Multicenter assessment and longitudinal study of the prevalence of antibodies and related adaptive immune responses to AAV in adult males with hemophilia.

Adeno-associated virus (AAV) based gene therapy has demonstrated effective disease control in hemophilia. However, pre-existing immunity from wild-type AAV exposure impacts gene therapy eligibility. The aim of this multicenter epidemiologic study was to determine the prevalence and persistence of preexisting immunity against AAV2, AAV5, and AAV8, in adult participants with hemophilia A or B. Blood samples were collected at baseline and annually for ≤3 years at trial sites in Austria, France, Germany, Italy, Spain, and the United States. At baseline, AAV8, AAV2, and AAV5 neutralizing antibodies (NAbs) were present in 46.9%, 53.1%, and 53.4% of participants, respectively; these values remained stable at Years 1 and 2. Co-prevalence of NAbs to at least two serotypes and all three serotypes was present at baseline for ~40% and 38.2% of participants, respectively. For each serotype, ~10% of participants who tested negative for NAbs at baseline were seropositive at Year 1. At baseline, 38.3% of participants had detectable cell mediated immunity by ELISpot, although no correlations were observed with the humoral response. In conclusion, participants with hemophilia may have significant preexisting immunity to AAV capsids. Insights from this study may assist in understanding capsid-based immunity trends in participants considering AAV vector-based gene therapy.

BAX 335 hemophilia B gene therapy clinical trial results: potential impact of CpG sequences on gene expression.

Gene therapy has the potential to maintain therapeutic blood clotting factor IX (FIX) levels in patients with hemophilia B by delivering a functional human F9 gene into liver cells. This phase 1/2, open-label dose-escalation study investigated BAX 335 (AskBio009, AAV8.sc-TTR-FIXR338Lopt), an adeno-associated virus serotype 8 (AAV8)-based FIX Padua gene therapy, in patients with hemophilia B. This report focuses on 12-month interim analyses of safety, pharmacokinetic variables, effects on FIX activity, and immune responses for dosed participants. Eight adult male participants (aged 20-69 years; range FIX activity, 0.5% to 2.0%) received 1 of 3 BAX 335 IV doses: 2.0 × 1011; 1.0 × 1012; or 3.0 × 1012 vector genomes/kg. Three (37.5%) participants had 4 serious adverse events, all considered unrelated to BAX 335. No serious adverse event led to death. No clinical thrombosis, inhibitors, or other FIX Padua-directed immunity was reported. FIX expression was measurable in 7 of 8 participants; peak FIX activity displayed dose dependence (32.0% to 58.5% in cohort 3). One participant achieved sustained therapeutic FIX activity of ∼20%, without bleeding or replacement therapy, for 4 years; in others, FIX activity was not sustained beyond 5 to 11 weeks. In contrast to some previous studies, corticosteroid treatment did not stabilize FIX activity loss. We hypothesize that the loss of transgene expression could have been caused by stimulation of innate immune responses, including CpG oligodeoxynucleotides introduced into the BAX 335 coding sequence by codon optimization. This trial was registered at www.clinicaltrials.gov as #NCT01687608.

An update on the US adult thalassaemia population: a report from the CDC thalassaemia treatment centres.

Thalassaemia is caused by genetic globin defects leading to anaemia, transfusion-dependence and comorbidities. Reduced survival and systemic organ disease affect transfusion-dependent thalassaemia major and thalassaemia intermedia. Recent improvements in clinical management have reduced thalassaemia mortality. The therapeutic landscape of thalassaemia may soon include gene therapies as functional cures. An analysis of the adult US thalassaemia population has not been performed since the Thalassemia Clinical Research Network cohort study from 2000 to 2006. The Centers for Disease Control and Prevention supported US thalassaemia treatment centres (TTCs) to compile longitudinal information on individuals with thalassaemia. This dataset provided an opportunity to evaluate iron balance, chelation, comorbidities and demographics of adults with thalassaemia receiving care at TTCs. Two adult cohorts were compared: those over 40 years old (n = 75) and younger adults ages 18-39 (n = 201). The older adult cohort was characterized by higher numbers of iron-related comorbidities and transfusion-related complications. By contrast, younger adults had excess hepatic and cardiac iron and were receiving combination chelation therapy. The ethnic composition of the younger cohort was predominantly of Asian origin, reflecting the demographics of immigration. These findings demonstrate that comprehensive care and periodic surveys are needed to ensure optimal health and access to emerging therapies.

