Clinical trial NCT01585428
Conditions: Cervical Cancer Oropharyngeal Cancer Vaginal Cancer Anal Cancer Penile Cancer
Keywords: Immunotherapy HPV-Associated Cancers Cervical Cancer Oropharyngeal Cancer HPV Infection
Sponsor: National Cancer Institute (NCI)
Start date: 2012-04-13; Completion date: 2016-08-01
Enrollment: 29 participants; Allocation: NON_RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Background:

The human papillomavirus (HPV) can cause a number of cancers, including cervical and throat cancers. The National Cancer Institute (NCI) Surgery Branch has developed an experimental therapy that involves taking white blood cells from patients' tumors, growing them in the laboratory in large numbers, and then giving the cells back to the patient. These cells are called Tumor Infiltrating Lymphocytes, or TIL and we have given this type of treatment to over 200 patients with melanoma. Researchers want to know if TIL shrink s tumors in people with human papilloma virus (HPV)-related cancer. In this study, we are selecting a specific subset of white blood cells from the tumor that we think are the most effective in fighting tumors and will use only these cells in making the tumor fighting cells.

Objective:

The purpose of this study is to see if these specifically selected tumor fighting cells can cause HPV-related cancers to shrink and to see if this treatment is safe.

Eligibility:

\- Adults age 18-66 with HPV-related cancer who have a tumor that can be safely removed.

Design:

Work up stage: Patients will be seen as an outpatient at the National Institutes of Health (NIH) clinical Center and undergo a history and physical examination, scans, x-rays, lab tests, and other tests as needed.

Surgery: If the patients meet all of the requirements for the study they will undergo surgery to remove a tumor that can be used to grow the TIL product.

Leukapheresis: Patients may undergo leukapheresis to obtain additional white blood cells. {Leukapheresis is a common procedure, which removes only the white blood cells from the patient.}

Treatment: Once their cells have grown, the patients will be admitted to the hospital for the conditioning chemotherapy, the TIL cells and aldesleukin. They will stay in the hospital for about 4 weeks for the treatment.

Follow up: Patients will return to the clinic for a physical exam, review of side effects, lab tests, and scans about every 1-3 months for the first year, and then every 6 months to 1 year as long as their tumors are shrinking. Follow up visits will take up to 2 days.

Detailed description:
Background:

* Metastatic or locally advanced refractory/recurrent human papillomavirus (HPV)-associated malignancies (cervical, vulvar, vaginal, penile, anal, and oropharyngeal) are incurable and poorly palliated by standard therapies.
* Administration of autologous tumor infiltrating lymphocytes (TIL) generated from resected metastatic melanoma can induce objective long-term tumor responses.
* Young TIL can be generated from HPV-associated tumors.

Objectives:

* To determine if autologous Young TIL infused in conjunction with high dose aldesleukin following a non-myeloablative lymphodepleting preparative regimen can mediate tumor regression in patients with metastatic or locally advanced refractory/recurrent HPV-associated cancer.
* To study immunologic correlates associated with Young TIL therapy for HPV-associated cancers.
* To determine the toxicity of this treatment regimen.

Eligibility:

\- Patients greater than or equal to 18 years old with a pathologically confirmed diagnosis of metastatic or locally advanced refractory/recurrent HIPV-16+ or HPV-18+ human papillomavirus-associated cancer.

Design:

* Patients will undergo biopsy or resection to obtain tumor for generation of autologous TIL cultures and autologous cancer cell lines.
* All patients will receive a non-myeloablative lymphocyte depleting preparative regimen of cyclophosphamide (60 mg/kg/day intravenous (IV)) on days -7 and -6 and fludarabine (25 mg/m(2)/day IV) on days -5 through -1.
* On day 0 patients will receive between 1 times 10 (9) to 2 times 10(11) young TIL and then begin high dose aldesleukin (720,000 IU/kg IV every 8 hours for up to 15 doses).
* Clinical and immunologic response will be evaluated about 4-6 weeks after TIL infusion.
* Initially, 18 evaluable patients will be enrolled in two cohorts; patients with cervical cancer and those with non- cervical cancer. For each cohort, if 0 to 2 of the 18 patients experience a clinical response, then no further patients will be enrolled. If 3 or more of the first 18 evaluable patients enrolled have a clinical response, then accrual will continue until a total of 35 evaluable patients have been enrolled in each cohort. Up to 73 patients may be enrolled over approximately 3-4 years.

Arms: arm 1: Patients will receive a non-myeloablative lymphocyte depleting preparative regimen consisting of cyclophosphamide and fludarabine followed by intravenous (IV) infusion of Young tumor infiltrating lymphocytes (TIL) plus high dose IV aldesleukin. arm 2: Patients will receive a non-myeloablative lymphocyte depleting preparative regimen consisting of cyclophosphamide and fludarabine followed by intravenous (IV) infusion of Young tumor infiltrating lymphocytes (TIL) plus high dose IV aldesleukin
Interventions: intervention 1: Fludarabine intervention 2: Cyclophosphamide intervention 3: Young TIL intervention 4: Aldesleukin
Intervention descriptions: intervention 1: Fludarabine 25 mg/m(2)/day intravenous piggyback (IVPB) daily over 30 minutes for 5 days. intervention 2: Cyclophosphamide 60 mg/kg/day X 2 days intravenous (IV) in 250 ml in 5% dextrose in water (D5W) over 1 hr. intervention 3: Cells will be infused intravenously (i.v.) on the Patient Care Unit over 20 to 30 minutes (between one and four days after the last dose of fludarabine). intervention 4: Aldesleukin 720,000 IU/kg intravenous (IV) (based on total body weight) over a 15 minute period approximately every eight hours (+/- 1 hour) beginning within 24 hours of cell infusion and continuing for up to 5 days (maximum of 15 doses).

