Pre-Treatment of Highly Suspicious Pigmented Skin Lesions With Interleukin-2
Phase: PHASE3
Conditions: Melanoma (Skin), Melanoma in Situ
Sponsor: Carman Giacomantonio

 
 
 
STUDY PROTOCOL 
Pre-Treatment of Highly Suspicious Pigmented Skin Lesions with 
Interleukin-2 
 
QEII Hospital 
Nova Scotia Health Authority 
Halifax 
Canada 
 
 
Date: October 2018 
NCT Number: NCT03233828 
 
 
Dr. Carman Giacomantonio 
Rm. 8-826, VG Site, 
1276 South Park St.,  
Halifax, NS, B3H 2Y9 
Ph: 902-473-6177 
Fax: 902-473-6178 
 
 
 

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   1 
 
 
Pre-Treatment of Highly Suspicious Pigmented Skin Lesions with Interleukin-2 
 
Lay Summary 
Skin cancer is the world’s most common form of cancer – with more diagnoses annually 
than breast, prostate, lung, and colon cancers combined. Melanoma, the most aggressive form of 
skin cancer, has killed an estimated 1250 Canadians in 2017 alone. When discovered early, 
melanoma can usually be cured with surgery alone, but once it metastasizes therapeutic options 
are limited. Up until 2011, only two therapies were approved by the US food and drug 
administration (FDA) for the treatment of advanced metastatic melanoma (MM) – Dacarbazine 
(DTIC) and systemic Interleukin-2 (IL-2). In the past 10 years of variety of immunological 
agents have been approved for the treatment of MM including direct injection (intralesional) of 
IL-2 into tumors. 
IL-2 naturally occurs in the immune system and has been used as a cancer 
immunotherapy for almost 40 years. Systemically administered melanoma immunotherapies, 
such as IL-2, show promise but are at high doses have morbid toxicity profiles and are costly. By 
administering IL-2 intralesionally via injection, local efficacy and toxicity profiles are improved, 
compared to systemic administration, as lower doses of IL-2 are needed and are administered 
directly into the lesion. IL-2 treats MM by engaging the immune system.  One of the potential 
benefits theoretically from this therapy is the creation of lasting tumor immunity that could 
protect the patient from further development of metastatic disease. The hallmark of an effective 
immune response is the presence of a robust population of immune cells, so called tumor 
infiltration lymphocytes (TILs), identified histologically in surgically removed tumors.  
When a patient is identified having a highly suspicious pigmented lesion, the diagnosis of 
melanoma is confirmed with an excisional biopsy (a biopsy that grossly includes a narrow rim of 
uninvolved tissue), part of the normal standard of care.  When present, demonstration of a robust 
population of TILs in the biopsied specimen is a strong indicator of a natural immune response. 
We aim to assess the utility of treating highly suspicious pre-metastatic lesions in a randomized, 
placebo-controlled, trial of intralesional IL-2. We hypothesize that a “proactive” pre-treatment 
strategy with IL-2 will boost local immune response to suspicious lesions by stimulating a robust 
TIL response that could ultimately prevent distant metastasis. We believe this to be the future of 
melanoma therapy. 
We aim to include 20 local participants over the next 24 months in a randomized, 
placebo-controlled, trial of intralesional IL-2 to assess the utility of treating highly suspicious 
pre-metastatic lesions. Patient consent will be obtained using an NS Health Authority, REB-
approved SOP. All patients will have lesions biopsied following standard surgical practice 
techniques, and will provide blood and urine for analysis. Tissue samples will be assessed for 
immune system activity, and blood and urine will undergo proteomic and metabolomic analysis.  
Background Information: 
Skin cancer is by far the world’s most common form of cancer – with more diagnoses annually 
than breast, prostate, lung, and colon cancers combined.1 Every year in Canada over 80,000 
cases of skin cancer are diagnosed, over 7,200 of which are Melanoma, the most deadly form of 
skin cancer.2 It is estimated that in 2017, over 1250 Canadians have died of Melanoma.2  
The incidence of melanoma in Canada is on the rise in both men and women. In women 
the incidence of melanoma has increased by 2.8% between 2001 and 2010, an annual percentage 

