The epitopes defined by HIV-1 broadly neutralizing antibodies (bNAbs) are valuable templates for vaccine design, and studies of the immunological development of these antibodies are providing ...insights for vaccination strategies. In addition, the most potent and broadly reactive of these bNAbs have potential for clinical use. We previously described a family of 12 V1V2-directed neutralizing antibodies, CAP256-VRC26, isolated from an HIV-1 clade C-infected donor at years 1, 2, and 4 of infection (N. A. Doria-Rose et al., Nature 509:55-62, 2014, http://dx.doi.org/10.1038/nature13036). Here, we report on the isolation and characterization of new members of the family mostly obtained at time points of peak serum neutralization breadth and potency. Thirteen antibodies were isolated from B cell culture, and eight were isolated using trimeric envelope probes for differential single B cell sorting. One of the new antibodies displayed a 10-fold greater neutralization potency than previously published lineage members. This antibody, CAP256-VRC26.25, neutralized 57% of diverse clade viral isolates and 70% of clade C isolates with remarkable potency. Among the viruses neutralized, the median 50% inhibitory concentration was 0.001 μg/ml. All 33 lineage members targeted a quaternary epitope focused on V2. While all known bNAbs targeting the V1V2 region interact with the N160 glycan, the CAP256-VRC26 antibodies showed an inverse correlation of neutralization potency with dependence on this glycan. Overall, our results highlight the ongoing evolution within a single antibody lineage and describe more potent and broadly neutralizing members with potential clinical utility, particularly in areas where clade C is prevalent.
Studies of HIV-1 broadly neutralizing antibodies (bNAbs) provide valuable information for vaccine design, and the most potent and broadly reactive of these bNAbs have potential for clinical use. We previously described a family of V1V2-directed neutralizing antibodies from an HIV-1 clade C-infected donor. Here, we report on the isolation and characterization of new members of the family mostly obtained at time points of peak serum neutralization breadth and potency. One of the new antibodies, CAP256-VRC26.25, displayed a 10-fold greater neutralization potency than previously described lineage members. It neutralized 57% of diverse clade viral isolates and 70% of clade C isolates with remarkable potency: the median 50% inhibitory concentration was 0.001 μg/ml. Our results highlight the ongoing evolution within a single antibody lineage and describe more potent and broadly neutralizing members with potential clinical utility, particularly in areas where clade C is prevalent.
Malaria continues to impose a global health burden. Drug-resistant parasites have emerged to each introduced small-molecule therapy, highlighting the need for novel treatment approaches for the ...future eradication of malaria. Herein, targeted drug delivery with peptide–drug conjugates (PDCs) was investigated as an alternative antimalarial therapy, inspired by the success of emerging antibody–drug conjugates utilized in cancer treatment. A synthetic peptide derived from an innate human defense molecule was conjugated to the antimalarial drug primaquine (PQ) to produce PDCs with low micromolar potency toward Plasmodium falciparum in vitro. A suite of PDCs with different design features was developed to identify optimal conjugation site and investigate linker length, hydrophilicity, and cleavability. Conjugation within a flexible spacer region of the peptide, with a cleavable linker to liberate the PQ cargo, was important to retain activity of the peptide and drug.
Human mitochondrial RNase P (mt-RNase P) is responsible for 5′ end processing of mitochondrial precursor tRNAs, a vital step in mitochondrial RNA maturation, and is comprised of three protein ...subunits: TRMT10C, SDR5C1 (HSD10), and PRORP. Pathogenic variants in TRMT10C and SDR5C1 are associated with distinct recessive or x-linked infantile onset disorders, resulting from defects in mitochondrial RNA processing. We report four unrelated families with multisystem disease associated with bi-allelic variants in PRORP, the metallonuclease subunit of mt-RNase P. Affected individuals presented with variable phenotypes comprising sensorineural hearing loss, primary ovarian insufficiency, developmental delay, and brain white matter changes. Fibroblasts from affected individuals in two families demonstrated decreased steady state levels of PRORP, an accumulation of unprocessed mitochondrial transcripts, and decreased steady state levels of mitochondrial-encoded proteins, which were rescued by introduction of the wild-type PRORP cDNA. In mt-tRNA processing assays performed with recombinant mt-RNase P proteins, the disease-associated variants resulted in diminished mitochondrial tRNA processing. Identification of disease-causing variants in PRORP indicates that pathogenic variants in all three subunits of mt-RNase P can cause mitochondrial dysfunction, each with distinct pleiotropic clinical presentations.
