Introducing Lu-1, a Novel Lactobacillus jensenii Phage Abundant in the Urogenital Tract

Bacteriophages (phages) play a key role in shaping microbial communities, including those of the human body. Phages are abundant members of the urogenital tract, most often persisting through the lysogenic life cycle as prophages integrated within the genomes of their bacterial hosts. While numerous studies of the urogenital microbiota have focused on the most abundant bacterial member of this niche–Lactobacillus species–very little is known about Lactobacillus phages. Focusing on Lactobacillus jensenii strains from the urinary tract, we identified numerous prophages related to the previously characterized Lv-1 phage from a vaginal L. jensenii strain. Furthermore, we identified a new L. jensenii phage, Lu-1. Evidence suggests that both phages are abundant within the urogenital tract. CRISPR spacer sequences matching to Lv-1 and Lu-1 prophages were identified. While first detected in urinary isolates, the Lu-1 phage was also discovered in L. jensenii isolates from vaginal and perineal swabs, and both phages were found in metagenomic data sets. The prevalence of these phages in the isolates suggests that both phages are active members of the urogenital microbiota

Development of Clinical Recommendations for Progressive Return to Activity After Military Mild Traumatic Brain Injury: Guidance for Rehabilitation Providers

Objective: Previously published mild traumatic brain injury (mTBI) management guidelines provide very general recommendations to return individuals with mTBI to activity. This lack of specific guidance creates variation in military rehabilitation. The Office of the Army Surgeon General in collaboration with the Defense and Veterans Brain Injury Center, a component center of the Defense Centers of Excellence for Psychological Health and Traumatic Brain Injury, convened an expert working group to review the existing literature and propose clinical recommendations that standardize rehabilitation activity progression following mTBI

Factor IX Expression within the Normal Range Prevents Spontaneous Bleeds Requiring Treatment Following FLT180a Gene Therapy in Patients with Severe Hemophilia B: Long-Term Follow- up Study of the B-Amaze Program

Introduction: FLT180a (verbrinacogene setparvovec) is an investigational, liver-directed AAV gene therapy for the treatment of patients with hemophilia B (HB). FLT180a consists of a novel, potent, engineered capsid (AAVS3) containing an expression cassette encoding a Factor IX (FIX) gain-of-function protein variant ('Padua'; FIX-R338L). The B-AMAZE study was designed to identify a dose of FLT180a that maintains FIX activity within the normal range (50-150%) and thereby protect patients with severe HB from spontaneous and traumatic bleeds. Methods: B-AMAZE was a multicentre, open-label Phase 1/2 clinical trial (NCT03369444; sponsored by UCL) that evaluated FLT180a dose levels using an escalating/descending adaptive design in patients with severe (FIX activity <1%) or moderately severe (FIX activity 1-2%) HB who were negative for AAVS3 neutralizing antibodies. A novel regimen of prophylactic corticosteroids with/without tacrolimus was implemented to mitigate the impact of vector-related transaminitis on FIX expression. Patients who completed the 26-week B-AMAZE study were eligible for the ongoing long-term follow-up study (NCT03641703; sponsored by Freeline). Results: Ten HB patients received a single dose of FLT180a. Four FLT180a doses ranging from 3.84e11 vg/kg to 1.28e12 vg/kg were assessed. As of the data cut-off date, all patients have been followed for ≥16 months. FLT180a demonstrated a favorable safety profile, without evidence of inhibitors against FIX, infusion-related or allergic reactions. The most common treatment-related adverse event was transient elevation in alanine aminotransferase. An event of AV fistula thrombosis occurred in a 67-year-old patient who received the highest dose of 1.28e12 vg/kg (total dose of 1.15e14 vg) and had supranormal FIX levels; this patient was treated with anticoagulants. While these FIX levels demonstrate the potency of our proprietary AAVS3 capsid, this dose will not be used in future hemophilia studies. At Week 26 after FLT180a administration, a dose-response relationship was observed with mean FIX activity of 45.0%, 35.5%, 141.5%, and 175.5% for 3.84e11, 6.4e11, 8.32e11, and 1.28e12 vg/kg doses, respectively (Table); FIX activity levels ≥50% were achieved in 7 of 8 patients treated with the three highest doses. One patient (Patient 4) who received 6.4e11 vg/kg lost transgene expression early due to transaminitis and resumed routine factor prophylaxis. The 8.32e11 vg/kg cohort received an extended immune management regimen (9-18 weeks) with prophylactic tacrolimus in addition to prednisolone to prevent breakthrough vector-related transaminitis. However, after cessation of the immune management regimen, transaminitis with concomitant reductions in FIX activity was observed in all patients in the 8.32e11 vg/kg cohort. The combination of prophylactic tacrolimus and prednisolone appeared to have suppressed immune-mediated transaminitis while administered, but recurrence of transaminitis developed soon after cessation. This unique and previously unreported observation suggests that the longer-duration prophylactic immune management regimen may have prevented tolerization to the vector because this was not observed in earlier cohorts where a brief course of tacrolimus was given reactively for breakthrough transaminitis. All patients (including the 8.32e11 vg/kg cohort) have achieved steady state. Patients in the earliest cohort who received the lowest dose (3.84e11 vg/kg) have shown stable FIX activity for >3 years. There were no spontaneous bleeds that required FIX supplementation in patients who maintained FIX activity above 50%; Patient 4 in the 6.4e11 vg/kg cohort experienced two bleeds (cause unknown) after he lost transgene expression, which were treated with exogenous FIX. One patient received exogenous FIX for treatment of a traumatic bleed, but his FIX activity level was 57% at the time of the event. Additional efficacy and safety results with >3.5 years of follow-up will be presented. Conclusions: B-AMAZE is the first HB gene therapy study to achieve normal levels of FIX activity using relatively low vector doses. Results suggest that a dose of 7.7e11 vg/kg, coupled with a short course of prophylactic immune management, has the potential to achieve durable FIX activity in the normal range (50-150%) and thereby prevent spontaneous bleeds and normalize hemostasis in the event of traumatic bleeds

