Modality Moves at Vertex

REPRINT FROM JANUARY 17, 2019

MELETIOS VERRAS/ISTOCK/GETTY IMAGES PRODUCT R&D MODALITY MOVES AT VERTEX By Lauren Martz, Associate Editor In licensing a suite of newly discovered gene editing tools, pulmonary delivery of mRNAs and develop mRNA therapies Vertex Pharmaceuticals Inc. is broadening its options for solving for CF. Vertex paid $40 million up front including $20 million in problems that its core small molecule technology can’t address. cash and a $20 million convertible note. Moderna also is eligible On Jan. 3, Vertex announced a deal with Arbor Biotechnologies for up to $275 million in milestones and royalties. Inc. that gives it access to novel CRISPR endonucleases — the A Vertex spokesperson told BioCentury the collaborations with workhorse enzymes of the technology — allowing the company CRISPR, Moderna and Arbor are part of the company’s ongoing to sidestep the complex IP landscape surrounding the more research into gene-based therapies, but did not disclose whether commonly used CRISPR-Cas9 (see “Plenty of CRISPR Pie.”) Vertex has internal discovery programs in this area. The same day, Arbor researchers published aScience study that Vertex’s small molecule approach in CF may eventually identified four new Type V CRISPR endonucleases — Cas12c, cover about 90% of patients by using combinations of CFTR Cas12g, Cas12h and Cas12i — with unique features that apply potentiators and correctors. Potentiators help open the mutated to diagnostics, base editing and high-specificity cleavage of a protein channel at the cell surface to increase chloride flow, and broad range of target sites. correctors help the mutated protein form the right shape to The partnership is the third in just over three years that indicate traffic to the cell surface. Vertex is branching out the modality base of its early pipeline. The company’s approved therapies — Symdeko tezacaftor/ “It’s a high order priority for us to allocate capital outside the ivacaftor, Orkambi lumacaftor/ivacaftor and Kalydeco ivacaftor company to do collaborations to give us opportunities with — are in several Phase II and Phase III trials studying triple- more modalities and more medicines for diseases that we would therapy combinations with Vertex’s investigational treatments be interested in,” COO and EVP Ian Smith told investors on the to address the 90% of CF patients who express some form of the company’s 3Q earnings call last year. CFTR channel. In October 2015, Vertex partnered with CRISPR Therapeutics Vertex told BioCentury that gene editing is one of several AG to gain rights to up to six CRISPR-Cas9-based therapies strategies it is using to address the 10% of patients who don’t for cystic fibrosis, sickle cell disease and other indications for make the CFTR protein. For example, the mRNA approach $105 million up front, including $75 million in cash and a $30 aims to produce functional copies of the CFTR channel. million equity investment. CRISPR is eligible for $420 million With gene editing, the goal would be to correct the CFTR in milestones, plus royalties, for each of the six. mutations in the lung epithelial cells that line the airways. Vertex In July 2016, Vertex added mRNA therapeutics to its repertoire declined to say whether it expects to ultimately extend these through a deal with Moderna Therapeutics Inc. to explore approaches to the full patient population. “Vertex is committed