Platelets and platelet-derived factor Va confer hemostatic competence in complete factor V deficiency.

Whole genome sequencing of an individual completely devoid of plasma- and platelet-derived factor V (FV) identified 167 variants in his F5 gene including previously identified and damaging missense mutations at rs6027 and Leu90Ser. Because the administration of fresh frozen plasma (FFP) prevents gastrointestinal bleeding in this individual, its effects on his plasma- and platelet-derived FV concentrations were assessed. The patient's plasma FV levels peaked by 2 hours following FFP administration and were undetectable 96 hours later. In contrast, increased platelet-derived FV/Va concentrations were observed within 6 hours, peaked at 24 hours, decreased slowly over 7 days, and originated from megakaryocyte endocytosis and intracellular processing of plasma FV. Ten days after transfusion, no thrombin was generated in a tissue factor-initiated whole blood clotting assay unless exogenous FV was added, consistent with the complete absence of plasma FV. In marked contrast, release of the patient's platelet-derived FV/Va (7% of normal) following platelet activation resulted in robust thrombin generation, similar to that in an individual with normal plasma- and platelet-derived FV concentrations. Thus, total FV deficiency can be corrected by plasma administration, which partially repletes and sustains the platelet cofactor pool, thereby highlighting the critical role of platelet-derived FV/Va in ensuring hemostatic competence.

Gender and duration of disease differentiate responses to rituximab-dexamethasone therapy in adults with immune thrombocytopenia.

Adults often develop chronic immune thrombocytopenia (ITP) for which treatment order is uncertain. Rituximab and three cycles of dexamethasone (4R + 3Dex) improve treatment responses and short-term disease control but long-term outcome is not known. In adults with ITP treated with 4R + 3D, we sought long-term outcome and associated prognostic variables. Forty-nine adults treated at Weill-Cornell received 4R + 3Dex. Their clinical characteristics were reviewed. Duration was median time to treatment failure; Kaplan-Meier estimates were developed. Vbeta Tcell receptor (VBTCR) repertoire was obtained after treatment in 36 patients. Patients were adults with ITP 18-64 years old, median age 37. The 27 females were twice as likely to have an ongoing response to 4R + 3Dex (44.1%) as males (19.6%; P = 0.009). For ITP duration <12 months, 52.7% of patients had continuing responses to 4R + 3Dex compared to 15.3% of patients with diagnosis >12 months (P = 0.02). Females with ITP duration of <12 months had continuing responses in 78.6%, compared to males with <12 months duration of ITP (21.2%). For patients with disease duration <12 months, 67% of females had continuing responses, compared to 31% of males (P = 0.004). Post-treatment polyclonal VBTCR was seen in 9/10 continuing responders (six female, three male) but only 13/26 relapsers/nonresponders (P = 0.068). Durable remissions after treatment with 4R + 3Dex were more frequent in female patients with <12 months of ITP duration and those with polyclonal VBTCR after treatment, emphasizing the roles of duration of disease, gender and T cells in chronic ITP. Differences in pathophysiology of ITP by gender and by duration of ITP require further study. Am. J. Hematol. 91:907-911, 2016. © 2016 Wiley Periodicals, Inc.

The role of complement activation in thrombosis and hemolytic anemias.

