Dr. Weeks’ Comment:  Difluoromethylornithine, or DFMO, is chemical which ODC (the supply chain for cancer) creating insufficient proteins (polyamides) for the racer to grow. It is being used to treat neuroblastoma brain cancer and more intriguingly, has been shown to stop cancer STEM cell production. Below is an introductory article but HERE is a recent paper published September 27 2018 in Scientific Reports shows the positive results of a phase II clinical trial using the oral medication DFMO to prevent relapse in children with High Risk Neuroblastoma (HRNB). According to Patrick Lacey of Beat NB Cancer Foundation, one of the childhood cancer parent-led foundations that funded this clinical trial. “Not only did this oral drug lead to a prolonged and stable remission for the children in this study, but the drug was extremely safe and well tolerated in this patient population.”  How does DFMO achieve long term remission? It shuts off the cancer STEM cells too.. “DFMO works by targeting specific cancer stem cell pathways and “turning off” the cells, thereby preventing the cancer from growing back.”

DFMO

March 31st, 2017   |   Posted in Relapse,Treatment   |   By: Antonia Palmer   | SOURCE

Introduction

High-risk neuroblastoma is an enigma,^[1] a puzzle of infuriating proportion that requires a myriad of different treatments.  For those children who are able to achieve remission, what can be done to try and keep the disease from returning?  It is not known if the extension of “maintenance” therapy could further improve the outcomes of children with neuroblastoma.  In Canada, there are currently no options available for enrolment on a clinical trial of extended maintenance that might destroy any lurking disease and prevent relapse.  However, US patients have options such as vaccines^[2] and DFMO (difluoromethylornithine).

DFMO is an interesting drug, and one that is currently being considered for a future clinical trial in Canada for children with neuroblastoma.  There is a great deal to understand about this drug – both in how it works and the process for opening up a clinical trial in Canada.

Before getting into the details of DFMO, there is some biology and science may help you understand how DFMO works.  If you are not the sciencey type, have no fear; just skim your way though this first bit to get the basic idea (there won’t be a test at the end).  So, here we go…

 

MYC and MYCN ^[3]

MYC is a family of proto-oncogenes – a normal gene that has the potential to become cancer because of mutations (errors in the genetic code) or some type of increased expression (expression is how information from a gene is utilized to create another genetic product such as a protein).

MYCN is a protein and member of the MYC family of proto-oncogenes.^[4]  In normal cellular function, MYCN plays a dual role in regulating cell growth by being able to both drive cellular proliferation and sensitizing cells to apoptosis (programmed cell death).^[5]  MYCN is able to simultaneously control how cells grow and how they die.^[6]

MYC proteins, including MYCN, play multiple roles in cancer “such as altering metabolic programs, supporting angiogenesis, promoting self-renewal and “stemness,” and driving proliferation while inhibiting differentiation.”^[7]  In normal cells, MYC activity is carefully regulated; however, in neuroblastoma cancer cells, MYC activity is not tightly regulated but is deregulated.  Even though this provides great evidence that MYCN plays a role in the initial formation of neuroblastoma tumours, it is thought that MYCN does not act alone, but that there are other events that also play a role in the tumourigenesis of neuroblastoma.

 

ODC and Polyamines

Cancer cells that are promoted by MYC, including tumours such as neuroblastoma, require large amounts of polyamines to function.  A polyamine is an organic compound that performs essential functions in all living cells.  Polyamines support the functions of MYC by promoting protein translation and ribogenesis.^[8]  Cells that are “non-proliferating” have reduced levels of polyamines.  Cells that are “proliferating”, both normal and cancerous, have higher levels of polyamines.

Ornithine decarboxylase (ODC) is a gene involved with the generation of polyamines.  ODC is regulated to ensure that cells have the appropriate amount of polyamines to survive and grow.

