Biology subjects

Fletcher, R. B.

Publications and source records attributed to Fletcher, R. B..

3 recordsLinked to original sources

bioRxiv2026

A targeted bispecific TGFBR2 antagonist antibody demonstrates cell selectivity and enhanced potency on human fibroblasts

Beyond its critical roles in development and tissue homeostasis, TGF{beta} signaling promotes key aspects of cancer progression and is a primary driver of fibrosis. Although blocking TGF{beta} signaling has great therapeutic potential for cancer and fibrotic diseases such as idiopathic pulmonary fibrosis (IPF), preclinical and clinical studies revealed that non-specific alteration of the pathway can have severe adverse consequences; therefore, inhibiting TGF{beta} signaling in a cell-type specific manner may avoid systemic toxic effects while preserving potential therapeutic effects. The parasitic helminth Heligmosomoides polygyrus has evolved cell-type-targeted modulators of TGF{beta} signaling. With insights from the development of other targeted signaling modulators and using the worm proteins as a guide, we sought to develop a human-fibroblast-targeted TGFBR2 antagonist. Here, we report mechanistic insights into the targeted worm TGFBR2 antagonist TGM6 and fusion proteins containing the TGM6 targeting domains. We created a bispecific antibody TGFBR2 antagonist that binds PDGFRA as a targeting receptor and demonstrates cell selectivity and enhanced potency in fibroblasts. Our findings suggest a viable path for developing targeted TGF{beta} signaling antagonists as therapeutics for cancer and tissue fibrosis.

cell biology↗

bioRxiv2024

Synthesis and characterization of chloroquine-modified albumin-binding siRNA-lipid conjugates for improved intracellular delivery and gene silencing in cancer cells

siRNA therapeutics have considerable potential as molecularly-targeted therapeutics in malignant disease, but identification of effective delivery strategies that mediate rapid intracellular delivery while minimizing toxicity has been challenging. Our group recently developed and optimized an siRNA conjugate platform termed "siRNA-L2," which harnesses non-covalent association with endogenous circulating albumin to extend circulation half-life and achieve tumor-selective delivery without the use of traditional cationic lipids or polymers for transfection. To improve intracellular delivery and particularly the endosomal escape properties of siRNA-L2 towards more efficient gene silencing, we report synthesis of siRNA-CQ-L2 conjugates, in which chloroquine (CQ), an endosomolytic quinoline alkaloid, is covalently incorporated into the branching lipid tail structure. We accomplished this via synthesis of a novel CQ phosphoramidite, which can be incorporated into a modular siRNA-L2 backbone using on-column solid-phase synthesis through use of asymmetric branchers with levulinyl-protected hydroxide groups that allow covalent addition of pendant CQ repeats. We demonstrate that siRNA-CQ-L2 maintains the ability to non-covalently bind albumin, and with multiple copies of CQ, siRNA-CQ-L2 mediates higher endosomal disruption, cellular uptake/retention, and reporter gene knockdown in cancer cells. Further, in mice, the addition of CQ did not significantly affect circulation kinetics nor organ biodistribution and did not produce hematologic or organ-level toxicity. Thus, controlled, multivalent conjugation of albumin-binding siRNA-L2 to endosomolytic small molecule compounds holds promise in improving siRNA-L2 knockdown potency while maintaining albumin-binding properties and overall safety.

bioengineering↗

bioRxiv2017

Slingshot: Cell lineage and pseudotime inference for single-cell transcriptomics

Single-cell transcriptomics allows researchers to investigate complex communities of heterogeneous cells. These methods can be applied to stem cells and their descendants in order to chart the progression from multipotent progenitors to fully differentiated cells. While a number of statistical and computational methods have been proposed for analyzing cell lineages, the problem of accurately characterizing multiple branching lineages remains difficult to solve. Here, we introduce a novel method, Slingshot, for inferring multiple developmental lineages from single-cell gene expression data. Slingshot is a uniquely robust and flexible tool for inferring developmental lineages and ordering cells to reflect continuous, branching processes.

bioinformatics↗