Topic: Cell Therapy

Stem cells have revolutionized the field of regenerative medicine, offering promising solutions for various medical conditions, including difficult-to-heal skin wounds. This review focuses on the background and manufacturing processes of skin cell-based therapies, particularly keratinocytes and adipose-derived mesenchymal stem cells (AD-MSCs), as highlighted in the thesis by Hady Shahin[1]. This article aims to provide a comprehensive overview for Contract Development and Manufacturing Organizations (CDMOs) involved in cell-based solution production.

Stem cell basics

Stem cells have unique abilities to self-renew and to recreate functional tissues. They can develop into many different cell types in the body during early life and growth[2]. Researchers study many different types of stem cells, including pluripotent stem cells (embryonic stem cells and induced pluripotent stem cells) and non-embryonic or somatic stem cells (commonly called adult stem cells)[2]. Pluripotent stem cells have the ability to differentiate into cells of the 3 main germ layers in the adult body[2].  Adult stem cells are found in specific anatomical locations and can differentiate to yield the specialized cell types of that tissue or organ[2]. They serve as an internal repair system that generates replacements for cells lost through normal wear and tear, injury, or disease.

Properties of stem cells

Stem cells have the remarkable potential to renew themselves and to differentiate into various specialized cell types[2].  When a stem cell divides, the resulting two daughter cells may be both stem cells, a stem cell and a more differentiated cell, or both more differentiated cells[2]. Discovering the mechanism behind self-renewal may make it possible to understand how cell fate is regulated during normal embryonic development and post-natally, or mis-regulated during aging or in the development of cancer[2].

The skin and regenerative therapies

The skin, the largest organ of the body, serves as a protective barrier against the external environment. It consists of three distinct layers: the epidermis, dermis, and subcutaneous adipose tissue[1]. The epidermis, primarily composed of keratinocytes, plays a crucial role in maintaining skin integrity and facilitating wound healing. Keratinocytes move from the basal layer to the surface, undergoing differentiation and forming a protective barrier[1].

Difficult-to-heal wounds, such as those caused by chronic diseases, trauma, or burns, pose significant challenges in clinical practice. These wounds often result in prolonged pain, infection, and impaired quality of life[1]. Regenerative advanced therapy medicinal products (ATMPs), including cell-based approaches, offer promising solutions for enhancing wound healing and improving patient outcomes[1].

Autologous vs allogenic skin ATMPs

Keratinocytes —the most abundant cell type in the epidermis— are instrumental in the re-epithelialization process during wound healing. They proliferate and migrate to cover the wound bed, forming new epidermal layers[1]. An autologous therapeutic approach involves harvesting skin biopsies from a patient’s healthy donor sites, isolating keratinocytes, expanding them, and reapplying them to the same patient after thorough characterization, quality control, and safety testing. The classical method for culturing keratinocytes includes enzymatic digestion of the epidermis, followed by expansion in culture media [1]. However, a major challenge in this process is the use of animal-derived products, which poses regulatory hurdles [1]. To address these challenges, Shahin’s thesis proposes a xeno-free workflow for keratinocyte isolation and expansion. The study validates the use of a xeno-free workflow to manufacture human keratinocytes as ATMP [1]. This approach ensures the production of keratinocytes that comply with regulatory standards, making them suitable for clinical applications [1].

The allogeneic approach, on the other hand, involves Adipose-Derived Mesenchymal Stem Cells (AD-MSCs) as a promising alternative to address the scalability challenges associated with keratinocytes, which are mature cells. AD-MSCs are multipotent stem cells isolated from adipose (fat) tissue. They possess the ability to differentiate into various cell types, including osteoblasts, chondrocytes, and adipocytes[1]. AD-MSCs are particularly attractive for regenerative therapies due to their ease of isolation, high yield, and immunomodulatory properties[1].

In the context of wound healing, AD-MSCs contribute to tissue repair by promoting angiogenesis, reducing inflammation, and enhancing collagen synthesis[1]. Shahin’s thesis explores the potential of AD-MSCs as an alternative to keratinocytes for treating difficult-to-heal wounds[1]. The study highlights the differentiation of AD-MSCs into keratinocyte-like cells through direct co-culture with keratinocytes[1]. This approach leverages the paracrine signaling between the two cell types to enhance the differentiation process[1].