In neuroblastoma, polyamine metabolism is upregulated so that polyamines are in abundance.  And, it is MYCN that activates the ODC gene expression.^[9]   ODC is not only activated by MYCN; however, it is one way that ODC is overexpressed in most high-risk neuroblastoma tumours.  In almost all high-risk neuroblastoma tumours, ODC1, the product of the ODC gene, is significantly higher.  Laboratory research strongly supports that polyamines are necessary (polyamine sufficiency) in the later stages of the process (downstream) of cancer development when initiated by MYCN, such as that with neuroblastoma.^[10]  The ODC gene is also found in breast, colorectal and prostate cancers.

With two genes implicated in high-risk neuroblastoma tumours^[11], this is how many express one or the other or both:

• Co-amplified ODC1 and MYCN genes: ~8%-20%^[12]
• MYCN amplification alone: ~32%-40%
• No amplification of either gene: ~60%

ODC overexpression, regardless of if the cancer is MYCN amplified or not, has been shown to be an important risk factor on its own for neuroblastoma.

Other research has shown that inhibiting the production of polyamines directly impacts the cancer by making it more sensitive to chemotherapy and by inducing tumour regression.^[13]  It is possible that it could also work in cooperation with immunotherapy to further improve patient outcomes.  However, we need to have a better understanding of how inhibiting polyamines (or polyamine depletion) actually functions in the stopping and killing of neuroblastoma cancer cells.

As you would expect, there is quite a bit more to the science and biology.  If you are interested in diving in some more, an archived presentation from the 2015 Children’s Neuroblastoma Cancer Foundation in which Dr. Giselle Sholler describes the science and DFMO.  https://www.youtube.com/watch?v=x_VF4yqSnZM

 

About DFMO

Difluoromethylornithine, or DFMO, was developed in the 1970s^[14] as a drug that might have the potential to inhibit ODC activity.  However, initial research showed only minimal therapeutic impact and interest in the drug diminished.  In 2004^[15], DFMO was identified as a possible agent that could be used to inhibit ODC activity in neuroblastoma.  It was hypothesized that by inhibiting ODC activity, this would inhibit the production of polyamines, which would inhibit neuroblastoma from growing. ^[16]

 

DFMO in mice:

There have been a number of different research studies on the impact of DFMO on neuroblastoma to date:

1. Mice with neuroblastoma tumours were treated with DFMO and celecoxib (anti-inflammatory drug) and the experiment found that the two drugs worked synergistically (1+1 = more than 2) with one another on a number of different NB tumour types (e.g., MYCN amplified, ALK mutated, etc.).^[17] In mice treated with DFMO and celecoxib, a phenomenon called “pseudo-progression” has been observed.  Pseudo-progression occurs when a tumour is treated, there is some tumour growth, and then the tumour shrinks (regression).  It is a delayed reaction to the treatment and has often been seen in tumours when treated with immunotherapy.  There is still much to be understood about pseudo-progression; however, it is an important phenomenon which necessitates a better understanding of the tumour microenvironment (TME).

2. DFMO was examined using cisplatin and cyclophosphamide in mice with neuroblastoma tumours. Tumour-free survival was longer for mice treated with cyclophosphamide and DFMO versus cyclophosphamide alone.  In addition, mice had a longer tumor-free survival when treated with DFMO and cisplatin versus cisplatin alone.^[18],[19]

3. DFMO and etoposide have been shown to work together to have an effect on leukemia and myeloma cancer cells in mice. DFMO and etoposide work better together at killing cancer cells versus either etoposide or DFMO alone.^[20]

4. DFMO has been shown to target an important cancer stem cell pathway, decreasing the energy of the cells (ATP) and targeting the LIN28/Let 7 pathway. Treatment with DFMO inhibits LIN28 and tumor formation in mice as a model for preventing relapse.^[21]

5. DFMO prevents tumor formation in mice when neuroblastoma cells are injected into mice after pretreatment with DFMO.^[22] This research suggests that DFMO may target neuroblastoma cancer stem cells.