OBJECTIVE: The objective of this study was to describe complement activation in hemostatic and pathologic states of coagulation and in the acquired and congenital hemolytic anemias. METHODS AND RESULTS: We review published and emerging data on the involvement of the classic, alternative and lectin-based complement pathways in coagulation and the hemolytic anemias. The alternative pathway in particular is always "on," at low levels, and is particularly sensitive to hyper-activation in a variety of physiologic and pathologic states including infection, autoimmune disorders, thrombosis and pregnancy, requiring tight control predicated on a variety of soluble and membrane bound regulatory proteins. In acquired hemolytic anemias such as paroxysmal nocturnal hemoglobinuria (PNH) and cold agglutinin disease (CAD), the complement system directly induces red blood cell injury, resulting in intravascular and extravascular hemolysis. In congenital hemolytic anemias such as sickle cell disease and ß-thalassemia, the complement system may also contribute to thrombosis and vascular disease. Complement activation may also lead to a storage lesion in red blood cells prior to transfusion. CONCLUSION: Complement pathways are activated in hemolytic anemias and are closely linked with thrombosis. In acquired disorders such as PNH and possibly CAD, inhibition of the alternative complement pathway improves clinical outcomes and reduces thrombosis risk. Whether complement inhibition has a similar role in congenital hemolytic anemias apart from the atypical hemolytic-uremic (aHUS)-type thrombotic microangiopathies remains to be determined.

Clinical presentations, risk factors, treatment and outcomes in patients with splanchnic vein thrombosis: a single-center experience.

Splanchnic vein thrombosis (SVT) is an uncommon form of venous thrombosis. Management can be challenging due to underlying conditions, increased bleeding risk, and lack of evidence from clinical trials. We sought to characterize the presentation and management of patients with SVT at a large tertiary hospital. A total of 43 patients' electronic medical records were reviewed. Median age at diagnosis was 43 (18-71). Sixteen patients had isolated portal vein thrombosis (37.2 %), and 16 (37.2 %) had thrombosis involving multiple splanchnic veins. Abdominal pain was the most common clinical presentation (67.4 %). Thrombophilia was present in 18 patients (41.9 %), nine had underlying liver disease (20.9 %) and seven had inflammatory bowel disease (16.3 %). Thirty-nine (90.7 %) patients were treated with anticoagulation, and 11(25.6 %) of these patients underwent interventional procedures. Thirty (69.8 %) patients remained on indefinite anticoagulation. Results of follow-up imaging at least 1 month after diagnosis were available for 29 patients; imaging showed chronic, stable thrombosis in 14 patients (48.3 %), resolution of thrombosis in 13 patients (44.8 %) and asymptomatic progression in two patients (6.9 %). Recurrent thrombosis occurred in four patients (9.3 %). Major bleeding occurred in eight patients who received anticoagulation (18.6 %), including fatal subdural hematoma in one patient. In this cohort of patients managed by hematologists and gastroenterologists, the majority of patients were treated with anticoagulation. Interventional procedures were higher than in previously reported series. Our study strongly supports the interdisciplinary management of splanchnic venous thrombosis.

Hematopoietic transplant-associated thrombotic microangiopathy: case report and review of diagnosis and treatments.

