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Development of a Multiplex Proteomics Assay for High-Throughput Newborn Screening of a New Set of Treatable Neonatal Diseases

Award Number: R01HD111410
ORGANIZATION: NATIONAL INSTITUTE OF CHILD HEALTH AND HUMAN DEVELOPMENT
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

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Development of a Multiplex Proteomics Assay for High-Throughput Newborn Screening of a New Set of Treatable Neonatal Diseases - PROJECT SUMMARY Newborn screening (NBS) is one of the most successful public health programs that provides cost-effective early diagnosis of treatable disorders. The basis of NBS is to quantify a biomarker that is indicative of disease in a dried blood spot (DBS). It was projected that 60 cell and gene therapies could be approved by the FDA within 10 years, which underscores an urgent need to modernize the NBS system. A major challenge for NBS of many treatable genetic disorders is the unavailability of a biomarker in DBS, because there is no accumulation of small molecules in patients. In this proposed studies, it is also not feasible to measure the biological function of the protein (i.e. enzyme assay or other functional assays). Instead, reduction in the abundance of a specific protein (encoded by the disease-causing gene) can be used for the diagnosis of genetic diseases as many genetic conditions are caused by mutations that lead to non-functional proteins that are quickly degraded, as exemplified in Wilson disease. Given the complexity of the DBS sample and the typical low-abundance nature of target proteins, a technique called Immuno-SRM (Immunocapture coupled to selected-reaction-monitoring, or SRM) had been successfully developed to quantify proteins in DBS. One drawback of our current Immuno-SRM for NBS is that the LC-MS/MS time is about 3-4 min per sample. In this study, we propose a novel idea of expanded multiplexing by using newborn-specific peptide mass tags to greatly increase the throughput of proteomic-based NBS. Tagging of peptides with mass-differentiated tags allows samples from multiple newborns to be analyzed in the same LC-MS/MS run. The objective of this application is to develop Immuno-SRM multiplex assay to quantify 19 peptides for NBS of 12 genetic conditions including tuberous sclerosis. Successful implementation of the project will enable the screening of 12 diseases simultaneously using a single DBS sample with a MS instrument time < 1 min per sample. The goal will be achieved through the following specific aims. (1) Develop an Immuno-SRM method to quantify TSC2 for tuberous sclerosis that is suitable for newborn screening. Considering that we have already developed monoclonal antibodies (mAb, which is used to enrich surrogate peptides for disease-specific proteins) for all conditions except TSC, the first aim is to raise mAb against TSC2 surrogate peptides and develop an Immuno-SRM for its quantification followed by optimization for multiplex analysis. (2) Development of peptide tagging for increasing the throughput of Immuno-SRM assays. Tagging of peptides with mass-differentiated tags allows samples from multiple newborns (>4 samples) to be analyzed in the same LC-MS/MS run (equivalent to individual barcode sequences for next-generation- sequencing). (3) Performance evaluation of the multiplexed Immuno-SRM method. First, the workflow will be tested on known patient samples for its sensitivity. Second, a mini-pilot study on 3000 random newborn DBS will be performed to test the robustness of the method.


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2025 ( Subtotal = $437,165 )
2025	2025	SEATTLE CHILDREN'S HOSPITAL	4800 SAND POINT WAY NE	SEATTLE	WA	98105	KING	USA	Child Health and Human Development Extramural Research	000	3	2/12/2025	NON-COMPETING CONTINUATION	$437,165
														Subtotal = $437,165

Issue Date FY: 2024 ( Subtotal = $553,803 )
2024	2024	SEATTLE CHILDREN'S HOSPITAL	4800 SAND POINT WAY NE	SEATTLE	WA	98105	KING	USA	Child Health and Human Development Extramural Research	000	2	2/7/2024	NON-COMPETING CONTINUATION	$503,458
2024	2024	SEATTLE CHILDREN'S HOSPITAL	4800 SAND POINT WAY NE	SEATTLE	WA	98105	KING	USA	Child Health and Human Development Extramural Research	001	2	5/1/2024	NON-COMPETING CONTINUATION	$50,345
														Subtotal = $553,803

Issue Date FY: 2023 ( Subtotal = $583,225 )
2023	2023	SEATTLE CHILDREN'S HOSPITAL	4800 SAND POINT WAY NE	SEATTLE	WA	98105	KING	USA	Child Health and Human Development Extramural Research	000	1	4/18/2023	NEW	$583,225
														Subtotal = $583,225

Grand Total All Awards = $1,574,193

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Development of a Multiplex Proteomics Assay for High-Throughput Newborn Screening of a New Set of Treatable Neonatal Diseases

Award Number: R01HD111410

ORGANIZATION: NATIONAL INSTITUTE OF CHILD HEALTH AND HUMAN DEVELOPMENT

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

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