 

DFMO for patients with relapsed and refractory neuroblastoma:

A study published in 2015 details the results of a Phase 1 clinical trial using DFMO and etoposide to treat relapsed and refractory neuroblastoma patients.  In total, 21 patients participated in the study.  DFMO was given daily by mouth for three weeks (cycle 1), plus additional three week cycles of DFMO with daily low dose oral etoposide chemotherapy (cycles 2-5).  Within the study, the dosing of DFMO was increased for different patient groups (no dose increases for etoposide) to determine the maximum tolerated dose of the drug; however, there were no dose-limiting toxicities (DLTs) or serious adverse events (SAEs) experienced by the patients at any dose level.

Of the 18 evaluable patients on this study, 11 of them completed 5 cycles or less, 7 patients completed 7 cycles or more.  The longest treatment was 43 cycles (~130 weeks).

Side-effects consisted of anemia, decreased white blood cell count, decreased platelet count, ALT increase, AST increase, anorexia, constipation, diarrhea, infection, and others.  A maximum tolerated dose (MTD) was not reached in the study.  8/18 patients were unknown if they had MYCN amplification, all others did not have MYCN amplified neuroblastoma.  In this phase 1 study, the impact on the disease was not a primary endpoint; however, the following summarizes the effect on patient disease:

After Cycle 1: 12 had stable disease, 6 had progressive disease

After Cycle 2+: 1 had a partial response (PR), 12 had stable disease, 5 had progressive disease

It is important to highlight that disease response could have been because of the addition of etoposide during cycles 2-5.  The average amount of time patients remained free of progressive disease was 80.5 days, or about 2 ½ months.

Three of these patients have long term survival after treatment with DFMO for 2-2.5 years, with etoposide in cycles 2-5, and are now 4-6 years off therapy without any further anti-cancer therapy after DFMO.^[23]  Two of these patients where in the first cohort with the lowest dose of DFMO.

 

Current Use of DFMO

For neuroblastoma therapy, there are currently different ways that DFMO is being researched:

1. Combination Therapy: DFMO and Chemotherapy to treat Relapsed and Refractory Disease
   1. DFMO with Bortezomib (Phase 1/2)
   2. DFMO with Celecoxib, Cyclophosphamide and Topotecan (Phase 1)
2. Combination Therapy: DFMO and Chemotherapy for Newly Diagnosed Patients (Frontline Therapy). In this study genomic targeted therapy is added to chemotherapy cycles 3-6 and DFMO is given with Antibody and Accutane for 6 months, as well as for 2 years after completion of antibody.
3. Pediatric Precision Laboratory Advance Neuroblastoma Therapy (PEDS-PLAN) (Phase 1)
4. First Phase II study: DFMO to Prevent Neuroblastoma Relapse (Phase 2 Study completed Feb 2016)Dr. Giselle Sholler (Study Chair, Spectrum Health Hospitals) is the Principal Investigator of the “Neuroblastoma Maintenance Therapy Trial (NMTT)” that is being run by the Neuroblastoma and Medulloblastoma Translational Research Consortium.  Patients in the clinical trial receive only DFMO, no other drugs (called a single agent study).  DFMO is taken twice daily for up to 730 days (as long as there are no adverse events or disease recurrence).  There are two cohorts, or groups, of patients. The first group of patients have had immunotherapy and are without any disease, and the second group of patients have relapsed, but are back in remission.  Interim results presented in 2016^[24] showed that the patients who enrolled on this study after the completion of immunotherapy (no relapse) had a 2 year event-free survival (EFS) rate of 89% and an overall survival (OS) rate of 98%.  The most common side-effect was elevated liver enzymes (experienced by more than 10% of patients).