Manufacturing cell therapies for clinical use

The manufacturing of stem cells for clinical applications involves several critical steps, including cell isolation, expansion, and quality control. Ensuring compliance with Good Manufacturing Practice (GMP) guidelines is essential to produce safe and effective cell-based therapies[1].

By saying something like at NorthX we foster the expertise that can support in bringing cell-therapy into manufacturing scale check out our services in the link

Proposed workflow for manufacturing a cell-based ATMP for wound healing

1. Cell isolation

Keratinocytes are typically isolated from skin biopsies using enzymatic digestion. Shahin’s thesis validates the use of a completely xeno-free keratinocytes extraction method, ensuring the production of GMP-compliant keratinocytes in a timely manner[1]. AD-MSCs are isolated from adipose tissue through enzymatic digestion and centrifugation[1]. The high yield of AD-MSCs from adipose tissue makes them a viable option for large-scale production[1].

2. Cell expansion

The expansion of keratinocytes and AD-MSCs requires optimized culture conditions to maintain cell viability and functionality. Shahin’s study demonstrates the use of xeno-free culture media for keratinocyte expansion, eliminating the need for animal-derived products[1]. For AD-MSCs, the co-culture with keratinocytes enhances their differentiation into keratinocyte-like cells, providing a scalable approach for producing epidermal cells [1].. Rigorous quality control is needed to ensure such in-vitro cell manipulation is safe and does not compromise the properties of the cells. Therefore, thorough characterization and stability testing are needed for cell-therapies to be considered safe for clinical use and to fulfil stringent regulatory requirements for ATMPs.

That can help you design a comprehensive analytics panel for your ATMP (consulting on release criteria with the regulatory bodies)

3. Cell transportation

As cell therapy manufacturing for clinical use must be conducted under strict control in a GMP facility, the final cell solution often needs to be transported from the production site to the treatment site, which may be several hours away. Shahin’s study demonstrated that the cell solution can be transported for up to 24 hours under controlled conditions while maintaining cell functionality and characteristics.

This finding allowed the research team to establish human serum albumin as the preferred carrier solution for keratinocytes in clinical treatments, ensuring their viability and functionality during transport. Additionally, it was validated as the final formulation solution for administration.

4.  Ensuring quality and safety in cell-based ATMP manufacturing

Ensuring the quality and safety of cell-based ATMPs is paramount. Shahin’s thesis emphasizes the importance of thorough characterization of keratinocytes and AD-MSCs, including the assessment of cell viability, differentiation potential, and functionality[1]. The use of cell and molecular characterization methods, including but not limited to immunophenotyping, gene and protein expression analyses are instrumental tools for monitoring and ensuring the quality of the produced cells [1].

Conclusion

The thesis by Hady Shahin offers valuable insights into manufacturing cell-based regenerative solutions for skin healing. The proposed xeno-free workflows and co-culture techniques present promising methods for producing GMP-compliant cell therapies. These advancements pave the way for effective treatments for difficult-to-heal wounds, ultimately improving patient outcomes.

References

1. Shahin, H. (2023). Keratinocytes and Adipose-derived mesenchymal stem cells: The heir and the spare to regenerative cellular therapies for difficult-to-heal skin wounds. Linköping University Medical Dissertation No. 1880.
2. https://liu.diva-portal.org/smash/get/diva2:1810734/FULLTEXT01.pdf
   National Institutes of Health. (2021). Stem Cell Basics. Retrieved from https://stemcells.nih.gov/info/basics/stc-basics.

Neoantigen cancer vaccines represent a groundbreaking advancement in personalized medicine, offering tailored cancer treatments designed for each patient’s unique tumor profile. These therapies require rapid turnaround from tumor identification to clinical delivery — a critical factor when days can mean the difference between life and death for cancer patients. NorthX Biologics is uniquely positioned to meet these demands, providing agile, small-volume, multi-batch production, in-house rapid analytics, and robust supply chain solutions to bring lifesaving therapies to patients faster.

