Reduced Volume and Faster Infusion Rate of Activated Prothrombin Complex Concentrate: A Phase 3b/4 Trial in Adults with Hemophilia A with Inhibitors.

Background Activated prothrombin complex concentrate (aPCC) is indicated for bleed treatment and prevention in patients with hemophilia with inhibitors. The safety and tolerability of intravenous aPCC at a reduced volume and faster infusion rates were evaluated. Methods This multicenter, open-label trial (NCT02764489) enrolled adults with hemophilia A with inhibitors. In part 1, patients were randomized to receive three infusions of aPCC (85 ± 15 U/kg) at 2 U/kg/min (the approved standard rate at the time of the study), in a regular or 50% reduced volume, and were then crossed over to receive three infusions in the alternative volume. In part 2, patients received three sequential infusions of aPCC in a 50% reduced volume at 4 U/kg/min and then at 10 U/kg/min. Primary outcome measures included the incidence of adverse events (AEs), allergic-type hypersensitivity reactions (AHRs), infusion-site reactions (ISRs), and thromboembolic events. Results Of the 45 patients enrolled, 33 received aPCC in part 1 and 30 in part 2. In part 1, 24.2 and 23.3% of patients with regular and reduced volumes experienced AEs, respectively; 11 AEs in eight patients were treatment related. AHRs and ISRs occurred in four (12.1%) and two (6.1%) patients, respectively. In part 2, 3.3 and 14.3% of patients with infusion rates of 4 and 10 U/kg/min experienced AEs, respectively; only one AE in one patient was treatment related; no AHRs or ISRs were reported. Most AEs were mild/moderate in severity. Overall, no thromboembolic events were reported. Conclusions aPCC was well tolerated at a reduced volume and faster infusion rates, with safety profiles comparable to the approved regimen.

Correction: Expert Opinions on the Management of Hemophilia A in India: The Role of Emicizumab.

[This corrects the article DOI: 10.7759/cureus.58941.].

Expert Opinions on the Management of Hemophilia A in India: The Role of Emicizumab.

Hemophilia A (HA) is a genetic disorder of hemostasis associated with a deficiency or reduced activity of clotting factor VIII (FVIII). This disorder remains unacceptably underdiagnosed in India. Early diagnosis and appropriate management of HA can substantially prevent morbidity and mortality. Currently, HA is managed with regular replacement therapy using standard or extended half-life FVIII concentrates or non-factor drug products. The challenges associated with FVIII concentrates include plateauing of drug effect, issues with its administration and adherence to treatment, breakthrough bleeds, and the development of inhibiting antibodies against administered clotting factors. Emicizumab is a bispecific antibody, launched in India in April 2019, for managing patients with HA. To investigate the role of emicizumab in Indian patients with HA, opinions were sought from 13 eminent hematologists and experts from India on the effectiveness of emicizumab in preventing all bleeds, spontaneous bleeds, perioperative bleeds, and intracranial hemorrhage; resolving target joints; and reducing the rate of hospitalizations and fatality associated with HA in children and adults, with or without inhibitors. The benefits of emicizumab over traditional FVIII concentrates include the subcutaneous route of delivery, less frequent dosing, and a lack of inhibitor development, in addition to providing sustained hemostasis without in-depth monitoring. It is a safe and effective management option for all HA patients, especially for patients with certain archetypes, such as those with inhibitors, those with high annualized bleed rates, those living far away from hemophilia care centers, pediatric patients and infants with intravenous access challenges, and those with a history of life-threatening bleeding events.

Management of pregnancy and delivery in congenital fibrinogen disorders: communication from the ISTH SSC Subcommittee on Factor XIII and Fibrinogen.