 

A Comment from Dr. Sholler:After a Phase I study, the following step is a Phase II study which is a single arm study (all children receive DFMO).  After completion of a Phase II study with encouraging results the next phase would normally be a randomized Phase III study.  This is one in which the one group of children would receive DFMO and one group would receive placebo. This is the most scientific way to show in a direct comparison of 2 groups the effect of a drug.   At the completion of the Phase II, the NMTRC physicians and parent advocates met with the FDA. Given the encouraging results based on historical experience with HRNB and published results by COG, MSKCC, SIOP, and there was enough evidence that the FDA recommended a confirmatory Phase II study due to ethical considerations/concerns in giving HRNB patients placebo with survival outcomes >98% with DFMO reaching out to 5 years.  The reasons a randomized study was not recommended at this point were ethical as well as the issue of feasibility: the fact that all children would be required to have exactly the same upfront therapy (which varies across the country/world in regards to number of cycles of chemo/number of transplants/MIBG and type of antibody) and then be randomized, from the statisticians review, the number of patients this would require to be enrolled, as well as follow up time to see effect, would be 10-15 years. This is unlikely to be possible to complete because of so many new advances happening in NB.  Since it is unclear that this study could be completed given the current varying state of upfront NB care and because half of the children would be receiving placebo it is not acceptable to do a randomized study in this group given the results to date.  A randomized study may be the best scientific choice, but not the best choice for the children.  A randomized study in a different group of NB children is under design.  Therefore a confirmatory study was started in Feb 2016 and the FDA will review after completion to evaluate the outcomes.

 

5. Second Phase II Study: Maintenance Therapy: DFMO to Prevent Neuroblastoma Relapse
   This study is a confirmatory Phase II study to replicate the results seen in the initial Phase II trial above, as well as to provide additional pharmacokinetic studies requested by the FDA and additional safety data (the safety package for drug approval requires 300-600 patients).

 

Liam’s Story:  by his Dad, AndrewWhen we completed up front therapy with NED (no evidence of disease) and were sent home with the odds of 50% relapse, we really didn’t know what to do. We had researched many things while we were sitting in the hospital during up front therapy and what potential options were out there after Immunotherapy. We found two options: one was the vaccine at Memorial Sloan Kettering Cancer Centre and the other was DFMO. After a lot of discussions with our Oncologist, and Liam’s success during upfront therapy, we really were unsure if we should pursue anything else or just get on with life. But that 50% chance of relapse kept us scared and driven. What we learned about DFMO was that it had been showing promise in the phase 1 & 2 trials (in Neuroblastoma patients) and that Oncologists felt it was safe based on previous use in the other trials (and for other diseases). Our Oncologist gave us all the information possible after speaking with the researcher leading the DFMO project, and supported our family decision to join the next phase trial that was opening in February 2016. This support from both medical teams gave us a great sense of hope that we were doing everything we could possibly do for Liam. The trial has been going very well and the follow-up is essentially the same as children receive after COG up front therapy (scans, labs etc).  Liam takes 3 tablets, twice per day with his meals.  There are some food restrictions that we follow but the reality is that we are living a very regular life.  While we were made aware of some possible side effects, Liam has not had any of them (13 months in).  Liam is indistinguishable from his peers and is active and happy.  The results for DFMO continue to show great promise, and we would never look back on our decision.  We have always held out great hope from the day this nightmare started (his diagnosis) and DFMO has really helped re-inforce that hope. Our goal is to make this trial available to all parents in Canada so they can make an informed decision.  The support of our medical teams in both Toronto and Michigan made this possible for us, but the current trial is not accessible for All Canadian families.  With cooperation of the home hospital in Canada, much of the scans and labs can be done at home alleviating tremendous costs.  The difficult part is that families need to travel to Michigan 7 times in 2 years, and depending on where they live, this can be financially difficult for them.  After 15-24 months of intense therapy to gain remission, many families are not only tapped emotionally, but financially as well.  We believe parents should have access, and the affordability to make the choice. When your child gets diagnosed the life of the parents completely changes.  You don’t have many choices, you have to trust your team (and we did).  My underlining message is for parents to have the choice, and not have to worry about losing their house over it.   An incredible parent story on the Lacey family was published online by the Boston Globe in 2016.  You can read about Will Lacey’s in this story titled “The Power of Will”

 

 

DFMO in Canada

DFMO is a drug that is not approved for use in Canada.^[25]  However, it is accessible within the context of a clinical trial for children with neuroblastoma.  Since this is the case, why can’t children with high-risk neuroblastoma just be given the drug?

