Here we report the results of a trial involving 61 ET patients treated with IFN-α, aimed to identify the baseline molecular and clinical parameters able to predict response to treatment.Methods.IFN treatment schedule implied an initial induction phase with 3MU/five times a week; in patients who reached a platelet count <400x109/L, IFN dose density was progressively reduced and patients considered Good-responders (Good-R). Patients who failed to achieve platelet normalization after induction, underwent an additional three-month period of IFN treatment at 3MU/five times a week. In case platelet count was still >600x109/L or platelet reduction was <50% the baseline level, the patient was considered resistant to the IFN therapy (Bad-R).Careful medical history, main laboratory data and spleen volume, assessed by ultrasonography scan were recorded in all patients at presentation and during follow-up. Complete hematological response (CHR) is defined as the normalization of both platelet and WBC counts (<400x109/L and <10x109/L, respectively) and the absence of disease-related symptoms.mRNA levels of JAK1, JAK2, STAT1, STAT3, SOCS1, SOCS3 and TYK2 were assayed in pre-treatment bone marrow specimens by Real-Time PCR using the SYBR Green method.Results.After a median follow-up of 41.2 months, 72% of patients achieved CHR and were considered Good-Rs for subsequent analysis, whereas the remaining 17 were considered Bad-Rs. Among the Good-Rs, 24 (54%) are still on therapy with standard IFN-α doses (i.e. 3 MU 3 or 2 times a week), whereas 10 (23%) are maintained in CHR by the administration of very low doses of IFN-α-2b (3 MU every 7 or 15 days), and 3 (7%) have maintained CHR after therapy discontinuation (up to a median time of 31 months).The initial univariate analysis indicated that the mRNA levels of JAK1, STAT3, SOCS3 were significantly lower in Good-R than in Bad-R patients. Interestingly, among the different genes involved in the IFN-α receptor pathway, the expression levels of JAK1, together with spleen volume and platelet count, were selected by the stepwise multivariate analysis as the variables that independently correlate with IFN-α response. We used the relative HRs and the optimal cut-offs for response calculated for each variable by the ROC analysis to develop a prognostic score able to predict IFN-α response. This score has an overall 87% diagnostic efficiency in discriminating IFN-α response and unambiguously identifies the response to IFN-α in most patients, avoiding treatment in those with no probability of gaining benefit from this therapy.In addition, this score is able to identify unambiguously the response to IFN in a sizeable proportion of patients: an IFN-R score of 3 or 4 (31 patients, corresponding to 70.4% of the Good-R) indicates a 100% odd to obtain CHR, while a score of 0 indicates no chance of achieving a response.Conclusion.In conclusion, this study shows for the first time that the use of three simples parameters predicts the response to IFN in ET patients. DisclosuresNo relevant conflicts of interest to declare.

10. Gene expression in interferon-α (IFN-α) induced persistent fatigue

Interferon-α (IFN-α) used to treat Chronic Hepatitis C Viral (HCV) infection induces fatigue, which can persist up to six months post-treatment. This study examined some putative biological mechanisms underlying the development of such side effect. We recruited 25 patients receiving IFN-α. Blood was collected using PAXgene tubes at baseline and treatment weeks 4, 12 and 24. Fatigue was assessed using the Chalder Fatigue Questionnaire (CFQ) administered at the same time points and six months post-treatment. Patients were stratified according to whether their fatigue levels at follow-up had improved or returned to baseline levels (Resolved Fatigue; RF, n = 15) or worsened (Persistent Fatigue; PF, n = 10). Assessing the effect of IFN-α, we found that at week 4, 663 genes were modulated in both groups, with a further 229 in the RF group, and 556 uniquely modulated in the PF group. Pathway analyses in the PF group reported alterations in IL-15 and Tec Kinase signaling, among others. Molecules of interest within those pathways included BCL2 (fold-change 1.50, p < .005), STAT1 (1.57, p < .0002) and FCER1A (−2.0, p < .0002), previously shown to be relevant to fatigue. Our data shows a differential modulation of inflammatory pathways by IFN-α in patients that develop persistent fatigue, suggesting a possible underlying role in the development of related symptoms.

THU0520 Interferon (IFN)-Lambda1/3, but not IFN-Alpha, is A Marker of Poor Response to Treatment in Lupus Nephritis

BackgroundLupus nephritis (LN) is a severe manifestation of systemic lupus erythematosus (SLE) affecting up to 60% of the patients. IFN-α is a central cytokine in the pathogenesis for LN, and...

Altered mRNA levels of MOV10, A3G, and IFN-α in patients with chronic hepatitis B.

To explore the relationship of the MOV10, A3G, and IFN-α mRNA levels with chronic hepatitis B virus (HBV) infection, Blood samples from 96 patients with chronic hepatitis B (CHB) and 21 healthy individuals as control were collected. HBV DNA load and aminotransferase in the serum were tested using real time PCR and velocity methods, respectively. The MOV10, A3G, and IFN-α mRNA levels in the peripheral blood mononuclear cells (PBMC) were examined through qRT-PCR. The MOV10, A3G, and IFN-α mRNA levels in CHB group was significantly lower than those in the control group (P<0.01, P<0.05, P<0.01, respectively). The A3G mRNA level in the high-HBV DNA load group was lower than that in the low-HBV DNA load group (P<0.05). However, no statistical difference was found in the MOV10 and IFN-α mRNA levels between the two HBV DNA load groups. Furthermore, the MOV10 mRNA level showed positive correlation with IFN-α in the control group. These results indicated that the expression of the innate immune factors MOV10, A3G, and IFN-α is affected by chronic HBV infection.