Protocol documents:
ICF_000.pdf: 
MEDICAL RECORD 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
INSTITUTE: 
National Cancer Institute 
STUDY NUMBER: 
12-C-0116 
PRINCIPAL INVESTIGATOR: Steven A. Rosenberg, M.D., Ph.D 
STUDY TITLE:  
A Phase II Study of Lymphodepletion followed by Autologous Tumor-
Infiltrating Lymphocytes and High-Dose Aldesleukin for Human Papillomavirus-
Associated Cancers 
Continuing Review Approved by the IRB on 01/11/16 
Amendment Approved by the IRB on 07/22/15  (J) 
                  Date posted to web:   02/05/16 
Standard 
PATIENT IDENTIFICATION 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
NIH-2514-1 (07-09) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent (1) 
 
INTRODUCTION 
 
We invite you to take part in a research study at the National Institutes of Health (NIH). 
 
First, we want you to know that: 
 
Taking part in NIH research is entirely voluntary. 
 
 
You may choose not to take part, or you may withdraw from the study at any time.  In 
either case, you will not lose any benefits to which you are otherwise entitled.  However, 
to receive care at the NIH, you must be taking part in a study or be under evaluation for 
study participation. 
 
 
You may receive no benefit from taking part.  The research may give us knowledge that 
may help people in the future. 
 
Second, some people have personal, religious or ethical beliefs that may limit the kinds of 
medical or research treatments they would want to receive (such as blood transfusions).  If you 
have such beliefs, please discuss them with your NIH doctors or research team before you agree 
to the study. 
 
Now we will describe this research study.  Before you decide to take part, please take as much 
time as you need to ask any questions and discuss this study with anyone at NIH, or with family, 
friends or your personal physician or other health professional. 
 
Why is this study being done?  
We have developed an experimental therapy that involves taking cells called lymphocytes from 
patients' tumors, growing them in the laboratory in large numbers, and then giving the cells back 
to the patient.  These cells are called Young Tumor Infiltrating Lymphocytes, or Young TIL and 
the therapy is called cell therapy.  Before receiving the cells, the patients receive 2 chemotherapy 
drugs to temporarily suppress the immune system to improve the chances that the tumor fighting 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
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NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 2 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
cells will be able to survive in the body. After the cells are given, the patients receive aldesleukin 
(IL-2) to help the tumor fighting cells stay alive longer.    
 
The purpose of this study is to see if these tumor fighting cells (young TIL) can cause tumors 
that are associated with the human papilloma virus (HPV) to shrink and to evaluate the toxicity 
of this treatment.  
 
Why are you being asked to take part in this study? 
You are being asked to participate in this study because you have been diagnosed with an HPV-
associated cancer such as cervical, vulvar, vaginal, penile, anal, or oropharyngeal cancer.  
 
How many people will take part in this Study?  
Up to 73 patients will be enrolled in this study. 
 
Description of Research Study 
The Cell protocol has several stages: 
Stage 
Timeframe 
Location 
Comments & Instructions 
Work up 
1-2 weeks 
Inpatient and 
out patient   
Scans, x-rays, labs leukapheresis other tests 
as needed 
Chemotherapy 
(day -7 to -1) 
1 week 
Inpatient 
Receive IV chemotherapy to prepare your 
immune system for the cells 
Cells and 
aldesleukin 
(Day 0-4) 
1-5 days 
Inpatient and 
possibly ICU 
Receive the TIL cells IV and then high dose 
aldesleukin about every 8 hours for up to 15 
doses 
Recovery 
1-2 weeks 
Inpatient unit Recover from the effects of treatment 
Follow -up 
Ongoing until 
disease 
progression 
Outpatient 
Return to clinic for physical exam, review 
of side effects, labs, scans every 1-6 
months. 
What will happen if you take part in this research study?  
 
Before you begin the study 
  
Cell harvest and growth 
We will be obtaining your cells for treatment from a biopsy of your tumor or during surgery, so 
we can grow Young TIL from your tumor cells in the laboratory.  In 1 out of 5 patients, we are 
not able to successfully grow the cells for this procedure.  If your cells do not grow or show HPV 
reactivity to certain cancer targets, you will not be able to receive the cell infusion.  If that 
happens, we will look at alternative treatments for you (best available care) or return you to the 
care of your referring physician.  We usually know after about 4 weeks whether the cells will 
grow well enough to be used as an experimental treatment on this protocol.  At the time we 
determine that your cells are not growing, we will inform you and discuss your options with you.  
 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 3 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
Work up 
Prior to receiving the experimental treatment you will undergo many tests. These include 
imaging procedures, heart and lung function tests, and laboratory tests.  If you are a woman, you 
will undergo a pregnancy test.  You will also have a large catheter inserted into a vein and 
leukapheresis will be performed (see below).  You may be admitted to the hospital for these 
tests. However, you will be allowed to leave on pass on the days that you are not having tests 
performed. 
 