First, it is important to understand a little about clinical trials.  Bear with us, it is hard to describe this complicated process in only a few words.

 

Clinical trials:

In its most basic form, a clinical trial is a research study that involves people with the purpose of testing a new drug, intervention (i.e., a new type of surgery), diagnostic test, medical device, or other health related tools to understand whether they provide benefit to a patient.  A clinical trial is one of the very last stages in a long journey of discovery that involves an idea, a hypothesis, a plan of action, the development of a team, sourcing funding, laboratory experimentation, running statistical analyses, writing reports, getting consortium approval, ethics clearance and many other steps.  The ultimate goal is to prove that a medical approach is more effective, less costly, less concerning, and/or less invasive for a patient than the current standard of care.  In the domain of medicine, clinical trials are an extremely important part of advancing science forward.  They are not only for patients with advanced disease; they are an integral part of all therapies, playing a role in helping develop better treatments for patients at all steps of their cancer treatment.  In the world of pediatric cancer, clinical trials have shaped treatment pathways and improved cure rates, over the last sixty years.

There are different phases and types of clinical trials.  We won’t get into that here but if you would like to know more, check out http://www.itstartswithme.ca.

 

Research oversight:

In Canada, clinical trials involving human subjects must be designed and executed according to Good Clinical Practices^[26] (GCP), regulations defined by Health Canada,^[27] possibly additional provincial guidelines, and stringent ethical standards.  GCPs are internationally agreed upon guidelines that work to ensure clinical trials are in patients’ best interests, that they are scientifically sound, conducted according to the protocol, are of high quality, approved by research ethics boards, have a qualified investigator and research staff, written informed consent is provided, and the therapy given is done in accordance with all required guidelines.^[28]  All clinical trials performed in Canada are reviewed by Health Canada when “testing new medications or testing known medications outside the approved indications – whether locally initiated, part of a clinical trial network or industry sponsored.”^[29]

In Canada, and most parts of the world, the design, associated documentation, and all details of a clinical trial must also be reviewed and approved by a Research Ethics Board (REB) before any patients are enrolled.  REBs, also called Institutional Review Boards (IRB) in the US, perform a number of different functions including monitoring clinical trials through their period of activity.  Canadian REBs must conform to the guidelines that are detailed in a document called the Tri-Council Policy Statement: Ethical Conduct for Research Involving Humans (TCPS), now in its second version called TCPS 2^[30].

The TCPS 2 contains the guiding principles that must be followed when conducting research with people in Canada, and outlines the necessities to have such research proposals reviewed by a Research Ethics Board (REB).^[31]  The REB is a committee that is composed of scientists, oncologists, bioethicists, lay representatives and other members with the purpose of ensuring participant safety through the identification of unnecessary risk while balancing the need for scientific advancement and therapeutic benefit.^[32],[33]  When followed, the guidelines^[34] are meant to help reduce unnecessary risk to the participant.  The principles and guidelines of the TCPS 2 are reflected in the tenets of research ethics that are established in the United States of America and other parts of the world.^[35]

 

The process to open a clinical trial:

To open up a DFMO clinical trial in Canada, there first needs to be someone (person or organization) to sponsor the study.  The sponsor of the study then works with C17, a non-profit organization that is responsible for the majority of academic clinical trials for childhood cancer in Canada.  C17 plays a critical role working to shepherd the necessary documentation through the process for the Principal Investigator (PI), or lead researcher, on the clinical trial.  Once the study is submitted to Health Canada, it has 30 days to review the study and make a decision. Once approved by Health Canada, the Investigator is given a “No Objection Letter” (NOL) and the trial is able to move on to the next stage of implementation.  If a clinical trial is to open at a number of different locations, each institution must go through the necessary REBs for approval of the clinical trial.

