The COVID-19 pandemic remains a global threat due to continued transmission of the SARS-CoV-2 virus despite the availability of several intramuscular vaccines. Vaccines administered intramuscularly via injection can elicit strong systemic responses and thus prevent the occurrence of severe disease. These vaccines don’t generally, however, lead to the development of mucosal immunity, which neutralizes the pathogen at the point of entry into the body. Mucosal immunity reduces viral replication at the level of the mucosa, which results in lower carriage by the vaccinated individual, hence reducing transmission.

Effective vaccines delivered via inhalation can lead to both strong mucosal and systemic immunity. The first such product against COVID-19 (Convidecia Air)—a collaborative effort between pharma company CanSino Biologics (CanSinoBIO, Tianjin, China) and respiratory device developer Aerogen (Galway, Ireland)—received regulatory approval in China in early September 2022 (1).

However, developing these vaccines can be challenging. Active substance formulation and delivery-device development must be pursued simultaneously. Once delivered, the vaccination agent must penetrate protective mucosal barriers and survive long enough to stimulate the desired immune response.

Mucosal membranes line the respiratory, digestive, and urogenital tracts and form an interconnected network of mucosal compartments (2–4). Vaccination that leads to mucosal immunity at one of these mucosal sites thus leads to immunity at distant mucosal sites. Mucosal immune responses involve activation of antigen-presenting cells (APCs) and the production of many different types of immune cells, antibodies, and inflammatory cytokines/chemokines.

Targeting mucosal layers in the upper and lower respiratory tract and lungs is an attractive approach to vaccination because most airborne pathogens enter the body through this route and infect these tissues (5). “Generation of mucosal immunity results in activation of many parts of the immune system, not just antibodies. In addition, replication of viruses is prevented in the respiratory system, enabling interruption of the transmission cycle,” explains J. Robert Coleman, CEO of Codagenix.

Numerous advantages to inhalation vaccines

Vaccines delivered via inhalation enable targeting of the respiratory tract mucosa and generation of both humoral and cell-mediated immunity, according to Pierre A. Morgon, executive vice president, Portfolio Strategy and Supranational Affairs and managing director, CanSinoBIO Europe. It is also possible to achieve good drug absorption, fast onset of action, and high bioavailability with inhaled vaccines (6).

In addition, they eliminate the need for needle-based administration, reducing the risk of needle-stick injuries and allowing avoidance of invasive injections, fear of which may keep some people from getting vaccinated (7).

In fact, inhaled vaccines delivered as liquids using nebulizers can potentially be administered with lower requirement for extensive healthcare infrastructure or as many trained healthcare personnel within an immunization center (2). Delivery by inhalation should, therefore, make large-scale mass vaccination easier to conduct, according to Ronan MacLoughlin, associate director of R&D, science and emerging technologies at Aerogen.

Importantly, less vaccine is needed for delivery via inhalation—typically less than 0.2 mL vs. 0.5 mL for injection, says Morgon. “Lower doses means more doses can be produced per batch, and mass vaccination can thus be achieved more cost-efficiently,” he says.

Furthermore, administration via inhalation can be theoretically achieved through the mouth or nose. The preferred route is, says MacLoughlin, entirely dependent on the target receptors for the vaccine. “If nasally expressed, then nasal should be the preferred route. Inhalation into the lung via the nose would allow for both nasal and lung deposition. Oral inhalation allows for a larger dose to be delivered to the lung, given that avoiding the nasal passages will increase the available fraction of aerosol,” he comments.Aerogen’s proprietary aerosol-generating technology converts the liquid vaccine into fine aerosol droplets that are suitable for delivery of the vaccine deep within the lung.

Some are of the opinion that vaccination through the mouth may result in enhanced induction of mucosal immunity. Researchers at McMaster University, for instance, have shown inhalation of a tuberculosis vaccine to be more effective than delivery via nasal sprays, because the vaccine penetrates much deeper into the airway (8).

Because inhaled vaccines provide local immunity to the respiratory tract, they are seen to be ideal solutions for interrupting the spread of viruses with high transmission rates and the potential to lead to global pandemics (9).