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   2 
 
 
change surpassed only by thyroid cancer.2 As mortality rates for almost all other forms of cancer 
decrease – the age-standardized mortality rate of melanoma is on the rise. Melanoma is the 
number one cancer killer of women aged 25 to 30.1 
As a whole, 5-year age-standardized survival rates for melanoma have been reported as 
high as 88%.2 When discovered early, melanoma can usually be cured with surgery alone, but 
once it metastasizes therapeutic options are limited. 5-15% of patients with melanoma will 
progress to stage IV MM.3,4 MM is one of the most aggressive malignancies, with 5-year 
survival rates of advanced disease between 5-23%, depending on the location of tumor 
dissemination.5,6 Although several new therapeutic interventions for metastatic melanoma are 
undergoing clinical trials, for the foreseeable future MM is a disease characterized by a worrying 
rate of mortality, and treatments of questionable efficacy.5,7  
Given the paucity of effective therapeutic interventions for MM, successful strategies in 
treating melanoma will include therapies or therapeutic adjuvants which limit, or altogether 
prevent metastases.   
 
Current Treatments: 
Limited therapeutic Options for Metastatic Melanoma 
Cure rates for melanoma are high when the disease is discovered before it has spread 
from its primary location; however, metastasis frequently occurs, presenting a clinical challenge 
that has frustrated physicians and researchers alike for decades. Up until 2011, only two 
therapies were approved by the US food and drug administration (FDA) for the treatment of 
advanced (stage III or stage IV) MM. DTIC, the only chemotherapeutic licensed to treat MM 
was approved in 1975, has limited therapeutic utility. Patients on DTIC can expect a one-in-eight 
chance of having tumors shrink.8 Systemic therapy utilizing high-dose IL-2 was approved by the 
FDA in 1998. IL-2 is a naturally occurring glycoprotein secreted by T cells to mediate cellular 
immune response, and has been used as a cancer immunotherapy for almost 40 years.9 IL-2 
mediates bystander activation and proliferation of CD4+ T-cell, CD8+ T-cell lymphocytes and to 
a lesser extent, NK cells.9 Although 4% of patients were cured using this immunological therapy, 
the side effects of systemic administration are significant and are fatal in 2% of patients.10 Even 
with these treatments, the outcome for patients with distant metastases is bleak, with median 
survival of 6 to 10 months and less than 5% of patients surviving for more than 5 years.6 
 
Melanoma Immunotherapies 
 
Indeed, immune control of melanoma is achievable in specific circumstances –  
elucidation of the molecular identity of several antigens that are recognized by the immune 
system of melanoma patients has led to the discovery of pathways affecting tumor immunity at a 
cellular and molecular level.11–13 Still MM has proven to be excessively difficult to treat, and the 
development of novel immunologically-mediated therapeutics has been slow. Many adjuvant 
vaccine trials in melanoma have been conducted including GMK vaccine and granulocyte-
monocyte colony-stimulating factor (GM-CSF), but these have been largely unsuccesful.14,15  
Similarly, trials assessing the treatment of MM with immune stimulants such as Bacillus 
Calmette–Guerin (BCG), Corynebacterium parvum and levamisole have yielded mixed 
inconsistent responses or been ineffective and even harmful to patients.16 

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   3 
 
 
 
Significant inroads have been made in recent years with the development of novel 
immunotherapies for the treatment of MM.  In 2011 the FDA approved the use of monoclonal 
antibody ipilimumab as a checkpoint immunotherapy for the treatment of advanced MM.17 
Ongoing studies are assessing whether adjuvant therapies such as chemotherapy or various 
immunologic therapies may improve on the antitumor effects achieved with ipilimumab.7 
Multiple systemic antibody-mediated immunotherapies for advanced MM – including nivolumab 
and pembrolizumab – are current under development, and have shown significant advances;18 
However response to these novel therapeutics have been tempered as these therapies are 
associated with serious (sometimes fatal) immune-mediated side effects and prohibitive 
pricing.19  
 