Congenital fibrinogen disorders (CFDs) are a heterogeneous group of rare congenital quantitative and/or qualitative fibrinogen deficiencies. The spectrum of molecular anomalies is broad, leading to several subtypes of fibrinogen disorders (ie, afibrinogenemia, hypofibrinogenemia, dysfibrinogenemia, and hypodysfibrinogenemia). Pregnancy in women with CFDs is a high-risk clinical situation, with an increased tendency for miscarriages, bleeding, and thrombosis. Even though it is well established that management of such pregnancies requires a multidisciplinary approach involving specialists (hematologists and maternal/fetal medicine experts with expertise in the management of inherited bleeding disorders), specific guidelines are lacking. In this International Society on Thrombosis and Haemostasis (ISTH) Scientific and Standardization Committee communication, we aim to propose an expert consensus opinion with literature evidence where available on the strategy for management of pregnancy, delivery, and puerperium in CFDs.

Efficacy and safety of fitusiran prophylaxis in people with haemophilia A or haemophilia B with inhibitors (ATLAS-INH): a multicentre, open-label, randomised phase 3 trial.

BACKGROUND: Fitusiran, a subcutaneous investigational small interfering RNA therapeutic, targets antithrombin to rebalance haemostasis in people with haemophilia A or haemophilia B, irrespective of inhibitor status. We evaluated the efficacy and safety of fitusiran prophylaxis in people with haemophilia A or haemophilia B with inhibitors. METHODS: This multicentre, randomised, open-label phase 3 study was done at 26 sites (primarily secondary or tertiary centres) in 12 countries. Men, boys, and young adults aged 12 years or older with severe haemophilia A or haemophilia B with inhibitors previously treated with on-demand bypassing agents were randomly assigned (2:1) to receive once-a-month 80 mg subcutaneous fitusiran prophylaxis (fitusiran prophylaxis group) or to continue with bypassing agents on-demand (bypassing agents on-demand group) for 9 months. The primary endpoint was mean annualised bleeding rate during the efficacy period in the intention-to-treat population estimated by negative binomial model. Safety was assessed as a secondary endpoint in the safety population. This trial is complete and is registered with ClinicalTrials.gov, NCT03417102. FINDINGS: Between Feb 14, 2018, and June 23, 2021, 85 participants were screened for inclusion, of whom 57 (67%; 57 [100%] men; median age 27·0 years [IQR 19·5-33·5]) were randomly assigned: 19 (33%) participants to the bypassing agent on-demand group and 38 (67%) participants to the fitusiran prophylaxis. Negative binomial model-based mean annualised bleeding rate was significantly lower in the fitusiran prophylaxis group (1·7 [95% CI 1·0-2·7]) than in the bypassing agents on-demand group (18·1 [10·6-30·8]), corresponding to a 90·8% (95% CI 80·8-95·6) reduction in annualised bleeding rate in favour of fitusiran prophylaxis (p<0·0001). 25 (66%) participants had zero treated bleeds in the fitusiran prophylaxis group versus one (5%) in the bypassing agents on-demand group. The most frequent treatment-emergent adverse event in the fitusiran prophylaxis group was increased alanine aminotransferase in 13 (32%) of 41 participants in the safety population; there were no increased alanine aminotransferase treatment-emergent adverse events in the bypassing agents on-demand group. Suspected or confirmed thromboembolic events were reported in two (5%) participants in the fitusiran prophylaxis group. No deaths were reported. INTERPRETATION: Subcutaneous fitusiran prophylaxis resulted in statistically significant reductions in annualised bleeding rate in participants with haemophilia A or haemophilia B with inhibitors, with two-thirds of participants having zero bleeds. Fitusiran prophylaxis might show haemostatic efficacy in participants with haemophilia A or haemophilia B with inhibitors; therefore, the therapeutic might have the potential to improve the management of people with haemophilia. FUNDING: Sanofi.

FUBP3 regulates chronic myeloid leukaemia progression through PRC2 complex regulated PAK1-ERK signalling.

