Profectus Biosciences (United States)

Somatic copy number alterations (sCNAs) drive cancer initiation, progression, resistance, and metastasis. Furthering our understanding of sCNAs requires substantially larger cohorts. Most tumors available for sequencing are preserved with formalin-fixed, paraffin-embedding (FFPE), which causes DNA cross-linking that distorts coverage profiles and challenges current sCNA estimation methods. Traditional methods denoise data using large panels of similar normal samples, which are impractical to obtain for FFPE cohorts. Here, HapASeg overcomes this limitation by leveraging haplotype phasing and unique covariates to accurately estimate sCNA segments across FFPE, fresh frozen, whole genome sequencing and whole exome sequencing sample types, outperforming current methods without requiring panel-of-normal correction.

BackgroundThe accumulation and aggregation of amyloid beta (Abeta)—a peptide fragment derived from the proteolytic processing of amyloid precursor protein (APP)—is a central pathological feature of Alzheimer’s disease (AD) and a current target for disease-modifying therapies. Mutations in APP can also drive early-onset AD. While the roles of alpha-, beta-, and gamma-secretases and their respective cleavage sites in APP processing are well characterized, much less is understood about the routine degradation of APP within sub-cellular compartments like the lysosome.MethodsWe applied Multiplexed Substrate Profiling by Mass Spectrometry (MSP-MS) to map cleavage sites within APP that may be targeted by lysosomal proteases, also known as cathepsins. We then employed cell-based and in vitro assays to examine the degradation of both wild-type and mutant APP by these enzymes.ResultsOur findings confirm that APP is enriched in the endo-lysosomal compartment, where it is processed by many cathepsins. Our experiments reveal that cleavages at several mapped APP sites are sensitive to both changes in pH and the presence of pathogenic variants E693G and E693Q. Additionally, we discovered that the large soluble domain of APP (sAPP) enhances tau cleavage by a specific cathepsin, CTSG, in vitro.ConclusionsCollectively, these results underscore the importance of lysosomal processing of APP, identify a link between APP and tau, and suggest new avenues for exploring AD pathogenesis. They also highlight potential therapeutic targets related to the lysosomal function of APP and its impact on neurodegenerative diseases.

Postexposure immunization can prevent disease and reduce transmission following pathogen exposure. The rapid immunostimulatory properties of recombinant vesicular stomatitis virus (rVSV)-based vaccines make them suitable postexposure treatments against the filoviruses Ebola virus and Marburg virus (MARV); however, the mechanisms that drive this protection are undefined. Previously, we reported 60–75% survival of rhesus macaques treated with rVSV vectors expressing MARV glycoprotein (GP) 20–30 minutes after a low dose exposure to the most pathogenic variant of MARV, Angola. Survival in this model was linked to production of GP-specific antibodies and lower viral load. To confirm these results and potentially identify novel correlates of postexposure protection, we performed a similar experiment, but analyzed plasma cytokine levels, frequencies of immune cell subsets, and the transcriptional response to infection in peripheral blood. In surviving macaques (80–89%), we observed induction of genes mapping to antiviral and interferon-related pathways early after treatment and a higher percentage of T helper 1 (Th1) and NK cells. In contrast, the response of non-surviving macaques was characterized by hypercytokinemia; a T helper 2 signature; recruitment of low HLA-DR expressing monocytes and regulatory T-cells; and transcription of immune checkpoint (e.g., PD-1, LAG3) genes. These results suggest dysregulated immunoregulation is associated with poor prognosis, whereas early innate signaling and Th1-skewed immunity are important for survival.

Characterizing the antigen‐binding and innate immune‐recruiting properties of the humoral response offers the chance to obtain deeper insights into mechanisms of protection than revealed by measuring only overall antibody titer. Here, a high‐throughput, multiplexed Fab‐Fc Array was employed to profile rhesus macaques vaccinated with a gp120‐CD4 fusion protein in combination with different genetically encoded adjuvants, and subsequently subjected to multiple heterologous simian immunodeficiency virus (SIV) challenges. Systems analyses modeling protection and adjuvant differences using Fab‐Fc Array measurements revealed a set of correlates yielding strong and robust predictive performance, while models based on measurements of response magnitude alone exhibited significantly inferior performance. At the same time, rendering Fab‐Fc measurements mathematically independent of titer had relatively little impact on predictive performance. Similar analyses for a distinct SIV vaccine study also showed that Fab‐Fc measurements performed significantly better than titer. These results suggest that predictive modeling with measurements of antibody properties can provide detailed correlates with robust predictive power, suggest directions for vaccine improvement, and potentially enable discovery of mechanistic associations.