The specific types of vaccines that can be formulated for inhalation delivery is not fully understood yet, according to Jean-Denis Shu, SVP of Medical Affairs with CanSinoBIO. “The critical factors are determined by the target receptor, the vaccine’s stability in the lung, and whether sufficient dose can be given to illicit the immune response,” he observes. CanSinoBIO’s inhaled COVID-19 vaccine, now approved in China, is based on an adenovirus-vectored vaccine (10).

“Inhalation delivery offers the potential advantage of bypassing anti-vector immunity for viral vector vaccines for which people may have already developed some immunity through, for instance, exposure to cold viruses,” Morgon adds. The formulation of CanSinoBIO’s inhaled vaccine (Convidecia Air) is the exact same as that of its intramuscular vaccine (Convidecia) and there is no need for an adjuvant to boost immunogenicity.

Codagenix’s inhaled COVID-19 vaccine candidate is a live, attenuated virus that, according to Coleman, safely and stably brings not only the SARS-CoV-2 spike protein, but many other proteins of the virus to the immune system, allowing for broad protection.

Beyond these two approaches, there are inhalation vaccines under development based on attenuated influenza virus, parainfluenza virus (PIV) 5, lentiviruses, Newcastle disease virus (NDV), and vesicular stomatitis virus (VSV); bacterium vectors, nucleic acids (messenger RNA, DNA), and protein subunits (3). Unlike the CanSinoBIO vaccine, the latter require formulation with adjuvants to ensure sufficient immunogenicity (4,11). A range of adjuvants have been explored, such as polyethyleneimine, N-[1-(2,3-Dioleoyloxy) propyl]-N,N,N-trimethylammonium chloride, chitosan, toxoids, cytokines, saponins, lipids, and toll-like receptor agonists.

The need for effective delivery technologies remains the main challenge for realizing successful mucosal vaccines, including those administered via inhalation (12). Not only are strong adjuvants required for many types of vaccines, it can be difficult to maintain the integrity of the vaccines. The vaccination agent must resist degradation in the harsh mucosal environment in order to have time to reach the immune inductive tissue and elicit the desired immune response while also ensuring that oral tolerance does not develop (11). Overall, therefore, appropriate formulation of inhalation vaccines is crucial for their success (13).

Inhalation vaccines can be formulated as liquids or powders. Dry powders have the potential for long-term storage without cold-chain requirements, while liquid formulations are generally simpler to develop and implement, notes MacLoughlin. Dry powder formulations can be generated by various methods, such as spray drying and thin-film freezing (4,6). It is crucial to control the particle size and particle size distribution to achieve the desired release profiles and distribution in the lung.

The aqueous nature of liquid formulations, meanwhile, allows for rapid interaction with the airway epithelium. “Such interactions may be subdued for dry powders due to local conditions in the airway surface lining fluid,” Morgon observes. He also points out that the need for cold-chain management is not an automatic disadvantage for liquid vaccines, because storage and handling capabilities at 3 to 8 °C are generally available at most locations.

Certain attributes are essential for successful inhalation delivery

The physicochemical attributes of vaccine actives delivered intranasally must allow the vaccine to mimic the wild type natural virus that comes in through the airway, according to Coleman. “Such behavior is essential to induce very broad immunity and stop onward transmission,” he contends. He adds that Codagenix chose to use an attenuated live virus rather than a viral vector or protein subunit for this reason. “Using an attenuated virus allows for presentation of multiple viral proteins and avoids any underlying immunity issues, affording increased vaccine uptake,” states Coleman.

Development of optimized vaccine/device combination essential

A consideration equally important to the design of the vaccine active and formulation is the development of an optimized vaccine/device combination, according to Shu. “Picking any delivery device off-the-shelf and applying that increases the risk that the immunization is not effective. A solid understanding of how the vaccine and device interact are critical,” Shu avers.

Morgon agrees. “Understanding formulation, device performance, delivered dose, distribution of dose within the airways, etc. all impact the
likelihood of success.”