Catheter insertion 
Prior to beginning the experimental treatment, you will have an intravenous (IV) catheter placed 
in your upper chest. The area will be numbed with an anesthetic before the catheter is put in.  
Although rare, putting these catheters in can sometimes cause collapse of a lung or cause 
bleeding.  Lung collapse is treated by putting a tube into your chest for a few days to allow your 
lung to expand.  Pressure is placed on any area that might bleed.  Other IVs may be needed in 
one or both of your arms if we need to give you extra fluids, medicines, or nutrition. 
 
Leukapheresis 
Leukapheresis is a procedure that allows us to remove certain types of blood cells from you and 
return the rest of your blood.  It is a very common procedure that is done routinely here at the 
NIH with very few risks.  During leukapheresis,  blood is removed from you through a needle in 
your arm, circulated through a machine that divides whole blood into red cells, plasma (the 
serum part), and lymphocytes (or white cells), and then the plasma and red cells are returned to 
you through a second needle in your other arm.  The white blood cells may be used to help grow 
the cells. In addition to the leukapheresis you will undergo as part of your work up, we will also 
ask you to undergo one additional pheresis procedure between 4 and 6 weeks after your cell 
treatment to see the impact of this therapy on the immune system and see if cells we gave you 
are still active. 
 
Chemotherapy Regimen (Day -7 through Day -1) 
After we have grown the Young TIL cells to large numbers in the laboratory, you will be 
admitted to the hospital to begin your experimental treatment.  You will be given two 
chemotherapy medicines, cyclophosphamide and fludarabine, to make space in your immune 
system so the Young TIL can work without any interference from the cells in your immune 
system. These medicines may cause your tumor to shrink some, but this shrinkage is anticipated 
to be only partial and of small duration.  The main purpose of the chemotherapy is to see if we 
can make the cells more effective in fighting cancer tumors.  Animal experiments have indicated 
that chemotherapy can make the cells more effective in fighting cancer tumors, but it is not 
known whether this is true in humans.  The cyclophosphamide will be given into your catheter 
over 1 hour for two days (Day -7 and Day -6) and the fludarabine will be given into your catheter 
for 30 minutes every day for the next five days (Day -5 through Day -1).  The side effects of 
these medicines are described on the following pages. 
 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 4 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
Cell Infusion and Aldesleukin Regimen (Day 0 through Day 5) 
All patients will be given the cells through their IV over 20-30 minutes one to four days after the 
last dose of chemotherapy.  Within 24 hours after your cell infusion you will be given high dose 
aldesleukin through one of the IVs.  It will be given as a 15-minute infusion about every 8 hours 
for up to five days after the cell infusion.  Aldesleukin is a cell growth factor and it is thought 
that it will help the cells live longer in your body.  
 
The day after your cells are infused, we will give you G-CSF (filgrastim) as a shot or injection 
under the skin every day to stimulate your blood cells until they increase to a sufficient number.  
We will watch you closely during this entire time for any side effects of this experimental 
regimen.  We will discuss the side effects below and we will include in your care all the 
medicines and treatments to prevent as many of these side effects as we can and to make you as 
comfortable as we can.   
 
Recovery 
You will recover in the hospital until you are well enough to go home.  This usually takes 7-12 
days after you have received cells or your last dose of aldesleukin; however, you may need to 
stay in the hospital for longer than this before you are well enough to go home. We will continue 
to give you support medications, do laboratory tests, and watch you closely for any side effects 
until we feel your condition is stable.  
 
In addition to the laboratory tests to monitor your condition, we will remove between 1 and 9 
teaspoons of blood daily to study the effects of this regimen on your immune system.  If you 
experience side effects in you kidneys, we will collect 1 additional teaspoon of blood and about 6 
teaspoons of urine to see if we can determine the cause of these side effects.  The maximum 
amount of blood for research is approximately 2.3 cups in 8 weeks. 
 
Follow up and Evaluation of Experimental Regimen  
You will need to continue to take Bactrim, an antibiotic, for at least 6 months following your 
treatment to prevent you from catching a certain type of pneumonia seen in patients who have 
low white blood cell counts. We will ask you to return to NIH 4-6 weeks after completing your 
regimen for evaluation.  This visit will probably take 2 days.   If your tumor shows evidence of 
shrinking, we will ask you to return for evaluation every 4-6 weeks for 1 year and then every six 
months after that for up to 5 years, and then yearly thereafter.  If your tumor appears to be 
growing, we will look for other investigational therapies you may be eligible for, or refer you 
back to the care of your local physician.  At some of your follow up visits, you may undergo 
leukapheresis so that we can see the effect this therapy has had on your immune system and if 
the cells we gave you are still alive.  
 
Retreatment 
If your tumor shrinks or disappears following the initial treatment and then recurs you may 
receive one additional treatment if you tolerated the treatment well and if all the side effects have 


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NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 5 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
resolved.  You will receive the same medications and cell infusion on the same schedule as with 
the first treatment. 
 
