Next Phase Newsletter

Juan Carlos Lopez

Andy Marshall

On September 11, the Lasker Foundation awarded the 2025 Lasker~DeBakey Clinical Medical Research Award to Michael Welsh, Jesús González and Paul Negulescu for discoveries that led to the development of Trikafta, a triple combination of cystic fibrosis transmembrane conductance regulator (CFTR) potentiators and correctors to treat cystic fibrosis. This award recognizes the contribution of Trikafta to improving the quality of life of ~90% of the 40,000 people living with this condition in the United States, reducing infection-related hospitalizations and lung transplants, among other benefits.

But what about the other 10% of patients who don’t respond to Trikafta, many of whom carry so-called Class I alleles that cannot be rescued by this drug combination? Although a lot of progress has been made, several obstacles lie in the path of effective medicines for people who produce no, or negligible amounts of, CFTR protein.

It should come as no surprise that the main therapeutic strategies for Class I alleles aim to put missing CFTR back into lung cells. Among these strategies, mRNA delivery is the most advanced. VX-522, an RNA therapeutic program from Vertex and Moderna currently in Phase 2, is an inhaled drug that aims to deliver full-length CFTR mRNA to the lung using lipid nanoparticles (LNPs). Two related, competing mRNA delivery programs are at a similar stage of clinical development: ARCT-032 by Arcturus Therapeutics using their LUNAR LNPs; and RCT-2100 by ReCode Therapeutics, which uses a lung-targeted SORT (selective organ-targeting) LNP.

A key feature of RNA-based therapies is that any therapeutic benefit would likely be transient, requiring periodic administration of the medicine to achieve sustained effects. Gene therapy and gene editing have the potential to be a curative, “one and done” procedure. Thus far, however, only gene therapy programs have advanced far enough to be in human testing.

Of these, 4D Molecular Therapeutics’ 4D-710 and Spirovants’ SP-101 use different AAV subtypes designed to optimize delivery to airway basal epithelial cells of a CFTR minigene that lacks the regulatory domain. Both projects are in Phase 1/2 of clinical development.

As the large size (6.2 kb) of the CFTR transgene exceeds the packaging capacity of AAV vectors, Krystal Biotech and Boehringer Ingelheim have launched Phase 1/2 clinical programs using viral vectors with a greater payload capacity: KB407 is a re-dosable herpes simplex virus (HSV)-1 vector with a cargo capacity >30 kb that delivers two copies of the CFTR gene to lung epithelial cells using a nebulizer. BI 3720931 is Boehringer’s inhaled lentiviral vector pseudotyped with Sendai virus F and HN envelope proteins (rSIV.F/HN) engineered to deliver a single copy of the CFTR gene. Further behind in the pipeline, Carbon Biosciences’ CGT-001 is a nebulized non-AAV parvovirus-based vector capable of delivering full-length CFTR gene. Thus far, it has been tested in nonhuman primates and in human bronchial cells in culture.

Companies are also pursuing oligonucleotide therapies to modify disease-causing mutations at the RNA level. SPL84 is an inhaled antisense oligonucleotide (ASO) addressing a splicing defect (cryptic exon; class V mutation) in the ~1,600 CF patients who carry the 3849+10kb C→T mutation. SpliSense has advanced the ASO into phase 2 testing, but it also has in preclinical development an exon-skipping ASO against the class I mutant W1282X. By masking the mutant premature termination codon in exon 23, SP23 induces the splicing machinery to skip exon 23 and stitch together exon 22 and exon 24, forming a partially functional CFTRΔex23 protein.

Gene editing is also beginning to appear on the therapeutic horizon. In July, Prime Medicine announced it had received $25 million in funding to advance prime editors, with a lead program focusing on G542X. Last year, Intellia Therapeutics and ReCode Therapeutics also announced a strategic collaboration to combine the CRISPR pioneer’s Cas9 DNA ‘writing’/insertion technology with Recode’s SORT LNPs. Academic groups have now shown that G542X correction is possible using inhaled LNP- or virus–like particle-delivered adenine base editors. And for RNA editing, at this year’s American Society of Gene & Cell Therapy Wave Life Sciences reported their oligo-based ADAR editors could achieve 21% correction (EC50 = 376nM) of CFTR W1282X nonsense mutations. This is likely a sliver of all the therapeutic activity underway; other programs are targeting mucus itself, which is much thicker than in healthy individuals. If we missed any drug-discovery projects in this space, please let us know!

