How long do mRNA in-vivo studies take? A case study with Matinas and BioNTech

In the rapidly evolving world of medical research, mRNA-based therapies and vaccines have emerged as powerful tools in the fight against various diseases, including COVID-19. As scientists continue to explore the potential applications of mRNA technology, understanding the timeframes involved in conducting mRNA in-vivo studies is crucial for researchers, investors, and the general public.

The duration of mRNA in-vivo studies can vary significantly depending on several factors, including the specific resfearch question, the experimental model being used, and the desired endpoints or readouts. Here are some general timeframes for different aspects of mRNA in-vivo studies:

1.  Study design and preparation: A few weeks to a few months This phase involves defining the research question, identifying the appropriate experimental model (e.g., cell culture, mouse, or non-human primates), and optimizing the experimental conditions.

2.  mRNA synthesis and formulation: A few days to a few weeks During this phase, researchers synthesize the mRNA, optimize the formulation, and perform quality control tests.

3.  In-vivo experiments: A few days to several months This phase encompasses the actual in-vivo experiments, including the administration of the mRNA to the experimental model, monitoring the animals, and evaluating the desired endpoints (e.g., protein expression, immune response, or therapeutic efficacy). The duration of this phase depends on the specific study and can range from a few days for short-term endpoints to several months for long-term endpoints.

4.  Data analysis and interpretation: A few weeks to a few months This phase involves processing and analyzing the data, drawing conclusions from the findings, and potentially conducting follow-up experiments.

mRNA in-vivo studies can take anywhere from a few months to over a year, depending on the complexity and scope of the research. Obviously, these timeframes are approximate and can vary significantly depending on the specific study and the resources available to the research team.

The GI tract and oral mRNA

Matinas BioPharma has commenced in-vivo mRNA studies with BioNTech in an effort to create oral mRNA vaccines. But how long until we know those are successful? Well, it depends. First, they have to formulate it and then feed it to a mouse and see if it survives the perilous journey through the GI tract.

To determine whether an orally administered mRNA has survived the gastrointestinal tract in a mouse model, they would need to assess both the integrity of the mRNA and its ability to enter target cells and produce the desired protein. Here are some approaches that can be used to evaluate this:

1.  Tissue and cell analysis: After administering the oral mRNA, collect tissue samples from various parts of the GI tract, including the stomach, small intestine, and large intestine, as well as the target organs or tissues where the mRNA is expected to exert its effects. Analyze these samples for the presence of the mRNA using techniques such as quantitative reverse transcription polymerase chain reaction (qRT-PCR) or in situ hybridization.

2.  Protein expression: Assess whether the administered mRNA leads to the production of the desired protein by analyzing the collected tissue samples for protein expression. This can be done using techniques such as enzyme-linked immunosorbent assay (ELISA), Western blot, or immunohistochemistry.

3.  Functional assays: In addition to analyzing the presence of the mRNA and the desired protein, you can assess the functionality of the mRNA by looking for evidence of the intended biological effect. For example, if the mRNA is intended to elicit an immune response, you could measure the production of relevant cytokines or the activation of immune cells.

4.  Intact mRNA analysis: To assess the integrity of the mRNA, you can use techniques such as gel electrophoresis, capillary electrophoresis, or RNA integrity number (RIN) analysis. These methods can help determine whether the mRNA remains intact after passing through the GI tract.

5.  Encapsulation strategies: To improve the chances of the mRNA surviving the harsh conditions of the GI tract, researchers often encapsulate the mRNA in protective vehicles such as lipid nanoparticles, polymer-based nanoparticles, or hydrogels. (Or this case, lipid nano-crystals.) You can assess the integrity and release of the mRNA from these vehicles using techniques such as dynamic light scattering (DLS), transmission electron microscopy (TEM), or confocal microscopy.

(From the latest Matinas BioPharma investor’s presentation.)

How long do functional assays take?

