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miR-SEQ: Highly efficient targeted quantification of extracellular miRNAs by Next Generation Sequencing

Award Number: R44GM115124
ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 03/15/2015
PERIOD OF PERFORMANCE END DATE: 10/31/2021

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miR-SEQ: Highly efficient targeted quantification of extracellular miRNAs by Next Generation Sequencing - Abstract: Recent discoveries highlight the importance of cell-free miRNAs (cf-miRNAs) as promising diagnostic and prognostic biomarkers for cancer and many other diseases. Biofluids such as plasma can be accessed with minimal invasiveness, unlike tissue biopsies. This, together with the high stability of cf-miRNAs in biofluids, makes them attractive for use in molecular diagnostics compared to other, more labile biomolecules. However, current techniques are inadequate for sensitive, specific and reliable quantification of miRNAs in biofluids. Microarrays and RT-qPCR are currently the preferred tools for expression profiling of cf-miRNAs, although each has major drawbacks. Microarrays suffer from low sensitivity, low dynamic range, and the inability to distinguish closely related miRNA sequences, while RT-qPCR has limited multiplexing capability and amplification biases. While next-generation sequencing (NGS) is superior in many of these respects, its reliability for cf-miRNA profiling in biofluids is limited due to bias (under- and over-detection) towards particular miRNA sequences, overwhelming amounts of unrelated sequencing data, the need for gel-purification of amplicons, and its high cost. Here we propose a new approach for constructing cf-miRNA sequencing libraries that addresses these problems. Called miR-SEQ, it incorporates a new combination of hybridization and enzymatic steps to simplify the preparation of miRNA sequencing libraries while significantly decreasing the sequencing bias. It involves a targeted sequencing approach allowing the quantification of all miRNAs of interest including rare tissue- (e.g., cancer-) derived miRNA species and their isomiRs representing the highest interest as biomarkers that would otherwise be represented by none or low numbers of sequencing reads, making their quantification problematic and expensive. In Phase I we demonstrated the feasibility of our approach by accurately detecting more than 100 cf-miRNAs with a targeted sequencing approach. In Phase II we will thoroughly optimize miR-SEQ to maximize its sensitivity and to allow sequencing of a larger variety of cf-miRNAs for commercial viability. In addition, we will streamline the protocol to facilitate its adoption by end users including academic labs, contract research facilities, corporate R&D and molecular diagnostic labs.


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: 2022 ( Subtotal = $0 )
2022	2020	REALSEQ BIOSCIENCES, INC.	2125 DELAWARE AVE STE B	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	000	6	7/11/2022	NON-COMPETING CONTINUATION	$0
														Subtotal = $0

Issue Date FY: 2020 ( Subtotal = $489,613 )
2020	2020	REALSEQ BIOSCIENCES, INC.	2161 DELAWARE AVE STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	002	6	5/15/2020	NON-COMPETING CONTINUATION	-$38,495
2020	2020	REALSEQ BIOSCIENCES, INC.	2161 DELAWARE AVE STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	001	6	1/27/2020	NON-COMPETING CONTINUATION	$564,588
2020	2019	REALSEQ BIOSCIENCES, INC.	2161 DELAWARE AVE STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	003	5	5/15/2020	CHANGE OF GRANTEE / TRAINING INSTITUTION / AWARDING INSTITUTION	-$36,480
2020	2019	SOMAGENICS, INC.	2161 DELAWARE ST STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	000	4	1/24/2020	NON-COMPETING CONTINUATION	$0
														Subtotal = $489,613

Issue Date FY: 2019 ( Subtotal = $583,682 )
2019	2019	SOMAGENICS, INC.	2161 DELAWARE ST STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	001	4	6/20/2019	NON-COMPETING CONTINUATION	-$291,842
2019	2019	SOMAGENICS, INC.	2161 DELAWARE ST STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	000	4	1/16/2019	NON-COMPETING CONTINUATION	$583,683
2019	2019	REALSEQ BIOSCIENCES, INC.	2161 DELAWARE AVE STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	002	5	6/21/2019	CHANGE OF GRANTEE / TRAINING INSTITUTION / AWARDING INSTITUTION	$291,841
														Subtotal = $583,682

Issue Date FY: 2018 ( Subtotal = $911,478 )
2018	2018	SOMAGENICS, INC.	2161 DELAWARE ST STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	001	3	8/24/2018	SUPPLEMENT FOR EXPANSION	$288,250
2018	2018	SOMAGENICS, INC.	2161 DELAWARE ST STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	002	3	8/27/2018	COMPETING CONTINUATION	-$8,935
2018	2018	SOMAGENICS, INC.	2161 DELAWARE ST STE E	SANTA CRUZ	CA	95060	SANTA CRUZ	USA	Biomedical Research and Research Training	000	3	1/23/2018	COMPETING CONTINUATION	$632,163
														Subtotal = $911,478

Grand Total All Awards = $1,984,773

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miR-SEQ: Highly efficient targeted quantification of extracellular miRNAs by Next Generation Sequencing

Award Number: R44GM115124

ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

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

PERIOD OF PERFORMANCE START DATE: 03/15/2015

PERIOD OF PERFORMANCE END DATE: 10/31/2021

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