Major infectious diseases, like COVID-19, often go undetected until they spread. Spotting the problem in a community isn’t easy, in part because infectious disease detection, which is the foundation for surveillance and contact tracing, can take hours, if not days. Compounding the problem is that traditional detection assays are designed a priori while newer bio-detection technologies have been slow to move from the lab into general use.
Pandemics may threaten both internal and external national security – the physical threat to U.S. citizens in terms of morbidity and mortality, and the decreased effectiveness of U.S. armed forces in protecting those citizens from external threats. The economic, political or social turmoil in adversary country is also potential threat. The U.S. military supports U.S. Government responses to public health emergencies such as Ebola, which can cause regional destabilization and spread through global travel, said DARPA. Eliminating pandemic outbreaks and mitigating the impact of a potential high threat biological agent release are national security priorities.
The DoD requires timely and comprehensive threat detection to support overall readiness, counter the spread of disease, and promote stabilization missions. State-of-the-art diagnostic and biosurveillance systems are unable to keep pace with disease outbreaks and fail to support decision-making at the time and place of need.
Now scientists are developing a portable device that is easily configurable for screening thousands of pathogens in minutes, not hours. The device will incorporate recent advances in biotechnology, imaging analysis and microfluidics. The aim is to improve the speed of treatment and enhance the standard of care for the public and to equip the military with new biosurveillance technology.
Rachel Fezzie, a biochemist and molecular biologist at Draper, says infectious disease detection needs to be faster, lower in cost and field-ready. “The development of a rapid, low-cost diagnostic device that is capable of detecting 1,000 biomarkers in one processing cycle is an important step forward in improving infectious disease detection,” Fezzie explains. “A device with the ability to detect pathogens sooner and in a greater variety would improve public health, add protections for military units and provide critical information to medical decision-makers within minutes.”
DARPA Detect It with Gene Editing Technologies (DIGET) program
DARPA launched the DIGET program at the end of 2019 to create diagnostic and biosurveillance systems that can keep pace with disease outbreaks and support decision-making at the time and place of need while taking advantage of advances in gene editing technologies. The “Detect It with Gene Editing Technologies” (DIGET) program aims to leverage advances in gene editing technologies to develop field-forward diagnostic and biosurveillance technologies that enable detection of any threat, anytime, anywhere.
The overarching goal of DIGET is to provide comprehensive, specific, and trusted information about health threats to medical decision-makers within minutes, even in far-flung regions of the globe, to prevent the spread of disease, enable timely deployment of countermeasures, and improve the standard of care after diagnosis.
“DARPA is pursuing the capability to detect and characterize any pathogen, regardless of when or where it emerges,” said Renee Wegrzyn, the DIGET program manager. “There are three primary reasons that we think gene editing systems can help us realize that vision, and they boil down to speed, accuracy, and precision. First, they’re programmable, which means we can easily adapt assays as needed to account for previously unknown threats. Second, their extreme sensitivity means they can identify pathogenic targets even when those targets are present in very low abundance. And third, they can be both broad and specific, meaning a user might, for instance, confirm the presence of influenza generally or identify a specific flu strain along with its characteristics.”
In a twist on how gene editing technology might be applied in the future, DARPA’s newest biotechnology funding opportunity aims to incorporate gene editors into detectors for distributed health biosurveillance and rapid, point-of-need diagnostics for endemic, emerging, and engineered pathogenic threats.
The distributed, effectively continuous detection capability DARPA is pursuing with DIGET does not currently exist. At present, biosurveillance relies on periodic shipping of samples from across broad geographic areas for analysis at centralized laboratories. The timeline for shipping, testing, and reporting results delays the return of actionable information anywhere from hours to days. If a central laboratory detects a threat, the finding may lead to distributed laboratories coming online to better track the spread of disease, but often that new surveillance capacity comes too late to halt an outbreak before its natural peak. With DIGET technology on the other hand, it could become possible to begin with a distributed biosurveillance model, then use the results from that network to inform the rapid creation and distribution of single-use diagnostics to accurately measure the spread of infection.
The DIGET vision incorporates two devices: a handheld, disposable point-of-need device that screens samples for at least 10 pathogens or host biomarkers at once, combined with a massively multiplexed detection (MMD) platform capable of screening clinical and environmental samples for more than 1,000 targets simultaneously. The MMD device will enable early threat detection, assess disease severity and improve situational awareness. Both pieces of the system could be quickly reconfigured to adapt to changing needs. Both devices must be simple to operate, low-cost and rapidly reconfigurable.
While DARPA seeks a variety of approaches, DIGET technologies could work by introducing an inactive gene editor paired with a programmable guide into a biological sample to identify a target protein. Upon detection, the gene editing enzymes would be activated, triggering them to cut the RNA or DNA at the exact target location. The activated enzyme would remain active and continue to cleave reporter molecules in close proximity to the reaction, providing an easily observed indication of a positive match. That boosting of the reporter signal removes the need to amplify targets prior to testing, simplifying the process and reducing time to result.
“Two hypothetical, but realistic scenarios reveal the potential impact of these tools and the unambiguous results they’d provide,” explained Wegrzyn. “First, consider the case of a service member presenting with flu-like symptoms. With DIGET, we think it will be possible to not only confirm that he or she has influenza, but also to determine the strain, the origin, and whether the strain is drug resistant. In addition, DIGET tools could assess the severity of disease to guide how the patient is triaged and treated.