Improving feed utilization in cattle is required to reduce input costs, increase production, and ultimately improve sustainability of the beef cattle industry. Characterizing metabolic differences between efficient and non-efficient animals will allow stakeholders to identify more efficient cattle during backgrounding. This study used an untargeted metabolomics approach to determine differences in serum metabolites between animals of low and high residual feed intake. Residual feed intake was determined for 50 purebred Angus steers and 29 steers were selected for the study steers based on low versus high feed efficiency. Blood samples were collected from steers and analyzed using untargeted metabolomics via mass spectrometry. Metabolite data was analyzed using Metaboanalyst, visualized using orthogonal partial least squares discriminant analysis, and p-values derived from permutation testing. Non-esterified fatty acids, urea nitrogen, and glucose were measured using commercially available calorimetric assay kits. Differences in metabolites measured were grouped by residual feed intake was measured using one-way analysis of variance in SAS 9.4. Four metabolites were found to be associated with differences in feed efficiency. No differences were found in other serum metabolites, including serum urea nitrogen, non-esterified fatty acids, and glucose. Four metabolites that differed between low and high residual feed intake have important functions related to nutrient utilization, among other functions, in cattle. This information will allow identification of more efficient steers during backgrounding.

The recombinant ALVAC vaccine coupled with the monomeric gp120/alum protein have decreased the risk of HIV and SIV acquisition. Ab responses to the V1/V2 regions have correlated with a decreased risk of virus acquisition in both humans and macaques. We hypothesized that the breadth and functional profile of Abs induced by an ALVAC/envelope protein regimen could be improved by substituting the monomeric gp120 boost, with the full-length single-chain (FLSC) protein. FLSC is a CD4-gp120 fusion immunogen that exposes cryptic gp120 epitopes to the immune system. We compared the immunogenicity and relative efficiency of an ALVAC-SIV vaccine boosted either with bivalent FLSC proteins or with monomeric gp120 in alum. FLSC was superior to monomeric gp120 in directing Abs to the C3 α2 helix, the V5 loop, and the V3 region that contains the putative CCR5 binding site. In addition, FLSC boosting elicited significantly higher binding Abs to V2 and increased both the Ab-dependent cellular cytotoxicity activity and the breadth of neutralizing Abs. However, the FLSC vaccine regimen demonstrated only a trend in vaccine efficacy, whereas the monomeric gp120 regimen significantly decreased the risk of SIVmac251 acquisition. In both vaccine regimens, anti-V2 Abs correlated with a decreased risk of virus acquisition but differed with regard to systemic or mucosal origin. In the FLSC regimen, serum Abs to V2 correlated, whereas in the monomeric gp120 regimen, V2 Abs in rectal secretions, the site of viral challenge, were associated with efficacy.

A guiding principle for HIV vaccine design has been that cellular and humoral immunity work together to provide the strongest degree of efficacy. However, three efficacy trials of Ad5-vectored HIV vaccines showed no protection. Transmission was increased in two of the trials, suggesting that this vaccine strategy elicited CD4+ T-cell responses that provide more targets for infection, attenuating protection or increasing transmission. The degree to which this problem extends to other HIV vaccine candidates is not known. Here, we show that a gp120-CD4 chimeric subunit protein vaccine (full-length single chain) elicits heterologous protection against simian-human immunodeficiency virus (SHIV) or simian immunodeficiency virus (SIV) acquisition in three independent rhesus macaque repeated low-dose rectal challenge studies with SHIV162P3 or SIVmac251. Protection against acquisition was observed with multiple formulations and challenges. In each study, protection correlated with antibody-dependent cellular cytotoxicity specific for CD4-induced epitopes, provided that the concurrent antivaccine T-cell responses were minimal. Protection was lost in instances when T-cell responses were high or when the requisite antibody titers had declined. Our studies suggest that balance between a protective antibody response and antigen-specific T-cell activation is the critical element to vaccine-mediated protection against HIV. Achieving and sustaining such a balance, while enhancing antibody durability, is the major challenge for HIV vaccine development, regardless of the immunogen or vaccine formulation.