“Co-development, such as that conducted by Aerogen and CanSinoBio, has enabled an effective immunization approach that uses a fraction of the injected dose by volume or mg of drug substance,” MacLoughlin concludes.

Devices use for the delivery of inhalation vaccines should, ideally, be patient agnostic. “Taking the variability out of the patient input in the process is a key requirement in order to ensure consistent, reproducible, and reliable dose across a population,” MacLoughlin observes. To this end, Aerogen has focused on development of devices that require the patient to make only a single inhalation for their entire vaccine dose. “This way, there is no requirement for coordination of breath with aerosol generation, which is a key failing of pressurized metered-dose inhalers, and every patient has the same dose available to them for inhalation,” he notes.

Development of mucosal vaccines, including inhalation vaccines, faces an additional hurdle that vaccines administered by other routes generally do not. “The historic body of work in vaccinology has focused on the production of antibodies in the bloodstream. Successful vaccines are determined to be so because they result in high levels of antibodies,” Coleman says.

Some mucosal vaccines, however, may generate lower levels of antibodies in the blood. That does not mean they are less effective, but regulators may not be aware of the differences and thus still compare the data for mucosal vaccines to those for injectable versions, Coleman adds. In the case of CanSinoBIO’s Convidecia Air, the humoral and cellular responses are higher with the inhaled booster than with the injectable one, Morgon comments.

One program designed to address this concern has been established by the World Health Organization (WHO). Its Solidarity Trial Vaccines program supports the development of second-generation COVID-19 vaccines with greater efficacy, greater protection against variants of concern, longer duration of protection, and improved storage and/or simplified delivery with needle-free administration (14). In the Phase III trial of Codagenix’s CoviLiv vaccine candidate, both antibody generation and actual efficacy (determined by comparing vaccinated and placebo groups) are being evaluated, so the actual efficacy rate will be measured. CanSinoBIO has also agreed to the request from WHO to include Convidecia Air in the Solidarity trial.

Several other inhaled COVID-19 vaccines in development

The same group of researchers at McMaster University that compared the delivery of a tuberculosis vaccine via nasal spray and inhalation also have an inhaled, adenoviral-vectored, aerosol COVID-19 candidate in a Phase I clinical trial (8,15).

Scientists at MIT have shown that peptide vaccines with albumin-binding lipid tails when formulated with the adjuvant CpG and administered via inhalation generate long-lasting immune responses (5). A group at University of North Carolina-Chapel Hill, meanwhile, used lung-derived exosomes, or nanoscale vesicles secreted from lung spheroid cells (LSC-Exo), to effectively deliver nucleotide and protein-based vaccines to the lungs via an inhaler (8,16).

In addition to the above inhalation vaccines under development for COVID-19, several groups are investigating the potential of intranasal vaccines intended to induce mucosal immunity. Some of these have been found wanting, most recently the intranasal version of the AstraZeneca viral-vector vaccine (17). These disappointing results, says Morgon, may drive a heightened focus toward the inhalation route.

At the same time, the approval of the CanSinoBIO inhaled COVID-19 vaccine provides evidence that inhalation delivery can be successful at generating mucosal immunity. “We now have significant data on the safety and efficacy of an inhaled vaccine. These data should give others confidence to proceed to trial with what is known to be a broad pipeline of vaccines that may have utility if inhaled,” Morgon says. “Indeed,” he adds, “significant activity has begun/restarted in the area, given that this new route of administration has been shown to be feasible, and on a global mass immunization scale.” Furthermore, if Convidecia Air is convincingly shown to materially reduce viral carriage in vaccinated individuals and therefore curb transmission, CanSinoBIO believes it will turn out to be a blockbuster product.

“The future of inhaled vaccines has just received a significant boost of more useful rapid methods, although further development is required to improve accuracy and repeatability,” he highlights. “Of the different ‘PAT [Process Analytical Technologies] methods’, ATP bioluminescence technology offers a higher sensitivity compared with colorimetric methods.”

