SBIR-STTR Award

Direct to Phase II

A low-input compatible, end-to-end kitted HiChIP workflow for concurrent analyses of transcriptionalprotein binding and chromatin interactions toward a mechanistic understanding of gene regulationArima Genomics/Precise regulation of gene expression is paramount to establishing cellular identities, and mis-regulation of genescauses human disease. Cells regulate gene expression using regulatory elements (REs), short DNA sequencesembedded throughout the genome, who are bound by transcriptional proteins to facilitate their regulatoryfunction. Molecular mapping tools, such as Chromatin immunoprecipitation and next gen sequencing (ChIP-seq), produce "maps" of REs along the genome and have been a ubiquitous approach towards understand generegulation and define cell types and states based on unique RE signatures. However, these locations of REs areonly understood in context of a linear genome. In reality, REs execute their gene control within a threedimensional (3D) genome. Therefore to truly understand gene regulation - gene regulation must be mapped in3D. Indeed, high throughput chromatin interaction capture (HiC) was developed to produce 3D interaction mapsof all 3 billion bases in the human genome. HiC has facilitated discovery of several fundamental principles DNAfolding in 3D, including cases where DNA mis-folding contributes to disease. However, HiC does measuretranscriptional protein binding, nor whether a chromatin interaction is regulatory, thus having limited utility inadvancing our understanding 3D gene regulation. Recently, novel approaches attempt to combine the molecularsteps of ChIP-seq and chromatin interaction capture to measure transcriptional protein binding and mediatedchromatin interactions in a single assay. However these approaches, termed ChIA-PET and HiChIP, do notefficiently capture chromatin interactions or transcriptional protein binding, respectively. Therefore, there is direneed for improve methods that truly facilitate mapping of gene regulation in 3D.We satisfy this unmet need via our highly optimized, first generation HiChIP solution, Arima-HiChIP (A-HiChIP),that demonstrates efficient and reproducible mapping of transcriptional protein binding and chromatininteractions in cell lines, with higher cellular inputs and a limited set of transcriptional proteins. Our team hasunmatchable expertise in the science of chromatin interaction capture and its commercialization. First, wecommercialized Arima-HiC kits in 2018 for studying general principles of chromatin interactions and generated$1.2M in revenue in the 1st year of commercialization and $2M in revenue in the 2nd year, with 500+ customers,and 100% growth from 2018 to 2019. Based on VOC analytics, we shifted our focus to develop a more relatableproduct to the gene regulation market - A-HiChIP - that customers wanted and that represented a larger marketopportunity. Indeed, after our self-funded phase-1 R&D and commercial developments, we launched our firstgeneration HiChIP solution into the market and have seen remarkable success - measured by HiChIP growingfrom 19% to 40% of our revenue contributions, increased quality of revenue, and traction with KOLs, largeconsortia, and COVID research. However, these kits are limited in terms of the capabilities - they are not robustto a range of transcriptional proteins, they are not optimized towards tissue samples, and they are not optimizedtowards lower sample input quantities. To enable broader adoption and discovery, we have shown thedevelopment towards our second-generation A-HiChIP solution, with advancement towards low sample inputs,tissues, and a broader range of transcriptional proteins. We validate the technology on internal samples providedby academic collaborators and externally in customer hands via prototype beta kits.As part of this direct-2-phase II program, we propose to further develop our technology into truly robust, low inputcompatible, end-to-end kitted HiChIP solution for concurrent analysis of transcriptional protein binding andchromatin interactions in tissue samples and across a host of important transcriptional proteins. We also proposerigorous and essential product development experiments, to ensure commercialization of a robust, premium-performance kit-based product that is optimally integrated into the ecosystem. Upon successful completion ofthe technical and commercial developments in Aims 1 & 2, we propose to benchmark and validate the our next-generation HiChIP solution through collaboration and prototype (beta) kit and bioinformatics evaluations with keyopinion leaders (KOLs) across customer segments.

Public Health Relevance Statement:
A low-input compatible, end-to-end kitted HiChIP workflow for concurrent analyses of transcriptional protein binding and chromatin interactions toward a mechanistic understanding of gene regulation Arima Genomics Project Narrative Precise gene regulation is essential to specify the unique functions of cells and tissues in the human body. When gene regulation goes awry, human disease can result. Cells utilize regulatory elements (REs), bound by specific transcriptional proteins, to control gene expression. To obtain a complete picture of gene regulation and understand basic biology and disease, one must map which regulatory elements are regulating which genes. Current methods fail to capture the specific details of gene regulation. For example, ChIP-seq maps REs based on transcriptional protein binding. However, the regulatory maps are viewed through the lens of a linear genome, whereas in fact the genome functions in a three dimensional (3D) context. Chromatin interaction capture (HiC) maps chromatin interactions and provides a snapshot of the 3D genome, but does not measure transcriptional protein binding, or decipher which chromatin interactions are mediated by which transcriptional proteins. ChIA-PET and HiChIP attempt to bridge the two, but suffer from inefficient capture of chromatin interactions and transcriptional protein binding, respectively. Our solution to this problem is a deeply optimized HiChIP kit for high quality mapping of transcriptional protein binding and chromatin interactions. We systematically identify and optimize key aspects of the protocol to vastly improve data quality and utility, develop more comprehensive and easy-to-use bioinfo, and validate our solution internally and externally in customer hands via prototype HiChIP kits.

Project Terms:
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