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Safety

Overview

  • Safety as a Top Priority: The Reneurish team has made safety a top priority in their work with induced pluripotent stem cells (iPSCs) for stroke treatment. Every step of the process has been carefully thought out to meet the highest safety standards, ensuring that both patients and the broader research environment are protected.
  • Data Protection: Patient privacy has been a key focus from the start. The team follows strict European and Spanish laws, such as the GDPR and LOPDGDD, to ensure all data is handled responsibly. They’ve taken extra care to pseudonymize patient information, so personal details are never exposed unnecessarily. The principle of data minimization has been applied, ensuring no more than the essential information is collected. Lastly, patients have been fully informed about how their data will be used and can withdraw their consent at any time.
  • Clinical Safety:
    • Avoiding teratoma formation: By designing a precise cell-sorting system as a second safety layer added to the current successful methods used in our lab, we ensure to safely tackle one of the main concerns of stem-cell therapies.
    • Avoiding Prolonged BDNF exposure: Our design aims for a transient modification and a patented-pending time-limited expression regulation to avoid the possible long-term negative effects of overexpression.
    • GMO concerns: We have selected AAV vectors to avoid the integration of our construct in the human genome, thus ensuring that all genetic alterations are temporary and well-controlled. Moreover, the transplanted cells are designed to be clean transgenes.
  • Laboratory Safety: In the lab, safety has been central to every decision. We have mapped out potential hazards ahead of time, such as the use of human neural progenitor cells and lentiviral vectors, for which we had the corresponding Check-In forms approved. We have always followed best practices to minimize risks related to hazardous chemicals, iPSCs, lentiviral vectors, and genetically modified organisms (GMOs).

Overall, we have taken every precaution, both in protecting patient privacy and ensuring clinical and laboratory safety.

Data Protection

Introduction

At our stroke project, we take data protection and privacy with the utmost seriousness. We are deeply committed to ensuring that all personal information is handled with the highest level of care, adhering to both ethical standards and legal requirements. Understanding the sensitive nature of the data we collect, particularly related to patients’ medical histories and experiences, we have implemented robust measures to guarantee confidentiality and protect their rights. By following strict protocols and ensuring full compliance with all relevant laws and regulations, we aim not only to safeguard personal data but also to build trust with all the individuals involved. This commitment is at the core of our approach to responsible and ethical research. The relevant legal frameworks include:


  • The Charter of Fundamental Rights of the European Union, which establishes that all Union citizens have the right to have their personal data protected.
  • Regulation (EU) 2016/679 of the European Parliament and of the Council of April 27, 2016 (General Data Protection Regulation, GDPR). [1]
  • Organic Law 3/2018, of December 5, on the Protection of Personal Data and Guarantee of Digital Rights (LOPDGDD) in Spain. [2]

In compliance with the University of Barcelona’s guidelines for handling personal data in research projects, we have ensured that all procedures are in line with the General Data Protection Regulation (GDPR) and the LOPDGDD. Specifically, the University holds responsibility for safeguarding the personal data used in scientific research and mandates that all research groups and students involved respect the fundamental rights and freedoms of individuals. [3]

As we are students processing the personal data for research purposes, we have been explicitly authorised by the principal investigator, and we have been thoroughly informed of their obligations under the data protection regulations. The data we process are pseudonymized whenever necessary to ensure additional privacy protection, following the University’s prescribed standards.

How did we anonymize the data?

As part of our dedication to protecting the privacy of all individuals involved, we have taken comprehensive steps to ensure that the personal information collected is anonymized. Given the sensitive nature of the medical data, particularly from patients who have experienced a stroke, it was essential for us to implement strict privacy measures to prevent any unintended identification. We approached this process carefully: only the interviewers have knowledge of the patients' real identities. Throughout all documentation and meeting minutes, we have used pseudonyms or coded names, which are also reflected in our project’s internal records and the wiki. According to the relevant data protection regulations, this process can be classified as either anonymization or pseudonymization, depending on the reversibility of the identification.

The key difference between these two concepts lies in whether or not the data can be linked back to the individuals:

In anonymization, it is not possible to identify the patients, and the process is irreversible. In pseudonymization, the data is anonymized, but there is a separate record that can identify the individuals, making it reversible.

