How the CRISPR Alphabet Soup Can Make You a Trillion Dollars

Genome editing is primed to develop the next-generation of crops, livestock, ingredients, and diagnostics, putting it at the leading edge of product development for markets in the trillions of dollars. Technologies including transcription activator-like effector nucleases (TALENs) and zinc finger nucleases (ZFNs) have already made a mark toward improving crop quality, but the scalability of these two technologies is limited by cost and flexibility. Clustered regularly interspaced short palindromic repeat (CRISPR) RNA-guided genome editing systems do not suffer from these constraints, and the non-regulated status of genome edits throughout most the world adds commercial attractiveness. CRISPR is now poised to lead the charge towards the next-generation of crops and livestock, and numerous companies, large and small, are actively engaged. However, the CRISPR landscape remains complex. In addition to the much-covered initial CRISPR-Cas9 discovery, there is a veritable alphabet soup of additional tools in development, each with special qualities and benefits.

Focusing in on just one of those trillion-dollar markets, let’s look at agriculture. CRISPR systems developed for use in agriculture can be divided among four capabilities: genome editing, control of gene expression, nucleic acid detection, and pathogen treatment. Follow along as we lead you through the space and identify the molecular tools that complement your competencies.

 

Genome Editing

Genome editing with CRISPR alters the DNA sequence of a specific gene resulting in its disfunction, referred to as a gene knockout. The canonical CRISPR-Cas9 system developed at the Broad Institute (Feng Zhang) and the University of California, Berkeley (Jennifer Doudna) effectively creates gene knockouts but suffers from low-efficiency specific nucleotide insertions or high-efficiency random nucleotide alterations. CRISPR-Cas-based editors (CRISPR-dCas9 or CRISPR-Cas9 nickase) have emerged to overcome these issues by fusing an engineered version of Cas9, which cannot cut DNA, to a nucleotide deaminase. The deaminase converts a targeted nucleotide to another (C to T, A to G), altering a single nucleotide pair. These applications have also been applied to multiple genes in a single organism with the Cas9 system, allowing developers to create advanced new varieties in a single generation.

While technology development has centered around the CRISPR-Cas9 system, alternative CRISPR systems have been discovered and developed, and are taking advantage of the IP battle for the CRISPR-Cas9 system to gain market share.

  • Benson Hill Biosystems (Lux Take: Strong Positive) adopted the internal development strategy, recently receiving patents for what it refers to as a CRISPR 3.0 system (Cms1). Cms1 nucleases (four identified) are smaller than other CRISPR nucleases, and do not require tracrRNA. In another move, Benson Hill petitioned the US patent office to review the Cpf1 CRISPR grant held by MIT, Harvard, and the Broad Institute, which interferes with a patent of its own. Cpf1 has an arguably simpler mechanism than Cas9 and increases the diversity of CRISPR regions available for genome editing.
     
  • Taking the latter strategy, Inscripta released an alternative CRISPR enzyme, MAD7. The company provides “the DNA sequence of E. coli-optimized MAD7 free for any academic or commercial R&D use ... to catalyze the testing, improvement, and adoption of MAD7.” The company receives only a small licensing fee for commercial products and is therefore more interested in changing the face of adopted CRISPR technology. 
     

Lux Research - CRISPR-Cas9 Molecular Structure

 

Control of Expression

Structurally mutated CRISPR base editors (dCas9) can also be fused to proteins other than deaminases to alter their functionality. Fused functionalities developed for dCas9 include the activation or repression of target genes and epigenetic modifiers – essentially ways to change how a gene is expressed. Control of gene expression has not been applied in the agriculture setting yet, but it greatly increases the potential to fine-tune gene expression to affect traits.   

 

Nucleic Acid Detection

A competition is underway between Mammoth Biosciences and Arbor Biotechnologies to be the first company to commercialize a nucleic acid detection product utilizing CRISPR proteins. The companies, with Jennifer Doudna and Feng Zhang among their founders, respectively, are utilizing the single-stranded DNase or RNase activity of the Cas12 and Cas13 proteins (alternative CRISPR proteins) to report the presence of pathogen or disease-state (e.g. cancer) DNA. These systems are being developed as point-of-care (POC) diagnostics and are estimated to cost as little as $1. While these companies currently target human health applications, there are obvious potential applications in agriculture, considering the immense effects that diseases have on crops and livestock production. Alternatively, the functionality of dCas9 can be further altered for fluorescent visualization of genomic loci. 

 

Pathogen Treatment

In a novel application of CRISPR, Locus Biosciences and Eligo Bioscience develop CRISPR-Cas3 phage (crPhage) systems. These systems utilize the highly-specific targeting of CRISPR proteins but apply this capability to Cas3, which identifies target DNA and then destroys it. Therefore, this system has potential to specifically target pathogenic microbes, especially bacteria.  

 

CRISPR technology provides an incredible opportunity to develop improved agriculture systems in the wake of regulatory (e.g., water use, pesticides), environmental (e.g., drought, sea level rise), and sociological (e.g., consumer perceptions) changes. Multiplying the potential for this technology is the improvement of next-generation sequencing efforts (i.e., sugar cane and wheat) and bioinformatics tools, which serve to improve the development of guide sequences for CRISPR enzymes. In the next several years, whether you are ready for it or not, commercialized CRISPR-edited commodities will be available, and will capture a significant portion of the trillion-dollar global food markets. Genome editing is also democratizing crop improvement, increasing the prevalence of trait developers and threatening the dominance of current incumbents. Explore the CRISPR alphabet soup and decide which can impact your business, be it through partnership, competition, licensing, or research and development. 

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