The development of resistance and heterogeneity in differential response towards tyrosine kinase inhibitors (TKI) in chronic myeloid leukaemia (CML) treatment has led to the exploration of factors independent of the Philadelphia chromosome. Among these are the association of deletions of genes on derivative (der) 9 chromosome with adverse outcomes in CML patients. However, the functional role of genes near the breakpoint on der (9) in CML prognosis and progression remains largely unexplored. Copy number variation and mRNA expression were evaluated for five genes located near the breakpoint on der (9). Our data showed a significant association between microdeletions of the FUBP3 gene and its reduced expression with poor prognostic markers and adverse response outcomes in CML patients. Further investigation using K562 cells showed that the decrease in FUBP3 protein was associated with an increase in proliferation and survival due to activation of the MAPK-ERK pathway. We have established a novel direct interaction of FUBP3 protein and PRC2 complex in the regulation of ERK signalling via PAK1. Our findings demonstrate the role of the FUBP3 gene located on der (9) in poor response and progression in CML with the identification of additional druggable targets such as PAK1 in improving response outcomes in CML patients.

Pharmacokinetics, efficacy and safety of a novel fibrinogen concentrate in pediatric patients with congenital afibrinogenemia.

INTRODUCTION: Congenital afibrinogenemia treatment with plasma-derived fibrinogen concentrates in pediatric patients is limited. This study investigated the pharmacokinetics, surrogate efficacy, and safety of a plasma-derived fibrinogen concentrate (FIB Grifols) in pediatric patients with congenital afibrinogenemia. METHODS: Patients aged <18 years old diagnosed with congenital afibrinogenemia were included in this prospective, multinational, phase 1-2, single-arm study. After a single dose of a plasma-derived fibrinogen concentrate (70 mg/kg body weight), pharmacokinetic parameters were determined from plasma fibrinogen activity (Clauss method) and antigen method (ELISA), and calculated by noncompartmental and population pharmacokinetic (popPK) models. Patients were followed up over 14 days. Efficacy variables were the mean change on thromboelastographic variables (maximum clot firmness [MCF], alpha angle [ α ]) and coagulation tests (prothrombin time, activated partial thromboplastin time, and thrombin time) 1 h postinfusion. Safety parameters were assessed. RESULTS: Eleven patients with a median (range) age 8.80 (3.7-12.7) years were treated with the plasma-derived fibrinogen concentrate. Using the popPK modeling, fibrinogen activity reached a mean (standard deviation) Cmax of 1.3 (0.225) g/l, half-life ( t1/2 ) of 60.6 (4.48) h and incremental in vivo recovery (IVR) of 1.86 (0.322) (mg/dl)/(mg/kg). Surrogate efficacy was demonstrated by significant increase in MCF (9.23 [3.94] mm; P  < 0.001; 95% confidence interval 6.58, 11.87). All coagulation times were significantly shortened after fibrinogen concentrate infusion. Adverse events were mild or moderate in severity, and unrelated to fibrinogen concentrate. CONCLUSIONS: In pediatric patients with congenital afibrinogenemia, plasma-derived fibrinogen concentrate revealed a favorable and specific pharmacokinetic profile, demonstrated efficacy in coagulation and was safe and well tolerated.

Immunogenicity and safety of quadrivalent and 9-valent human papillomavirus vaccines in Indian clinical trial participants.

The quadrivalent human papillomavirus (qHPV; HPV6/11/16/18) and 9-valent HPV (9vHPV; HPV6/11/16/18/31/33/45/52/58) vaccines have demonstrated efficacy, immunogenicity, and safety in international clinical trials. We report outcomes from three completed clinical trials in India: a single-arm study (V501-029 [NCT00380367]) in Indian girls (aged 9-15 years; N = 110) evaluating qHPV vaccine immunogenicity and safety; a subgroup analysis (n = 225) of Indian girls/boys (9-15 years) and women (16-26 years) from a global study (V503-002 [NCT00943722]) evaluating 9vHPV vaccine immunogenicity and safety; and a qHPV vaccine post-marketing safety surveillance study (V501-125) in Indian females (aged 9-45 years; N = 188) vaccinated during routine care. In V501-029 and V503-002, HPV vaccines were administered as 3 doses (Day 1, Month 2, Month 6). Serum HPV antibodies were evaluated by competitive Luminex immunoassays at Day 1 and Month 7 (both studies) and Months 12, 24, and 36 (V503-002 only). Adverse events (AEs) were collected by Vaccination Report Card. In V501-125, participants were actively surveilled for serious AEs (SAEs) within 30 days post-qHPV vaccination. In per-protocol analyses, qHPV and 9vHPV vaccines induced robust anti-HPV6/11/16/18 (V501-029) and HPV6/11/16/18/31/33/45/52/58 (V503-002) responses, respectively; ≥97% of participants seroconverted at Month 7 for each vaccine HPV type in both studies, and antibody responses persisted through 36 months in V503-002. The most common AEs were injection-site-associated. Most AEs were mild/moderate; no deaths, vaccine-related SAEs, or discontinuations due to AEs were reported. In V501-125, no SAE was reported. Overall, the qHPV and 9vHPV vaccines elicited robust antibody responses and were generally well tolerated in Indian participants.