Authors: Ola Tuvesson, Chief Technology Officer, and Isa Lindgren, Ph.D., Head of Analytics, NorthX Biologics

Neoantigen cancer vaccines: a personalized approach gaining momentum

Personalized medicines are revolutionizing treatment paradigms by tailoring therapies to each patient’s unique genetic and biological characteristics. A particularly promising area is neoantigen-based cancer vaccines, which target antigens specific to an individual’s tumor microenvironment, offering a precise and highly individualized approach to oncology.

Although no neoantigen cancer vaccines have received regulatory approval yet, the field is advancing rapidly, with several candidates progressing to phase II and later-stage clinical trials. Both established pharmaceutical companies and agile startups are actively developing these therapies. Messenger RNA (mRNA) remains the most common modality due to its versatility and rapid production capabilities, though DNA-based vaccines are also gaining traction.

While the science behind neoantigen cancer vaccines is robust, critical challenges remain, particularly in the realm of analytics, process development, and chemistry, manufacturing, and control (CMC). For these therapies to advance through clinical trials and reach commercialization, comprehensive characterization and validated processes are essential. Developers must overcome hurdles such as the need for rapid analytics, platform processes, and stringent sterility testing under accelerated timelines. For a patient battling cancer, delays in treatment can mean the difference between life and death, underscoring the urgency of streamlined processes from tumor identification to clinical delivery.

Overcoming regulatory hurdles for neoantigen therapies

The evolution of personalized medicine, including neoantigen-based cancer vaccines, depends on innovative companies willing to pave the way and progressive contract development and manufacturing organizations (CDMOs) capable of supporting these groundbreaking therapies. However, regulatory uncertainty remains a significant hurdle. Current regulatory frameworks, designed for traditional therapies like small molecules and biologics, are not adapted to the needs for personalized medicines, where each product is tailored to an individual patient.

To address this challenge, a paradigm shift in regulatory evaluation is required. Reviewing every individualized therapy on a case-by-case basis is impractical. Instead, a platform-based approach — validating the overall manufacturing process rather than individual batches — is the most feasible path forward. Such a shift would allow regulators to focus on standardizing key processes while permitting minor, patient-specific variations in raw materials.

Critical to this transition is the development of robust, reliable platform analytical methods. These methods must be qualified using a bracketing principle, ensuring that key attributes remain consistent across therapies, regardless of genetic sequence variations. This platform approach can be supplemented with sequence-dependent analytical techniques to confirm the product’s identity, balancing regulatory rigor with the flexibility required for personalized treatments.

Scaling down for personalization

Personalized medicines, such as neoantigen cancer vaccines, require a fundamental shift in biomanufacturing approaches. For decades, the industry has focused on scaling up — boosting titers and increasing batch sizes to efficiently produce biologics for large patient populations. Personalized therapies, however, demand the exact opposite: scaling down to produce one batch per patient. This shift introduces new complexities, requiring innovative solutions to maintain productivity and efficiency and keep cost of goods (COGs) manageable.

With each batch tailored to an individual patient, large-scale bioreactors and processes become impractical and cost-prohibitive. Instead, manufacturers must embrace smaller, highly efficient systems capable of running multiple batches with short product turnovers or even in parallel under GMP conditions. These systems must also integrate streamlined supply chains and pre-positioned raw materials to meet accelerated production timelines.

Orchestrating supply chains for individualized therapies

In personalized medicines like neoantigen cancer vaccines, organizational precision across supply chains and manufacturing operations is paramount. Unlike therapies produced in bulk for large patient populations, personalized treatments require multiple small batches to be manufactured simultaneously, each tailored to a specific patient. This introduces significant logistical challenges, demanding seamless coordination of material inflows, production processes, and product deliveries.

For neoantigen cancer vaccines, the process begins with the collection and analysis of a patient’s tumor sample to identify relevant neoantigens. This analysis triggers a narrow production window, during which the drug product must be manufactured and delivered. For patients with life-threatening cancers, every day matters. Personalized therapies must transition from concept to clinic with unprecedented speed, necessitating CDMOs that excel in agile, efficient production, and robust in-house analytics.