Transplant-associated thrombotic microangiopathy (TA-TMA) refers to inflammatory and thrombotic diseases of the microvasculature characterized by hemolytic anemia, thrombocytopenia, and evidence of organ damage, particularly acute renal failure. This syndrome occurs in 10% to 20% of patients with allogeneic hematopoietic stem cell transplants (HSCTs). It is much less frequent in the autologous setting. TA-TMAs present diagnostic challenges because they may not clearly fall into one of the categories of the 2 major TMAs: atypical hemolytic uremic syndrome (aHUS) and thrombotic thrombocytopenic purpura (TTP). In addition, complications of the transplant itself, including infection, graft-versus-host disease, and disseminated intravascular coagulation, as well as the side effects of immunosuppressive drugs, can mimic a TMA. Because the pathophysiology of TA-TMA is poorly understood, current treatment options are suboptimal, and the condition carries a very high mortality rate. In 3 recent case summaries, the median acute response rate to plasma exchange was as high as 55%, but this therapy failed to alter underlying disease pathology and had little impact on overall mortality, which was approximately 80%. Indeed, the vast majority of TA-TMA patients lack suppression of ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) activity to less than 5% to 10% of normal and do not have a complete response to plasma exchange, characteristics indicating that a TTP-like disorder is not involved. Recent advances in the treatment of aHUS may offer a therapeutic option in the aHUS-like TMAs associated with HSCTs. These issues are discussed in the context of a patient recently evaluated and treated at our institution; the case serves to illustrate the difficulties associated with the diagnosis and treatment of TA-TMA.

Similarities Between Somatosensory Cortical Responses Induced via Natural Touch and Microstimulation in the Ventral Posterior Lateral Thalamus in Macaques.

Lost sensations, such as touch, could be restored by microstimulation (MiSt) along the sensory neural substrate. Such neuroprosthetic sensory information can be used as feedback from an invasive brain-machine interface (BMI) to control a robotic arm/hand, such that tactile and proprioceptive feedback from the sensorized robotic arm/hand is directly given to the BMI user. Microstimulation in the human somatosensory thalamus (Vc) has been shown to produce somatosensory perceptions. However, until recently, systematic methods for using thalamic stimulation to evoke naturalistic touch perceptions were lacking. We have recently presented rigorous methods for determining a mapping between ventral posterior lateral thalamus (VPL) MiSt, and neural responses in the somatosensory cortex (S1), in a rodent model (Choi et al., 2016; Choi and Francis, 2018). Our technique minimizes the difference between S1 neural responses induced by natural sensory stimuli and those generated via VPL MiSt. Our goal is to develop systems that know what neural response a given MiSt will produce and possibly allow the development of natural "sensation." To date, our optimization has been conducted in the rodent model and simulations. Here, we present data from simple non-optimized thalamic MiSt during peri-operative experiments, where we used MiSt in the VPL of macaques, which have a somatosensory system more like humans, as compared to our previous rat work (Li et al., 2014; Choi et al., 2016). We implanted arrays of microelectrodes across the hand area of the macaque S1 cortex as well as in the VPL. Multi and single-unit recordings were used to compare cortical responses to natural touch and thalamic MiSt in the anesthetized state. Post-stimulus time histograms were highly correlated between the VPL MiSt and natural touch modalities, adding support to the use of VPL MiSt toward producing a somatosensory neuroprosthesis in humans.

Upstander Intervention and Parenting Styles.

Findings from a survey of children and adolescents (N = 645) documents that students witness and experience a range of abuse at home and at school. Participants freely acknowledged pushing or shoving (46%) and slapping or hitting peers (40%). The study contributes to the literature by focusing on upstanding (active versus passive bystander intervention) and parenting styles. Findings reveal an interesting disconnect between those who say they will intervene when confronted by friends' or peers' bullying behaviors and those who actually have intervened. Children and adolescents with authoritarian parents are more likely to say they would intervene to help peers, but when asked if they actually have done so, they are the least likely to follow-through. In contrast, children with authoritative or permissive parents show the opposite pattern: No significant difference in their intent to intervene, but they are more likely to become upstanders, rather than passive bystanders when actually confronted with bullying behavior.

Peri- and Postpartum Management of Patients With Factor XI Deficiency.