BIOCENTURY INNOVATIONS ©2019 BIOCENTURY PUBLICATIONS, INC. to bringing effective treatments to all patients with CF,” the homing, engineered form of Cas13d could edit pathological tau company told BioCentury in an emailed statement. isoforms responsible for frontotemporal dementia. The deal with Arbor gives Vertex an array of gene editing tools Arbor said its deal with Vertex allows it to realize the translational with different properties, to complement the existing pipeline. potential of the newly discovered isoforms. Vertex told BioCentury the deal includes a $30 million upfront “We’re excited to see how the newly published Cas12 family payment to Arbor, a $15 million convertible note and mid- enzymes can potentially treat serious human diseases such as single digit royalties. cystic fibrosis,” said an Arbor spokesperson. The partners will work together to discover proteins and tools All four new enzymes have a collateral cleavage property ideal that Vertex may use to develop new therapies, with a focus for diagnostic applications; Cas12h is about 33% smaller than on discovering novel programmable DNA endonucleases or Cas9 — an advantage for delivery, Cas12i creates double- nickase and transduction approaches. Most of the discovery stranded DNA nicks for base editing, and Cas12c offers a broad activities will be conducted by Arbor and funded by Vertex. range of target sites (see Figure: “CRISPR Enzyme Lineup”). Under the terms of the deal, Vertex has an exclusive license to In the Science study, the Arbor team demonstrated that the new develop gene editing therapy products for five diseases including endonucleases join Cas13, Cas14 and other Cas12 family members CF, using CRISPR DNA endonucleases discovered through the as diagnostic tools with collateral cleavage properties. When collaboration. Cas12g encounters a target RNA sequence, and when the others encounter a target DNA sequence, they cleave the target sites then ENZYME DIVERSITY indiscriminately cleave RNA and/or single-stranded DNA. Arbor emerged from stealth mode last year with $15.6 million Two companies, Mammoth Biosciences and Dahlia Biosciences in series A funding, a paper highlighting a novel CRISPR Inc., have disclosed technologies to create DNA- and RNA-based enzyme, and a novel biodiscovery platform. Arbor’s scientific CRISPR diagnostics that rely on a combination of enzymes with founders include Feng Zhang, one of the pioneers of CRISPR- collateral cleavage activity to detect DNA or RNA sequences. based gene editing, and David Walt, a co-founder of Illumina Inc. and Quanterix Corp. The paper also identified Cas12h, which “represents the smallest double-stranded DNA targeting system discovered to date,” said Zhang, a core institute member at the Broad Institute of MIT an Arbor spokesperson. and Harvard and co-founder of Editas Medicine Inc., sits on the opposite side of the CRISPR-Cas9 patent dispute from While the most commonly used Cas9 orthologs are in the 1,300 CRISPR Therapeutics, which joins the University of California amino acid range, Cas12h is between 870 and 933 amino acids in disputing the inventorship of the technology. long. According to Arbor, the smaller size may enable a wider range of delivery options for gene editing. But Arbor’s technology is one of several discoveries that offer paths around that IP dispute, which focuses on CRISPR-based Cas12h is also able to target sites in the genome with a different gene editing using Cas9. Several other CRISPR enzymes have protospacer adjacent motif (PAM) sequence than those targeted been identified that expand the possible applications and offer by Cas9. A PAM is a short nucleotide sequence required at a different editing properties from Cas9 (see“ Cutting Edge target site for the CRISPR endonuclease to bind. Diagnostics”). The team also demonstrated that Cas12i is the first example Using its protein biodiscovery platform to mine bacterial of a Type V enzyme with predominant double-stranded DNA genomes, Arbor is expanding the collection of CRISPR nickase activity. enzymes. The company has not disclosed immediate plans to The DNA nickase preferentially cuts the DNA strand that is non- develop therapies in-house. complementary to the guide RNA. Creating double-stranded The newly described enzymes add to Cas13d, the first enzyme breaks requires that paired nickases each hit the correct site, Arbor disclosed upon its launch. Cas13d is small compared driving down off-target double-stranded edits. with other Cas enzymes at just over 900 amino acids, and has Other benefits of nickases are increased gene insertion accuracy properties well suited for diagnostics. A group at Salk Institute due to the long overhangs created by the nicks, and utility in base for Biological Studies discovered in parallel that a nucleus- editing, which involves using deactivated Cas9 or Cas nickases plus deaminase enzymes to convert one target base to another.

