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Acquired hemophilia A (AHA) is an autoimmune bleeding disorder caused by neutralizing antibodies against coagulation factor VIII. Immunosuppressive therapy (IST) is standard of care to eradicate autoantibody production and protect from further bleeding but carries a risk of severe infection and mortality in frail patients with AHA. Recently, emicizumab has been studied for its potential to reduce the need for early and aggressive IST.

The root cause of autoantibody formation against factor VIII (FVIII) in acquired hemophilia A (AHA) remains unclear. We aimed to assess whether AHA is exclusively associated with autoantibodies toward FVIII or whether patients also produce increased levels of autoantibodies against other targets. A case-control study was performed enrolling patients with AHA and age-matched controls. Human epithelial cell (HEp-2) immunofluorescence was applied to screen for antinuclear (ANA) and anticytoplasmic autoantibodies. Screening for autoantibodies against extractable nuclear antigens was performed by enzyme immunoassay detecting SS-A/Ro, SS-B/La, U1RNP, Scl-70, Jo-1, centromere B, Sm, double-stranded DNA, and α-fodrin (AF). Patients with AHA were more often positive for ANA than control patients (64% vs 30%; odds ratio [OR] 4.02, 1.98-8.18) and had higher ANA titers detected than controls. Cytoplasmic autoantibodies and anti-AF immunoglobulin A autoantibodies were also more frequent in patients with AHA compared with controls. Autoantibodies against any target other than FVIII were found in 78% of patients with AHA compared with 46% of controls (OR 4.16, 1.98-8.39). Results were similar preforming sensitivity analyses (excluding either subjects with autoimmune disorders, cancer, pregnancy, or immunosuppressive medication at baseline) and in multivariable binary logistic regression. To exclude that autoantibody staining was merely a result of cross-reactivity of anti-FVIII autoantibodies, we tested a mix of 7 well-characterized monoclonal anti-FVIII antibodies. These antibodies did not stain HEp-2 cells used for ANA detection. In conclusion, a diverse pattern of autoantibodies is associated with AHA, suggesting that a more general breakdown of immune tolerance might be involved in its pathology.

•Seventy-eight percent of patients with AHA were found to have autoantibodies against targets other than factor VIII.•The data suggest that a generalized breakdown of self-tolerance mechanisms is involved in the pathology of AHA.

Despite current therapies, there remains an unmet need for treatment for patients with hemophilia. The main parts of two phase 2 trials established clinical proof-of-concept for once-daily, subcutaneous concizumab prophylaxis in patients with hemophilia A/B with inhibitors (HAwI/HBwI; explorer4) and severe hemophilia A without inhibitors (HA; explorer5). Here, we present results from extension parts of these trials, included to evaluate longer term safety and efficacy. Both trials included main (≥24 weeks) and extension (52-102 weeks) parts, with patients receiving concizumab 0.15 mg/kg with potential dose escalation to concizumab 0.20 or 0.25 mg/kg if they experienced ≥3 treated spontaneous bleeding episodes within 12 weeks. Endpoints included annualized bleeding rate (ABR), adverse events (AEs), and occurrence of antidrug antibodies. Thromboembolic events were AEs of special interest. Thirty-six patients with HA, 15 with HAwI, and 10 with HBwI were exposed to concizumab. Estimated ABRs during the main + extension parts at last dose level were 4.8 (95% confidence interval [CI], 3.2-7.2) and 6.4 (95% CI, 4.1-9.9) in explorer4 and explorer5, respectively (spontaneous ABRs were 1.8 [95% CI, 1.2-2.6] and 2.1 [95% CI, 1.3-3.3]). Most AEs were mild, with no deaths, events leading to withdrawal, or thromboembolic events. Anti-drug antibodies developed in 25% of patients and were low titer and transient, with no observed clinical effect in most cases. Results of the main + extension parts of these trials were consistent with results of the main parts. Ongoing phase 3 trials will further evaluate concizumab as a once-daily, subcutaneous treatment across hemophilia subtypes. These trials were registered at www.clinicaltrials.gov as #NCT03196284 and #NCT03196297.