Factor XI (FXI) deficiency is an uncommon autosomal disorder with variable bleeding phenotype, making peripartum management challenging. We describe our experience in pregnant women with FXI deficiency and identify strategies to minimize the use of hemostatic agents and increase utilization of neuraxial anesthesia. Electronic records of 28 pregnant women with FXI deficiency seen by a hematology service in an academic medical center from January 2006 to August 2018 were reviewed. Data on bleeding, obstetric history, peripartum management, and FXI activity were collected. Partial FXI deficiency was defined as >20 IU/dL and severe <20 IU/dL. Median FXI activity was 42 IU/dL (range <1-73 IU/dL), and median activated partial thromboplastin time was 32.2 seconds (range: 27.8-75 seconds). There were 64 pregnancies: 53 (83%) live births and 11 (17%) pregnancy losses. Postpartum hemorrhage occurred in 9 (17%) pregnancies. Antifibrinolytic agents and fresh frozen plasma were used only in women with severe deficiency (42% with bleeding and 17% with no bleeding phenotype, respectively). Neuraxial anesthesia was successfully administered in 32 (59%) deliveries. Most women with FXI deficiency have uncomplicated pregnancies and deliveries with minimal hemostatic support. Neuraxial anesthesia can be safely administered in most women.

Creatine monohydrate in ALS: effects on strength, fatigue, respiratory status and ALSFRS.

Our objective was to determine the effect of creatine monohydrate on disease progression in patients with amyotrophic lateral sclerosis (ALS). One hundred and seven patients with the diagnosis of probable or definite ALS, of less than five years duration from symptom onset, were randomized to either treatment with daily creatine monohydrate (5 g/d) or placebo. In this multicenter, double-blinded study we followed changes in disease progression: using quantitative measures of strength via maximal isometric voluntary contraction, forced vital capacity, ALSFRS, quality of life, fatigue and survival. Patients were followed for nine months. The results showed that creatine monohydrate did not significantly improve motor, respiratory or functional capacity in this patient population. The drug was well tolerated and the study groups well balanced, especially considering the absence of forced vital capacity criteria for entrance into the study. There was a trend toward improved survival in patients taking daily creatine monohydrate and this was identical to the trend seen in another recently published report of creatine in ALS patients 1. In conclusion, creatine monohydrate (5 g/d) did not have an obvious benefit on the multiple markers of disease progression measured over nine months. We measured fatigue during isometric contraction and found no significant improvement despite anecdotal patient reports prior to and during the study. The trend toward improved survival was also found in another recently completed blinded trial using creatine monohydrate. Further investigation on the possible survival benefit of creatine in this patient population is ongoing.

A novel, efficient, randomized selection trial comparing combinations of drug therapy for ALS.

Combining agents with different mechanisms of action may be necessary for meaningful results in treating ALS. The combinations of minocycline-creatine and celecoxib-creatine have additive effects in the murine model. New trial designs are needed to efficiently screen the growing number of potential neuroprotective agents. Our objective was to assess two drug combinations in ALS using a novel phase II trial design. We conducted a randomized, double-blind selection trial in sequential pools of 60 patients. Participants received minocycline (100 mg)-creatine (10 g) twice daily or celecoxib (400 mg)-creatine (10 g) twice daily for six months. The primary objective was treatment selection based on which combination best slowed deterioration in the ALS Functional Rating Scale-Revised (ALSFRS-R); the trial could be stopped after one pool if the difference between the two arms was adequately large. At trial conclusion, each arm was compared to a historical control group in a futility analysis. Safety measures were also examined. After the first patient pool, the mean six-month decline in ALSFRS-R was 5.27 (SD=5.54) in the celecoxib-creatine group and 6.47 (SD=9.14) in the minocycline-creatine group. The corresponding decline was 5.82 (SD=6.77) in the historical controls. The difference between the two sample means exceeded the stopping criterion. The null hypothesis of superiority was not rejected in the futility analysis. Skin rash occurred more frequently in the celecoxib-creatine group. In conclusion, the celecoxib-creatine combination was selected as preferable to the minocycline-creatine combination for further evaluation. This phase II design was efficient, leading to treatment selection after just 60 patients, and can be used in other phase II trials to assess different agents.

Using multi-neuron population recordings for neural prosthetics.