In terms of management of cleaning validation processes, software developments are piquing Sandle’s interest, including some that are employing artificial intelligence. “Such packages can help to sort equipment into types where a matrix approach is preferred, as well as enabling scheduling and for setting re-validation targets,” he states.

At its very essence, cleaning validation will require greater consideration, simply as a result of the evolving therapeutic landscape, Kolbert asserts. “The increased use of biological therapeutic agents in pharmaceutical manufacturing will require increased complexity of analytical techniques as activity of the residue is as important as chemical presence,” he summarizes.

Pharmaceutical Cleaning Validation Market: Industry Analysis and Forecast (2021–2027) by Product Type, Validation Test, and Region. Report

CFR Title 21, 210 Part 133.4 (Government Printing Office, Washington, DC).

Validation of Cleaning Processes (7/93), Inspection Guide. 

Cynthia A. Challener, PhD has been a freelance technical writer for over 20 years, leveraging her education from Stanford University (BS) and University of Chicago (PhD) and 10+ years of industry experience. She currently focuses on pharma/biopharma topics, writing technical articles, white papers, blogs, and other content for a variety of clients in addition to contributing regularly to 


Vol. 47, No. 1
January 2023
Page: 26–29, 39

When referring to this article, please cite it as C. Challener. Inhalation Vaccine Development. 

Articles

Development

Inhaled vaccines must resist degradation and penetrate the mucosal lining in the airways and lungs. 

Inhalation Vaccine Development

Of the variety of FDA warning letters sent to bio/pharmaceutical manufacturing and distribution companies at the end of 2022, a good amount seemed to be targeted at those companies trying to market unadulterated COVID-19 treatments. But the agency did have some feedback for bio/pharma manufacturers producing legitimate medicines.

As expected, quality assurance and control were the top issues FDA pointed out to companies. Investigating deviations through corrective actions and preventive actions (CAPA), establishing robust quality control units (QU), and having written procedures in place were mentioned in a variety of warning letters sent to drug manufacturers in 2022.

The following are some notable comments from FDA.

In a warning letter (1) regarding an inspection done in March and April of 2022 at a Phoenix, Ariz. facility, the agency reproached the company for not thoroughly investigating batch discrepancies, failures, and out-of-specification results.

“You failed to effectively implement corrective actions and preventive actions (CAPA) and return your aseptic processing line and manufacturing operations to a state of control. Multiple media fill failures occurred between April and October 2021, when simulating aseptic processing operations on the filling line used for your drug product Abraxane (a sterile (b)(4) drug),” the letter stated.

According to FDA, the company did not adequately perform an evaluation and remediation of the loss of control of aseptic operations and released batches that may have been impacted by the batch failures. “Your investigations into the following media fill failures were inadequate in that they lacked sufficient rigor in determining root causes and scope of impact after the media fills revealed serious non-sterility risks in your aseptic process operations,” the letter stated.

After the agency’s inspection and findings, the company took steps that included a Field Alert Report, batch rejections, and suspension of operations. However, FDA stated these actions were not sufficient. “However, you continued to observe contaminated units in subsequent media fills. The recurring incidents of contaminated units in media fills are an indicator of an adverse trend in your aseptic filling line. You did not provide a comprehensive assessment of contamination risk factors in your aseptic processing operations that support robust and holistic remediations to your aseptic processing operation design. Systemic remediations are essential to address a pattern of recurring contamination events and ensure sustainable control of your aseptic processing operations,” the letter stated.

In response, the company was required to provide FDA with a comprehensive risk assessment of contamination hazards in its aseptic facility and processes. The company must also provide a remediation plan and a CAPA plan to implement routines, operations, and management. An evaluation of the company’s aseptic processing operations was also required by the agency.

Following good manufacturing practices (GMPs) helps produce a quality product that is effective and safe. Having written standard operating procedures (SOPs) is a staple of GMPs. These written procedures not only ensure that employees know the required steps to perform manufacturing operations, they also assure FDA that a company’s processes and methods are sound and efficient.