Intralesional Therapies  
 
The serious toxicity profile associated with systemic immunotherapies can be altogether 
avoided by treating instead with intralesional injections. Studies have shown lowers rates of 
toxicity are associated with intralesional injections when compared with systemic administration; 
furthermore, by delivering an increased concentration of the drug at the site of action, increased 
rates of efficacy are observed.20 Intralesional IL-2 is associated with modest flulike symptoms 
alone – a vast improvement on the toxicity profile associated with systemic administration of IL-
2.10,21 Intralesional injections of IL-2 have an added bonus of causing a so-called “bystander 
effect”, whereby cancerous cells that are not immediately adjacent to the local injection site, also 
are effectively treated through development of an adaptive regional immune response.20 
 
“Proactive” vs “Reactive” Treatment: The Future of Melanoma Therapy 
IL-2 has been effectively used to treat MM when administered intralesionally.21 This 
mode of treatment using IL-2 is being effectively delivered by Dr. Carman Giacomantonio to 
treat patients with advanced cutaneous MM at the QEII HSC, NSHA. IL-2 used in this capacity 
is a “reactive” treatment to MM. Herein we propose the use of IL-2 as a “proactive” treatment to 
pre-metastatic melanoma. 
 
We plan to treat patients with highly suspicious lesions – as diagnosed by qualified 
dermatologists – with intralesional IL-2 in an effort to generate an adaptive immune response 
with activation and proliferation of CD8+ T-cell effector lymphocytes and immune sensitized 
CD8+ T memory cells to address the potential risk of subsequent melanoma metastasis.9 
Moreover, after treatment with IL-2, the proliferation of CD8+ T-memory lymphocytes may 
allow cells of immune surveillance to mount an immune response to new, de novo pre-cancerous 
and/or cancerous melanoma cells. 
 
With 1250 estimated deaths in Canada in 2017 due to metastatic melanoma, finding 
preventative treatments is crucial. We believe that a “Proactive” pre-treatment with IL-2 is the 
future of melanoma therapy.  
 
Hypothesis: 

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   4 
 
 
Hypothesis: Pre-treatment with IL-2 in patients with highly suspicious pigmented lesions will 
reduce the incidence of metastatic melanoma compared to the PBS placebo group by initiating an 
adaptive immune response to native melanoma protein and peptide. 
Major Aims: 
1) To conduct a pilot study to assess the number of patients needed to analyze in order to 
achieve a statistically significant differentiation between treatment and control outcomes 
in study measures including tumor infiltrating lymphocytes (TILs) and circulating 
immunomodulators. 
2) To assess the efficacy of intralesional pre-treatment of IL-2 on highly suspicious lesions 
in generating an adaptive immune response and preventing melanoma metastasis.  
3) Sub-study – To assess if there a difference in specific immune modulators in patients 
receiving IL-2 pre-treatment compared to patients receiving a placebo. 
 
 
Subject Selection:  
 
Qualified dermatologists will identify and approach potential participants to recruit them 
to become part of the study. The participant population will include patients characterized by: 
nodular/polypoid features, bleeding/ulcerated lesions, excluding face and vulvo-genital lesions; 
and who are not: currently immunocompromized, on immuno-therapy for other diagnosis, have 
known inflammatory or autoimmune diseases or are otherwise incapacitated, between the ages of 
18 and 80 years of age. We aim to include a minimum of 20 (up to 60) local participants over the 
next 24 months. Patients will be randomized and will receive a subcutaneous injection of either 
IL-2 treatment (treatment group) or the placebo PBS treatment (placebo group).  
 