Classical single-neuron recording methods led to 'neuron-centric' concepts of neural coding, whereas more recent multi-neuron population recordings have inspired 'population-centric' concepts of distributed processing in neural systems. Because most neocortical neurons code information coarsely, sensory or motor processing tends to be widely distributed across neuronal populations. Dynamic fluctuations in neural population functions thus involve subtle changes in the overall pattern of neural activity. Mathematical analysis of neural population codes allows extraction of 'motor signals' from neuronal population recordings in the motor cortices, which can then be used in real-time to directly control movement of a robot arm. This technique holds promise for the development of neurally controlled prosthetic devices and provides insights into how information is distributed across several brain regions.

Youth perceptions of their school violence risks.

In order to gauge youth perceptions of school violence, this study links two perceptual bias literatures: third-person perception and optimistic bias. The intersection of the two literatures may be especially beneficial in understanding how adolescents process and interpret public health messages and subsequently engage in risk behaviors or self-protective behaviors in health contexts. Participants were 350 urban adolescents in school-based violence prevention sessions who completed a survey. Findings indicate shared predictors of third-person perception and optimistic bias (age, self-esteem) as well as differences (knowledge). The findings also provide insight into understanding how adolescents process and interpret public health messages and subsequently engage in risk behaviors or self-protective behaviors in health contexts.

Von Willebrand disease in the elderly: clinical perspectives.

Von Willebrand disease (VWD) is an inherited bleeding disorder that affects up to 1% of the population. In most cases, VWD results from a mutation in the von Willebrand Factor (VWF) gene, which alters the amount and function of VWF, a key glycoprotein in both primary and secondary hemostasis. A comprehensive analysis of patients with VWD should include VWF activity, antigen levels, platelet function, and a careful bleeding history. Treatment options include antifibrinolytics, desmopressin, and VWF replacement therapy. VWF levels fluctuate due to age, stress, environmental exposures, and pharmacologic treatment. Treatment guidelines exist to treat and prevent bleeding for patients undergoing surgery and medical procedures, but often these must be reevaluated in the setting of age-related comorbidities including cardiovascular events, venous thrombosis, and malignancy. In addition, many age-related complications are associated with a secondary acquired von Willebrand syndrome (AVWS), including malignancies, hypothyroidism, cardiovascular diseases, and cardiac replacement devices. The current literature is limited by a lack of older patients in clinical trials. Larger studies are needed to determine if age-related comorbidities affect VWD patients at different frequencies than the general elderly population. There is also a significant need for registry-based studies to evaluate many age-related comorbidities in VWD patients.

Gene Therapy for Hemophilia: Progress to Date.

Hemophilia is a congenital bleeding disorder that affects nearly half a million individuals worldwide. Joint bleeding and other co-morbidities are a significant source of debilitation for this population. Current therapies are effective but must be given lifelong at regular intervals, are costly, and are available to only about 25% of the hemophilia population living in resource-rich countries. Gene therapy for hemophilia has been in development for three decades and is now entering pivotal-stage clinical trials. While many different technology platforms exist for gene therapy, all current clinical trials for hemophilia employ adeno-associated vector (AAV)-based cell transduction. This small viral particle is capable of packaging modified F8 or F9 transgenes, can be generated robustly from cell lines, and transduces several relatively end-differentiated target tissues such as the liver with high efficiency. While pre-existing neutralizing antibodies to the AAV capsid are recognized to limit current therapy, other challenges have been identified in human studies that were not seen in preclinical studies. Both liver transaminase elevations and immune-mediated loss of transgene expression have been observed in clinical trials. Toll-like receptors, cytotoxic T cells, and other components of the immune response have been implicated in the loss of factor expression, but a full understanding of the immune response awaits clarification. Despite these challenges, many patients enrolled in gene therapy trials have attained long-term expression of factors VIII and IX. This emerging technology now represents a cure for the severe bleeding and joint damage associated with hemophilia.