In a warning letter dated Sept. 27, 2022 (2), FDA inspectors found a company in India had not established adequate written procedures for equipment cleaning and for monitoring control of processing steps that may cause variability in quality characteristics. This lack of written procedures resulted in the agency asking the company to provide an independent, retrospective assessment of effectiveness of their cleaning processes to evaluate the possibility of cross-contamination. A CAPA plan and improvements in the company’s cleaning validation program were also required. The agency also asked for a procedure for a risked-based review of all of the company’s products.

A China facility also received notification from FDA that they lacked written procedures for equipment cleaning (3). “You manufacture drug products using the same equipment that you use to manufacture non-drug industrial products such as (b)(4) and (b)(4). Inadequate removal of residues from manufacturing equipment during cleaning can lead to contamination of drug products subsequently manufactured on the non-dedicated equipment … In your response, you provided a document outlining your procedures for cleaning and disinfecting manufacturing equipment. It is unacceptable as a matter of CGMP to continue manufacturing drugs using the same equipment that you use to manufacture industrial products due to the risk of cross-contamination,” FDA stated in the letter.

In addition to a risk assessment of the products manufactured on the shared equipment, the agency requested a summary of updated SOPs that showed an appropriate validation and verification program for cleaning procedures.

The creation of an adequate quality control unit has been something FDA has continually mentioned in warning letters to drug manufacturing companies. In October 2022, a Mexico facility (4) was given an FDA warning letter that pointed out the company’s quality control unit did not ensure the products manufactured met CGMP compliance or established specifications.

The letter stated the quality unit did not provide adequate oversight and did not have adequate procedures for handling vendor qualification, batch release, reserve samples, investigations, complaints, and training. The QU also did not properly test incoming components or adhere to a stability program.

In another FDA warning letter (5), a China facility failed to fully investigate API impurities in associated batches. “Though you identified at least four lots that failed your (b)(4) impurity specification, you did not extend your investigation to all lots manufactured prior to August 2018 after you identified (b)(4) had a higher risk of forming,” FDA stated in the letter. The firm also did not establish an impurity profile for identified and unidentified potential impurities. “Your firm failed to perform a systematic evaluation of the manufacturing process of (b)(4) API and have robust procedures to identify the potential formation of (b)(4) impurity,” the agency stated.

Computer and systems controls were mentioned in an FDA warning letter to a German facility (6), where the company did not have appropriate controls ensuring changes to master production and control records were limited to authorized personnel. “You failed to have adequate controls in place for your Fourier Transform Infrared Spectrometer (FTIR) system. For example, you did not establish unique usernames and passwords for each analyst. In addition, the audit trail function was disabled even though this Fourier Transform Infrared Spectrometer is routinely used for raw material and finished product testing … Your quality system does not adequately ensure the accuracy and integrity of data to support the safety, effectiveness, and quality of the drugs you manufacture,” the agency stated.

The issues included herein have been seen in a variety of FDA warning letters this year and in the past. It is apparent that the agency is still focused on ensuring that manufacturers are following GMPs and taking product quality seriously.

Warning Letter to Beijing Xinggu Lvsan Technology Co., Ltd. Formerly known as Beijing Lvsan Technology Co.,Ltd

Warning Letter to Eksa Mills S.A. de C.V.

Warning Letter to Zhejiang Tianyu Pharmaceutical Co.. Ltd

Warning Letter to System Kosmetik Produktionsgesellschaft fur kosmetische GmbH


Vol. 47, No. 1
January 2023
Page: 38–39

When referring to this article, please cite it as S. Haigney. Write it Down,
Investigate, and Control. 

Quality/Regulations

FDA sent a variety of warning letters to drug manufacturers at the end of 2022 highlighting myriad CGMP and quality control violations.