Acquired hemophilia A (AHA) is due to autoantibodies against coagulation factor VIII (FVIII) and most often presents with unexpected bleeding. In contrast to congenital hemophilia, the patient's residual FVIII activity does not seem to correlate with the risk of bleeding as suggested from previous studies. Risk factors for bleeding have not been described. We used data from the prospective GTH-AH 01/2010 study to assess the risk of bleeding and the efficacy of hemostatic therapy. FVIII activity was measured at baseline and weekly thereafter. Bleeding events were assessed by treating physicians. A total of 289 bleeds were recorded in 102 patients. There were 141 new bleeds observed starting after day 1 in 59% of the patients, with a mean rate of 0.13 bleed per patient-week in weeks 1 to 12, or 0.27 bleed per patient-week before achieving partial remission. Weekly measured FVIII activity was significantly associated with the bleeding rate, but only achieving FVIII activity ≥50% abolished the risk of bleeding. A good World Health Organization performance status assessed at baseline (score 0 vs higher) was associated with a lower bleeding rate. Hemostatic treatment was reportedly effective in 96% of bleeds. Thus, the risk of new bleeds after a first diagnosis of AHA remains high until partial remission is achieved, and weekly measured FVIII activity may aid in assessing the individual risk of bleeding. These results will help to define future strategies for prophylaxis of bleeding in AHA.

Acquired hemophilia A (AHA) is due to autoantibodies against coagulation factor VIII (FVIII) and most often presents with unexpected bleeding. In contrast to congenital hemophilia, the patient’s residual FVIII activity does not seem to correlate with the risk of bleeding as suggested from previous studies. Risk factors for bleeding have not been described. We used data from the prospective GTH-AH 01/2010 study to assess the risk of bleeding and the efficacy of hemostatic therapy. FVIII activity was measured at baseline and weekly thereafter. Bleeding events were assessed by treating physicians. A total of 289 bleeds were recorded in 102 patients. There were 141 new bleeds observed starting after day 1 in 59% of the patients, with a mean rate of 0.13 bleed per patient-week in weeks 1 to 12, or 0.27 bleed per patient-week before achieving partial remission. Weekly measured FVIII activity was significantly associated with the bleeding rate, but only achieving FVIII activity ≥50% abolished the risk of bleeding. A good World Health Organization performance status assessed at baseline (score 0 vs higher) was associated with a lower bleeding rate. Hemostatic treatment was reportedly effective in 96% of bleeds. Thus, the risk of new bleeds after a first diagnosis of AHA remains high until partial remission is achieved, and weekly measured FVIII activity may aid in assessing the individual risk of bleeding. These results will help to define future strategies for prophylaxis of bleeding in AHA.

Recombinant porcine factor VIII (rpFVIII, OBI‐1, susoctocog alfa) is used for the treatment of acute bleeds in patients with acquired hemophilia A (AHA). Inhibitors in AHA can sometimes cross‐react with rpFVIII.

Concizumab is an anti-tissue factor pathway inhibitor monoclonal antibody in development as a once-daily, subcutaneous prophylaxis for patients with haemophilia A or haemophilia B with or without inhibitors. We aimed to assess the efficacy and safety of concizumab in patients with haemophilia A or B without inhibitors. Here we report the results from the confirmatory analysis cutoff.

Results from the main parts (24 weeks) of 2 concizumab phase 2 trials are presented: explorer4 in hemophilia A (HA) or B (HB) with inhibitors (HAwI/HBwI) and explorer5 in HA. The trials aimed to evaluate the efficacy of daily subcutaneous concizumab prophylaxis (evaluated as annualized bleeding rate [ABR] at last dose level), with secondary objectives being safety and immunogenicity (assessed as number of adverse events [AEs] and antidrug antibodies [ADAs]). Patients received 0.15 mg/kg concizumab, with potential dose escalation to 0.20 and 0.25 mg/kg (if =3 spontaneous bleeding episodes within 12 weeks of concizumab treatment). Relevant pharmacokinetic/pharmacodynamic (PK/PD) parameters were assessed. Thirty-six HA, 9 HAwI, and 8 HBwI patients were exposed to concizumab. Most inhibitor patients (15 of 17; 88.2%) did not escalate the dose; all patients chose to continue to the extension phase of the trials. Clinical proof of concept for prevention of bleeding episodes was demonstrated in both trials. Estimated ABRs in HAwI and HBwI were lower vs HA: 3.0 (95% confidence interval [CI], 1.7; 5.3) and 5.9 (95% CI, 4.2; 8.5) vs 7.0 (95% CI, 4.6; 10.7), respectively. PK/PD results were as expected, with no difference between hemophilia subtypes for concizumab exposure, free tissue factor pathway inhibitor, thrombin generation, prothrombin fragment 1+2, and d-dimers. Concizumab was safe and well tolerated (no severe AEs, AE-related withdrawals, or thromboembolic events). Three patients had (very low to medium titer) ADA+ tests in each trial, with no observed clinical effect. These results support further development of concizumab as a daily prophylactic treatment in all hemophilia patients. These trials were registered at www.clinicaltrials.gov as #NCT03196284 and #NCT03196297.