Birth Control 
 
If you are a woman who is breast feeding or pregnant, you may not participate in the study 
because we don’t know how this medicine would affect your unborn child or your baby.  If you 
are a woman who can become pregnant, or are the partner of a woman who can become 
pregnant, you will need to practice an effective form of birth control before starting study 
treatment, during study treatment, and for four months after you finish study treatment.  If you 
think that you or your partner is pregnant, you should tell your study doctor or nurse at once. 
 
Effective forms of birth control include: 
• abstinence 
• intrauterine device (IUD) 
• hormonal [birth control pills, injections, or implants] 
• tubal ligation 
• vasectomy 
 
Alternative Approaches or Treatments 
 
What other choices do I have if I do not take part in this study?  
 
If there are effective salvage regimens (regimens used when standard regimens have failed), you 
will be directed to undergo these regimens before participating in this protocol. 
 
Other options for treatment of your cancer include: 
• Standard chemotherapies, including targeted therapies; 
• experimental vaccines;  
• experimental chemotherapies; or biotherapies;  
• other combination therapies; or 
• getting no treatment; or getting comfort care which is also called palliative care. This 
type of care helps reduce pain, tiredness, appetite problems, and other problems caused 
by cancer. It does not treat the cancer directly, but instead tries to improve how you feel. 
Comfort care tries to keep you as active and comfortable as possible. 
 
Please talk to your doctor about these and other options. 
 
 
 
 
 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 6 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
Risks or Discomforts of Participation 
 
What side effects or risks can I expect from being in this study? 
The risks and discomforts of this research study can be significant.  This experimental treatment 
can lead to long-term decrease in your immune function.  If you have received radiation therapy, 
you will need to recover completely (usually at least 4 weeks) from the side effects of the 
radiation before beginning treatment on this protocol.  It is also possible that you may lose your 
fertility following this experimental treatment. It is possible, although unlikely, that this 
experimental treatment may cause your death. 
 
We will discuss the side effects of this experimental treatment with you. You will be given 
medicines, transfusions, and treatments to prevent or treat the side effects including drugs to 
prevent and/or treat different types of infections.  We will try to make you as comfortable as 
possible. 
 
Leukapheresis 
 
During the leukapheresis procedure, you may have some tingling in your face and lips due to the 
medicine used to keep your blood from clotting during the procedure.  The nurses may give you 
a calcium-containing antacid to chew that takes away this tingling.  Rarely, people may 
experience lightheadedness or dizziness.  We ask that you eat prior to the procedure to prevent 
this.  Rare complications of this procedure are lowered blood pressure, bleeding or bruising 
where the needles are put in your arms. 
 
Young TIL Cell Infusion  
All Young TIL cells will be given through your catheter while we keep you in the patient care 
unit so we can watch you closely. 
 
Potential risks include:  
• Fever, chills and shortness of breath, which are common and may last for a few hours 
• Lung congestion causing shortness of breath 
• Immune-type reaction, such as pharyngitis and vaginitis. Note: In similar clinical 
trials with cells targeting a melanoma protein, we have observed the following 
immune-mediated toxicities: loss of skin pigment (known as vitiligo), and 
inflammation of the eye (uveitis).  The skin and the eye are all sites where that 
targeted melanoma protein is known to exist.   
• As this is a new experimental therapy, side effects that we do not anticipate that may 
cause your condition to deteriorate may be encountered.  Any new information that 
becomes available during the course of this study will be shared with you. 
 
 
 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
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NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 7 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
Medications 
 
The side effects of cyclophosphamide, fludarabine, high dose aldesleukin and some of the other 
medications you will receive are listed below:  
 
 
Cyclophosphamide and Fludarabine side effects 
Common 
Less Common 
Rare 
 
Changes in blood 
counts including: 
low red cell count 
(causing fatigue and 
shortness of breath), 
low platelet count 
(increasing the risk 
of bleeding and 
bruising), decrease 
in white blood cells 
(increasing the risk 
of infection and the 
need for treatment 
with antibiotics or 
other treatment)  
 
Loss of appetite, 
nausea, vomiting,  
 
Diarrhea, stomach 
pain 
 
Mouth sores 
 
Hair loss 
 
Fatigue 
 
Muscle or joint 
aches  
 
Bleeding 
 
Infection 
 
Bladder irritation with 
bloody urine 
 
Severe allergic reaction 
(difficulty 
breathing/swelling) 
 
Headache or dizziness 
 
Sweating 
 
Swelling of arms or legs 
 
Skin changes, rash, 
blisters  
 
Weakness 
 
Hearing loss 
 
Heart damage 
 
Lung damage 
 
Kidney damage 
 
Inflammation of the eye resulting 
in blindness 
 
Inflammation of nervous system 
resulting in death 
 
Epstein Barr Virus Lymphoma.  
This can be fatal (Two patients 
on other studies in the Surgery 
Branch developed EBV 
lymphoma, and one died as a 
result of this disease.) 
 
Loss of fertility 
 
Two out of the first 81 patients 
enrolled on the initial young TIL 
study died from complications 
resulting from suppression of the 
immune function which resulted 
in a severe infection (one of 
these patients also received 
radiation as part of their 
treatment regimen).   
 