In our case, while we do not keep a formal registry of data that could directly identify the patients, the interviewers still have the ability to recognize them. As a result, this could be classified as a form of anonymization, but it is not entirely complete. To ensure compliance with best practices and regulatory standards, we have opted to follow the procedures for pseudonymization. Below, we outline the key steps taken in this process.

Data Minimization:


We have applied the principle of data minimization, which states that personal data should be adequate, relevant, and limited to what is necessary in relation to the purposes for which they are processed (Article 5(1c) GDPR).

Data Collected and Why:


Age: Stroke is strongly correlated with age, and we aim to illustrate this while also highlighting exceptions. Experience during stroke and treatment: As we are developing new treatments, we need to understand where previous treatments may have failed to improve them, as well as identify successful aspects to preserve them. After-effects: This helps us understand the social stigma surrounding stroke and how it impacts patients' lives.

Informing Patients about Data Processing:


When patients signed the consent form for data processing, they were fully informed about how their data would be handled and assured that they could withdraw their consent at any time. However, they were also made aware that once the project’s wiki is finalised and frozen, no further modifications will be possible. This clause was clearly communicated to ensure transparency and understanding of the limitations once the data becomes part of the finalised documentation.

Measures to Safeguard Rights and Freedoms:


We ensured that only selected team members have access to information that can reidentify individuals. This way, it is impossible for other members of the team to identify them, maintaining a strict separation between personal identifiers and research data. These measures are designed to enhance privacy and minimise any potential risk of reidentification, aligning with the highest standards of data protection and confidentiality. These precautions guarantee that personal identifiers are kept entirely separate from research data, further reinforcing the privacy and security of the participants, and ensuring that no external parties can reidentify them under any circumstances.

For more details on the consent process and the rights granted to participants, you can access the Consent Form Document here . This document outlines the scope of data usage, withdrawal procedures, and the guarantees for safeguarding personal information.

Clinical safety

Risk 1: Teratomas

A main issue encountered with iPSCs-based cell-therapies is the teratome and teratocarcinoma formation. Teratomas are tumours containing tissues from all three different germ layers (ectoderm, mesoderm and endoderm) as they are formed from embryonic stem cells or, in this case, induced Pluripotent Stem Cells. Despite inducing differentiation of cells before injection, a residual cell population can remain undifferentiated which, despite being very few cells, can form teratomas. Teratoma formation implies some risks for the patient apart from limiting the cell therapy, such as the interference with the normal function of the target tissues and the malignant potential of developing into more aggressive teratocarcinomas [4].

Our PI, Daniel Tornero, has been working with Neural Progenitor Cells for over 20 years and has developed some techniques to minimise the teratoma formation risk. The NPC line used (derived from UBI001-A hiPSCs cell line) was approved by the Health Department of Generalitat de Catalunya in Tornero’s project to be injected in mice, thus, assuring its safety features at least for its use in animals. After submitting the cell cultures to the priming conditions used by Dr. Tornero, the NPCs so efficiently into neural cells that in over 200 mice injected, none has presented teratomes. In his words, “2 months after the transplant, only 5% of cells were dividing, and after 6 months, none” [5].

In order to reduce even more the teratoma formation risk, we designed an individual-cell-sorting system to select only the differentiated cells for the transplant (see: engineering branch 3.3). The transfected cells will have a selection cassette (which will be removed by Cre-LoxP systems before transplant) encoding for a membrane protein. This protein will only be expressed in transfected cells and under the regulation of a specific neural progenitor promotor. Cells will be marked with fluorescent antibodies against the expressed protein and then selected via Fluorescence-Activated Cell Sorting. This cell-sorting system performs at a 95-100% purifying rate, and submitting the cell populations several times through it, we will ensure that only neural progenitor cells will be selected for the transplant.

By using this two-layer safety system, we are dramatically reducing the teratoma risk which is one of the major set-backs for hiPSC-based therapies.

Risk 2: Prolonged exposure to BDNF

During the first stage after stroke, BDNF overexpression can be a game-changing factor of the recovery prognosis. Thanks to its neuroprotective role, high levels of BDNF favour the rescue of different types of neurons; it also enhances neuroplasticity, supports the survival and differentiation of neural progenitors. However, those features are only wanted during the first critical stage after stroke; in the mid and long term, BDNF overexpression should be gradually suppressed. Prolonged exposure to BDNF has several downsides such as a possible homeostatic shift that pushes the neural network into an attenuated state which is related to reduced neuronal excitability and disruptions of neural circuit functions. On the other hand, BDNF binds to tyrosine kinase receptors TrkB which activates several signalling pathways that have proven to be upregulated in various cancers including neuroblastoma. [6] [7] [8]

We developed a two-layer safety approach to avoid prolonged exposure to BDNF while assuring the first stage overexpression.

