Background Given the high mortality rate for those with end-stage kidney d sease on dialysis and the efficacy and safety of current hepatitis C virus (HCV) treatments, currently-discarded kidneys from HCV-infected (HCV+) donors may be a neglected public health resource. Objective To determine the tolerability and feasibility of kidney transplantation (KT) from HCV+ donors to HCV-uninfected recipients (HCV D+/R−) in combination with direct-acting antivirals (DAAs) as pre- and post-transplant prophylaxis. Design Open-label, non-randomized trial. (ClinicalTrials.gov: NCT02781649) Setting Single-center. Participants 10 HCV-uninfected KT candidates over the age of 50 years with no available living donors. Intervention KT from deceased donors ages 13–50 years with a positive HCV RNA and HCV antibody test. All recipients received a dose of grazoprevir 100 mg/elbasvir 50 mg (GZR/EBR) immediately prior to transplant. For genotype 1 donors, recipients continued GZR/EBR for 12 weeks post-transplant; for genotype 2 or 3 donors, sofosbuvir 400 mg was added to GZR/EBR for 12 weeks of triple-therapy. Measurements The primary safety outcome was the incidence of adverse events related to GZR-EBR. The primary efficacy outcome was the proportion recipients with HCV RNA less than the lower limit of quantification 12 weeks after prophylaxis. Results Among 10 HCV D+/R− there were no treatment-related adverse events and HCV RNA was not detected in any recipient 12 weeks after treatment. Limitations Nonrandomized study design and small number of patients. Conclusions Pre- and post-transplant HCV treatment was safe and prevented chronic hepatitis C in HCV D+/R− KT. If confirmed in larger studies, this strategy should markedly expand organ options and reduce mortality for HCV− KT candidates.

Although late cases occur, the first year after SOT is the period of highest risk for histoplasmosis. In patients who survive the first month after diagnosis, treatment with an amphotericin formulation followed by an azole for 12 months is usually successful, with only rare relapse.

SummaryBackgroundInfluenza causes significant morbidity and mortality despite currently available treatments. Anecdotal reports suggest plasma with high antibody titers towards influenza may be of benefit in the treatment of severe influenza.MethodsWe conducted a randomized, open-label, multicenter phase 2 trial at 29 academic medical centers in the United States to assess the safety and efficacy of anti-influenza plasma with hemagglutination inhibition (HAI) antibody titers of ≥ 1:80 to the infecting strain. Hospitalized children and adults (including pregnant women) with severe influenza A or B (defined as hypoxia or tachypnea) were randomly assigned to receive either 2 units (or pediatric equivalent) of anti-influenza plasma plus standard care (P+S), versus standard care alone (S), and were followed for 28 days. The primary endpoint was time to normalization of patients’ respiratory status (respiratory rate of ≤ 20 for adults or age defined thresholds of 20–38 for children), and a room air saturation of oxygen ≥ 93%. ClinicalTrials.gov Identifier: NCT01052480FindingsBetween January 13, 2011 and March 2, 2015, 113 participants were screened, and 98 were randomized. Of the participants with confirmed influenza, 28 of 42 (67%) of P+S participants normalized their respiratory status by Day 28, as compared to 24 of 45 (53%) of S participants (p=0·069). The estimated hazard ratio comparing P+S to S was 1·71 (95% CI: 0·96 to 3·06). Six participants died, 1 (2%) and 5 (10%) from the P+S and S arms respectively (p=0·093). P+S participants had non-significant reductions in days in hospital (median 6 vs. 11 days, p=0·13) and days on mechanical ventilation (median 0 vs. 3 days, p=0·14), and significantly improved clinical status at Day 7 (p=0·020). Fewer P+S participants experienced SAEs compared to S recipients (20% vs. 38%, p= 0·041), the most frequent of which were acute respiratory distress syndrome (1 [2%] vs 2 [4%]) and stroke (1 [2%] vs 2 [4%]).InterpretationResults from this Phase II randomized trial of immune plasma for the treatment of severe influenza provides support for a possible benefit of immunotherapy across the primary and secondary endpoints. A Phase III randomized trial is now underway to further evaluate this intervention.