Write it Down, Investigate, and Control

The coming year promises to embroil federal health agencies, providers, and biomedical research organizations in a range of contentious issues, as promising research on breakthrough therapies confronts policymakers looking to limit government spending on public health, R&D, and regulatory operations. Politics also will shape global health initiatives to combat antimicrobial resistance and to prepare for future pandemics, while drug coverage and costs remain a hot topic. FDA’s accelerated approval program will be in the spotlight, with manufacturers facing more pressure to withdraw from the market products that fail to confirm efficacy in timely postapproval studies. Drug shortages will also continue
to draw attention and proposed remedies, as rising infection rates send patients to hospitals and drive a surge in demand for treatments.

An anticipated end in 2023 to the public health emergency adopted two years ago to facilitate the approval of new treatments and preventives to COVID-19 will limit FDA emergency use authorizations (EUAs) that have helped speed promising treatments to patients. The change also may limit telehealth flexibilities and broader healthcare coverage for lower income patients.

At the same time, FDA faces tight deadlines for implementing a host of new programs and policies established by recently reauthorized user fee legislation for drugs and medical products. These include initiatives to modernize manufacturing processes and for testing innovative therapies and monitoring their effects. Looking ahead, 2023 promises to be a big year for cell and gene therapies, as seen in FDA’s formation of an expanded Office of Therapeutic Products to handle the surge in regenerative, genetic, and messenger RNA therapies.

Additionally, FDA and industry face a November 2023 deadline for full implementation of the Drug Supply Chain Security Act. This ten-year-old program requires all prescription drugs to be traceable from manufacturers to distributors and pharmacies, but it has been difficult to establish and has been subject to multiple delays.

These and other FDA initiatives require the agency to attract hundreds of new scientists and analysts able to evaluate innovative therapies, handle complex regulatory requirements, and monitor manufacturing operations needed to ensure quality standards. The likely retirement of dozens of senior staffers, moreover, will further challenge agency capabilities.

Notable changes in Congress promise to alter the tone and substance of the legislative debate and put pressure on federal funding of healthcare and biomedical research. New GOP committee chairs in the House are readying probes of Biden administration mismanagement of the pandemic and failure to fully assess the Chinese origins of COVID-19. Republican promises to reduce federal spending also threaten anticipated increases in budgets for FDA, the National Institutes of Health, and Medicare and Medicaid. Cost-cutting for Medicare includes an increase in the eligibility age for seniors, reduced payments to providers and hospitals, and further revisions in Medicare reimbursement and coverage of prescription drugs.

Although Democrats continue to control the Senate, there will be notable changes at the Senate Health, Education, Labor, and Pensions (HELP) Committee, as Sen. Bernie Sanders of Vermont is in line to become chair. Sanders has long championed universal health care and attacked high drug costs, and it remains to be seen where he can gain support from Republicans on the panel.

The battle over abortion drugs will continue to escalate, with pro-life organizations challenging FDA’s approval of mifepristone based on alleged inadequate safety testing. While the suit appears weak to legal experts, this attack strategy will consume considerable agency attention and resources. The fight is slated to go global, moreover, as some advocates work to increase access to abortion pills in other countries, while others seek curbs on abortion access.

Continued Democratic control of the Senate makes it unlikely that Congress will revise the provisions in the Inflation Reduction Act (IRA) that authorized Medicare price negotiations, curbs on certain drug price increases, and a price cap on insulin for Medicare beneficiaries. Democrats still want to extend the insulin cap and price hike limits to all patients, but that may be too costly to advance. Sky-high launch prices on new treatments, though, draw criticism, as seen in the $3.5 million list price for CSL’s new hemophilia B gene therapy (1). FDA leaders are exploring if less costly clinical testing strategies could reduce development costs for such bespoke therapies.

In the name of reducing outlays for medicines, some states will continue to push for FDA authorization of drug import plans, as permitted by policies adopted during the Trump administration. Several states have devised such Canadian drug import programs, but so far, FDA has not approved any due to continued concerns that the administrators cannot prevent distribution of unsafe or unregulated products.

Pharmacy benefit managers (PBMs) also will be in the spotlight, as Congressional leaders move to examine their role in drug access and pricing, issues already under investigation by the Federal Trade Commission (2). Both parties are reviewing the role PBMs play in determining patient outlays for drugs and in influencing Medicare drug coverage and costs.