The patient’s journey demands a biomanufacturing process that prioritizes speed without compromising quality. Ensuring the availability of raw materials — pre-positioned and ready for use — is critical. Likewise, platform-based manufacturing processes that allow for rapid initiation and parallel execution are essential to meeting these time-sensitive demands.

From tumor analysis to delivery: breaking down analytical roadblocks

For neoantigen cancer vaccines, the journey begins with identifying the specific genetic mutations within each patient’s tumor — a highly individualized process that forms the foundation for these personalized therapies. This critical first step relies on next-generation sequencing (NGS) and machine learning algorithms to pinpoint relevant neoantigens. While this analysis typically falls under the responsibility of the therapy developer, the subsequent steps in the process require efficient manufacturing and rigorous testing to ensure the final drug product meets quality standards and can be delivered to the patient on time.

Once the relevant antigens have been defined, the focus shifts to ensuring rapid and reliable production and testing. At this critical stage, NorthX Biologics provides comprehensive support, offering streamlined CMC processes and rapid product release testing to minimize delays. NorthX Biologics’ integrated in-house analytics capabilities, including advanced sterility testing solutions, enable efficient product release, ensuring therapies move swiftly from manufacturing to the clinic.

In the case of highly personalized, short-lifespan therapies like neoantigen cancer vaccines, sterility testing can present a significant challenge. While rapid sterility tests have been developed, the process remains time-consuming. NorthX Biologics addresses this challenge by employing closed systems and stringent aseptic controls, reducing the risk of contamination and ensuring product quality. Additionally, the company collaborates with innovators in advanced testing solutions to remain at the forefront of analytical capabilities, minimizing delays that could impact patient outcomes.

A trusted partner for neoantigen cancer vaccine development

NorthX Biologics is uniquely positioned to support the development and manufacturing of personalized neoantigen-based cancer vaccines. The company’s fully integrated in-house capabilities — including specialized analytics for process development and product release — enable efficient, end-to-end support. Recognizing the importance of rapid turnaround, NorthX Biologics collaborates closely with experts in advanced testing solutions, such as accelerated sterility testing, to minimize delays and keep timelines on track.

With decades of experience manufacturing both technical and therapeutic proteins, NorthX Biologics combines its proven expertise with the scientific innovation of its Innovation Hub. This powerful combination has enabled the establishment of a highly agile manufacturing organization and a streamlined, adaptable supply chain. Effective quality control, quality assurance, and product release processes ensure NorthX Biologics can meet the rigorous demands of personalized medicine while maintaining the highest standards.

NorthX Biologics stands out through its “Beyond CDMO” approach, extending beyond traditional manufacturing services to act as an innovation partner, enabler, and strategic guide for its clients. By fostering strong collaborations with suppliers and customers, the company provides customized solutions that address the specific needs of each project. This forward-thinking philosophy reflects a commitment to advancing therapeutic development and delivering personalized medicines faster. Guided by the principle “small enough to care and big enough to deliver,” NorthX Biologics delivers decisions quickly, adapts readily to change, and leverages its deep expertise to help drug developers bring life-changing therapies to patients in need. By combining agility, flexibility, and strategic insight, NorthX Biologics empowers its clients to navigate the complexities of this evolving landscape, ensuring that innovation reaches patients without delay.

Ola Tuvesson
Chief Technology Officer
NorthX Biologics

As CTO, Ola is leading NorthX Biologic’s development and project organization, focusing on delivering technologies and strategies to ensure high-end services within bioprocessing and analytics. He has more than 20 years’ experience from the pharma and biotech industry, ranging from early development to commercial GMP manufacturing. Ola has worked in several fields, including ATMP products, vaccines, and other biologicals. He has the essential knowledge to take a product from early pre-clinical development into clinical trials and to the market.

Isa Lindgren, Ph.D.
Head of Analytics
NorthX Biologics

Isa Lindgren, Ph.D., is Head of Analytics at NorthX Biologics, leading the QC and Analytical Development teams across the Matfors and Stockholm sites. With a background of 15+ years in life sciences research and experience from preclinical work at Chiesi Pharma in biologics and ATMPs, Isa has extensive expertise in analytics. Six years in the US have equipped her with valuable international experience for global communication and high-level customer care. Known for her technological acumen, she ensures NorthX Biologics remains a front-runner in analytics to deliver biologics at the highest quality. 