In the first place, the construct is transfected episomically so that it ensures a transient overexpression. On one hand, the cell proliferation will continue for a short period of time after the transplant; on the other hand, nor the construct or the AAV that contains it are capable of replication. Therefore, the constructs will dilute amongst the cells in each division and the resulting BDNF will be diluted too in the growing population of neural cells. Moreover, the episomal constructs are degraded in the cells in the mid and long-term.

In the second place, the BDNF expression is controlled by a patent-dependent regulatory system that ensures the expression only during the first period of time after transplant. The regulatory system is being tested to optimise both overexpression and self downregulation in the wanted periods of time.

Risk 3: Patient’s genome modification

Gene-modified cell therapy is a new revolutionary approach in medicine based on altering the genetic material of cells ex vivo to treat inherited and acquired diseases. It has proven to be a success in areas like blood disorders or cancer (with CAR-T cell therapy) and has opened the doors for the research of many others. Nevertheless, it has raised a brand new bioethical concern: genetically engineering human cells. At Reneurish, we have approached our gene-modified cell therapy to achieve top level biosafety standards and gain the trust of the most reticent patients.

Firstly, starting from the point that we are modifying somatic human cells, we avoid the greater bioethical concerns. The modifications in our transplanted neural progenitors are not capable of transferring to the germ line so the key international agreements such as the Oviedo Convention (European treaty) are guaranteed.

Secondly, a key point of our therapy design that has been discussed with patients and experts is to perform an episomal modification rather than a genomic one in the transplanted cells.

As some experts exposed (see: integrated human practices, loop 2) it would be more economically viable to integrate our construct to the genome of the neural progenitor cells, we would have a stable transgenic cell line ready to thaw and be transplanted into the patient's brain. However, patients and their families have repeatedly preferred not to be treated with permanently modified cells, even if the medical safety being guaranteed (see: integrated human practices, loop 3). Thirdly, we will use AAV as they are amongst the safest vectors to genetically modify human cells. One of the key points on using AAVs is that they are inherently replication-deficient, so we make sure the construct is not reencapsidised and transfected to other unwanted host cells after the transplant.

Fourthly, the transgent load is reduced to the bare minimum necessary. We are eliminating the cell-sorting cassette containing H2B-mCherry using the Cre-LoxP system, resulting in a clean transgene that we are looking for.

Safe lab practices

Biosafety and biosecurity are two key elements when performing an experiment. There are lots of things to consider and keep in mind when working in the lab. Consequently, it is essential to learn and know about risks, measures of prevention and permissions needed in order to assure a safe project not only for team members but for the whole world, including people, animals, plants, environment…

Reneurish team is very aware that small actions in the laboratory can have big consequences for the world, whether for better or worse. That is why we have carefully studied and followed an adequate process to guarantee the safety and security of our project.

Establish the lab work

First of all, it is very important to plan the experiments that will be done in the lab and identify the elements or components that may be dangerous, complicated to manipulate or require permits. This includes organisms, chemicals, engineering parts…

Organisms and parts

It is very important to mention all organisms that the experiment will use or require, and also a brief explanation of the process or the modification you are planning to do to these organisms.

lt-NES C4 (human neural progenitors), derived from UBI001-A hiPSCs: it is the organism that we are creating: genetically engineered human neural progenitor cells. Our goal is to express a growth factor in transplanting human induced Pluripotent Stem Cells (hiPSC) in order to enhance cell integration in a stroke-damaged brain area. Shortly, we will transfect It-NES C4 cells with our lentivirus containing the plasmid EF1a_BDNF_mCh so they overexpress our desired growth factor. The growth factor's expression will have safety regulators to minimise its adverse effects. Then, we will test neuron integration by using the modified neurons by using a culture chip from XonaMicrofluidics. On the other hand, we will test 3 different constructs to find the best promoter for expressing the growth factor in our neural progenitor cells (It-NES C4).