 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
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NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 8 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
 
IL-2 (aldesleukin) side effects 
Common 
Less common  
Rare 
 
Fever, chills, and 
fatigue   
 
Lowered platelet and 
red blood cell levels 
that may require 
transfusions  
 
Significant fluid 
retention causing 
weight gain (as much as 
20 pounds).  
 
Low blood pressure 
 
Increased heart rate 
 
Low urine output 
 
Swelling in your 
extremities,  
 
Fluid in your lungs that 
can require oxygen  
 
Dry mouth, nausea, 
vomiting and diarrhea; 
 
Rash, itching; and 
changes in skin or hair 
pigmentation, called 
vitiligo;   
 
Changes in mental 
status, including 
confusion, difficulty 
sleeping or vivid 
dreams; this can be 
severe and require 
sedation and 
monitoring in the ICU 
 
 
Decrease in thyroid function 
that may require daily 
thyroid hormone 
replacement; 
 
Abnormal kidney and liver 
function that can be severe;   
 
Abnormal heartbeats or low 
blood pressure that may 
require treatment in the 
ICU.  
 
Breathing problems which 
may need monitoring in 
ICU and insertion of a 
breathing tube.   
 
 
Bowel perforation (a hole) 
requiring longer 
hospitalization or surgery. 
 
Autoimmune disease, where 
your immune system attacks 
cells in organs of your body.  
Should this occur, you will 
be treated with steroids to 
stop the immune response. 
 
Damage to the heart muscle 
or heart attack 
 
Loss of blood flow to the 
extremities due to medicines 
used to treat very low blood 
pressure and shock. In one 
instance a patient had to 
have her lower arm 
amputated after treatment 
with these medicines. 
 
IL-2 is mixed with human 
albumin which could cause 
an allergic reaction or 
potentially transmit viral 
infections, although we have 
not had this occur. 
 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 9 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
 
Support Medications – side effects 
Common 
Less common 
Rare 
Filgrastim (To increase production of white blood cells) 
 
Bone Pain 
 
Severe headache 
 
Severe breathing problems 
 
Rupture of your spleen 
Bactrim (To prevent a specific type of pneumonia) 
 
 
Fever 
 
Nausea, vomiting,  
 
Skin rash with itching 
 
reduced number of 
white blood cells  
 
Allergic reaction 
 
Fluconazole:  (To prevent fungal infections)  
 
Headache 
 
Nausea, vomiting, 
diarrhea, abdominal 
pain 
 
Itching  
 
 
A skin disorder called Stevens 
Johnson Syndrome, which can be 
fatal 
 
Liver damage which may be 
permanent 
Acyclovir and Valacyclovir 
 
 
Temporary decrease in 
kidney function which 
may not cause any 
symptoms 
 
Nausea, vomiting, 
diarrhea, constipation  
 
Pain and irritation at 
place of injection 
 
Skin rash, hives, itching  
 
Tremors, dizziness, Confusion, 
seizures 
 
Fatigue  
 
Blood in the urine 
 
Potential Benefits of Participation 
 
Are there benefits to taking part in this study?  
The aim of this study is to see if this new experimental treatment will cause your tumors to 
shrink. We do not know if you will receive personal, medical benefit from taking part in this 
study. These potential benefits could include shrinking of your tumor or lessening of your 
symptoms, such as pain, that are caused by the cancer.  Because there is not much information 
about the Young TIL therapy effect on your type of cancer, we do not know if you will benefit 
from taking part in this study, although the knowledge gained from this study may benefit others 
in the future who have cancer.   
 
 
 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 10 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
Research Subject’s Rights 
 
What are the costs of taking part in this study?  
If you choose to take part in the study, the following will apply, in keeping with the NIH policy: 
• 
You will receive study treatment at no charge to you. This may include 
surgery, medicines, laboratory testing, x-rays or scans done at the 
Clinical Center, National Institutes of Health (NIH), or arranged for you 
by the research team to be done outside the Clinical Center, NIH if the 
study related treatment is not available at the NIH.  
• 
There are limited funds available to cover the cost of some tests and 
procedures performed outside the Clinical Center, NIH.  You may have 
to pay for these costs even if they are not covered by your insurance 
company. 
• 
Medicines that are not part of the study treatment will not be provided or 
paid for by the Clinical Center, NIH. 
• 
Once you have completed taking part in the study, medical care will no 
longer be provided by the Clinical Center, NIH. 
 
Will your medical information be kept private? 
 
We will do our best to make sure that the personal information in your medical record will be 
kept private.  However, we cannot guarantee total privacy.  Organizations that may look at 
and/or copy your medical records for research, quality assurance, and data analysis include: 
• The National Cancer Institute (NCI) and other government agencies, like the Food and 
Drug Administration (FDA), which are involved in keeping research safe for people. 
• NCI Institutional Review Board 
• The study Sponsor (Steven A. Rosenberg, M.D., Ph.D.) 
• Prometheus Laboratories, Inc. 
 
A description of this clinical trial will be available on http://www.Clinicaltrials.gov, as required 
by U.S. Law.  This Web site will not include information that can identify you.  At most the Web 
site will include a summary of the results.  You can search this Web site at any time. 
 