Diacylglycerol pyrophosphate (DGPP) is involved in a putative novel lipid signaling pathway. DGPP phosphatase (DGPP phosphohydrolase) is a membrane-associated 34-kDa enzyme from Saccharomyces cerevisiae which catalyzes the dephosphorylation of DGPP to yield phosphatidate (PA) and then catalyzes the dephosphorylation of PA to yield diacylglycerol. Amino acid sequence information derived from DGPP phosphatase was used to identify and isolate the DPP1 (diacylglycerol pyrophosphate phosphatase) gene encoding the enzyme. Multicopy plasmids containing the DPP1 gene directed a 10-fold overexpression of DGPP phosphatase activity in S. cerevisiae. The heterologous expression of the S. cerevisiae DPP1 gene in Sf-9 insect cells resulted in a 500-fold overexpression of DGPP phosphatase activity over that expressed in wild-type S. cerevisiae. DGPP phosphatase possesses a Mg 2؉ -independent PA phosphatase activity, and its expression correlated with the overexpression of DGPP phosphatase activity in S. cerevisiae and in insect cells. DGPP phosphatase was predicted to be an integral membrane protein with six transmembrane-spanning domains. The enzyme contains a novel phosphatase sequence motif found in a superfamily of phosphatases. A dpp1⌬ mutant was constructed by deletion of the chromosomal copy of the DPP1 gene. The dpp1⌬ mutant was viable and did not exhibit any obvious growth defects. The mutant was devoid of DGPP phosphatase activity and accumulated (4-fold) DGPP. Analysis of the mutant showed that the DPP1 gene was not responsible for all of the Mg 2؉ -independent PA phosphatase activity in S. cerevisiae. Diacylglycerol pyrophosphate (DGPP)1 is a novel phospholipid that contains a pyrophosphate group attached to diacylglycerol (DG) (Fig. 1) (1). DGPP has been found in a variety of plants (2, 3) and in the yeast Saccharomyces cerevisiae (4). This phospholipid is synthesized from phosphatidate (PA) and ATP via the reaction catalyzed by the membrane-associated enzyme PA kinase (1) and is dephosphorylated to PA via the reaction catalyzed by the membrane-associated enzyme DGPP phosphatase ( Fig. 1) (4). The amounts of DGPP in wild-type S. cerevisiae and in plants are barely detectable (3, 4). For example, DGPP accounts for only 0.18 mol % of the major phospholipids in S. cerevisiae (4). The low abundance of DGPP is reminiscent of lipid signaling molecules such as the inositol-containing phospholipids (5-9). Recent studies indicate that the metabolism of DGPP is involved in a novel lipid signaling pathway. DGPP accumulates in plant tissues upon G protein activation through the stimulation of PA kinase activity (3), and metabolic labeling studies with Catharanthus roseus cells have shown that DGPP is metabolized rapidly to PA and then to DG (10). It has been suggested that DGPP may function as a signaling molecule (3, 4). Alternatively, the formation of DGPP may serve to attenuate the signaling functions of PA (11,12).DGPP phosphatase activity has been identified in S. cerevisiae, C. roseus, Escherichia coli, rat liver, pig l...

