# New Congenital Disorders of Glycosylation: Therapy and Models

> **NIH NIH R01** · SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE · 2024 · $809,566

## Abstract

PROJECT SUMMARY
AIM1 studies how a therapeutic sugar is mobilized for glycosylation. Sugars taken up from outside the cell are
handled differently than those produced inside the cell. We found that activated fucose (GDP-Fuc) exists in
multiple cytoplasmic pools that differentially contribute to glycosylation., There is no homogenous, feely mixing
pool of sugars. This segregation requires a previously unimagined level of cellular organization to distinguish
fucose from different origins. We measured simultaneous contributions of the de novo, exogenous, and recycled
(salvaged) fucose into multiple glycoproteins. Their contributions varied based on the fucose origin, glycoprotein
acceptor, cellular compartment (ER or Golgi), and fucosyltransferase. These multiple, separate, yet conversant,
GDP-fucose pools require regulation, since their combined amount remains constant. We will use multiple 13C-
sugar labeling and quantitative lectin binding as functional readouts to probe this cellular organization and
control. Now we will determine 1.) how the metabolic origin of fucose affects its ultimate incorporation into
different glycoproteins. 2.) basis of fucose segregation 3). the effect of relocating fucosyltransferses from the ER
to the Golgi. 4.) the genes and transporters involved in fucose segregation, degradation, and recycling. 5.) Use
lectin-based siRNA knockdown and whole genome CRISPR screens to search for genes that impact GDP-Fuc
pool utilization. Discovery of multiple GDP-Fucose pools led us to find evidence for separate GDP-Mannose
pools. We will measure functional effects using 13C-monosaccharide incorporation into bulk N- and O-linked
glycans or into single glycoproteins using lectin binding. Finding the players and mechanisms that segregate
pools will likely apply to other monosaccharides and precursors for other macromolecules.
In AIM 2, we will continue to identify and validate new CDGs linking clinical medicine to cell biology and
glycosylation. We already identified 25 types of CDG. Each candidate requires individualized biochemical and
functional validation. Our decades-long clinical pipeline provides a steady stream of potential candidate genes.
We prioritize them based on inheritance pedigrees, nature of the variants, interactome and structural analysis of
the protein. UGGT1, GOLG2A, MGAT1, and GET1 are currently in our pipeline. We design functional assays to
test pathogenicity of variants using patient-derived cells or by functional complementation of KO lines with
patient-variant constructs. Pathogenic variants do not complement, showing they are legitimate CDGs. Assays
include intracellular protein trafficking, Golgi morphology and dynamics, glycomic analysis and lectin binding.
The clinical discoveries enhance our insights into basic science and sometimes provide therapy for patients.

## Key facts

- **NIH application ID:** 10980334
- **Project number:** 2R01DK099551-10A1
- **Recipient organization:** SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
- **Principal Investigator:** Hudson H. Freeze
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $809,566
- **Award type:** 2
- **Project period:** 2014-05-01 → 2029-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10980334

## Citation

> US National Institutes of Health, RePORTER application 10980334, New Congenital Disorders of Glycosylation: Therapy and Models (2R01DK099551-10A1). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10980334. Licensed CC0.

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