Phase 1: research and development

The initial phase in the production of cell-based therapies is where innovative ideas are explored and potential therapies are identified. In the dynamic and rapidly evolving field of cell therapy, thorough research and development play a vital role in bringing safe and effective treatments to patients.

During this phase, extensive laboratory work is conducted to understand the underlying mechanisms of action and optimize the manufacturing process. Scientists investigate various cell types, their behavior in different conditions, and their potential therapeutic applications. They also evaluate different techniques for cell isolation, expansion, and characterization.

The goal of Phase 1 of the cell therapy manufacturing process is to establish proof-of-concept and gather sufficient data to support future clinical development. Process controls are put in place to ensure consistency and reproducibility of the manufacturing process. In addition, quality control measures are implemented to meet the strict standards set by regulatory authorities.

Phase 2: preclinical testing

Phase 2 oversees the effectiveness and safety of cell therapy through the use of animal models. This allows researchers to gather valuable data before proceeding to human trials. During Phase 2, extensive characterization studies are conducted on cell products, including functional assays and downstream processing optimization. All these activities adhere to strict guidelines set by organizations like the International Organization for Standardization (ISO).

Here is what you need to know about Phase 2:

• Preclinical animal models: In this phase, various animal species are used to mimic human diseases and evaluate the therapeutic potential of cell-based treatments.
• Release assays: These tests assess the quality and potency of the cells manufactured for therapy. They ensure that only safe and effective products move forward in the development process.
• Process development challenges: Developing robust manufacturing process controls for cell therapies can be complex due to factors like scalability, reproducibility, and regulatory compliance.

Phase 3: clinical trials – phase I

Phase 3 is where the safety and efficacy of the cell therapy will be evaluated in humans. This phase is commonly known as Phase I of clinical trials, which are a crucial step in cell therapy production. In phase I of clinical trials, small groups of individuals receive the experimental treatment for the first time. These trials help researchers determine how cells behave within the human body and evaluate any potential risks associated with their administration. 

Researchers must closely monitor participants for any adverse effects or unexpected responses. The scale of Phase I clinical trials may vary depending on factors such as available funding, study design, and regulatory requirements. Furthermore, meticulous documentation and analysis are essential to gather valuable data regarding dose optimization and patient response.

Phase 4: clinical trials – phase II

Phase II focuses on a larger group of participants, typically ranging from several dozen to a few hundred individuals. During this phase, researchers closely monitor critical process parameters such as cell expansion, ensuring that batches of cell therapy products are manufactured consistently using pluripotent stem cells as raw materials.

Here’s what can be expected during this phase:

• Increased sample size: Phase II trials involve a larger number of participants to gather more data on how the treatment affects different individuals.
• Dose determination: Researchers refine the dosing parameters established in Phase I by testing various doses or schedules to determine the optimal therapeutic effect.
• Control groups: Some participants may receive a placebo or standard treatment as a control group for comparison purposes.
• Efficacy assessment: The primary goal is to assess whether the treatment is effective in treating the specific condition or disease being targeted.

Phase 5: clinical trials – phase III

Phase III clinical trials are the final evaluation of the treatment’s safety and effectiveness. The goal is to ensure that the therapy can be produced consistently and reliably, meeting strict quality standards.

To achieve this, various factors need to be considered:

• Cell sources must be carefully selected to ensure their suitability for therapeutic use. This involves assessing characteristics such as differentiation potential and immune compatibility.
• Optimizing cell densities is crucial to achieving optimal therapeutic outcomes. Researchers aim to determine the ideal number of cells per dose to maximize efficacy while minimizing any potential adverse effects.
• Cell viability is another critical parameter assessed during Phase III clinical trials. It refers to the percentage of viable cells within a given sample and serves as an indicator of product quality. High cell viability is essential for ensuring that patients receive a potent and effective therapy.
• Disposable bioreactors are often used due to their ease of use and scalability. These systems allow for the efficient expansion of cells while maintaining strict control over environmental conditions.