Rare Complication of a Rare Malignancy: Case Report of Cardiac Amyloidosis Secondary to Waldenstrom Macroglobulinemia.

Cardiac amyloidosis is a rare disorder caused by the myocardial deposition of abnormal fibrils. A 52-year-old man was referred to our center with clinical features of heart failure, after cardiac magnetic resonance imaging showed restrictive cardiomyopathy. Abdominal fat pad biopsy showed features of amyloidosis, and after hematological workup, he was diagnosed with Waldenstrom macroglobulinemia (WM). He was initiated on a rituximab-based chemotherapy regimen, and his cardiac function was assessed serially. Because of non-response, he was switched to a bortezomib-based regimen. Unfortunately, three days into this regimen, the patient died. WM is a rare plasma cell dyscrasia with a nonspecific presentation. It uncommonly presents with sequelae of amyloidosis-the IgM subtype of amyloid-light chain (AL) amyloidosis. Diagnostic delays are common, contributing to an already poor prognosis. Amyloidosis in WM requires urgent treatment - clonal chemotherapy, and supportive cardiac care in heart involvement. Bortezomib-based regimens are commonly recommended, with diuretics as the mainstay for cardiac treatment. However, in most advanced cases, the prognosis is poor; thus, a high degree of suspicion is necessary for early diagnosis. This case illustrates the possible presentation of cardiac amyloidosis as a rare malignancy.

Autoimmune Hemolytic Anaemias in Pregnancy: Experience in a Tertiary Care Hospital in South India.

BACKGROUND: Autoimmune hemolytic anaemia is very rare and there is limited data regarding their pregnancy outcomes. Hence we aimed to study the maternal and perinatal outcomes in pregnancies with autoimmune hemolytic anaemias (AIHA). METHODS: A retrospective descriptive study of pregnant women with AIHA, who delivered at SJMCH between January 2011 and January 2016 was carried out. Their antenatal and labour records were reviewed and demographic details noted.The primary outcome measures studied were-the prevalence of AIHA, gestational age at delivery, antepartum, intrapartum and postpartum complications, mode of delivery and requirement of transfusion of blood and blood products. The secondary outcome measures studied included neonatal outcomes such as low birth weight, intrauterine growth restriction and need for intensive care. The data is presented as descriptive statistics, including means and percentage. RESULTS: The prevalence of AIHA was (18/12,420) 0.14%. The mean gestational age at delivery was 34 weeks; 100%, 77% and 50% had antenatal, intra partum or postpartum complications, respectively. 44% had preeclampsia, 38% intrauterine growth restriction and 16% preterm labour. 83% required additional drugs for treatment of AIHA.72% had vaginal delivery; 28% had caesarean delivery; 33% were transfused antenatally and 22% postnatally; 50% of the babies were preterm and required intensive care, 66% had low birth weight. There was no maternal mortality. CONCLUSION: Multidisciplinary approach, early diagnosis and detection of autoimmune hemolytic anaemia and complications, good antenatal care, judicious transfusions and delivery at tertiary care centre are the keys to successful outcomes.

Innate Immune Cytokine Profiling and Biomarker Identification for Outcome in Dengue Patients.