CTP synthetase (EC 6.3.4.2, UTP:ammonia ligase (ADP-forming)) activity in Saccharomyces cerevisiae is allosterically regulated by CTP product inhibition. Amino acid residue Glu 161 in the URA7-encoded and URA8-encoded CTP synthetases was identified as being involved in the regulation of these enzymes by CTP product inhibition. The specific activities of the URA7-encoded and URA8-encoded enzymes with a Glu 161 3 Lys (E161K) mutation were 2-fold greater when compared with the wild-type enzymes. The E161K mutant URA7-encoded and URA8-encoded CTP synthetases were less sensitive to CTP product inhibition with inhibitor constants for CTP of 8.4-and 5-fold greater, respectively, than those of their wild-type counterparts. Cells expressing the E161K mutant enzymes on a multicopy plasmid exhibited an increase in resistance to the pyrimidine poison and cancer therapeutic drug cyclopentenylcytosine and accumulated elevated (6 -15-fold) levels of CTP when compared with cells expressing the wild-type enzymes. Cells expressing the E161K mutation in the URA7-encoded CTP synthetase exhibited an increase (1.5-fold) in the utilization of the Kennedy pathway for phosphatidylcholine synthesis when compared with control cells. Cells bearing the mutation also exhibited an increase in the synthesis of phosphatidylcholine (1.5-fold), phosphatidylethanolamine (1.3-fold), and phosphatidate (2-fold) and a decrease in the synthesis of phosphatidylserine (1.7-fold). These alterations were accompanied by an inositol excretion phenotype due to the misregulation of the INO1 gene. Moreover, cells bearing the E161K mutation exhibited an increase (1.6-fold) in the ratio of total neutral lipids to phospholipids, an increase in triacylglycerol (1.4-fold), free fatty acids (1.7-fold), and ergosterol ester (1.8-fold), and a decrease in diacylglycerol (1.3-fold) when compared with control cells. These data indicated that the regulation of CTP synthetase activity by CTP plays an important role in the regulation of phospholipid synthesis.CTP synthetase (EC 6.3.4.2, UTP:ammonia ligase (ADPforming)) is a cytosolic-associated glutamine amidotransferase that catalyzes the ATP-dependent transfer of the amide nitrogen from glutamine to the C-4 position of UTP to form CTP (1, 2). This enzyme plays an essential role in the synthesis of all membrane phospholipids in eukaryotic cells (3, 4). Its reaction product CTP is the direct precursor of the activated, energyrich phospholipid pathway intermediates CDP-DG 1 (5), CDPcholine (6), and CDP-ethanolamine (6) (Fig. 1). CDP-DG is the source of the phosphatidyl moiety of PS, PE, and PC synthesized by the CDP-DG pathway as well as PI, phosphatidylglycerol, and cardiolipin (3, 4). CDP-choline and CDP-ethanolamine are the sources of the hydrophilic head groups of PC and PE synthesized by the Kennedy pathways, respectively (3, 4). Our laboratory utilizes the yeast Saccharomyces cerevisiae as a model eukaryote to study the regulation of CTP synthetase and its impact on phospholipid metabolism. CTP synthetase is encoded by ...

Background COVID-19 associated pulmonary aspergillosis (CAPA) occurs in critically ill COVID-19 patients. Risks and outcomes remain poorly understood. Methods A retrospective cohort study of adult mechanically ventilated COVID-19 patients admitted to five Johns Hopkins hospitals was conducted between March and August 2020. CAPA was defined using composite clinical criteria. Fine and Gray competing risks regression was used to analyze clinical outcomes and multilevel mixed-effects ordinal logistic regression was used to compare longitudinal disease severity scores. Results Amongst the cohort of 396 people, 39 met criteria for CAPA. Compared to those without, patients with CAPA were more likely to have underlying pulmonary vascular disease (41% vs 21.6%, p=0.01), liver disease (35.9% vs 18.2%, p=0.02), coagulopathy (51.3% vs 33.1%, p=0.03), solid tumors (25.6% vs 10.9%, p=0.017), multiple myeloma (5.1% vs 0.3%, p=0.027), corticosteroid exposure during index admission (66.7% vs 42.6%, p=0.005), and had a lower BMI (median 26.6 vs 29.9, p=0.04). People with CAPA had worse outcomes as measured by ordinal severity of disease scores, requiring longer time to improvement (adjusted odds ratio 1.081.091.1, p<0.001), and advancing in severity almost twice as fast (subhazard ratio, sHR 1.31.82.5, p<0.001). People with CAPA were intubated twice as long as those without (sHR) 0.40.50.6, p<0.001) and had a longer hospital length of stay [median (IQR) 41.1 (20.5, 72.4) vs 18.5 (10.7, 31.8), p<0.001]. Conclusion CAPA is associated with poor outcomes. Attention towards preventative measures (screening and/or prophylaxis) is warranted in people with high risk of developing CAPA.