Stopping Therapy  
Your doctor may decide to take you off this study under the following circumstances: 
• if he/she believes that it is in your best interest 
• if your disease comes back during treatment 
• if you experience side effects from the treatment that are considered too severe 
• if new information becomes available that shows that another treatment would be better for 
you 
 


MEDICAL RECORD 
CONTINUATION SHEET for either: 
NIH 2514-1, Consent to Participate in A Clinical Research Study 
NIH 2514-2, Minor Patient’s Assent to Participate In A Clinical Research Study 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 11 of 13 pages 
 
PATIENT IDENTIFICATION 
CONTINUATION SHEET for either: 
NIH-2514-1 (07-09) 
NIH-2514-2 (10-84) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
In this case, you will be informed of the reason for that decision. 
 
You can stop participating at any time. However, if you decide to stop participating in the study, 
we encourage you to talk to the study doctor and your regular doctor first. If you refuse to 
participate or withdraw from the protocol or at the completion of the protocol, we will attempt to 
offer you participation in other NIH protocols if these are available, or will refer you to your 
home physician for further management. 
 
If you decide at any time to withdraw your consent to participate in the trial, we will not collect 
any additional medical information about you. However, according to FDA guidelines, 
information collected on you up to that point may still be provided to the Sponsor. If you 
withdraw your consent and leave the trial, any samples of yours that have been obtained for the 
study and stored at the NCI can be destroyed upon request. However, any samples and data 
generated from the samples that have already been distributed to other researchers or placed in 
the research databases cannot be recalled and destroyed. 
 
You should understand that this study involves research and that your participation is voluntary. 
Unexpected or unforeseeable side effects may also occur. Your participation in this protocol may 
be terminated without your consent if your physician feels that it would not be safe for you to 
continue.  Any significant new findings that relate to this protocol will be discussed with you. 
 
Conflict of Interest 
 
The National Institutes of Health reviews NIH staff researchers at least yearly for conflicts of 
interest. This process is detailed in a Protocol review Guide.   You may ask your research team 
for additional information or a copy of the Protocol Review Guide or for more information.  
Members of the research team who do not work for NIH are expected to follow these guidelines 
but they do not need to report their personal finances to the NIH. 
 
Members of the research team working on this study may have up to $15,000 of stock in the 
companies that make products used in this study.  This is allowed under federal rules and is not a 
conflict of interest.  
   
It is possible that the information obtained from your participation on this study may become 
valuable for commercial research and development purposes (including patentable inventions), 
which may be of significant benefit to society, the sponsor of this study, individual researchers or 
other third parties.  You will not receive direct financial benefit from such research and 
development.  The NIH and 2 of the investigators on the research team have developed the cell 
process being used in this research and have a patent pending.  This means that it is possible that 
the results of this study could lead to payments to NIH scientists and to the NIH.  By law, 
government scientists are required to receive such payments for their inventions.   
 


MEDICAL RECORD 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 12 of 13 pages 
 
PATIENT IDENTIFICATION 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH 
STUDY (Continuation Sheet) 
• 
Adult Patient or 
• Parent, for Minor Patient 
NIH-2514-1 (07-09) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
 
OTHER PERTINENT INFORMATION 
1. 
Confidentiality.  When results of an NIH research study are reported in medical journals 
or at scientific meetings, the people who take part are not named and identified.  In most cases, 
the NIH will not release any information about your research involvement without your written 
permission.  However, if you sign a release of information form, for example, for an insurance 
company, the NIH will give the insurance company information from your medical record.  This 
information might affect (either favorably or unfavorably) the willingness of the insurance 
company to sell you insurance. 
The Federal Privacy Act protects the confidentiality of your NIH medical records.  However, you 
should know that the Act allows release of some information from your medical record without 
your permission, for example, if it is required by the Food and Drug Administration (FDA), 
members of Congress, law enforcement officials, or authorized hospital accreditation 
organizations. 
2. 
Policy Regarding Research-Related Injuries.  The Clinical Center will provide short-
term medical care for any injury resulting from your participation in research here.  In general, 
no long-term medical care or financial compensation for research-related injuries will be 
provided by the National Institutes of Health, the Clinical Center, or the Federal Government.  
However, you have the right to pursue legal remedy if you believe that your injury justifies such 
action. 
3. 
Payments.  The amount of payment to research volunteers is guided by the National 
Institutes of Health policies.  In general, patients are not paid for taking part in research studies at 
the National Institutes of Health.  Reimbursement of travel and subsistence will be offered 
consistent with NIH guidelines. 
4. 
Problems or Questions.  If you have any problems or questions about this study, or 
about your rights as a research participant, or about any research-related injury, contact the 
Principal Investigator, Steven A. Rosenberg, M.D., Ph.D., Building 10 CRC, Room 3-3940, 
Telephone:  301-496-4164. You may also call the Clinical Center Patient Representative at 301-
496-2626.  
5. 
Consent Document.  Please keep a copy of this document in case you want to read it 
again. 