 
Participant consent: 
 
Consent will be obtained using an approved NS Health Authority REB protocol. It will be 
clear to the patient that it is important that they tell their surgeon about any drugs or medicines 
they are taking or wish to take and that they must also tell their surgeon if they are having any 
adverse effects. Patients will be allowed to withdraw from the study protocol at any time. It will 
also be clear that if they withdraw consent, the information about them and their donated tumor 
tissue that was collected before they left the study will still be used. No new information about 
them will be collected (and no further testing of donated tumor tissue, blood or urine will be done) 
without permission. 
 
 
Research Plan: 
Clinical Research Plan: Is intralesional pre-treatment of IL-2 in highly suspicious pigmented 
lesions effective in preventing future melanoma lesion spread and metastasis?  
This study is primarily designed to determine if tumor specific immunity can be generated 
in patients with highly suspicious pigmented lesions in response to intralesional IL-2, and whether 

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   5 
 
 
that immunity can confer resistance to melanoma metastasis. Patients will be identified by 
qualified dermatologists and interviews will be held at the surgery clinic (4th floor Dickson Center) 
QEII HSC, NSHA.  
The standard wait time from consultation to surgical biopsy is up to 4 weeks. We will 
ensure that patients are seen immediately upon notification from participating dermatologist and 
all research components of the study are completed within the normally anticipated wait time.  
Through utilization of this standard wait time, we can potentially affect an immune response using 
intralesional IL-2, that otherwise could not be achieved after the biopsy is completed. Given that 
the study is conducted within the normal wait time, it doesn't deviate from the normal standard of 
care. Following our protocol, patients will receive intralesional injections on Day 1(Week 1 Visit) 
and Day 8 (Week 2 Visit), and excisional biopsies will be performed on Day 15 (Week 3 Visit), 
well within the accepted wait time from consultation to biopsy. The intralesional injections and 
collection of biospecimens beyond the biopsy deviate from but does not delay the normal standard 
of care. Two additional visits are required in addition to the initial consultation. 
This study is a randomized, controlled, double-blind study. Patients with highly suspicious 
pigmented lesions will be randomly assigned by an algorithm to one of two groups: 1) treatment 
group patients will be treated intralesionally with IL-2 (Proleukin (Aldesleukin), Novartis 
Pharmaceuticals Canada Inc.) at a dose of 500,000 International Units (IU) (0.1ml) for 2 treatment 
cycles 1 week apart (Day 1 and Day 8) Figure 1; 2) control group patients will be treated 
intralesionally with sterilized sodium chloride (0.9%, m/v) (0.1ml) Figure 1. Randomization will 
be done at the pharmacy for each patient (generated by a Random Block algorithm). IL-2 
(Proleukin) will be obtained from the Provincial Drug Distribution Program by the pharmacy, paid 
for by Dr. Carman Giacomantonio's research services account (please see budget attached). The 
pharmacy will prepare either IL-2 treatment (supplied as 1 x 0.8ml (4 million units) or injectable 
placebo (0.9% sodium chloride) supplied as 0.8ml). The syringes will be prepared by following 
aseptic technique in a biohazard hood. A 22-million unit vial of IL-2 will be reconstituted with 
1.2ml of sterile water for injection and mixed gently. After drawing up 3.2ml of 5% dextrose in 
water and transferring to the IL-2 vial, the resulting concentration will be 5 million units/ml. 0.8ml 
of the IL-2 solution will then be drawn into a 3ml syringe. Expiry time of the reconstituted 
solutions is 48 hours at 4C. The pharmacy will assign a codified ID which will be provided along 
with the syringe with the patient name labeled as "Interleukin-2/Placebo" (thus blinding study to 
clinician and patient). As a pilot study, the principle of the design is to test the feasibility of 
proceeding to a larger and more expensive trial following the methodology and protocol proposed. 
As such there will be a minimum of 20 (10 treatment, 10 placebo) participants (up to a maximum 
of 60). It is estimated that statistical significance will be reached with 20 patients, however it is 
not reached after 20 patients, an additional 10 (5 treatment, 5 control) patients will be enrolled. If 
significance is still not reached after 30 patients, an additional 10 patients will be enrolled and so 
on up to a maximum of 60 patients. If after 60 patients, significance is not achieved then further 
recruitment under the current protocol would not be logical and therefore the methodology would 
need to be revised or study discontinued. 
On Day 1 (first treatment) and 15 (excisional biopsy), all patients will have blood (4 vials) 
and urine (25-50 ml) samples taken.  Local reactions to injections will be monitored for non- 
specific signs such as bleeding, arythema, infection, or irritation. We don't anticipate specific 
changes in the pigmented lesion related to the injection in this short time period, however all 
changes will be noted.     