Background Antiviral-resistant or refractory CMV infection is challenging, and salvage therapies, foscarnet and cidofovir, have significant toxicities. Several investigational anti-CMV agents are under development, but more information is needed on outcomes of current treatments, to facilitate clinical trial design for new drugs. Methods Records of solid organ (SOT) and hematopoietic cell transplant (HCT) recipients at a single center over a 10-year period were reviewed retrospectively to characterize those who had received foscarnet treatment for ganciclovir-resistant or refractory CMV infection. Data were collected on virologic responses, mortality, and nephrotoxicity. Results Of 39 patients (22 SOT, 17 HCT), 15 had documented ganciclovir resistance mutations and 11/39 (28%) had tissue-invasive CMV. Median duration of foscarnet was 32 days. Virologic failure occurred in 13/39 (33%) and relapses of viremia occurred in 31%. Mortality was 12/39 (31%) and was higher in HCT than SOT (p=0.001), although ganciclovir resistance was more common in SOT (p = 0.003). Doses of ganciclovir or valganciclovir were low in 10/39 (26%) at some time prior to switching to foscarnet. Renal dysfunction occurred in 20/39 (51%) by end of treatment and in 7/25 (28%) after 6 months. Conclusions Outcomes of existing treatment for ganciclovir-resistant or refractory CMV are suboptimal, in terms of virologic clearance, renal dysfunction, and mortality. These data should provide background information for future clinical trials of newer antiviral agents.

Background The epidemiology of invasive mold infections (IMI) in transplant recipients differs, based on geography, hosts, preventative strategies, and methods of diagnosis. Methods We conducted a retrospective observational study to evaluate the epidemiology of proven and probable IMI, using prior definitions, among all adult hematopoietic stem cell (HSCT) and solid organ transplant (SOT) recipients in the era of “classic” culture-based diagnostics (2000–2009). Epidemiology was evaluated before and after an initiative was begun to increase bronchoscopy in HSCT recipients after 2005. Results In total, 106 patients with one IMI were identified. Invasive aspergillosis (IA) was the most common IMI (69; 65.1%), followed by mucormycosis (9; 8.5%). The overall rate of IMI (and IA) was 3.5% (2.5%) in allogeneic HSCT recipients. The overall incidence for IMI among lung, kidney, liver, and heart transplant recipients were 49, 2, 11, and 10 per 1000 person-years, respectively. The observed rate of IMI among HLA-matched unrelated and haploidentical HSCT recipients increased from 0.6% annually to 3.0% after bronchoscopy initiation (P<0.05). The 12-week mortality among allogeneic HSCT, liver, kidney, heart, and lung recipients with IMI was 52.4%, 47.1%, 27.8%, 16.7%, and 9.5%, respectively. Among allogeneic HSCT (odds ratio [OR]: 0.07, P=0.007) and SOT (OR: 0.22, P=0.05) recipients with IA, normal platelet count was associated with improved survival. Male gender (OR: 14.4, P=0.007) and elevated bilirubin (OR: 5.7, P=0.04) were significant predictors of mortality for allogeneic HSCT and SOT recipients with IA, respectively. Conclusions During the era of culture-based diagnostics, observed rates of IMI were low among all transplants except lung transplant recipients, with relatively higher mortality rates. Diagnostic aggressiveness and host variables impact the reported incidence and outcome of IMI and likely account for institutional variability in multicenter studies. Definitions to standardize diagnoses among SOT recipients are needed.