Despite the plethora of programs, developing genetic therapies against cystic fibrosis patients with class I CFTR mutations faces some stiff translational challenges. For starters, targeted delivery of drugs to lung tissue remains a work in progress. The optimal cell type to be targeted by gene therapy/editing remains an open question, especially as the community continues to identify new cell types in the lung; is it enough to target the more prevalent epithelial cells (alveolar type 2 cells), or will it be necessary to target rarer stem cells (alveolar type 1 cells) to see a long-lasting therapeutic effect? What about the contribution of genetic modifiers and other ion channels known to affect airway dysfunction in CF airway epithelial cells? Also, how to figure out the pharmacokinetics and pharmacodynamics of these disease-modifying therapies in lungs and measure delivery in patients? Specifically, establishing protein expression levels after inhaling a DNA- or RNA-based product would likely require a bronchial biopsy, which is impractical particularly in this fragile patient population.

Last, not unlike most pathologies, new animal and in vitro models with predictive value need to be developed. The use of human bronchial epithelium culture is not as predictive of the efficacy of genetic therapies as it has been for small molecules. At present, the ferret is the gold standard disease model. But it is a time-consuming, challenging animal model, which is only supported by a few groups. All of which slows the path to clinical translation.

Six years after the approval of Trikafta, patient foundations like the CF Foundation​, Emily’s Entourage, and the Cystic Fibrosis Trust are devoting increasing resources to translational research to push forward treatments for patients with CFTR Class I mutations who do not respond to potentiators and correctors. The Lasker recognition of the science that led to Trikafta will surely inspire researchers working on those projects to overcome the remaining hurdles.

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Juan Carlos Lopez

Andy Marshall

It is now nearly a decade since Dublin-based startup ​Inflazome burst onto the scene with the description of MCC950, the first nanomolar selective inhibitor of the inflammasome. Inflammasome-mediated low-grade inflammation has been associated with cancers, numerous chronic complex diseases—including inflammatory bowel disease, arthritis, metabolic dysfunction-associated steatohepatitis (MASH), atherosclerosis, Alzheimer’s, Parkinson’s and ALS—as well as rare autoinflammatory diseases, such as cryopyrin-associated periodic syndromes (CAPS). There is a wealth of compelling human genetic evidence from Muckle-Wells syndrome and other autosomal dominant familial genetic diseases showing gain-of-function mutations in NLRP3 are causative of autoinflammatory disease.

Given the ‘pipeline in a product’ potential of drugs targeting this pathway, big pharma has shown considerable interest, with Genentech/Roche snapping up Jecure Therapeutics for an undisclosed amount, and both Novartis and Roche splashing out hundreds of millions of dollars for pioneer companies IFM Tre and Inflazome, respectively. In 2022, Novo Nordisk licensed Ventus Therapeutics’ peripherally restricted NLRP3 inhibitor in a deal worth up to $703 million, lending weight to pharmacological inhibition of NLRP3 as a complement to glucagon-like peptide-1 agonists (GLP-1s) in cardiometabolic disease. And with several programs now entering the clinic, investment activity in the area has continued, with Enveda’s announcement last week of a $150 million series D round to fund a phase 1 trial for ENV-6946, an orally delivered gut-restricted small molecule targeting the NLRP3/tumor necrosis factor-like cytokine 1A (TL1A) pathway in inflammatory bowel disease.

Today, Haystack counts at least 17 independent companies pursuing inflammasome therapeutics (AC Immune, Azome Therapeutics, BioAge Labs, Cardiol Therapeutics, EpicentRx, Enveda, Halia Therapeutics, InflammX Therapeutics, Insilico Medicine, Neumora Therapeutics, NodThera, Olatec Therapeutics, Shaperon, Ventus Therapeutics, Ventyx Biosciences, ZyVersa Therapeutics and Zydus LifeSciences) and 8 programs now in clinical testing specifically targeting the key inflammasome component NLRP3.