MEDICAL RECORD 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH STUDY 
• 
Adult Patient or 
• Parent, for Minor Patient 
STUDY NUMBER: 12-C-0116 
CONTINUATION: page 13 of 13 pages 
 
PATIENT IDENTIFICATION 
CONSENT TO PARTICIPATE IN A CLINICAL RESEARCH 
STUDY (Continuation Sheet) 
• 
Adult Patient or 
• Parent, for Minor Patient 
NIH-2514-1 (07-09) 
P.A.:  09-25-0099 
File in Section 4:  Protocol Consent 
 
 
 
 
COMPLETE APPROPRIATE ITEM(S) BELOW: 
A. Adult Patient’s Consent 
B. Parent’s Permission for Minor Patient. 
I have read the explanation about this study 
and have been given the opportunity to discuss 
it and to ask questions.  I hereby consent to 
take part in this study. 
I have read the explanation about this study 
and have been given the opportunity to discuss 
it and to ask questions.  I hereby give 
permission for my child to take part in this 
study.   
(Attach NIH 2514-2, Minor’s Assent, if 
applicable.) 
 
  
  
 
 
 
Signature of Adult Patient/ 
Legal Representative 
Date 
Signature of Parent(s)/ Guardian Date 
 
 
Print Name 
 
 
Print Name 
C. Child’s Verbal Assent (If Applicable) 
The information in the above consent was described to my child and my child agrees to 
participate in the study. 
 
  
  
 
Signature of Parent(s)/Guardian 
Date 
Print Name 
THIS CONSENT DOCUMENT HAS BEEN APPROVED FOR USE  
FROM JANUARY 11, 2016 THROUGH JULY 10, 2016 
 
 
 
 
 
 
 
 
 
 
 
 
Signature of Investigator 
Date 
Signature of Witness 
Date 
 
 
Print Name 
 
 
Print Name 
 

Prot_001.pdf: 
Amendment
Protocol Number:
12C0116-K
Principal Investigator: Steve Rosenberg 
NCI 
SB 
301.496.4164 
sar@mail.nih.gov
(NIH Employee Name, Institute/Branch, Telephone and e-mail)
Reference Number:
359203
Protocol Title:
A Phase II Study of Lymphodepletion followed by Autologous Tumor-Infiltrating Lymphocytes and High-Dose Aldesleukin for Human 
Papillomavirus-Associated Cancers
SIGNATURES
Principal Investigator (*): 
Jim Yang - applied signature on 09/12/2016 1:13 PM EDT
Accountable Investigator:
PI is the Accountable Investigator
Branch Chief/CC Department Head (**):
Jim Yang - applied signature on 09/12/2016 1:13 PM EDT
Medical Advisory Investigator (if applicable):
N/A
Lead Associate Investigator signature:
N/A
Referral Contact signatures:
N/A
Associate Investigators signatures:
Debbie Nathan - applied signature on 09/12/2016 12:05 PM EDT 
Debbie Nathan - applied signature on 09/12/2016 2:09 PM EDT
For Institute/Center Scientific Review Committee:
N/A
Other IC Clinical Director signatures:
N/A
APPROVALS
IRB Chair:
Michael Hamilton - applied signature on 09/12/2016 7:15 PM EDT
Clinical Director:
N/A
CONCURRENCE
OPS Protocol Specialist:
Signature
Print Name
Date
* Signature signifies that investigators on this protocol have been informed that the collection and use of personally identifiable information at
the NIH are maintained in a system of record governed under provisions of the Privacy Act of 1974. The information provided is mandatory for 
employees of the NIH to perform their assigned duties as related to the administration and reporting of intramural research protocols and 
used solely for those purposes.  Questions may be addressed to the Protrak System Owner. 
** I have reviewed this research project and considered the NIH Policy for Inclusion of Women and Minorities in Clinical Research. Taking into 
account the overall impact that the project could have on the research field involved, I feel the current plans adequately includes both sex/
gender, minorities, children, and special populations, as appropriate. The current enrollment is in line with the planned enrollment report for 
inclusion of individuals on the basis of their sex/gender, race, and ethnicity and is appropriate and of scientific and technical merit. 
IRB Meeting Date:
Expedited
DEC Clearance Date:
NA
Protocol Version Date:09/08/2016
GPS1*
Andrea N. Kelley 
 
 
 
   AM K  
 
 
   09/14/2016


Abbreviated Title: HPV TIL 
Version Date: 09/08/2016 
Confidential 
1
NCI Protocol Number: 12-C-0116 K 
Version Date: 09/08/2016 
 
PROTOCOL TITLE 
A Phase II Study of Lymphodepletion followed by Autologous Tumor-Infiltrating Lymphocytes 
and High-Dose Aldesleukin for Human Papillomavirus-Associated Cancers 
 
NCI Principal Investigator: 
Steven A. Rosenberg, M.D., Ph.D. A, B, C, D, E 
           
 
 
 
Chief of Surgery, CCR, NCI 
 
 
 
 
 
 
Building 10, CRC, Room 3-3940 
 
 
 
9000 Rockville Pike, Bethesda, MD 20892 
 
 
 
Phone: 301-496-4164 
 
 
 
Email: Steven.Rosenberg@nih.gov 
 
NIH Associate Investigators: 
Christian S. Hinrichs, MD, ETIB, NCI A, B, C, D, E 
 
 
 
Robert Somerville, Ph.D., Surgery Branch, NCI E 
 
 
 