The duration of functional assays in the context of mRNA studies depends on the specific assay being performed, the desired readouts, and the experimental model used. Here are some general timeframes for various functional assays:

1.  Cytokine measurement: A few hours to a few days Cytokines can be measured using techniques such as enzyme-linked immunosorbent assay (ELISA) or multiplex bead-based assays. The actual assay time is usually in the range of a few hours, but sample preparation and data analysis can take additional time.

2.  Cellular immune response assays: A few days to a few weeks Assays that evaluate the activation of immune cells, such as T cells or B cells, can take longer because they often involve cell culture and expansion. Examples include enzyme-linked immunospot (ELISPOT) assays, intracellular cytokine staining, and flow cytometry-based assays. Depending on the assay, the time for cell culture, assay execution, and data analysis can range from a few days to a few weeks.

3.  In vivo functional readouts: A few days to several months In vivo functional readouts, such as assessing the therapeutic efficacy of an mRNA vaccine or treatment, can take a variable amount of time depending on the specific endpoint being evaluated. For example, measuring tumor regression in a mouse model may take a few weeks to several months, while assessing protection from viral infection could take a few days to a few weeks.

4.  Functional genomics assays: A few days to a few weeks Assays that evaluate the downstream effects of the expressed protein on gene expression, such as RNA sequencing or microarray analysis, can take a few days to a few weeks, depending on sample preparation, sequencing or hybridization time, and data analysis.

While we don’t know exactly which drug that BioNTech and Matinas will be testing, we will have to assume it’s the Covid-19 vaccine. Which makes sense because they know it works already.

So how long will it take to get some meaningful results?

If researchers are testing an oral COVID-19 vaccine using lipid nano-crystals in a mouse model, the experimental timeline would involve several key steps. The duration of each step may vary depending on the specific experimental conditions, resources, and desired endpoints. Here's a general overview of the process and the time required:

1.  Vaccine preparation: A few days to a few weeks This step involves formulating the mRNA vaccine with lipid nano-crystals and optimizing the formulation for oral administration. This process may include quality control tests, such as assessing the encapsulation efficiency, size, and stability of the lipid nano-crystals.

2.  In vivo vaccination and sample collection: A few weeks after the vaccine formulation is prepared, the next step is to orally administer it to the mouse model. The mice would probably receive multiple doses, with a few weeks between doses to allow for an immune response to develop. Following the final dose, the researchers would wait for an appropriate duration (e.g., 1-2 weeks) to allow the immune system to respond to the vaccine before collecting samples, such as blood or tissue, for analysis.

3.  Immunogenicity assessment: A few days to a few weeks to determine the efficacy of the oral COVID-19 vaccine in the mouse model, researchers would evaluate the immune response generated by the vaccine. This may include measuring the levels of neutralizing antibodies using ELISA or other assays, which could take a few days to a week. Additionally, they might assess T-cell responses using methods such as ELISPOT or flow cytometry, which can take a few days to a few weeks.

4.  Protection from viral challenge: A few weeks to further confirm the vaccine's efficacy, researchers could perform a viral challenge experiment, in which the vaccinated mice are exposed to the SARS-CoV-2 virus (or a surrogate virus). Researchers would then monitor the mice for signs of infection, disease severity, and viral replication. This process can take a few weeks, depending on the duration of the disease course and the time required for sample collection and analysis.

In summary, the entire process of testing the oral COVID-19 vaccine in a mouse model, from vaccine preparation to evaluating its efficacy, could take anywhere from a month to several months. Although the timeline may vary based on the specific experimental design, the resources available, and the desired endpoints.

Why in-vivo data is more valuable than in-vitro data:

In-vivo data is considered more valuable than in-vitro data because it provides a more comprehensive understanding of the biological response in a living organism. While in-vitro studies offer valuable insights into specific cellular or molecular mechanisms, they may not always translate to the complex physiological environment of a living organism. Some key advantages of in-vivo data include:

1.  Complex physiological environment: In-vivo studies involve whole organisms, taking into account the complex interactions between cells, tissues, organs, and systems. This allows for a better understanding of how a treatment or intervention, such as an oral mRNA vaccine, performs in a biologically relevant context.