So your next question might be, just open the clinical trial already then.  Why is this taking so long to open a DFMO clinical trial in Canada?

Running clinical trials is an enormous undertaking requiring significant resources, time, immense responsibility, and flawless collaboration.  It requires a tenacity and determination of monolithic proportion.  It also requires financing, regulatory management, legal co-ordination, various layers of approvals, and many other facets that must be realized before the clinical trial is opened.  All of these decisions need to be made while asking if the research question of the clinical trial is the best question to ask for those children who are affected and for the intervention being proposed.  In Canada, discussions are currently underway to determine if and how a DFMO clinical trial could be opened.  The process has started.

 

Open Questions About DFMO

It is important to know that there are many questions that Oncologists and Parents have about DFMO.  These questions are all a part of the discussions that are currently happening as the possibility of bringing a DFMO clinical trial to Canada are being explored.  The following are just a few of the important ones:

1. Could it be possible that DFMO is being given to children who have already been cured by front-line treatment and immunotherapy?

2. If a randomized clinical trial was opened which had some patients take DFMO and some patients not take DFMO, would this be ethical? Would this approach be supported by Canadian patients and their families?  Would patients enroll in a randomized clinical trial when the clinical trial in the USA gives DFMO to all qualified patients without any randomization?

3. Over the many years that the maintenance DFMO trial has been open, the front-line treatment protocol for neuroblastoma has changed (e.g., tandem and single transplants). How is this accounted for in the different cohorts of the current trial in the US and how the data are analyzed?

4. What do the data look like for relapsed patients who are on DFMO with 2 years of follow-up?

5. The classification of NED (no evidence of disease) is an important achievement for neuroblastoma patients; however, there is still the potential of minimal residual disease (MRD) that could be lurking in the body. After immunotherapy, how can this be addressed and what if something like DFMO could deal with MRD effectively over time?

 

Conclusions

Discussions are already happening about bringing a DFMO trial to Canada.  There are still many hurdles to overcome; however, Oncologists and Clinicians understand that neuroblastoma patients and families are very interested in having a DFMO trial at Canadian hospitals.

We welcome your feedback, comments and interest in being a part of the discussion. Running clinical trials is not easy, it takes time, energy, funding and people to get the work done and there are always competing studies and protocols.  However, the goal is always to develop a treatment that leads to better outcomes for children with cancer – for both the children in cancer treatment today, and for the children who will be diagnosed with neuroblastoma in the future.  The ultimate goal is to have a better proven therapy as the standard of care for children with neuroblastoma.

Pediatric Oncologists are always trying to answer the next questions that science puts in front of them.  Their ultimate goal is to understand the disease and how to treat it with the least possible impact on the patient, but with the greatest effect on the cancer.  This is an immense challenge that is realized through the skilled machinery of a clinical trial.  There is still a great deal of work to be done for children with cancer and we must all work together to help realize the ultimate dream of cure.

 

 

Neuroblastoma Canada would like to thank Dr. Giselle Sholler, Dr. Doug Strother, Kathy Brodeur-Robb (C17), Patrick Lacey (BeatNB) and Kyle Matthews (BeatNB) for their valuable feedback and comments on this article. Thank you also to Andrew Cuttle for sharing Liam and his family’s story.

 

 

References:

^[1] Brodeur, G.M. (2003). Neuroblastoma: Biological Insights into a Clinical Enigma.  Nature Rev Cancer, Vol. 3, No. 3, pps. 203-216.

^[2] https://clinicaltrials.gov/ct2/show/NCT00911560

^[3] This section is taken from a former post written by Antonia Palmer on MYCN: http://inbraced.org/nbglobe/2013/04/11/mycn-amplification-in-neuroblastoma/

^[4] Slamon, D.J., Boone, T.C., Seeger, R.C., Keith, D.E., Chazin, V., Lee, H.C., Souza, L.M. (1986). Identification and Characterization of the Protein Encoded by the Human N-myc Oncogene. Science, 232, pps. 768-772. http://www.ncbi.nlm.nih.gov/pubmed/?term=3008339 [First characterization of the MYCN protein.]