Results from the main parts (24 weeks) of 2 concizumab phase 2 trials are presented: explorer4 in hemophilia A (HA) or B (HB) with inhibitors (HAwI/HBwI) and explorer5 in HA. The trials aimed to evaluate the efficacy of daily subcutaneous concizumab prophylaxis (evaluated as annualized bleeding rate [ABR] at last dose level), with secondary objectives being safety and immunogenicity (assessed as number of adverse events [AEs] and antidrug antibodies [ADAs]). Patients received 0.15 mg/kg concizumab, with potential dose escalation to 0.20 and 0.25 mg/kg (if ≥3 spontaneous bleeding episodes within 12 weeks of concizumab treatment). Relevant pharmacokinetic/pharmacodynamic (PK/PD) parameters were assessed. Thirty-six HA, 9 HAwI, and 8 HBwI patients were exposed to concizumab. Most inhibitor patients (15 of 17; 88.2%) did not escalate the dose; all patients chose to continue to the extension phase of the trials. Clinical proof of concept for prevention of bleeding episodes was demonstrated in both trials. Estimated ABRs in HAwI and HBwI were lower vs HA: 3.0 (95% confidence interval [CI], 1.7; 5.3) and 5.9 (95% CI, 4.2; 8.5) vs 7.0 (95% CI, 4.6; 10.7), respectively. PK/PD results were as expected, with no difference between hemophilia subtypes for concizumab exposure, free tissue factor pathway inhibitor, thrombin generation, prothrombin fragment 1+2, and d-dimers. Concizumab was safe and well tolerated (no severe AEs, AE-related withdrawals, or thromboembolic events). Three patients had (very low to medium titer) ADA+tests in each trial, with no observed clinical effect. These results support further development of concizumab as a daily prophylactic treatment in all hemophilia patients. These trials were registered at www.clinicaltrials.govas #NCT03196284 and #NCT03196297.

Introduction:AHA is a serious autoimmune bleeding disorder caused by neutralizing antibodies against coagulation factor VIII (FVIII). Standard of care involves immunosuppressive therapy (IST) to suppress formation of anti-FVIII autoantibodies. Recently, prophylaxis with emicizumab (EMI) has been used instead of IST to manage patients with AHA.

Introduction

ABSTRACTRecent findings strongly support a role for small regulatory RNAs in the regulation of human lifespan yet little information exists about the precise underlying mechanisms. Although extensive studies on model organisms have indicated that reduced activity of the nutrient response pathway, for example as a result of dietary restriction, can extend lifespan through the suppression of the protein kinase mechanistic target of rapamycin (mTOR), it still is subject of debate whether this mechanism is operative in humans as well. Here, we present findings indicating that human microRNA (miR)-496 targets 2 sites within the human mTOR3′UTR. Coexpression of miR-496 with different fusion transcripts, consisting of the luciferase transcript and either wild-type mTOR3′UTR or mTOR3′UTR transcript with the miR-496 binding sites singly or combined mutated, confirmed this prediction and revealed cooperativity between the 2 binding sites. miR-496 reduced the mTOR protein level in HeLa-K cells, and the levels of miR-496 and mTOR protein were inversely correlated in Peripheral Blood Mononuclear Cells (PBMC), with old individuals (n=40) harbouring high levels of miR-496 relative to young individuals (n=40). Together, these findings point to the possibility that miR-496 is involved in the regulation of human aging through the control of mTOR.