2.  Immune response: In-vivo studies provide a more accurate representation of the immune response to a vaccine or therapy, as the entire immune system is present and functional in the experimental model. This is particularly important when evaluating the efficacy of a vaccine, which relies on a coordinated immune response for protection.

3.  Pharmacokinetics and pharmacodynamics: In-vivo studies allow researchers to assess how the intervention is absorbed, distributed, metabolized, and excreted within the organism (pharmacokinetics), as well as its biological effects (pharmacodynamics). This information is critical for understanding the safety, efficacy, and optimal dosing of the intervention.

4.  Safety and toxicity: In-vivo studies provide a more accurate assessment of the safety and potential side effects of an intervention, as they can reveal adverse effects that may not be apparent in in-vitro settings.

5.  Translational potential: In-vivo data often has a higher translational potential, as it more closely mimics the conditions in humans. This can lead to a higher likelihood of success in clinical trials and eventual therapeutic applications.

In-vitro and in-vivo studies are complementary and should not be considered mutually exclusive. In-vitro studies are often more cost-effective, faster, and easier to perform, and they can provide crucial insights into the molecular and cellular mechanisms underlying an intervention. Ideally, researchers would use a combination of in-vitro and in-vivo studies to build a strong foundation of evidence for the safety and efficacy of an intervention, such as an oral mRNA vaccine.

How much more valuable would an oral mRNA covid-19 vaccine be?

An oral mRNA COVID-19 vaccine, assuming successful delivery using lipid nano-crystals, could offer several advantages over the current intramuscular mRNA vaccines. The value of such a vaccine would depend on various factors, including its safety, efficacy, and ease of administration. Some potential benefits include:

1.  Ease of administration: Oral vaccines are generally easier to administer than injections, as they do not require trained healthcare professionals for administration, nor do they generate biohazardous sharps waste. This could facilitate mass vaccination efforts, particularly in remote or resource-limited settings.

2.  Increased public acceptance: Some individuals are hesitant to receive injections due to needle phobia or other concerns. An oral vaccine could improve vaccine acceptance and coverage by providing a more appealing alternative.

3.  Improved immune response: Oral vaccines can stimulate both systemic and mucosal immunity, which is particularly relevant for respiratory viruses like SARS-CoV-2. Mucosal immunity at the site of viral entry (e.g., the respiratory tract) could potentially offer better protection against infection and transmission.

4.  Cost-effectiveness and scalability: If the production and storage requirements for the oral vaccine are more straightforward than those for the injected mRNA vaccines, it could lead to cost savings and improved scalability. For example, if the oral vaccine is more thermostable, it might not require the ultra-cold storage that some current mRNA vaccines do, simplifying the logistics of vaccine distribution.

5.  Potential for broader application: If the lipid nano-crystal technology proves successful for oral administration of mRNA vaccines, it could open the door for the development of oral vaccines for other diseases, expanding the potential applications of mRNA-based therapies.

Potential challenges to consider with an oral mRNA COVID-19 vaccine:

1.  Efficacy and safety: The oral vaccine would need to demonstrate safety and efficacy comparable to or better than the existing mRNA vaccines. Ensuring that the mRNA survives the harsh environment of the gastrointestinal tract and effectively enters target cells could be a significant challenge.

2.  Regulatory approval: The development of a new oral vaccine would require clinical trials and regulatory approval, which could take time and resources.

So, while it may take a few months to get a complete picture on the in-vivo work that Matinas and BioNTech are working on, it’s important to note that the exclusive research collaboration has expired. Matinas is free to talk to any other company about their oral mRNA work. Which means that BioNTech will probably be under pressure to cut a deal before the in-vivo work is completed. Especially if the oral vaccine proves more effective at preventing Covid-19 symptoms than the classic vaccine. Thanks for reading and don’t forget to follow us on Twitter.