While FDA avoids direct involvement in drug pricing issues, the agency will continue to support the development and marketing of generic drugs to promote market competition, particularly for products with complex formulations and dosage forms. Some members of Congress look to ensure that generic makers can access innovator products needed for testing and support efforts to advance the development of biosimilars by simplifying testing requirements for interchangeable therapies. The growth in follow-on treatments, though, promises to generate even more patent disputes, as will efforts to limit international patent protections strongly opposed by industry.

CBS News. FDA Approves Most Expensive Drug Ever, a $3.5 Million-per-dose Gene Therapy for Hemophilia B. 

FTC. FTC Launches Inquiry into Prescription Drug Middlemen Industry. Press Release. June 7, 2022.

Jill Wechsler is Washington reporter for 


Vol. 47, No. 1
January 2023
Pages: 14–15

When referring to this article, please cite it as J. Wechsler. A Rocky Road Ahead for FDA and Industry. 

Regulatory Watch

Politics and drug shortages will continue to impact FDA and drug manufacturers in 2023.

A Rocky Road Ahead for FDA and Industry 

For more than a decade, manufacturers have been exploring new ways to improve the efficiency of cell therapy production to make it more cost-effective and accessible to those in need. Many companies have been working in this field, including US-based CAMP4 Therapeutics and Stealth Biotherapeutics’ and France-based TreeFrog Therapeutics SAS, and innovative approaches have now been developed to provide viable stem cells, at scale. 

In 2018, Erwan Bezard, Jean-Luc Treillou, Kevin Alessandri, Laurent Cognet, Maxime Feyeux, and Pierre Nassoy co-founded Bordeaux-based TreeFrog Therapeutics with the sole mission to establish a secure cell production process based on their proprietary 3D C-Stem scalable technology to improve the accessibility and affordability of induced pluripotent stem cell (iPSC)-based therapeutics (1).

In July 2022, Peter Nell, PhD, joined TreeFrog as an independent board member. He previously worked at Bayer Healthcare and has extensive knowledge about business development and licensing and clustered regulatory interspaced short palindromic repeats (CRISPR) technology (2).

In January 2022, the company was recognized as one of the top 25 BioTech start-ups (1). Since its conception, it has attracted US$86.9 million (€80.8 million) in series A and B funding from private equity investors, including Leonard Green and Partners, XAnge, AQUITI Gestion, BNP Paribas Développement, and Bpifrance, as well as Bristol Myers Squibb, Aquitaine Science Transfer, and the European Commission (1).

The company has utilized this funding to leverage its C-Stem technology to generate a pipeline of innovative products which include iPSC dopaminergic neurons to treat Parkinson’s diseases (enter Phase I in 2024); and preclinical programmes using hematopoietic stems cells for bone marrow transplant, cardiomyocytes for myocardial infarction, iPSCs platform and NK and T cells for immune‑oncology (3).

TreeFrog’s lead programme targets Parkinson’s disease via the generation of three-dimensional (3D) neurospheres which contain mature dopaminergic neurons. Following promising pre-clinical studies, the company is expecting to enter first-in-human (FIH) trials in 2024 (3).

On 11 Oct. 2022, TreeFrog’s collaboration with the contract development and manufacturing organization (CDMO) Invetech, came to fruition, and the first good manufacturing practice (GMP)-grade cell encapsulation device was installed and will be used to produce TreeFrog’s PD cell therapy candidate.

The device combines microfluidics and C-Stem cell biology to generate biomimetic alginate capsules seeded with iPSCs. The encapsules iPSCs rapidly expand and differentiate in 3D culture to produce ready-to-transplant microtissues. The capsule helps to reduce the impact impeller shear stresses exerted on iPSCs cells in large-scale bioreactors which can generate more than 1000 capsules per second, and are suited to seed reactors from 200 mL to 10 L (4).