Despite current treatments, there remains an unmet need for patients with hemophilia. The main parts of two phase 2 trials established clinical proof-of-concept for once-daily, subcutaneous concizumab prophylaxis in patients with hemophilia A/B with inhibitors (HAwI/HBwI; explorer4) and severe hemophilia A without inhibitors (HA; explorer5,). Here, we present results from extension parts of these trials, included to evaluate longer-term safety and efficacy. Both trials included main (≥24 weeks) and extension parts (52–102 weeks), with patients receiving 0.15 mg/kg concizumab with potential dose escalation to 0.20 or 0.25 mg/kg if they experienced ≥3 treated spontaneous bleeds within 12 weeks. Endpoints included annualized bleeding rate (ABR), adverse events (AEs), and anti-drug antibody (ADA) occurrence. Thromboembolic events were AEs of special interest. Thirty-six patients with HA, 15 with HAwI and 10 with HBwI were exposed to concizumab. Estimated ABRs during the main+extension parts at last dose level were 4.8 (95% confidence interval [CI]: 3.2–7.2) and 6.4 (95% CI: 4.1–9.9) in explorer4 and explorer5, respectively (spontaneous ABRs were 1.8 [95% CI: 1.2–2.6] and 2.1 [95% CI: 1.3–3.3]). Most AEs were mild, with no deaths, events leading to withdrawal or thromboembolic events. ADAs developed in 25% of patients and were low-titer and transient with no observed clinical effect in most cases. Results of the main+extension parts of these trials were consistent with the main parts. Ongoing phase 3 trials will further evaluate concizumab as a once-daily, subcutaneous treatment across hemophilia subtypes. Trials registered at www.clinicaltrials.gov(NCT03196284; NCT03196297).

BACKGROUND: This prospective study was conducted to gain experience as to whether it is technically possible to produce autologous RBCs in additive solution from cord blood (CB), to optimize the blood supply for preterm infants. STUDY DESIGN AND METHODS: CB was collected from 47 infants with a mean (± SD) birth weight of 1717 (± 699) g. Whenever possible, RBC components were prepared by standard centrifugation using a six-bag system. All samples were put in sterility testing quarantine for 5 days, and a maximum storage of 14 days from collection to transfusion was specified. The babies were given either the autologous RBCs or standard allogeneic RBC concentrates, if autologous blood was not available. RESULTS: In 81 percent of the samples, autologous RBC components could be processed (vol, 7-87 mL; Hct, 31-82%). But within the group of extremely low birth weight infants (body weight <1000 g), a mean CB net volume of only 37 mL was collected, and the RBC preparation was successful only in exceptional cases. Three CB samples (8.6%) tested positive in sterility testing. Of the 47 infants, 21 were treated with a total of 62 allogeneic and 4 autologous RBC transfusions. Most infants with a body weight over 1400 g did not need any RBC transfusion. CONCLUSION: The preparation of autologous RBCs from the CB of preterm infants is technically possible in principle. However, major concerns must be raised as to whether such preparations are of benefit in ensuring safe care of neonates with blood components, with respect to the high rate of bacterial contamination and the limited availability in babies with low birth weight.

Introduction:Concizumab is an anti-tissue factor pathway inhibitor (TFPI) monoclonal antibody in phase 3 clinical development as a once-daily, subcutaneous prophylaxis across all hemophilia subtypes. The phase 2 trials explorer4 (NCT03196284) and explorer5 (NCT03196297) investigated the efficacy and safety of daily subcutaneous concizumab prophylaxis in patients with hemophilia A/B with inhibitors and in patients with severe hemophilia A without inhibitors, respectively. Minor surgeries and diagnostic procedures were permitted during both trials, allowing for the assessment of conducting such procedures during concizumab prophylaxis.