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   6 
 
 
On Day 15, following the second intralesional injection an excisional biopsy will be 
performed following standard surgical techniques, as follows: ellipse of skin encompassing the 
pigmented lesion extending into the subcutaneous fat will be performed to achieve a grossly clear 
but narrow margin of excision. This small defect will be closed primarily with interrupted sutures. 
The biopsied specimen will be subsequently evaluated using standard histological techniques to 
confirm diagnoses, assess margin status (clear or involved) and depth of lesion invasion. The depth 
of lesion invasion will dictate the extent of subsequent surgical excisions and margin selection 
according to standard National Comprehensive Cancer Network (NCCN) guidelines. Pre-biopsy 
intralesional injections of 0.1ml into the lesion will not alter dimensions or architecture of the 
lesion or impact the extent the subsequent biopsy required. Lesions that are felt to be too large for 
closure without tissue manipulation or creation of flaps, will be biopsied using a punch biopsy 
sampling technique whereby 4mm diameter discs of tissue from representative areas of the lesion 
will be taken to confirm the diagnosis and the depth of invasion. The depth of invasion will dictate 
the extent and complexity of subsequent surgery required to remove the lesion and achieve clear 
margins according to NCCN guidelines. The biopsied specimen will be processed as follows: using 
a 22G needle a fine needle aspiration biopsy will be performed (the needle will be passed through 
the center of the lesion two times) for RNA analysis. The remainder of the tissue will be sent to 
pathology for standard histological assessment. The pathologist will also report the extent of 
observed tumor infiltrating lymphocytes (TILs) associated with treatment compared to control 
injections. Blood and urine will be assessed using metabalomic and proteomic methods to assess 
systemic immune response to treatment.  
All blood, urine, and lesion biopsy samples will be labeled with a codified number (will 
not contain any patient identifying information) and will be immediately transported to Dr. Carman 
Giacomantonio laboratory (Sir Charles Tupper Medical Building, 11F11) at Dalhousie University 
for storage. Fine needle aspiration of excised samples will be assessed for tumor genetic and 
epigeneitic profile (Figure 2). Blood and urine will be assessed for proteomic and metabolomic 
profiles, respectively (Figure 2).  
A database containing patient IDs and codified numbers will be restricted to the office of 
Dr. Giancomantonio (11th floor, VG). This is a locked office, and the data will only be accessible 
to the investigators named on this application. This database for will contain patient IDs, and info 
on disease status, treatment/control, follow up visits, and sample analysis – only info pertinent to 
the outcome measures. All sample data will be assessed using codified descriptors and will contain 
no patient ID info. Once the RNA, proteomic, metabolomic and pathology data has been analyzed, 
unmasking treatment and control groups will be conducted at the VG office of Dr. Giacomantonio, 
by the named investigators. If the data reaches statistically significance, the study will not accept 
any more patients, if not, a further 10 patients will be incorporated – to a maximum of 60 patients. 
Any publications of study results will be completed devoid of any information that could be used 
to identify patients included in the study. 
All study participants will receive biannual assessments for 5 years after the initial 
intervention to assess disease progression, or the development of new melanoma, to compare 
between both treatment and control groups. There is no standardize test or measurable biomarker 
to assess established or lasting immunity. This aspect of the study is identical to the patient 
assessment conducted as per standard of care for melanoma patients. Again, any publications of 
study results will be completed devoid of any information that could be used to identify patients 
included in the study. 
 

