A comprehensive platform for analyzing longitudinal multi-omics data

The analysis of longitudinal bulk and single-cell multi-omics data is a highly complex task. Here, the authors introduce PALMO, a software platform with five modules to analyse longitudinal bulk and single-cell multi-omics data, which is extensively tested in external datasets that include multiple omics modalities

Rational design of a fibroblast growth factor 21-based clinical candidate, LY2405319.

Fibroblast growth factor 21 is a novel hormonal regulator with the potential to treat a broad variety of metabolic abnormalities, such as type 2 diabetes, obesity, hepatic steatosis, and cardiovascular disease. Human recombinant wild type FGF21 (FGF21) has been shown to ameliorate metabolic disorders in rodents and non-human primates. However, development of FGF21 as a drug is challenging and requires re-engineering of its amino acid sequence to improve protein expression and formulation stability. Here we report the design and characterization of a novel FGF21 variant, LY2405319. To enable the development of a potential drug product with a once-daily dosing profile, in a preserved, multi-use formulation, an additional disulfide bond was introduced in FGF21 through Leu118Cys and Ala134Cys mutations. FGF21 was further optimized by deleting the four N-terminal amino acids, His-Pro-Ile-Pro (HPIP), which was subject to proteolytic cleavage. In addition, to eliminate an O-linked glycosylation site in yeast a Ser167Ala mutation was introduced, thus allowing large-scale, homogenous protein production in Pichia pastoris. Altogether re-engineering of FGF21 led to significant improvements in its biopharmaceutical properties. The impact of these changes was assessed in a panel of in vitro and in vivo assays, which confirmed that biological properties of LY2405319 were essentially identical to FGF21. Specifically, subcutaneous administration of LY2405319 in ob/ob and diet-induced obese (DIO) mice over 7-14 days resulted in a 25-50% lowering of plasma glucose coupled with a 10-30% reduction in body weight. Thus, LY2405319 exhibited all the biopharmaceutical and biological properties required for initiation of a clinical program designed to test the hypothesis that administration of exogenous FGF21 would result in effects on disease-related metabolic parameters in humans

Simultaneous trimodal single-cell measurement of transcripts, epitopes, and chromatin accessibility using TEA-seq

Single-cell measurements of cellular characteristics have been instrumental in understanding the heterogeneous pathways that drive differentiation, cellular responses to signals, and human disease. Recent advances have allowed paired capture of protein abundance and transcriptomic state, but a lack of epigenetic information in these assays has left a missing link to gene regulation. Using the heterogeneous mixture of cells in human peripheral blood as a test case, we developed a novel scATAC-seq workflow that increases signal-to-noise and allows paired measurement of cell surface markers and chromatin accessibility: integrated cellular indexing of chromatin landscape and epitopes, called ICICLE-seq. We extended this approach using a droplet-based multiomics platform to develop a trimodal assay that simultaneously measures transcriptomics (scRNA-seq), epitopes, and chromatin accessibility (scATAC-seq) from thousands of single cells, which we term TEA-seq. Together, these multimodal single-cell assays provide a novel toolkit to identify type-specific gene regulation and expression grounded in phenotypically defined cell types

Persistent serum protein signatures define an inflammatory subcategory of long COVID

Abstract Long COVID or post-acute sequelae of SARS-CoV-2 (PASC) is a clinical syndrome featuring diverse symptoms that can persist for months following acute SARS-CoV-2 infection. The aetiologies may include persistent inflammation, unresolved tissue damage or delayed clearance of viral protein or RNA, but the biological differences they represent are not fully understood. Here we evaluate the serum proteome in samples, longitudinally collected from 55 PASC individuals with symptoms lasting ≥60 days after onset of acute infection, in comparison to samples from symptomatically recovered SARS-CoV-2 infected and uninfected individuals. Our analysis indicates heterogeneity in PASC and identified subsets with distinct signatures of persistent inflammation. Type II interferon signaling and canonical NF-κB signaling (particularly associated with TNF), appear to be the most differentially enriched signaling pathways, distinguishing a group of patients characterized also by a persistent neutrophil activation signature. These findings help to clarify biological diversity within PASC, identify participants with molecular evidence of persistent inflammation, and highlight dominant pathways that may have diagnostic or therapeutic relevance, including a protein panel that we propose as having diagnostic utility for differentiating inflammatory and non-inflammatory PASC

LY2405319 bioactivity is comparable to wild type FGF21.

<p>LY2405319 and FGF21 bioactivities were compared in mouse 3T3-L1/Klb fibroblasts (A) and differentiated mouse 3T3-L1 adipocytes (B) by monitoring 2-Deoxy-D-[U-¹⁴C]-glucose accumulation and in human HepG2 hepatoma cells by examining relative expression of GLUT1 mRNA (C) after incubation for 3 h or 1 h, respectively, with various concentrations of FGF21 (black squares) or LY2405319 (red triangles). Mean values ± SEM are presented as fold relative to treatment with vehicle alone.</p

Treatment of <i>ob/ob</i> mice with either FGF21 or LY2405319 improves metabolic dysfunction.

<p>Male <i>ob/ob</i> mice were treated with FGF21 or LY2405319 at various concentrations, as indicated, by constant infusion for 7 days. Blood glucose is reported as daily levels (A, B) or cumulatively for the treatment period (C). Plasma insulin was measured on the final day of treatment (D). Body weights were measured daily and are presented as body weight change (E) over the course of treatment. Plasma FGF21 and LY2405319 levels on the final day of treatment were measured by ELISA (F). Values are reported as mean ± SEM. (*), (**), and (***), p<0.01, p<0.001, and p<0.001, respectively, as compared to vehicle.</p

Treatment of DIO mice with either FGF21 or LY2405319 improves metabolic dysfunction.

<p>Male DIO mice were treated with FGF21 or LY2405319 at various concentrations, as indicated, by constant infusion for 14 days. Changes in body weight (A), fat mass (B) cumulative food intake (C), and blood glucose (D) were determined for the duration of treatment. Values are reported as mean ± SEM. (*), p<0.01 as compared to vehicle.</p

Dynamic light scattering assessment of FGF21 and LY2405319.

<p>Formulation stability of FGF21 (squares) and LY2405319 (diamonds) was assessed via dynamic light scattering at 37°C at 10 mg/mL in phosphate buffered saline at pH 7.4 containing 30 mM m-cresol. Protein concentration was 10 mg/ml.</p