In 2023, the GMP-grade cell encapsulation device will be transferred from TreeFrog to the CDMO to manufacturer clinical grade batches for Treefrog Parkinson’s Disease programme, and three additional GMP encapsulation devices will be delivered to Invetech to support TreeFrog’s pipeline programme.

TreeFrog taps into new technologies through strategic partnerships

TreeFrog has forged several partnerships to tap into new gene editing technologies by leveraging its C-Stem technology.

In June 2022, Treefrog teamed up with iPSC engineering Umoja Biopharma, USA, to use it synthetic rapamycin-activated cytokine receptor (RACR) technology in combination with TreeFrogs C-Stem platform to generate off-the-shelf iPSC-derived allogenic products that target immune cells in solid and haematological malignancies (5).

Geographical expansion to drive C-Stem adoption

TreeFrog has ambitious plans to drive adoption of its C-Stem technology across the globe. The company has started expanding its workforce and facilities; in 2022, the company aimed to double its laboratory capacity for the development and production of iPSC therapies.

During 2022, it opened technological hubs in Boston in the United States and Kobe, Japan, and aimed to establish strategic alliances with leading academic and biopharma players.

In 2022, TreeFrog awarded several Stem Cell SpaceShot grants to leading academic institutions including the University of Sheffield, UK; MCR Laboratory of Molecular Biology, Cambridge, UK; Harvard University, USA; University of California, USA; R&D Center for Cell Therapy, Kobe, Japan; and CNRS, France, which brought together experts in stem cell, gene editing and biophysics to drive further innovations in this field (6).

Investors will be watching this unicorn stock closely to see if the company delivers on its business goals, providing three additional GMP-grade devices to Invetech in 2023 and initiating a FIH trial for iPSC targeting Parkinson’s Diseasein 2024.

Additionally, it is anticipated to bring more news during 2023/2024 regarding further partnerships/licensing deals across the globe, as the company leverage its C-Stem technology with leading academic institutes and biopharma companies.

Competition in this field will increase as new technologies come to the forefront and investment in competitor companies such as CAMP4 Therapeutics (7) and Stealth Biotherapeutics’ (8) have products nearing the market.

Crunchbase. TreeFrog Therapeutics, Company Profile & Funding. TreeFrog.fr (accessed 13 Dec. 2022).

TreeFrog Therapeutics. Peter Nell joins TreeFrog Therapeutics as Independent Board Member. Press Release, 25 July 2022 (accessed 13 Dec. 2022).

TreeFrog Therapeutics. Cell therapy pipeline. TreeFrog.fr (accessed 13 Dec 2022).

TreeFrog Therapeutics. Treefrog Therapeutics Moves to GMP Manufacturing with Cell Encapsulation Device Delivered by Invetech. Press Release. 11 Oct. 2022.

Umoja Biopharma. Umoja Biopharma and TreeFrog Therapeutics Announce Collaboration to Address Current Challenges Facing Ex Vivo Allogeneic Therapies in Immuno-Oncology. Press Release. 10 June 2022.

TreeFrog Therapeutics. 3 PhD Students from King’s College London Win the $100,000 Stem Cell SpaceShot Grant. Press Release. 20 Oct. 2022.

CAMP4. CAMP4 Secures $100 Million Series B Financing to Accelerate Expansion of its Novel Regulatory RNA-Targeting Platform and Advance Lead Programs into the Clinic. Press Release. 20 July 2022.

Stealth BioTherapeutics. Programs and Pipeline. Stealthbt.com. (accessed 13 Dec. 2022). 

Pharmaceutical Technology Europe
Volume 47, No.1
January 2023
Pages 17-18

When referring to this article, please cite it as Barton, C. 

Frontrunners in the Latest Stem Cell Technology

. Pharmaceutical Technology Europe 47 (1) 2023.

European Frontrunners

TreeFrog Therapeutics is leaping ahead in cell therapies through 
resources such as new technologies and investor partnerships.


Pharmaceutical Technology Europe, January 2023

Inhalation Vaccine Development

Write it Down, Investigate, and Control

A Rocky Road Ahead for FDA and Industry

Frontrunners in the Latest Stem Cell Technology