Pancreas and islet cell transplantation

Currently, for the patient with type 1 diabetes, a definitive treatment without resorting to the use of exogenous insulin can be achieved only with pancreas or islet cell transplantation. These means of restoring β-cell mass can completely maintain essentially normal long-term glucose homeostasis, although the need for powerful immunosuppressive regimens limits their application to only a subgroup of adult patients. Apart from the shortage of donors that has limited all kinds of transplantation, however, the widespread use of β-cell replacement has been precluded until recently by the drawbacks associated with both organ and islet cell transplantation. Although the study of recurrence of diabetes has generated attention, the fundamental obstacle to pancreas and islet transplantation has been, and remains, the alloimmune response. With a better elucidation of the mechanisms of alloengraftment achieved during the last 3 years, the stage has been set for further advances

Current status of islet xenotransplantation

Cell therapy for Type 1 diabetes (T1D) utilizing islet cell transplantation can successfully restore endogenous insulin production in affected patients. Islet cell engraftment and survival are conditional on the use of efficacious anti-rejection therapies and on the availability of healthy donor cells. The scarcity of healthy human donor pancreata is a limiting factor in providing sufficient tissue to meet the demand for islet transplantation worldwide. A potential alternative to the use of cadaveric human donor pancreases is the use of animal sourced islets.Pancreatic islets obtained from pigs have emerged as an alternative to human tissues due to their great availability, physiological similarities to human islets, including the time-tested use of porcine insulin in diabetic patients and the ability to genetically modify the donor source.The evolution of refined, efficacious immunosuppressive therapies with reduced toxicity, improvements in donor management and genetic manipulation of the donor have all contributed to facilitate long-term function in pre-clinical models of pig islet grafts in non-human primates.As clinical consideration for this option is growing, and trials involving the use of porcine islets have begun, more compelling experimental data suggest that the use of pig islets may soon become a viable, safe, effective and readily available treatment for insulin deficiency in T1D patients

Pig-to-Nonhuman Primates Pancreatic Islet Xenotransplantation: An Overview

The therapy of type 1 diabetes is an open challenging problem. The restoration of normoglycemia and insulin independence in immunosuppressed type 1 diabetic recipients of islet allotransplantation has shown the potential of a cell-based diabetes therapy. Even if successful, this approach poses a problem of scarce tissue supply. Xenotransplantation can be the answer to this limited donor availability and, among possible candidate tissues for xenotransplantation, porcine islets are the closest to a future clinical application. Xenotransplantation, with pigs as donors, offers the possibility of using healthy, living, and genetically modified islets from pathogen-free animals available in unlimited number of islets. Several studies in the pig-to-nonhuman primate model demonstrated the feasibility of successful preclinical islet xenotransplantation and have provided insights into the critical events and possible mechanisms of immune recognition and rejection of xenogeneic islet grafts. Particularly promising results in the achievement of prolonged insulin independence were obtained with newly developed, genetically modified pigs islets able to produce immunoregulatory products, using different implantation sites, and new immunotherapeutic strategies. Nonetheless, further efforts are needed to generate additional safety and efficacy data in nonhuman primate models to safely translate these findings into the clinic

Genomic Survey of E. coli From the Bladders of Women With and Without Lower Urinary Tract Symptoms

Urinary tract infections (UTIs) are one of the most common human bacterial infections. While UTIs are commonly associated with colonization by Escherichia coli, members of this species also have been found within the bladder of individuals with no lower urinary tract symptoms (no LUTS), also known as asymptomatic bacteriuria. Prior studies have found that both uropathogenic E. coli (UPEC) strains and E. coli isolates that are not associated with UTIs encode for virulence factors. Thus, the reason(s) why E. coli sometimes causes UTI-like symptoms remain(s) elusive. In this study, the genomes of 66 E. coli isolates from adult female bladders were sequenced. These isolates were collected from four cohorts, including women: (1) without lower urinary tract symptoms, (2) overactive bladder symptoms, (3) urgency urinary incontinence, and (4) a clinical diagnosis of UTI. Comparative genomic analyses were conducted, including core and accessory genome analyses, virulence and motility gene analyses, and antibiotic resistance prediction and testing. We found that the genomic content of these 66 E. coli isolates does not correspond with the participant’s symptom status. We thus looked beyond the E. coli genomes to the composition of the entire urobiome and found that the presence of E. coli alone was not sufficient to distinguish between the urobiomes of individuals with UTI and those with no LUTS. Because E. coli presence, abundance, and genomic content appear to be weak predictors of UTI status, we hypothesize that UTI symptoms associated with detection of E. coli are more likely the result of urobiome composition