NeuBase chairman, CEO, and founder Dietrich A. Stephan, PhD

A Pittsburgh developer of treatments says it will spend as much as the next two years bringing its lead next-generation antisense oligonucleotide (ASO) therapies into the clinic, and growing its pipeline beyond those candidates, after announcing positive preclinical data it said validates its proprietary Peptide-Nucleic Acid Antisense Oligos (PATrOL™) platform.

NeuBase Therapeutics says the data—generated from a pharmacokinetic study in non-human primates (NHPs) and in vitro pharmacodynamics studies in patient-derived Huntington’s disease (HD) cell lines—supports its decision to advance the development of its lead programs in HD and myotonic dystrophy type 1 (DM1), and expand its pipeline into other indications.

“Now that we have the pharmacokinetic data in hand that tells us where the drugs go after systemic administration, we can in an informed way start to pick new indications,” NeuBase chairman, CEO, and founder Dietrich Stephan, PhD, told GEN Edge. “The two sets of indications that we will be evaluating are monogenic diseases—the 5,000 to 7,000 of the dominant rare x-linked disease. But also oncology indications.”

Would NeuBase consider expanding into development of a COVID-19 therapeutic, as more than 150 companies have done, according to GEN research to be published soon in an upcoming A-List?

That’s a great question. Our technology is perfectly suited to address the virus, in that it’s a single-stranded RNA virus. And we’ve got drugs that target single strands of RNA. It can’t be more perfectly suited from that perspective,” Stephan replied. “Secondly, the biodistribution of the drug is that it goes everywhere. Then, the question is, Is that an appropriate strategic priority for the company at this point in time? We have not made that decision yet.”

Several steps

In the meantime, he added, the company will carry out several steps over the next 18 to 24 months.

“The first is to continue to optimize our candidate compounds in both indications, so that we have a high potency, high selectivity drug. Concurrently, we want to understand the durability of our compounds in animal models well out past a week,” Stephan said. “The third thing that we will do is, define the durability of the therapeutic effect. It’s likely that that will go hand in hand with the presence of the drug, but we need to define that.”

NeuBase also plans to initiate IND-enabling toxicology packages, Stephan continued, then use the NHP pharmacokinetics data it collected to expand into new pipeline programs and organ systems—including into dominant and recessive monogenic diseases, as well as oncology indications.

“We are absolutely continuing to develop therapies on our own and intend to bring them into the marketplace,” Stephan said. “We are open to partnerships in non-core areas, given how many different diseases this platform can potentially address.”

Stephan founded NeuBase in 2019, based on intellectual propertygen developed by the lab of Carnegie Mellon University researcher Danith H. Ly, PhD. He previously chaired the Department of Human Genetics at the University of Pittsburgh from 2013 to 2018, and founded or co-founded 14 biotechnology companies and advised an additional 12 companies.

Among companies he co-founded is Navigenics, the personal genomics pioneer acquired in 2012 by Life Technologies (itself bought by Thermo Fisher Scientific). Stephan co-founded Navigenics with David B. Agus, MD, during a sabbatical from TGen, where he was a senior Investigator and founding Director of TGen’s Neurogenomics Division, and oversaw discovery research.

NeuBase says its PATrOL platform is designed to improve upon current gene silencing technologies by combining the specificity of antisense approaches with the intracellular penetration and broad organ distribution capabilities of small molecule therapeutics.

PATrOL uses short sequences of engineered ASOs, peptide-nucleic acids (PNAs) that are designed to bind mutant RNA to prevent translation or alter splicing to remove harmful protein. While traditional ASOs have a sugar backbone, NeuBase’s PNA therapies are designed with a peptide backbone that according to NeuBase enables the PATrOL platform to be truly modular and highly scalable, permitting rapid development of PATrOL-enabled therapies for many different genetic diseases.

In the NHP study of the HD candidate, quantitative whole-body autoradiography was carried out on Macaca fascicularis NHPs. 5 mg/kg of a Huntington’s disease candidate compound was injected into the tail vein of NHPs, which were sacrificed and flash-frozen at 4 hours, 12 hours and 7 days post-injection. Plasma, whole blood, urine, and feces were collected and assayed.

Remaining in vivo

Following systemic intravenous administration of the HD compound, NeuBase said, a rapid uptake out of circulation was seen, with a half-life in plasma of approximately 1.5 hours. Nearly all (96%) of the drug remained in vivo after a one-week period, while 1.5% of the drug was excreted in urine, and 1.7%, excreted in feces.

“The drug gets into every organ system, including into the brain, and into skeletal muscle, which are the two systems that we’re interested in,” Stephan said.

The HD compound crossed the blood-brain barrier and into the most important deep brain structures, including the caudate. That outcome, according to NeuBase, supports a key capability for the development of the company’s ASO for HD.

The DM1 compound found its way into skeletal muscle, the primary organ system that is affected in the disorder. About 90% of DM1 compound concentrations were achieved in skeletal muscle over the course of one-week post-single-dose administration.

NeuBase says the broad biodistribution of the compounds may enable a potential whole-body therapeutic solution in both DM1 and HD, since both have manifestations outside of the primary affected organ:

Cell line study

In the HD cell line study, multiple candidate compounds were incubated with HD-derived cells and studied for their toxicity and their ability to selectively knock down mutant huntingtin protein (mHTT) expression by engaging with the CAG repeat expansion in the huntingtin (HTT) gene transcript.

Cells were grown for three days and thereafter assayed for cell death. Cell pellets were also collected, lysed, and run on gradient SDS-PAGE gels. Following the transfer of the proteins to a membrane, the membrane was probed with anti-huntingtin and anti-beta-actin antibodies, with secondary antibodies employed to image the immunoblots.

The beta-actin bands were used to normalize the amount of protein across the wells. The amounts of mutant and wild type huntingtin protein in treated cells were compared to untreated cells to determine the level of knockdown.

NeuBase’s study showed the HD candidates successfully engaged target disease-causing transcripts and knocked down resultant malfunctioning mHTT protein levels preferentially over normal HTT protein.

Dose limiting toxicities were not seen compared to a control, either at or above the doses demonstrating activity in human cells in vitro.

As NeuBase picks up the pace of its R&D activity, Stephan said, the company expects to grow its workforce, which now stands at 12 full-time equivalent (FTE) employees based at its Pittsburgh headquarters, plus another six contract FTEs based in India.

“We’re probably going to double in the next I’d say six to nine months,” Stephan projected, “and keep expanding from there.”