EF1a_BDNF_mCh: lentivirus used to transfect It-NES C4 (only for the growth factor).

AAV2-7m8: we will insert the best-performing construct of point 3 via AAV to It-NES C4 cells and conduct again the testing of neuron integration.

E. coli VB UltraStable™ from Vector Builder: cloning host strain.

E. coli XL-1 Blue Competent Cells: transformed cells for plasmid clonning.

HEK293T cells: transfected cell culture to observe plasmid expression. We will design a cotransfection reporter system to target fluorescent proteins to specific cellular compartments and test it on HEK293T cells. The transfection is by lipofectamine and assessment of fluorescence through microscopy and flow cytometry.

Chemicals

Every chemical in the lab has its own properties. Some of these can be dangerous if the chemical is not manipulated correctly, so it is very important to know the possible risks and how to manage them. Dangerous chemicals identified:

Paraformaldehyde: it can produce formaldehyde fumes, which are irritant, toxic and carcinogenic.

Chelating agents such as azide: the contact with acid can produce hydrazoic acid, which is toxic and volatile.

Concentrated NaOH: extremely corrosive. If it is mixed with other substances, it can liberate fumes.

Concentrated HCl: it liberates fumes that are very corrosive and irritating.

In order to safely manipulate these chemicals, it is useful to count with a chemical fume hood. We also identified SYBR Safe as a mutagenic, but it does not require to work in the hood.

Identify possible risks in the lab


Once the idea of the project is developed, the possible risks and how they may happen must be identified. This is useful for us but also if we have to ask for any permission.

  • Human iPSC cells: they may carry human pathogens during our experiments, which pose a risk of infection to lab personnel. When working with human cells, if they get accidentally contaminated with a human pathogen and they are not properly handled in the class II safety cabinet, there is a risk of exposure to a team member or other laboratory workers. This contamination may occur if a team member has not properly followed the safety and PPE measures accordingly, specially when preparing culture media.
  • Lentiviral vectors: designed to integrate into human cell genome, to transfect the cells we will be working with. This integration can lead to unintended mutagenesis or oncogenesis. If lentiviral vectors are not properly handled and protocols are not stringently followed, they may contact directly with the skin of a team member or laboratory worker and there is a potential risk of causing somatic mutations.
  • GMOs: we will genetically modify the aforementioned organisms, such as lentiviral vectors, neural progenitor cells and E. coli. There may be a biological hazard for the environment if any of them get out of the laboratory. There is a risk of accidental release of any of the GMOs that we are using in the laboratory, specially for lentiviral vectors and Escherichia coli due to their higher robustness to environmental conditions. These organisms could potentially disrupt local ecosystems, transfer genetic material to other organisms, or proliferate uncontrollably outside the lab environment. This accidental release may occur when a team member has used a glycerol stock of a genetically-modified organism but hasn't followed the waste-treatment protocols established by the laboratory to dispose of the aliquot (for instance, they might not have used bleach to neutralise any remaining organism in the container, but rather regular hand soap).
  • Transportation: we shall transport some material from one laboratory to the other, including GMOs. This increases the risk of accidental release. For instance, the transportation box might not be properly sealed and cause a spillage. Additionally, some inactivated organisms might remain after DNA purification if the protocol is not followed meticulously, which might reach the environment if the sample is sent to sequencing.
  • Hazardous chemicals: some experiments may require the use of hazardous chemicals, like paraformaldehyde for sample preservation or chelating agents when running electrophoresis gels, as well as concentrated NaOH or HCl for pH calibration. Exposure to volatile hazardous chemicals might result from not using the chemical fume hood properly, specially when dealing with paraformaldehyde. Regarding chelating agents and NaOH or HCl, direct skin exposure might result from not using gloves or handling the material carelessly.


In summary, we believe that our project may lead to:

  • Harm to human health and safety
  • Harm to the environment, including wild plants and animals Social inequities


However, we do not believe that it can lead to:

  • Breaking norms about engineering biology
  • Reducing global, national or health security
  • Harm to materials, equipment, and infrastructure
  • Harm to agricultural animals, crops, or domesticated animals


Moreover, if our project was fully developed, any of our engineered organisms or parts could spread autonomously in the environment:

  • AAV and Lentiviral vectors that we are using are incompetent so they cannot replicate by themselves.
  • Any hiPSC or derived cells are not able to survive out of ideal conditions in culture plates.
  • The E. coli strains that will be used present an auxotrophy for leucine. Additionally, stringent containment measures will be maintained throughout the experimental part of our project.