The original CRISPR-based gene editing technology was based on the has a minimal PAM sequence of either one (TN) or two (TG) nucleotides, endonuclease Cas9, which sits at the center of the disputed IP. An assortment creating fewer restrictions on target sites than enzymes with more complex of alternative enzymes have since been discovered. Vertex Pharmaceuticals PAMs. The figure includes PAM sequences, which can vary by species of the Inc. has partnered with Arbor Biotechnologies Inc. to access its CRISPR toolkit, most commonly used orthologs. which includes five enzymes the startup discovered: Cas12c, Cas12g, Cas12h, The type of cut impacts the types of repair mechanisms used. Staggered cuts Cas12i and Cas13d. used by Cas12a improve homology-directed repair efficiency needed for gene Each of the enzymes has different features, lending to use in gene knockdown insertion, while Cas12i's nicks, which are created by preferentially cutting only or correction, base editing, or diagnostics. The enzyme properties vary by the one of the DNA strands, triggers base excision repair ideal for editing a single type of nucleic acid it targets, size, protospacer adjacent motif (PAM), type of nucleotide position. cut and collateral cleavage capability. With the exception of Cas9, the CRISPR endonucleases have a collateral The figure shows the landscape of reported Cas enzymes with translational cleavage property useful for diagnostics. The enzymes can indiscriminately relevance. Cas enzymes (green circles) cleave a target DNA or RNA site cleave ssDNA and/or RNA after cleaving the target site. determined by a guide RNA, which is made up of a crRNA and sometimes Cas14 targets sites on single stranded DNA (ssDNA), which may be useful for a tracrRNA. To bind a target, most Cas enzymes require a short nucleotide targeting pathogenic ssDNA from bacteria and viruses without the risk of off- sequence called a protospacer adjacent motif (PAM) at the site. target effects on human dsDNA. At least six Cas enzymes target double-stranded DNA (dsDNA), including At least four enzymes (Cas12g, Cas13a, Cas13b and Cas13d) target specific Cas9 and Cas12a, which are being used by drug developers to create CRISPR- sites on single-stranded DNA and could be useful for targeting or detecting based therapeutics. certain viruses. For DNA-targeting CRISPR tools, smaller sizes mean easier packaging into Cas9 - CRISPR-associated protein 9; Cas12a (Cpf1) - CRISPR from Prevotella vectors for delivery. Arbor’s Cas12h is the smallest published DNA-targeting and Francisella 1; Cas12c - CRISPR-associated protein 12c; Cas12g - CRISPR- Type V CRISPR endonuclease. Enzyme sizes are measured in amino acids (aa) associated protein 12g; Cas12h - CRISPR-associated protein 12h; Cas12i - and reflect the lengths of the most commonly used and studied orthologs. CRISPR-associated protein 12i; Cas13a (C2c2) - CRISPR-associated protein 13a; Because each enzyme uses a distinct PAM sequence, having a collection of Cas13b - CRISPR-associated protein 13b; Cas13d - CRISPR-associated protein different CRISPR endonucleases creates more potential target sites. Cas12c 13d; Cas14 - CRISPR-associated protein 14

dsDNA

NGG TTN TN RTR TTN

crRNA crRNA crRNA crRNA crRNA tracrRNA tracrRNA

Cas9 Cas12a, Cas12b Cas12c Cas12h Cas12i

SIZE: 1368-1424aa 1353, 1129aa 1209-1330aa 870-933aa 1033-1093aa PAM: NGG TTN TN or TG RTR TTN CUT: Blunt Staggered Blunt Blunt Nicks COLLATERAL No Yes Yes Yes Yes CLEAVAGE:

ssDNA ssRNA

crRNA crRNA crRNA

tracrRNA

tracrRNA Cas14 Cas12g Cas13a, Cas13b, Cas13d

SIZE: 400-700aa 720-830aa 930-1250aa PAM: None None None CUT: Single strand Single strand Single strand COLLATERAL Yes Yes Yes CLEAVAGE:

BIOCENTURY INNOVATIONS ©2019 BIOCENTURY PUBLICATIONS, INC. Base editing company Beam Therapeutics has a platform that Homology Medicines Inc. also is taking an ex vivo approach to uses deactivated Cas9 and Cas13. sickle cell, only with adeno-associated viral (AAV) vectors that The last enzyme, Cas12c, was previously described but never induce gene editing via recombination rather than DNA cutting. characterized due to incomplete genomic data. Arbor showed The program is in discovery and partnered with Novartis. one Cas12c ortholog requires a minimal, single nucleotide PAM While Vertex hasn’t disclosed any gene editing programs in sequence next to the target site in order to bind. Cas12c should AAT deficiency or pain — the two other disease areas in its therefore be able to access a broader range of target sites than pipeline — Intellia Therapeutics Inc. and Editas both havein Cas9, which requires a three-nucleotide PAM. vivo CRISPR gene editing programs for AAT deficiency. Arbor told BioCentury the company has identified other CRISPR- At last year’s Morgan Stanley Global Health Care Conference associated enzymes that have not yet been published. It has not in New York, Ian Smith, EVP and COO of Vertex, said the announced any other research collaborations or licensing deals. company is considering how multiple modalities could be used to approach Vertex’s newly validated pain target NaV1.8; the APPLYING GENE EDITING company hasn’t reported any gene editing programs for the Vertex is using its gene editing technologies in three disclosed indication. pipeline areas — CF, sickle cell disease and β-thalassemia — but has rights to CRISPR technologies in additional undisclosed COMPANIES AND INSTITUTIONS MENTIONED indications, and has hinted that it might explore gene editing Arbor Biotechnologies Inc., Cambridge, Mass. for pain. Beam Therapeutics, Cambridge, Mass. In CF, Vertex is using CRISPR-based gene editing to correct mutant Broad Institute of MIT and Harvard, Cambridge, Mass. CFTR and address the patients unserved by its small molecules. CRISPR Therapeutics AG (NASDAQ:CRSP), Zug, Switzerland Dahlia Biosciences Inc., San Francisco, Calif. At least two other companies are also exploring gene editing for CF. Editas Medicine Inc. (NASDAQ:EDIT), Cambridge, Mass. Editas has a CRISPR-based CF program in its pipeline; and Etagen Etagen Pharma, Cambridge, Mass. Pharma is using its genome editing oligomers for the indication. Homology Medicines Inc. (NASDAQ:FIXX), Bedford, Mass. CRISPR did not respond in time for publication. Illumina Inc. (NASDAQ:ILMN), San Diego, Calif. Intellia Therapeutics Inc. (NASDAQ:NTLA), Cambridge, Mass. Vertex is using its CRISPR Therapeutics deal to test gene editing Mammoth Biosciences, Santa Clara, Calif. for sickle cell disease and β-thalassemia. Moderna Inc. (NASDAQ:MRNA), Cambridge, Mass. The companies are co-developing and co-commercializing Quanterix Corp. (NASDAQ:QTRX), Lexington, Mass. CRISPR Therapeutics’ lead candidate CTX001, a gene edited University of California, Oakland, Calif. stem cell therapy in Phase I/II testing for sickle cell disease in Salk Institute for Biological Studies, La Jolla, Calif. the U.S. and β -thalassemia in Europe. CTX001 has fast track Sangamo Therapeutics Inc. (NASDAQ:SGMO), Richmond, Calif. designation in the U.S. Sanofi(Euronext:SAN; NASDAQ:SNY), Paris, France Vertex Pharmaceuticals Inc. (NASDAQ:VRTX), Boston, Mass. Sickle cell and β-thalassemia are a primary focus of gene editing companies because the technology can restore hemoglobin TARGETS production in two ways: through ex vivo engineering of stem Cas12a (Cpf1) - CRISPR from Prevotella and Francisella 1 cells to them produce functional hemoglobin before delivery Cas12c - CRISPR-associated protein 12c back to patients, and through in vivo correction of the point Cas12g - CRISPR-associated protein 12g mutations responsible for the diseases. Cas12h - CRISPR-associated protein 12h Cas12i - CRISPR-associated protein 12i Vertex and CRISPR Therapeutics are taking the former approach. Cas13d - CRISPR-associated protein 13d Sangamo Therapeutics Inc. and Editas are doing the same, using Cas9 - CRISPR-associated protein 9 zinc finger nucleases and CRISPR, respectively, while Etagen CFTR - Cystic fibrosis transmembrane conductance regulator is employing in vivo gene correction for the two indications. NaV1.8 (PN3; SCN10A) - Sodium voltage-gated channel α subunit 10 Sangamo’s program is partnered with Sanofi and is in Phase I/II, tau (MAPT; FTDP-17) - Microtubule-associated protein τ Editas’ and Etagen’s programs are in discovery.

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