Phase 6: regulatory approval

Regulatory approval ensures that the treatment meets all necessary standards for widespread use. This phase involves obtaining the necessary approvals from regulatory agencies to bring a cell therapy product to market.

Regulatory agencies carefully evaluate the data generated during clinical trials to assess whether the benefits of the treatment outweigh any potential risks. To obtain regulatory approval, several key processes must be followed to demonstrate the safety, efficacy, and quality of the cell therapy product.

The challenges of cell therapy regulatory approval are multifaceted:

• Regulatory agencies require evidence-based documentation on various aspects of cell therapy including manufacturing processes, dosing regimens, adverse event monitoring, and follow-up protocols.
• Ensuring product quality throughout manufacturing and distribution is essential for regulatory compliance.
• Logistical challenges such as cold chain management and timely delivery also need to be addressed.

Phase 7: manufacturing and scale-up

The manufacturing and scale-up phase involves increasing production capacity to meet the demand for cell therapy products. During this phase, the focus is on efficiently producing large quantities of cell therapy materials while maintaining their quality and safety. The first step in this process is selecting the appropriate cell type for the therapy. In autologous cell therapy, the cells are derived from the patient themselves. When cells are sourced from a donor, the process is known as allogeneic cell therapy.

Cell isolation techniques are employed to separate desired cell populations from other components. Process decisions such as media formulation and culture conditions are considered to ensure optimal growth and functionality of the cells. These decisions need to comply with regulations specific to regenerative medicines. Scaling up production involves increasing batch sizes and implementing scalable production methods that can reliably produce a consistent drug product. It is crucial to continually monitor and control critical parameters such as temperature, pH levels, and nutrient supply.

Phase 8: quality control and assurance

During this phase, the quality of the medicine is assessed to meet the required standards for safety and efficacy. Quality control and assurance are crucial in the production of cell therapies to ensure that they are effective and safe for patients.

Here are three important aspects to consider during this phase:

• Quality Control Assays: To verify the identity, purity, potency, and safety of the cell therapies, various assays are performed. These assays include testing for microbial contamination, measuring cell viability and functionality, and confirming genetic stability.
• Chain of Custody: Maintaining a clear chain of custody is essential in ensuring traceability and accountability throughout the production process. This involves proper documentation at each step, from sourcing cells to final product distribution.
• Contract Manufacturing Organization (CMO): Collaborating with cell therapy manufacturers streamlines quality control processes. CMOs have well-established quality systems in place to meet regulatory requirements.

Phase 9: distribution and patient access

A reliable distribution system ensures that patients have access to cell therapy products. A robust supply chain is essential for the large-scale production and distribution of these therapies. The distribution process begins after the cellular product manufacturing is completed and quality control has been performed. Allogeneic products, such as CAR-T cell therapy, require careful handling and storage to maintain their efficacy. 

The downstream process involves packaging, labeling, and shipping the products in accordance with regulatory guidelines. As part of this phase, proper cell collection and counting techniques are employed to ensure that each patient receives the appropriate dose of cells. Quality assurance protocols are implemented throughout the distribution process to guarantee consistent product quality at every step.

Key takeaways

• The phases of cell therapy production include research and development, preclinical testing, and clinical trials in multiple phases.
• Cell characteristics and selection are important factors in the production of cell therapies, including the selection of cell sources and optimizing cell densities.
• Manufacturing processes and scale-up are crucial steps in cell therapy production, including regulatory approval and ensuring manufacturing and scale-up processes are in place.
• Quality control and assurance are essential in cell therapy production, including maintaining product quality, implementing a distribution system, and employing accurate cell collection and counting techniques.

Cell therapy manufacturing services

Collaboration between academic institutions, biotech companies, and cell therapy manufacturers is crucial in the production of life-saving medicines. It promotes knowledge sharing and accelerates advancements in the cell therapy industry.

At NorthX Biologics, we specialize in providing a full suite of services to transform the way cell therapies are developed, produced, and delivered. Thanks to our state-of-the-art technological systems and expertise, you can rest assured knowing that your cell therapy products will be efficient, consistent, and of the highest quality.