Background: Early biomarkers of progression to severe dengue are urgently required to enable effective patient management and control treatment costs. Innate immune cells, which comprise the earliest responders to infection and along with the cytokines and chemokines they secrete, play a vital role in orchestrating the subsequent adaptive immune response and have been implicated in the enhancement of infection and "cytokine storm" associated with dengue severity. We investigated the early innate immune cytokine profile of dengue patients during acute phase of disease in a prospective blinded study that included subjects with acute dengue and febrile controls from four major hospitals in Bengaluru, India along with healthy controls. We used intracellular cytokine staining and flow cytometry to identify innate immune biomarkers that can predict progression to severe dengue. Results: Dengue infection resulted in enhanced secretion of multiple cytokines by all queried innate immune cell subsets, dominated by TNF-α from CD56+CD3+ NKT cells, monocyte subsets, and granulocytes along with IFN-γ from CD56+CD3+ NKT cells. Of note, significantly higher proportions of TNF-α secreting granulocytes and monocyte subsets at admission were associated with mild dengue and minimal symptoms. Dengue NS1 antigenemia used as a surrogate of viral load directly correlated with proportion of cytokine-secreting innate immune cells and was significantly higher in those who went on to recover with minimal symptoms. In patients with secondary dengue or those with bleeding or elevated liver enzymes who revealed predisposition to severe outcomes, early activation as well as efficient downregulation of innate responses were compromised. Conclusion: Our findings suggested that faulty/delayed kinetics of innate immune activation and downregulation was a driver of disease severity. We identified IFN-γ+CD56+CD3+ NKT cells and IL-6+ granulocytes at admission as novel early biomarkers that can predict the risk of progression to severity (composite AUC = 0.85-0.9). Strong correlations among multiple cytokine-secreting innate cell subsets revealed that coordinated early activation of the entire innate immune system in response to dengue virus infection contributed to resolution of infection and speedy recovery.

Microarray analysis reveals distinct immune signatures in BCR-ABL positive and negative myeloproliferative neoplasms.

BACKGROUND: BCR-ABL mutation on the Philadelphia chromosome is the key driver of chronic myeloid leukemia (CML) pathogenesis. However, there are certain cases of myeloproliferative neoplasms (MPN) wherein no inherent driver mutation is detected resulting in clinical phenotype. It is important to identify key genes and pathways in driving the disease. The aim of the study was to use a gene-based omics approach to molecularly characterize these mutation-positive and negative cases to further strengthen diagnostics and precision medicine. METHODS: A microarray profiling was done on CD34 positive cells isolated from two BCR-ABL positive and five BCR-ABL negative samples. JAK2V617F mutation testing was also done to rule out the presence of any other mutation in the latter group. The fold change cut-off was taken as ±1.5 with p≤0.5 for significant genes. The gene network and pathway analysis were done using DAVID and STRING software. RESULTS: The genes upregulated in BCR-ABL negative samples were shown to be involved in immune regulation, signal transduction and T- and B-cell signalling. The protein-protein interaction network of upregulated genes in these samples were enriched for various immunomodulatory genes such as HLADP, HLADQ, IL7R, CCR7, CD3 subtypes. These genes further formed a network with signal transduction genes such as LCK, FYN, RAG1, DOCK1, AKT3, SMAD3, LEF1. CONCLUSION: The results suggested a modulation of immune response genes and its subsequent effect on oncogenic signalling in BCR-ABL negative samples as compared to BCR-ABL positive samples. The protein network analysis was enriched for genes involved in Src, TGF-beta and PI3K-AKT pathway contributing to the proliferation of neoplastic clone.

Gene expression profiling of CD34(+) cells from patients with myeloproliferative neoplasms.