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   7 
 
 
 
       Figure 1 – Clinical procedure  
 
 
 
      Figure 2 Clinical and Sub-study Procedures 
 
Analysis of Data 
Outcome measure/study objectives: 

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   8 
 
 
Assessment of Number of Patients Needed to Obtain Significance [Time Frame: 1 year] – Data 
will be analyzed in order to achieve a statistically significant differentiation between treatment 
and control outcomes in study measures including tumor infiltrating lymphocytes (TILs) and 
circulating immunomodulators. 
   
Assessment of Metastasis [Time Frame: 5 years] - All patients will receive assessments every 4 
months for 2 years and then biannual assessments for years 3-5 after the initial intervention to 
assess disease metastasis in treatment and control groups. Both number of new metastases (integer 
value) and thickness (mm) will be measured as a part of this assessment. 
 
Sub-Study Outcome Measure: 
 
Assessment of RNA genetic profile [Time Frame: 5 years] - RNA analysis of excised tissue will 
be compared to unaffected patient tissue obtained from the clear margins of the excisional biopsy 
to assess genetic changes resulting from the melanoma and treatment/placebo injections.  RNA 
analysis will occur at Dr. Giacomantonio's laboratory (Sir Charles Tupper Medical Building, 
11F11) at Dalhousie University.  
 
Assessment of Systemic Immune Response: Proteomic Analysis [Time Frame: 5 years] - 
Proteomic analysis will be conducted on blood samples to assess systemic immune response to 
both treatment and control groups. Serum collected from patient blood samples will be used for 
proteomic analysis to assess protein expression, including circulating immunomodulators 
(cytokines and chemokines) before, and after, treatment. Proteomic analysis will occur at 
Dalhousie’s Proteomics and Mass Spectrometric Core Facility located in the Life Sciences 
Research Institute. This study may serve to help develop diagnostic protocols and methods of 
assessing response to treatments.  
 
Assessment of Systemic Immune Response: Metabolomic Analysis [Time Frame: 5 years] - 
Metabolomic analysis will be conducted on urine samples to assess systemic immune response to 
both treatment and control groups. All tumor and tissue produce by products in waste that are 
excreted by the kidneys. Urine samples can be evaluated using techniques, such as Mass 
Spectrometric, to determine if biological compounds can be identified in association with the 
presence of a malignant process that would not be produced by normal tissue. Metabolome analysis 
will occur at Dalhousie’s Proteomics and Mass Spectrometric Core Facility located in the Life 
Sciences Research Institute. This study may serve to help develop diagnostic protocols and 
methods of assessing response to treatments. 
 
How will the data be analyzed: 
All study practices and statistical methods are based on the International Conference on 
Harmonization (IHC) document “Statistical Principles for Clinical Trials.”  Data will be 
summarized by treatment group. Baseline characteristic, safety outputs and a total overall 
column will be included to summarize all subjects.  For all baseline, demographic, safety and 
efficacy outputs data will be summarized by treatment group.  

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   9 
 
 
 
In summary tables of continuous variables, the minimum and maximum statistics, the 
arithmetic mean and median, the 95% confidence interval, standard deviation, and standard error 
will be presented will to the same number of decimal places as the original data.  In summary 
tables of categorical variables, counts and percentages will be used. The denominator for each 
percentage will be the number of subjects within the population treatment group unless otherwise 
specified.  All hypothesis testing will be carried out at the 5% (2-sided) significance level unless 
otherwise specified.  P-values will be rounded to three decimal places. P-values less than 0.001 
will be reported as <0.001 in tables. 
 
The treatment label for all tables, listings and figures will be: 
 
Treatment 
Label  
2 treatment cycles of 
500,000 IU of IL-2 in 0.1 
mL 
IL-2 
Treatment 
0.1 mL of sterile saline 
(0.9% m/v)  
Placebo 
All Treatments 
Total 
 
Where that any of the statistical methods described herein prove unsuitable during 
analysis, more appropriate methods will be used. All changes in methodology will be 
documented in the clinical study report.  Additional ad-hoc analyses may be conducted as 
deemed suitable.  
 