Second, think about an Ebola triage center in a low-resource country. If you’re screening someone for a deadly disease, you know that every minute of potential exposure counts, so you need a fast-acting assay. You also need absolute certainty in the diagnosis, because if a test comes back with a false positive, you risk that the patient will contract the disease based on proximity to actual infected individuals during treatment. If a test returns a false negative, the individual is released back into the community to unintentionally spread the virus. These are life and death scenarios that DARPA believes DIGET technology can resolve.”
DIGET researchers will face a range of challenges that will demand expertise in gene editing, synthetic biology, assay development, nucleic acid technologies, bioinformatics, and diagnostic device engineering and development. In the computational design space, research teams will need to identify a minimal set of targets from a robust bioinformatics pipeline. These targets will inform the design of programmable probes and guides for broad target sequences present across a number of strains, as well as for more specific target sequences that identify subtypes of pathogens, rare mutations, and even co-infections.
In the gene editing space, researchers will need to design assays, enzymes, and reporters for identification and signaling of positive results. And in the engineering space, teams will need to integrate their detection and reporting capabilities into user-friendly, field-ready prototype systems that go from sample to answer in under 15 minutes and provide unambiguous results. Scheduled capability demonstrations and planned independent validation and verification will test researchers’ prototype systems, assess their ability to screen clinical and environment samples, and gauge the speed with which the systems can be reconfigured.
If the DIGET program succeeds, the resulting platforms will provide a wealth of information for global health monitoring and rapid response, and support characterization of circulating pathogens to inform the development of medical countermeasures, including the class of nucleic acid-based countermeasures DARPA is pursuing under its Pandemic Prevention Platform, Nucleic Acids On-Demand Worldwide (NOW), and PReemptive Expression of Protective Alleles and Response Elements (PREPARE) programs. Additionally, although DIGET is focused on biological threats, similar technologies could be applied to chemical and radiological threats by detecting biomarkers of exposure.
“DARPA is already transforming nucleic acid technologies into fast-acting tools for preserving or restoring health, and now we’re extending their value to rapid detection and characterization of health threats to better direct how those prophylactic and therapeutic tools are used,” said Wegrzyn. “DIGET fills in a major piece of the puzzle for DARPA’s vision of defending against any biological threat, anytime, and anywhere.”
The “Detect It with Gene Editing Technologies” (DIGET) program could help the Department of Defense maintain force readiness by informing rapid medical response and increasing the standard of care for troops, and preserve geopolitical stability by preventing the spread of infectious disease from becoming a driver of conflict.
The Defense Advanced Research Projects Agency (DARPA) has awarded a contract worth up to $36.7 million over four years to MRIGlobal and a team of five other organizations under the auspices of its Detect It with Gene Editing Technologies (DIGET) program.
DIGET aims to provide next-generation detection for the DOD by combining gene-editing technology with fieldable diagnostics and biosurveillance. DIGET will apply CRISPR technologies to develop a disposable point of care device for detection of at least 10 targets and a massively multiplexed device for at least 1,000 targets. MRIGlobal received an initial contract award for $12.7M with a total contract value of up to $36.7M over a four-year period of performance.
MRIGlobal will lead a world-class team of partners: Mammoth Biosciences, Draper, IDbyDNA, University of California, San Francisco (UCSF), and Toolbox Medical Innovations to develop chemistry and devices. This program is timely and addresses the COVID-19 pandemic which is an application the program intends to address for emerging and novel biothreats.
Draper to Develop Biosurveillance Technology for DARPA
Fezzie and her colleagues at Draper are developing just such a technology as part of a team recently selected by the Defense Advanced Research Projects Agency (DARPA). Led by MRIGlobal, the team is developing a massively multiplexed device (MMD) for DARPA’s “Detect It with Gene Editing Technologies” (DIGET) program. The aim, says DARPA, is to incorporate gene editors into detectors for distributed health biosurveillance and rapid, point-of-need diagnostics for endemic, emerging and engineered pathogenic threats.
The foundation for the MMD is DETECTR™, a CRISPR-based detection platform developed by Mammoth Biosciences, which is a member of the MRIGlobal team. The platform has been proven as a rapid detection diagnosis technology that uses a gene editing molecular tool to find a specific genetic sequence within a sample.
The MMD will incorporate several innovative technologies developed by Draper. Fluid routing throughout the disposable cartridge will be controlled by Draper’s proprietary electromagnetic pump and valve technology, ideal for applications with low size, weight and power requirements. Draper’s demonstrated experience in functionalizing glass substrates and printing compact, high-density DNA arrays will directly inform the development path to scale Mammoth’s DETECTRTM chemistry to 1,000-plex via a microarray of Cas/gRNA complexes. Each Cas/gRNA complex is designed to detect a unique pathogen.
The MMD will also use a new approach to optical detection through development of a miniaturized, simple, lens-free system that has no moving parts, making it attractive for field deployment. Reconfiguration of the microarray will be enabled by printing array chips with new Cas/gRNA complexes, a process expected to take 24 hours or less.
“The MMD”—according to John Julias, Draper’s program manager for DIGET—”seeks to fundamentally alter the concept of operations for biosurveillance and diagnostic detection by increasing throughput, sensitivity and usability to levels unmatched by existing technologies. A single run of the MMD promises to provide not only species- or strain-level identification but also information on genetically encoded characteristics such as antimicrobial resistance, as well as gene expression data to indicate disease severity in a person.”
The projected sample-to-answer time of 15 minutes with the MMD will enable more assays to be run in a day, allowing appropriate action to be taken in near real-time, Fezzie explains. “Currently, you would need next-generation sequencing to identify such a broad a range of pathogens, and that typically takes days to perform by highly skilled laboratory staff.” Draper’s DIGET work is supported by the company’s Bioengineering, Microsystems and Advanced Material divisions, and it is managed by its Special Programs Office.
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