^[5] Bell, E., Chen, L., Liu, T., Marshall, G.M., Lunec, J., and Tweddle, D.A. (2010). MYCN Oncoprotein Targets and Their Therapeutic Potential. Cancer Letters, 293, pps. 144-157. http://www.ncbi.nlm.nih.gov/pubmed/20153925

^[6] Westermark, U.K., Wilhelm, M., Frenzel, A., and Henriksson, M.A. (2011). The MYCN Oncogene and Differentiation in Neuroblastoma. Seminars in Cancer Biology; October, 21(4), pp. 257. http://www.sciencedirect.com/science/article/pii/S1044579X11000496

^[7] Hogarty, M.D. and Maris, J.M. (2012). PI3King on MYCN to Improve Neuroblastoma Therapeutics. Cancer Cell. 21, pp. 145.  http://ac.els-cdn.com/S1535610812000487/1-s2.0-S1535610812000487-main.pdf?_tid=f66a5158-8ab8-11e2-a512-00000aab0f6c&acdnat=1363053958_96be81f9c97ebadcaf21c52c63b6814f

^[8] Bassiri, H., Benavides, A., Haber, M.,Gilmour, S.K., Norris, M.D., and Hogarty, M.D. (2015). Translational Development of Difluoromethylornithine (DFMO) for the Treatment of Neuroblastoma.  Translational Pediatrics, vol. 4, no. 3, pps. 228.

^[9] Sholler, G.L.S., Gerner, E.G., Bergendahl, G., MacArthur, R.B., et al. (2015). A Phase 1 Trial of DFMO Targeting Polyamine Addiction in Patients with Relapsed/Refractory Neuroblastoma.  PLOS One, May 27, 2015, pp. 3.

^[10] Hogarty, M.D., Norris, M.D., Davis, K., et al. (2008), ibid.

^[11] Bassiri, H. et al. (2015). Pg. 232.

^[12] Evageliou, N.F., Haber, M., Vu, A., Laetsch, T.W., et al. (2016). Polyamine Antagonist Therapies Inhibit Neuroblastoma Initiation and Progression. Clinical Cancer Research, Vol. 22, No. 17, pps. 4391-4401.

^[13] Hogarty, M.D., Norris, M.D., Davis, K., et al. (2008). ODC1 is a Critical Determinant of MYCN Oncogenesis and a Therapeutic Target in Neuroblastoma.  Cancer Research, 68, pps, 9735-45.

^[14] Meyskens, F.L. Jr. And Gerner, E.W. (1999). Development of difluoromethylornithine (DFMO) as a Chemoprevention Agent.  Clinical Cancer Research, Vol. 5, No. 5, pps. 945-951.

^[15] Bachmann, A.S., (2004). The Role of Polyamines in Human Cancer: Prospects for Drug Combination Therapies.  Hawaii Medical Journal, Vol. 63, No. 12, pps. 371-374.

^[16] Sholler, G.L.S., Gerner, E.G., Bergendahl, G., MacArthur, R.B., et al. (2015). A Phase 1 Trial of DFMO Targeting Polyamine Addiction in Patients with Relapsed/Refractory Neuroblastoma.  PLOS One, May 27, 2015, pps. 1-20.

^[17] Evageliou, N.F., Haber, M., Vu, A., Laetsch, T.W., et al. (2016). Ibid.

^[18] Hogarty, M.D., Norris, M.D., Davis, K., et al. (2008), ibid.

^[19] Rounbehler, R.J., Li, W., Hall, M.A., Yang, C., Fallahi, M., Cleveland, J.L.  (2009). Targeting Ornithine Decarboxylase Impairs Development of MYCN-Amplified Neuroblastoma. Cancer Research, 69(2), pps. 547-53.