While drugmakers have traditionally targeted downstream extracellular mediators of the inflammasome pathway (canakinumab or rilonacept against IL-1β or anakinra to block IL-1 receptor), NLRP3 represents a key upstream intracellular signaling hub, activated by innate immune pattern-recognition receptor (Toll like receptors 2/4) signaling via MyD88 and NFkappaB. Once activated, NLRP3 monomers unfold and associate into a massive 1.2 MDa oligomeric supracomplex with three other proteins: ASC, NEK7 and caspase 1. The mature complex then cleaves and activates proinflammatory cytokines interleukin (IL)-1β and IL-18 and primes gasdermin D to instigate cell pore formation and cell death via pyroptosis.

Discovering effective drugs against NLRP3 has proven challenging. The first NMR structure was obtained in 2016, but the structural basis for how NLRP3 ring-like oligomers associate with intracellular membranes and how its pyrin domains associate with ASC to orchestrate speck formation and caspase activation have only recently been elucidated. Thus far, the majority of small-molecule inhibitors (e.g., Inflazyme’s archetypal MCC950 and inzolemid; Zydus’s (ZYIL1), Olatec’s OLT117 and Jecure Therapeutics’ GDC-2394) form hydrogen bonds via a sulfonylurea group to NLRP3’s NACHT domain nucleotide-binding motifs, thereby obstructing ATP hydrolysis. Other companies are taking a different tack: thus, Halia Therapeutic’s small-molecule inhibitor orniflast and Monte Rosa Therapeutics’ MRT-8102 molecular glue target NEK7 rather than NLRP3.

But it has been less than straightforward to identify compounds with sufficient potency to target this pivotal innate immune signaling pathway without debilitating off-target effects. Indeed, several of the first wave of compounds entering the clinic have been dogged by serious toxicities, including liver problems (MCC950 and GDC-2394) and hypoglycemia (glyburide). Now, a team led by Rebecca Coll (Queen’s University Belfast) and Kevin Wilhelmsen (of BioAge Labs) reports in The Journal of Experimental Medicine the discovery and characterization of BAL-0028, a novel and selective small-molecule inhibitor of the human NLRP3 inflammasome.

Unlike previously studied inhibitors, BAL-0028 acts through a unique mechanism of action; it binds NLRP3’s NACHT domain at a site distinct from other inhibitors that act by directly interfering with ATPase activity. BAL-0028 has nanomolar potency against human and primate NLRP3 but, remarkably, has weak activity against the mouse target, highlighting species-specific differences.

As BAL-0028 showed very high plasma protein binding in mice, limiting its use in vivo, the team developed a derivative, BAL-0598, with improved pharmacokinetic properties. In a humanized NLRP3 mouse peritonitis model, BAL-0598 effectively reduced IL-1β and IL-6 production, confirming its anti-inflammatory activity in vivo. Importantly, both BAL-0028 and BAL-0598 inhibited hyperactive NLRP3 mutants associated with autoinflammatory diseases, in some cases more effectively than Vertex’s VX-765, a caspase 1 inhibitor, and compounds like MCC950, one of the best characterized NLRP3 inhibitors available.

The novel mechanism of action of BAL-0028 and BAL-0598 would suggest their off-target effects may be different from those associated with other NLRP3 inhibitors blocking ATP hydrolysis. The concern that such compounds might also bind other members of the NOD/NLR family (e.g., NLRP1, NLRP4 or AIM2 inflammasomes) is mitigated by most published studies indicating that NLRP3’s unique fold around the ATP binding site makes small-molecule binders selective for this family member alone. The most likely explanation from trials published to date is that the observed toxicities are associated with small molecule chemotype rather than any NLRP3 class-specific problem. In any case, the findings from this study support further investigation of these compounds as candidates for treating inflammatory and age-related diseases where NLRP3 plays a role. The race to develop a safe and effective NLRP3 inhibitor is on, with big pharma billion-dollar bets and startups jostling to create best-in-class assets across cancer, cardiovascular, neurodegenerative and metabolic disease.

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