Wheeler: Novo Nordisk A/S: Consultancy; Bayer: Consultancy; BioMarin: Consultancy; HEMA Biologics: Consultancy; Spark: Consultancy; Takeda: Consultancy; UniQure: Consultancy. Benson: Novo Nordisk A/S: Consultancy, Speakers Bureau; Takeda: Speakers Bureau; BMS: Speakers Bureau; Sobi: Speakers Bureau. Eichler: Pfizer: Research Funding; Novo Nordisk: Consultancy, Research Funding; BioMarin: Consultancy, Research Funding; Bayer: Consultancy, Research Funding; Takeda: Consultancy, Honoraria; Roche: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; CSL Behring: Consultancy, Honoraria; Biotest: Consultancy, Honoraria. Marie Tønder: Novo Nordisk Health Care AG: Current Employment. Cepo: Novo Nordisk A/S: Current Employment. Jimenez Yuste: Novo Nordisk A/S: Consultancy, Research Funding; Bayer: Consultancy; Takeda: Consultancy, Research Funding; Roche: Consultancy, Research Funding; Grifols: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Sobi: Consultancy, Research Funding; CSL Behring: Consultancy; BioMarin: Consultancy. Kavakli: Novo Nordisk A/S: Consultancy, Other: Clinical Trial Support; Takeda: Consultancy, Other: Clinical Trial Support; Roche: Consultancy, Other: Clinical Trial Support. Wong: Astellas Pharma, INc.: Research Funding. Matsushita: Baxalta/Shire/Takeda: Consultancy, Honoraria; Bayer: Consultancy; Novo Nordisk A/S: Consultancy, Honoraria, Other: educational and investigational support ; Chugai: Consultancy, Honoraria, Other: educational and investigational support ; Pfizer: Consultancy; Bioverative/Sanofi: Honoraria; CSL: Honoraria; JB: Honoraria; KMB: Honoraria; Kirin: Honoraria; Nichiyaku: Honoraria; Octapharm: Honoraria; Sysmex: Honoraria.

Pipe: Genventiv: Consultancy; Regeneron/ Intellia: Consultancy; uniQure: Consultancy, Other; Spark Therapeutics: Consultancy; Takeda: Consultancy; Sanofi: Consultancy, Other; Sangamo Therapeutics: Consultancy; Roche/Genentech: Consultancy, Other; Pfizer: Consultancy; Novo Nordisk: Consultancy; Freeline: Consultancy, Other: Clinical trial investigator; HEMA Biologics: Consultancy; CSL Behring: Consultancy; Catalyst Biosciences: Consultancy; Biomarin: Consultancy, Other: Clinical trial investigator; Bayer: Consultancy; ASC Therapeutics: Consultancy; Apcintex: Consultancy; Grifols: Consultancy; Octapharma: Consultancy; Shire: Consultancy. Sheehan: BioMarin: Consultancy, Research Funding; Roche: Consultancy, Research Funding; Bayer: Consultancy, Research Funding. Coppens: Portola/Alexion: Research Funding; CSL Behring: Consultancy, Research Funding; Bayer: Consultancy, Research Funding; Roche: Research Funding; Daiichi Sankyo: Research Funding; Sanquin Blood Supply: Research Funding; uniQure: Research Funding; Medcon International: Consultancy; MEDtalks: Consultancy; Novo Nordisk: Consultancy; Pfizer: Consultancy; Sobi: Consultancy. Eichler: Takeda: Consultancy, Honoraria; BioMarin: Consultancy, Research Funding; Roche: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; CSL Behring: Consultancy, Honoraria; Pfizer: Research Funding; Novo Nordisk: Consultancy, Research Funding; Biotest: Consultancy, Honoraria; Bayer: Consultancy, Research Funding. Linardi: Bayer: Current Employment. Wiegmann: Bayer: Current Employment. Hay: Pfizer: Consultancy, Research Funding; Inspiration: Consultancy, Honoraria; BioMarin: Consultancy, Honoraria; Roche: Consultancy, Research Funding; Bayer: Consultancy, Research Funding; Takeda: Consultancy, Research Funding; Spark Therapeutics: Consultancy, Research Funding. Lissitchkov: Bayer: Other: Principal Investigator of Clinical Trials; Catalist: Other: Principal Investigator of Clinical Trials; Grifols: Other: Principal Investigator of Clinical Trials.

Zusammenfassung Die Azofarbstoffe des Phenolphthaleins vereinigen in sich eine grosse Empfindlichkeit mit einem leicht wahrnehmbaren Farbenumschlag und sind je nach der Zusammensetzung auch zur Bestimmung schwacher Basen in vollkommenerem Maße verwendbar. Man kann also mit II oder III Säuren und Basen von verschiedenem Leitungsvermögen möglichst fehlerfrei bestimmen.