^[20] Saulnier Sholler, G., Currier, E., Bachmann, A.  (2010). Synergistic Inhibition of Neuroblastoma Tumor Development by Targeting Ornithine Decarboxylase and Topoisomerase II. June 2010 Advances in Neuroblastoma (ANR), Stockholm, Sweden.

^[21] Lozier, A.M., Rich, M.E., Grawe, A.P., Peck, A.S., Zhao, Pl, Chang, A.T.T., Bond, J.P., and Saulnier Sholler, G. (2014). Targeting Ornithine Decarboxylase Reverses the LIN28/Let-7 Axis and Inhibits Glycolytic Metabolism in Neuroblastoma.  Oncotarget, Vol., 6, No., 1, pps. 196-206.

^[22] Avequin, T., Zhao, P., Nagulapally, A.B., Bond, J., Azizi, E., Wicha, M., Saulnier Sholler, G. (2016). DFMO Targets Cancer Stem Cells in Neuroblastoma. American Association of Cancer Research (AACR) Annual Meeting, 2016.  New Orleans, LA

^[23] Communications with Dr. Sholler, March 27, 2017.

^[24] Saulnier-Sholler, G., Ferguson, W., Bergendahl, G., et al. (2016).  Maintenance DFMO Increases Survival in High-Risk Neuroblastoma.  ANR 2016.  Abstract PD-087, pg. 351.  http://www.siop2016.kenes.com/landing/Documents/PBC_Abstracts.pdf

^[25] A database that lists all drugs approved in Canada and their indications can be searched on the Health Canada website at http://www.hc-sc.gc.ca/dhp-mps/prodpharma/databasdon/index-eng.php

^[26] Devine, S., Dagher, R.N, Weiss, K.D. and Santana, V.M. (2008). Good Clinical Practice and the Conduct of Clinical Studies in Pediatric Oncology.  Pediatric Clinics of North America, Vol 55, pps. 187-209.

^[27] Food and Drug Regulations, Part C, C.05.010.  Accessed online on December 14, 2014 at http://laws-lois.justice.gc.ca/eng/regulations/C.R.C.%2C_c._870/page-285.html#docCont

^[28] Food and Drug Regulations, Part C, C.05.010.  Accessed online on December 14, 2014 at http://laws-lois.justice.gc.ca/eng/regulations/C.R.C.%2C_c._870/page-285.html#docCont

^[29] Bond, M.C. and Pritchard, S. (2006). Understanding Clinical Trials in Childhood Cancer.  Paediatric Child Health, Vol. 11, No. 3, pps. 148-150. Pp. 150.

^[30] Canadian Institutes of Health Research, Natural Sciences and Engineering Research Council of Canada, and Social Sciences and Humanities Research Council of Canada.  (2010). Tri-Council Policy Statement: Ethical Conduct for Research Involving Humans.  Accessed online at: http://www.pre.ethics.gc.ca/eng/policy-politique/initiatives/tcps2-eptc2/Default/

^[31] Fernandez, C. (2008). Ethical Issues in Health Research in Children.  Paediatric Child Health, Vol. 13, No. 8, pps. 707-712. Pp. 708.

^[32] Fernandez, C. (2008). Ethical Issues in Health Research in Children.  Paediatric Child Health, Vol. 13, No. 8, pps. 707-712. Pp. 708.

^[33] Fernandez, C. (2008). Ethical Issues in Health Research in Children.  Paediatric Child Health, Vol. 13, No. 8, pps. 707-712. Pp. 709.

^[34] Within Canada, health care is under the jurisdiction of the provinces and territories which means that additional guidelines may also need to be followed that are either provincial or territorial based (for example, when conducting research in Quebec, researchers must also take into account the provisions of article 21 of the Quebec Civil Code).

^[35] Fernandez, C. (2008). Ethical Issues in Health Research in Children.  Paediatric Child Health, Vol. 13, No. 8, pps. 707-712. Pp. 708.