Subject inclusion/exclusion criteria will be determined at baseline visit, and subjects who 
do not meet all criteria will not be entered into the study. Those subjects deemed eligible to 
participate will be allocated a 3-digit number at randomization prior to the initial treatment.  If a 
subject is discontinued at any time after entering the study, the Investigator will ensure this does 
not affect the patient’s standard of care. At the patients request all unused biological samples 
(blood, urine, and core biopsies) will be immediately destroyed. The reasons for withdrawal will 
be recorded on the CRF and will be included in the final report. Failure to complete both (2) 
treatment cycles will result in patient removal from the trial.  
Data resulting from our RNA, metabolomic and proteomic studies will be analyzed using 
specific statistical software available to us at Dalhousie University, including Bio-Rad qPCR 
machine and CFX manager statistical software for RNA analysis, and liquid chromatography-
tandem mass spectrometry (LC-MS/MS) systems (QTRAP5500 by ABSciex) and bioinformatic 
analysis for metabolomic and proteomic studies. 
What are the proposed benefits and potential harms of this research and how does the 
benefits outweigh the harms?: 
Potential benefit: Generate an effective immune response to the patient’s primary melanoma. 

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   10 
 
 
Risk-Breach of confidentiality; As with all research and sub-studies, there is a chance that 
confidentiality could be compromised; however, we are taking precautions to minimize this risk. 
All visits will be conducted in a private room to keep participant privacy. 
Risk-Drug injections: Patients will be receiving an intralesional injection which may cause 
irritation and swelling at the site of injection. These discomforts are minimal and brief 
Risk-Side effect of IL-2: Patients may feel mild flu like symptoms after receiving treatment, 
including fever, fatigue, headaches or muscle cramps. These symptoms are brief and general 
resolve within 48 hours 
Risk-Drawing Blood; There is a possibility of pain, bruising, swelling or infection related to giving 
blood.  These discomforts are minimal and brief. 
Pregnancy/Fertility: FDA Pregnancy Category C. The effects of IL-2 on fertility and pregnancy 
is unknown and therefore pregnant women will be excluded from the trial and men and woman 
will be asked to use contraceptive protection during the two-week injection period. 
Breastfeeding: it is not known whether IL-2 is excreted in human milk, therefore breastfeeding is 
contraindicated. 
 
Confidentiality: 
 
Non-identifying codes will be assigned to patient biological samples and personal health 
information (PHI) by the research offices at the QEII. The codes will not contain any identifiable 
information that can be traced back to the patient. The only file that links identifying codes with 
the patient will be kept on a password protected computer. A hard copy file will be stored 
separately in a secured location. Personnel who have access to identifiable patient information 
will be kept limited.  Patient records and research results will be kept indefinitely. 
 
Compensation: 
 
Participants will be compensated $25 for each additional visit (2 visits) that is not part of their 
normal standard of care. 
 
References 
 
1. 
Canadian Skin Cancer Foundation. About Skin Cancer. 2017 Available at: 
http://www.canadianskincancerfoundation.com/about-skin-cancer.html.  
2. 
Statistics Canada. Canadian Cancer Statistics 2017.  
3. 
Ranieri, J. M., Wagner, J. D., Wenck, S., Johnson, C. S. & Coleman, J. J. The prognostic 
importance of sentinel lymph node biopsy in thin melanoma. Ann. Surg. Oncol. 13, 927–
32 (2006). 
4. 
Kalady, M. F., White, R. R., Johnson, J. L., Tyler, D. S. & Seigler, H. F. Thin Melanomas. 
Trans. ... Meet. Am. Surg. Assoc. 121, 221–230 (2003). 