Pseudomonas aeruginosa is a ubiquitous organism that is the focus of intense research because of its prominent role in disease. Due to its relatively large genome and flexible metabolic capabilities, ...this organism exploits numerous environmental niches. It is an opportunistic pathogen that sets upon the human host when the normal immune defenses are disabled. Its deadliness is most apparent in cystic fibrosis patients, but it also is a major problem in burn wounds, chronic wounds, chronic obstructive pulmonary disorder, surface growth on implanted biomaterials, and within hospital surface and water supplies, where it poses a host of threats to vulnerable patients (Peleg and Hooper, N Engl J Med 362:1804–1813, 2010; Breathnach et al., J Hosp Infect 82:19–24, 2012). Once established in the patient, P. aeruginosa can be especially difficult to treat. The genome encodes a host of resistance genes, including multidrug efflux pumps (Poole, J Mol Microbiol Biotechnol 3:255–264, 2001) and enzymes conferring resistance to beta-lactam and aminoglycoside antibotics (Vahdani et al., Annal Burns Fire Disast 25:78–81, 2012), making therapy against this gram-negative pathogen particularly challenging due to the lack of novel antimicrobial therapeutics (Lewis, Nature 485: 439–440, 2012). This challenge is compounded by the ability of P. aeruginosa to grow in a biofilm, which may enhance its ability to cause infections by protecting bacteria from host defenses and chemotherapy. Here, we review recent studies of P. aeruginosa biofilms with a focus on how this unique mode of growth contributes to its ability to cause recalcitrant infections.
A worldwide estimate of over one million STIs are acquired daily and there is a desperate need for effective preventive as well as therapeutic measures to curtail this global health burden. Vaccines ...have been the most effective means for the control and potential eradication of infectious diseases; however, the development of vaccines against STIs has been a daunting task requiring extensive research for the development of safe and efficacious formulations. Nanoparticle-based vaccines represent a promising platform as they offer benefits such as targeted antigen presentation and delivery, co-localized antigen-adjuvant combinations for enhanced immunogenicity, and can be designed to be biologically inert. Here we discuss promising types of nanoparticles along with outcomes from nanoparticle-based vaccine preclinical studies against non-viral STIs including chlamydia, syphilis, gonorrhea, and recommendations for future nanoparticle-based vaccines against STIs.
Signaling pathways targeting mitochondria are poorly understood. We here examine phosphorylation by the cAMP-dependent pathway of subunits of cytochrome c oxidase (COX), the terminal enzyme of the ...electron transport chain. Using anti-phospho antibodies, we show that cow liver COX subunit I is tyrosinephosphorylated in the presence of theophylline, a phosphodiesterase inhibitor that creates high cAMP levels, but not in its absence. The site of phosphorylation, identified by mass spectrometry, is tyrosine 304 of COX catalytic subunit I. Subunit I phosphorylation leads to a decrease of Vmax and an increase of Km for cytochrome c and shifts the reaction kinetics from hyperbolic to sigmoidal such that COX is fully or strongly inhibited up to 10 μm cytochrome c substrate concentrations, even in the presence of allosteric activator ADP. To assess our findings with the isolated enzyme in a physiological context, we tested the starvation signal glucagon on human HepG2 cells and cow liver tissue. Glucagon leads to COX inactivation, an effect also observed after incubation with adenylyl cyclase activator forskolin. Thus, the glucagon receptor/G-protein/cAMP pathway regulates COX activity. At therapeutic concentrations used for asthma relief, theophylline causes lung COX inhibition and decreases cellular ATP levels, suggesting a mechanism for its clinical action.
Abstract Covalent organic frameworks (COFs) are an attractive class of materials for sensing applications due to their inherent crystallinity, high surface area, and designable framework ...functionalities. While the majority of COFs have high electrical resistance, making it difficult to harness these materials for electronic sensors, real‐time impedance spectroscopy can enable gas and vapor sensing of a much wider range of COFs. Herein, a set of post‐synthetically modified (PSM) COFs are explored though a straightforward one‐step substitution reaction by reacting phenol moieties on the pore wall, with alkyl bromides, to embed alkyl, aryl, or alcohol groups into the framework. This modular approach provides access to improved sensor properties toward the detection of volatile organics, acids, and potentially harmful gases. Sensor results indicate that post‐synthetically modified COFs offer better sensitivity toward NO 2 and acetic acid, with the aryl functionalized COF having an NO 2 detection limit of 10 ppm. Furthermore, modified COFs also show higher selectivity toward isopropanol and toluene. This work highlights the importance of methods that facilitate post‐synthetic modification of COFs so that functional groups and COF properties can be tuned.
CASK is a multi-domain scaffolding protein that interacts with the transcription factor TBR1 and regulates expression of genes involved in cortical development such as RELN. Here we describe a ...previously unreported X-linked brain malformation syndrome caused by mutations of CASK. All five affected individuals with CASK mutations had congenital or postnatal microcephaly, disproportionate brainstem and cerebellar hypoplasia, and severe mental retardation.
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