Myeloproliferative neoplasms (MPN) are clonal disorders characterized by the increased proliferation of hematopoietic stem cell precursors and mature blood cells. Mutations of Janus kinase 2 (JAK2), Calreticulin (CALR) and MPL (myeloproliferative leukemia virus) are key driver mutations in MPN. However, the molecular profile of triple negative MPN has been a subject of ambiguity over the past few years. Mutations of, methylcytosine dioxygenase TET2, polycomb group protein ASXL1 and histone-lysine N-methyltransferase EZH2 genes have accounted for certain subsets of triple negative MPNs but the driving cause for majority of cases is still unexplored. The present study performed a microarray-based transcriptomic profile analysis of bone marrow-derived CD34(+) cells from seven MPN samples. A total of 21,448 gene signatures were obtained, which were further filtered into 472 upregulated and 202 downregulated genes. Gene ontology and protein-protein interaction (PPI) network analysis highlighted an upregulation of genes involved in cell cycle and chromatin modification in JAK2V617F negative vs. positive MPN samples. Out of the upregulated genes, seven were associated with the hematopoietic stem cell signature, while forty-seven were associated with the embryonic stem cell signature. The majority of the genes identified were under the control of NANOG and E2F4 transcription factors. The PPI network indicated a strong interaction between chromatin modifiers and cell cycle genes, such as histone-lysine N-methyltransferase SUV39H1, SWI/SNF complex subunit SMARCC2, SMARCE2, chromatin remodeling complex subunit SS18, tubulin ß (TUBB) and cyclin dependent kinase CDK1. Among the upregulated epigenetic markers, there was a ~10-fold increase in MYB expression in JAK2V617F negative samples. A significant increase in total CD34 counts in JAK2V617F negative vs. positive samples (P<0.05) was also observed. Overall, the present data showed a distinct pattern of expression in JAK2V617F negative vs. positive samples with upregulated genes involved in epigenetic modification.

Pharmacokinetics, surrogate efficacy and safety evaluations of a new human plasma-derived fibrinogen concentrate (FIB Grifols) in adult patients with congenital afibrinogenemia.

BACKGROUND AND AIMS: Congenital afibrinogenemia is a rare coagulation disorder resulting from a deficiency in fibrinogen. This study assessed the pharmacokinetics, surrogate efficacy and safety of FIB Grifols, a new human plasma-derived fibrinogen concentrate, to treat congenital afibrinogenemia. METHODS: Eleven adult patients from a multinational, phase 1-2, prospective, open-label, single-arm, uncontrolled clinical study received a single infusion of FIB Grifols, 70 mg/kg bw. Fibrinogen pharmacokinetics (fibrinogen activity: Clauss method; antigen plasma concentrations: ELISA) and efficacy parameters were determined over 14 days after infusion. Efficacy endpoints were the mean change on plasma maximum clot firmness (MCF) on viscoelastic testing and coagulation tests 1-hour post-infusion, and correlation with fibrinogen levels throughout. Safety parameters were also assessed. RESULTS: For the Clauss method, (mean [standard deviation]) baseline adjusted Cmax was 1.99 (0.40) g/L, reached 1.76 (1.00) h after infusion, and half-life was 76.94 (20.21) h. Using ELISA, Cmax after FIB Grifols infusion was 2.88 (0.86) mg/mL, with a tmax of 3.06 (2.24) h. Fibrinogen activity and antigen concentrations showed statistically significant correlation of 0.9120 (P < 0.001). Surrogate efficacy was demonstrated by a significant increase of 12.35 (3.85) mm in MCF. Prothrombin time, activated partial thromboplastin time and thrombin time, returned to normal ranges over time, indicating restoration of functionally active fibrinogen. There were no treatment-related adverse events, allergic reactions, serious adverse events, or discontinuations. CONCLUSIONS: The pharmacokinetic profile of functionally active FIB Grifols was established, hemostasis was restored, and FIB Grifols was safe and well tolerated in fibrinogen-deficient patients.

Clinical phenotype, fibrinogen supplementation, and health-related quality of life in patients with afibrinogenemia.