Study Protocol  
Version 3 
October 2018 
 
1022767 
 
 
 
 
   11 
 
 
5. 
Sandru, A., Voinea, S., Panaitescu, E. & Blidaru, A. Survival rates of patients with 
malignant melanoma. J. Med. Life 7, 31–35 (2014). 
6. 
Tsao, H., Atkins, M. B. & Sober, A. J. Management of Cutaneous Melanoma. N. Engl. J. 
Med. 351, 998–1012 (2004). 
7. 
Smyth, E. C. & Carvajal, R. D. Treatment of Metastatic Melanoma: A New World Opens. 
(2016). Available at: http://www.skincancer.org/skin-cancer-
information/melanoma/melanoma-treatments/treatment-of-metastatic-melanoma.  
8. 
Chapman, B. P. B. et al. Phase III Multicenter Randomized Trial of the Dartmouth 
Regime Versus Dacarbazne in Patients With Metastatic Melanoma. J. Clin. Oncol. 17, 
2745–2751 (1975). 
9. 
Eklund, J. W. & Kuzel, T. M. A review of recent findings involving interleukin-2-based 
cancer therapy. Curr. Opin. Oncol. 16, 542–546 (2004). 
10. 
Atkins, B. M. B. et al. High-Dose Recombinant Interleukin 2 Therapy for Patients With 
Metastatic Melanoma: Analysis of 270 Patients Treated Between 1985 and 1993. J. Clin. 
Oncol. 17, 2105–2016 (1999). 
11. 
Ramirez-Montagut, T., Turk, M. J., Wolchok, J. D., Guevara-Patino, J. a & Houghton, A. 
N. Immunity to melanoma: unraveling the relation of tumor immunity and autoimmunity. 
Oncogene 22, 3180–3187 (2003). 
12. 
Tsai, K. K. & Daud, A. I. Nivolumab plus ipilimumab in the treatment of advanced 
melanoma. J. Hematol. Oncol. 8, 123 (2015). 
13. 
Tsai, K. K., Zarzoso, I. & Daud, A. I. PD-1 and PD-L1 antibodies for melanoma. Hum. 
Vaccin. Immunother. 10, 3111–3116 (2014). 
14. 
Kirkwood, J. M. et al. High-dose interferon alfa-2b significantly prolongs relapse-free and 
overall survival compared with the GM2-KLH/QS-21 vaccine in patients with resected 
stage IIB-III melanoma: Results of intergroup trial E1694/S9512/C509801. J. Clin. Oncol. 
19, 2370–2380 (2001). 
15. 
Lawson, D. H. et al. Randomized, Placebo-Controlled, Phase III Trial of Yeast-Derived 
Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) Versus Peptide 
Vaccination Versus GM-CSF Plus Peptide Vaccination Versus Placebo in Patients With 
No Evidence of Disease After Compl. J. Clin. Oncol. 33, JCO.2015.62.0500- (2015). 
16. 
Eggermont, A. M. M. & Gore, M. Randomized Adjuvant Therapy Trials in Melanoma: 
Surgical and Systemic. Semin. Oncol. 34, 509–515 (2007). 
17. 
Eggermont, A. M. M. et al. Prolonged Survival in Stage III Melanoma with Ipilimumab 
Adjuvant Therapy. N. Engl. J. Med. 375, 1845–1855 (2016). 
18. 
Hersey, P. & Gallagher, S. Intralesional immunotherapy for melanoma. J. Surg. Oncol. 
109, 320–326 (2014). 
19. 
Fellner, C. Ipilimumab ( Yervoy ) Prolongs Survival In Advanced Melanoma Serious Side 
Eff ects and a Hefty Price Tag May Limit Its Use. P&T 37, 503–512 (2012). 
20. 
Sloot, S., Rashid, O. M., Sarnaik, A. A., Zager, J. S. & Moffitt, L. Developments in 
Intralesional Therapy for Metastatic Melanoma. Cancer Control 23, 12–20 (2016). 
21. 
Boyd, K. U., Wehrli, B. M. & Temple, C. L. F. Intra-lesional interleukin-2 for the 
treatment of in-transit melanoma. J. Surg. Oncol. 104, 711–717 (2011).