Find a proper lab work space

When all the experiment is planned it is important to find or adapt an adequate space for lab work that can provide the safety measures that must be followed to assure a safe project. When looking for the lab where the experiment will be performed, it is not only significant to look for a lab related to the topic of the project, but also for a lab that is equipped with the necessary material and instruments in order to do the experiment safely.

We are conducting our experiments in two separate laboratories:

  • Laboratory 1: level 2 lab, focused solely on cell culture work, equipped for handling moderate-risk biological agents.
  • Laboratory 2: this lab has designated areas for both Level 1 microbiological work (with clean benches) and Level 2 cell cultures (with Class II, type B2 biosafety cabinets), as well as a designated GMO room.

These laboratories can afford our experiment in terms of resources and measures of prevention. Also, our host laboratory adheres to institutional biosafety protocols, ensuring compliance with both national and international standards. The host laboratory is equipped with biosafety level 2 (BSL-2) facilities, including Class II biological safety cabinets, autoclaves for sterilisation and personal protective equipment (PPE) such as lab coats, gloves, and safety goggles.

Permissions for lab work, legal basis and supporting organisations

The law of each country covers the work done at the lab, along with several organisations that ensure the safe and moral use of the scientific and technological advances available nowadays. The law varies between countries, so it is very important to consult the law of the country where the experiment will be performed. In order to guarantee biosecurity in the laboratory, iGEM UB 2024 team will work under the risk assessment and guidance provided in the Appendix 2 of the (Spanish) Royal Decree 664/1997, which is necessary when working with human cells culture. According to national legislation, «biosafety level 2 for the manipulation of genetically manipulated organisms (GMOs)» is required for the experimental part of our project. Thus, biosafety guidelines established by National Law 9/2003 together with Royal Decree 178/2004 will be followed.

Moreover, it is really useful to have the support of some organisations for the managing of risks. Also because although applying preventive measures, the risk does not never disappear and accidents can occur, and it is important to know who must be informed about it. Reneurish team has the support of The Bioethics Committee and the Office of Health, Safety and Environment of the University of Barcelona, as well as several professors and technicians from the Faculties of Biology and Medicine.We would go to all of them in the event of an accident.

Finally, we submitted the corresponding Check-In forms for both lentiviral vectors and neural progenitor cells that we will use. Both of them were approved.

Team training for security and safety

Any member of the team who gets inside the lab should have access without keeping in mind the basic safety measures that must be followed. Moreover, Wet Lab members must know the safety and prevention measures needed to perform the experiments and they must follow them. That is why it is very important to do previous training on safety and security before the lab work starts.

The Office of Health, Safety and Environment of the University of Barcelona (OSSMA) has given us training on how to conduct experiments in the university's laboratories. The aspects studied are the following:

  • Lab access and rules
  • Responsible individuals
  • Differences between biosafety levels
  • Biosafety equipment
  • Good microbial technique
  • Disinfection and sterilization
  • Emergency procedures
  • Rules for transporting samples between labs or shipping between institutions
  • Physical biosecurity
  • Personnel biosecurity
  • Chemical, fire and electrical safety

Performance of safety measures in the lab

Once in the lab, safety measures must be performed. To manage the risks associated with our gene-modified cell therapy research project, we have taken comprehensive actions and leveraged expert support, adhering to strict rules and training protocols, and implementing rigorous procedures and practices:

  • Accident reporting: The accident reporting system at the UB is managed by the OSSMA. The system consists of filling out an online notification form, which collects essential details about the incident, such as the nature of the accident, location, and any injuries sustained. The form can be found here. Once submitted, the report is reviewed by OSSMA, which then initiates an investigation to determine the cause of the incident and assess any potential risks.
  • Accident performing: In case of an accident, nearby the GMO room are eyewash units, sinks, and safety showers that are functional and properly maintained. The door contains a sheet with a list of the infectious agent(s) in use, entry requirements, and emergency and medical contact information. In case of spills, the GMO room contains a spill kit with absorbent material, plastic bags, and a protocol on how to proceed. Spills are addressed according to the procedures outlined by OSSMA.
  • Personal Protective Equipment / PPE: All team members operating in the GMO room wore individual dedicated lab coats and single-use disposable gloves. Lab coats were kept in the room at personal hooks and regularly washed. Splash goggles were worn when handling liquid culture. No open toe shoes were allowed. Hand washing occurred after removing gloves and prior to exiting the laboratory.
  • Inventory, data and physical access controls: Access to the lab was controlled with a key, including access to inventory and databases.
  • Lone Worker or Out of Hours policy
  • Medical surveillance
  • Waste management system: Used material is washed with bleach and autoclaved. Liquid waste is collected in containers with bleach, and solid waste containing GMOs is disposed of in designated containers according to the waste management protocol. The waste management company contracted by the University of Barcelona is PreZero.
  • Additional containment
  • Project-specific safety or security training: The OSSMA has provided training on laboratory experiments. All team members received training on Good Microbiological Techniques and handling of GMOs, maintaining continuous contact with experts and local safety committees.
  • Other consulting with iGEM about managing risks: Due Check-In forms for the organisms in use have been submitted.
  • Consulting with other experts and stakeholders about managing risks: Consultations have occurred with professors and technicians from the University of Barcelona and other institutions.
  • Crafting a responsible communication plan
  • Modifying the experimental design or methodology: Experiments are designed to minimize cellular use by using specialized culture chips.
  • Deciding not to do an activity: Specialized culture chips also avoid the need to work with animals.
  • Performing an adequate transportation: Biological materials are transported either frozen or securely contained, with no risk of loss. Materials are transferred directly from lab to lab and will remain within university premises.
  • Proper use of the safety facilities: Storage units and biohazardous materials are securely stored in clearly labelled containers. The protocol for using Biosafety Cabinets includes specific exposure and handling procedures.

By integrating these comprehensive measures, we are confident in our ability to manage the risks associated with our gene-modified cell therapy project. Our commitment to strict adherence to rules, thorough training, well-equipped facilities, and meticulous waste management practices ensures a safe and controlled research environment.

References


[1] La protección de datos en la UE. (s. f.). Comisión Europea. https://commission.europa.eu/law/law-topic/data-protection/data-protection-eu_es
[2] Normativa sobre datos personales: Ministerio de Hacienda. (s. f.). https://www.hacienda.gob.es/es-ES/El%20Ministerio/Paginas/DPD/Normativa_PD.aspx
[3] Recerca - Protecció de dades - UB. (s. f.). Protecció de Dades. https://web.ub.edu/ca/web/proteccio-dades/recerca
[4] Martin, R. M., Fowler, J. L., Cromer, M. K., Lesch, B. J., Ponce, E., Uchida, N., Nishimura, T., Porteus, M. H., & Loh, K. M. (2020). Improving the safety of human pluripotent stem cell therapies using genome-edited orthogonal safeguards. Nature Communications, 11(1). https://doi.org/10.1038/s41467-020-16455-7
[5] Palma-Tortosa, S., Tornero, D., Hansen, M. G., Monni, E., Hajy, M., Kartsivadze, S., Aktay, S., Tsupykov, O., Parmar, M., Deisseroth, K., Skibo, G., Lindvall, O., & Kokaia, Z. (2020). Activity in grafted human iPS cell–derived cortical neurons integrated in stroke-injured rat brain regulates motor behavior. Proceedings Of The National Academy Of Sciences, 117(16), 9094-9100. https://doi.org/10.1073/pnas.2000690117
[6] Liu, W., Wang, X., O’Connor, M., Wang, G., & Han, F. (2020). Brain-Derived Neurotrophic Factor and Its Potential Therapeutic Role in Stroke Comorbidities. Neural Plasticity, 2020, 1-13. https://doi.org/10.1155/2020/1969482
[7] O’Neill, K. M., Anderson, E. D., Mukherjee, S., Gandu, S., McEwan, S. A., Omelchenko, A., Rodriguez, A. R., Losert, W., Meaney, D. F., Babadi, B., & Firestein, B. L. (2023). Time-dependent homeostatic mechanisms underlie brain-derived neurotrophic factor action on neural circuitry. Communications Biology, 6(1). https://doi.org/10.1038/s42003-023-05638-9
[8] Radin, D. P., & Patel, P. (2017b). BDNF: An Oncogene or Tumor Suppressor? Anticancer Research, 37(8). https://doi.org/10.21873/anticanres.11783