Due to the low prevalence of afibrinogenemia, epidemiologic data on afibrinogenemia are limited, and no data are available on health-related quality of life (HRQoL). We conducted a cross-sectional international study to characterize the clinical features, the fibrinogen supplementation modalities, and their impact on HRQoL in patients with afibrinogenemia. A total of 204 patients (119 adults and 85 children) from 25 countries were included. The bleeding phenotype was severe: 68 (33.3%) patients having at least one bleed per month and 48 (23%) a history of cerebral bleeding. About 35% (n = 72) of patients were treated with fibrinogen concentrates or cryoprecipitates as prophylaxis, 18.1% (n = 37) received ≥1 injection per week, and 16.6% (n = 34) were on home treatment. A thrombotic event was reported in venous and/or arterial territories by 37 (18.1%) patients. Thrombosis occurred even in young patients, and recurrence was frequent (7.4%). The total HRQoL was lower in children than in adults. Discomfort linked to treatment and limitations to sports and leisure were the main concerns. Women and children were particularly affected in family relationships. In multivariate analyses, younger age, residence in Asia or Africa, and a previous thrombotic event were statistically correlated with a worse HRQoL. In summary, our study underlines the severe bleeding and thrombotic phenotype and their impact on HRQoL in afibrinogenemia. The optimal strategy for fibrinogen supplementation needs to be determined. This trial was registered at www.clinicaltrials.gov as #NCT03484065.

Turoctocog alfa is safe for the treatment of Indian patients with hemophilia A: Guardian 10 trial results.

BACKGROUND: Hemophilia A is an X chromosome-linked bleeding disorder caused by the deficiency of coagulation factor VIII (FVIII). The majority of the Indian population with hemophilia A use plasma-derived clotting factors and, in some instances, fresh frozen plasma and cryoprecipitate. Safer and more efficient treatment options are needed for this group of patients. OBJECTIVES: To assess the safety of turoctocog alfa, a third-generation recombinant FVIII molecule, for the treatment and prophylaxis of bleeding episodes in previously treated Indian patients with moderate or severe hemophilia A. PATIENTS/METHODS: This single-country, multicenter, open-label, nonrandomized trial enrolled 60 patients who received prophylactic treatment with turoctocog alfa for 8 weeks, which corresponded to a minimum of 20 exposure days. Confirmed development of FVIII inhibitors during the 8-week treatment period was evaluated. Other assessments included frequencies of adverse drug reactions (ARs), serious adverse reactions, drug-related allergic reactions, and infusion reactions during the 12-week period after the first treatment; hemostatic effect of turoctocog alfa for the treatment of bleeding episodes; and total annualized dose of turoctocog alfa administered during the 8-week treatment period. RESULTS: No incidence of FVIII inhibitors was detected. No safety concerns such as ARs, serious ARs, or drug-related allergic reactions were noted. The hemostatic success rate for the treatment of bleeding episodes with turoctocog alfa was 81.6%. CONCLUSIONS: The trial results demonstrated that turoctocog alfa is a safe treatment option for the prophylaxis and treatment of bleeding episodes in previously treated adolescent and adult patients with hemophilia A in the Indian population.

Effect of point mutation on structure-function correlation of hemoglobin variants, HbE and HbD Punjab.

Hemoglobinopathies are examples of autosomal recessive disorders of human hemoglobin. Hemoglobin E (HbE) and Hemoglobin D Punjab (HbD Punjab) are two of the most common hemoglobin variants geographically spread across Asian continent. These two variants differ from normal human hemoglobin (HbA) at a single amino acid residue caused by the point mutation of ß globin gene. The presence of the mutated amino acid residue causes perturbation in the function of both variants. However, the structure-function correlation of these variants has not been established yet. In the present study, we analyzed the conformational changes associated with oxygenation of hemoglobin variants using hydrogen/deuterium exchange-based mass spectrometry of backbone amide hydrogens of α and ß globin chains in the tetrameric hemoglobin molecule. We also performed the functional assay of these variants using oxygen dissociation equilibrium curve. Compared to HbA, both variants showed reduced oxygen affinity, as reported earlier. The functional perturbations exhibited by these variants were correlated well with their structural alterations with respect to the reported changes in the residue level interactions upon oxygenation of normal hemoglobin, monitored through the hydrogen/deuterium exchange kinetics of several peptic peptides originated from the isotopically exchanged oxy and deoxy forms of HbE and HbD Punjab.

Fibrinogen concentrate for treatment of bleeding and surgical prophylaxis in congenital fibrinogen deficiency patients.

BACKGROUND: Congenital fibrinogen deficiency is an ultra-rare disorder in which patients can experience severe and/or frequent bleeding episodes (BEs). Here, we present the largest prospective study to date on the treatment of this disorder. METHODS: Hemostatic efficacy of human fibrinogen concentrate (HFC; FIBRYGA® , Octapharma AG) for treatment of bleeding or surgical prophylaxis was assessed by investigators and adjudicated by an independent data monitoring and endpoint adjudication committee (IDMEAC) according to a four-point scale, using objective criteria. Thromboelastometry maximum clot firmness (MCF) was also determined. RESULTS: Twenty-five afibrinogenemia patients were treated with HFC: 24 for on-demand treatment of 89 BEs, and nine as prophylaxis for 12 surgeries. For BEs, treatment success (rating of excellent or good) evaluated by investigators was 96.6% (90% confidence interval [CI], 0.92-0.99; two missing ratings, classified as failures) and by the IDMEAC was 98.9% (90% CI, 0.95-0.999). Mean ± standard deviation (SD) increase in MCF was 5.8 ± 2.5 mm one hour after the first HFC infusion (mean ± SD dose, 61.88 ± 11.73 mg/kg). For the 12 surgeries (median [range] HFC dose/surgery, 85.80 mg/kg [34.09-225.36]), intraoperative and postoperative treatment success were both rated 100% (90% CI, 0.82-1.00) by investigators and the IDMEAC. Three adverse events were possibly treatment related, including a moderate case of thrombosis. There were no deaths, no severe allergic or hypersensitivity reactions, and no clinical evidence of neutralizing antifibrinogen antibodies. CONCLUSIONS: Human fibrinogen concentrate was efficacious for on-demand treatment of bleeding and as surgical prophylaxis, with a favorable safety profile, in patients with congenital afibrinogenemia.

RhoC regulates radioresistance via crosstalk of ROCK2 with the DNA repair machinery in cervical cancer.

BACKGROUND: Radioresistance remains a challenge to the successful treatment of various tumors. Intrinsic factors like alterations in signaling pathways regulate response to radiation. RhoC, which has been shown to modulate several tumor phenotypes has been investigated in this report for its role in radioresistance. In vitro and clinical sample-based studies have been performed to understand its contribution to radiation response in cervical cancer and this is the first report to establish the role of RhoC and its effector ROCK2 in cervical cancer radiation response. METHODS: Biochemical, transcriptomic and immunological approaches including flow cytometry and immunofluorescence were used to understand the role of RhoC and ROCK2. RhoC variants, siRNA and chemical inhibitors were used to alter the function of RhoC and ROCK2. Transcriptomic profiling was performed to understand the gene expression pattern of the cells. Live sorting using an intracellular antigen has been developed to isolate the cells for transcriptomic studies. RESULTS: Enhanced expression of RhoC conferred radioprotection on the tumor cells while inhibition of RhoC resulted in sensitization of cells to radiation. The RhoC overexpressing cells had a better DNA repair machinery as observed using transcriptomic analysis. Similarly, overexpression of ROCK2, protected tumor cells against radiation while its inhibition increased radiosensitivity in vitro. Further investigations revealed that ROCK2 inhibition abolished the radioresistance phenotype, conferred by RhoC on SiHa cells, confirming that it is a downstream effector of RhoC in this context. Additionally, transcriptional analysis of the live sorted ROCK2 high and ROCK2 low expressing SiHa cells revealed an upregulation of the DNA repair pathway proteins. Consequently, inhibition of ROCK2 resulted in reduced expression of pH2Ax and MRN complex proteins, critical to repair of double strand breaks. Clinical sample-based studies also demonstrated that ROCK2 inhibition sensitizes tumor cells to irradiation. CONCLUSIONS: Our data primarily indicates that RhoC and ROCK2 signaling is important for the radioresistance phenotype in cervical cancer tumor cells and is regulated via association of ROCK2 with the proteins of DNA repair pathway involving pH2Ax, MRE11 and RAD50 proteins, partly offering insights into the mechanism of radioresistance in tumor cells. These findings highlight RhoC-ROCK2 signaling involvement in DNA repair and urge the need for development of these molecules as targets to alleviate the non-responsiveness of cervical cancer tumor cells to irradiation treatment.