James C. Yang, M.D., Surgery Branch, NCI A, B, C, D, E 
 
Richard Sherry, M.D., Surgery Branch, NCI A, B, C, D, E 
 
Udai S. Kammula, M.D., Surgery Branch, NCI A, B, C, D, E 
 
Stephanie Goff, M.D., Surgery Branch, NCI A, B, C, D, E 
 
Sanja Stevanovic, Ph.D., ETIB, NCI E 
Sarah Helman, ETIB, NCI E 
 
Nicholas P. Restifo, M.D., Surgery Branch, NCI E 
 
Donald E. White, M.S., Surgery Branch, NCI E 
 
Seth M. Steinberg, Ph.D., BDMS, CCR, NCI E 
 
Daniel Zlott, PharmD., Pharmacy Dept., NIH Clinical Center E 
 
Monica Epstein, R.N., OCD, CCR, NCI A, B, C, D, E 
 
Marie Statler, R.N., OCD, CCR, NCI A, B, C, D, E 
 
Abigail Johnson, R.N., OCD, CCR, NCI A, B, C, D, E 
 
Lori McIntyre, R.N., OCD, CCR, NCI A, B, C, D, E 
 
Study Coordinator: 
 
Monica Epstein, R.N., OCD, CCR, NCI A, B, C, D, E 
 
 
 
Phone: 301-435-2375 
 
 
 
Email: Monica.Epstein@nih.gov 
 
 Referral Contacts: 
Jessica Yingling, R.N. OCD, CCR, NCI A, B, D 
 
 
 
Ellen Bodurian, R.N., OCD, CCR, NCI A, B, D 
 
Collaborator: 
 
Steven Fischkoff, MDF 
 
 
 
Chief Medical Officer 
 
 
 
Lion Biotechnologies, Inc. 
 
A. Obtain information by intervening or interacting with living individuals for research purposes 
B. Obtaining identifiable private information about living individuals 
C. Obtaining the voluntary informed consent of individuals to be subjects 
D. Makes decisions about subject eligibility 
E. Studying, interpreting, or analyzing identifiable private information or data/specimens for 
research purposes 
F. Studying, interpreting, or analyzing de-identified data or specimens for research purposes 
 


Abbreviated Title: HPV TIL 
Version Date: 09/08/2016 
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Investigational Agents: 
 
Drug Name: 
Young TIL 
IND Number: 
IND 14355 
Sponsor: 
Center for Cancer Research 
Manufacturer: Surgery Branch Cell Processing Facility 
 
Commercial Agents: Cyclophosphamide, Fludarabine, and Aldesleukin 
 
 


Abbreviated Title: HPV TIL 
Version Date: 09/08/2016 
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PRECIS 
Background: 
• Metastatic or locally advanced refractory/recurrent human papillomavirus (HPV)-
associated malignancies (cervical, vulvar, vaginal, penile, anal, and oropharyngeal) are 
incurable and poorly palliated by standard therapies. 
• Administration of autologous tumor infiltrating lymphocytes (TIL) generated from 
resected metastatic melanoma can induce objective long-term tumor responses. 
• Young TIL can be generated from HPV-associated tumors. 
 
Objectives: 
• To determine if autologous Young TIL infused in conjunction with high dose aldesleukin 
following a non-myeloablative lymphodepleting preparative regimen can mediate tumor 
regression in patients with metastatic or locally advanced refractory/recurrent HPV-
associated cancer. 
• To study immunologic correlates associated with Young TIL therapy for HPV-associated 
cancers. 
• To determine the toxicity of this treatment regimen. 
Eligibility:  
• Patients greater than or equal to 18 years old with a pathologically confirmed diagnosis of 
metastatic or locally advanced refractory/recurrent HPV-16+ or HPV-18+ human 
papillomavirus-associated cancer.  
Design: 
• Patients will undergo biopsy or resection to obtain tumor for generation of autologous 
TIL cultures and autologous cancer cell lines.  
• All patients will receive a non-myeloablative lymphocyte depleting preparative regimen 
of 
cyclophosphamide (60 mg/kg/day IV) on days -7 and -6 and fludarabine (25 mg/m2/day 
IV) on days -5 through -1. 
• On day 0 patients will receive between 1x109 to 2x1011 young TIL and then begin high 
dose 
aldesleukin (720,000 IU/kg IV every 8 hours for up to 15 doses). 
• Clinical and immunologic response will be evaluated about 4-6 weeks after TIL infusion. 
• Initially, 18 evaluable patients will be enrolled in two cohorts; patients with cervical 
cancer and those with non cervical cancer  For each cohort, if 0 to 2 of the 18 patients 
experience a clinical response, then no further patients will be enrolled.  If 3 or more of 
the first 18 evaluable patients enrolled have a clinical response, then accrual will continue 
until a total of 35 evaluable patients have been enrolled in each cohort. Up to 73 patients 
may be enrolled over approximately 3-4 years. 


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TABLE OF CONTENTS  
PRECIS .......................................................................................................................................... 3 
Table of Contents ............................................................................................................................ 4 
1. 
INTRODUCTION .................................................................................................................. 6 
1.1. 
STUDY OBJECTIVES ......................................................................................................... 6 
1.2. 
BACKGROUND AND RATIONALE: .................................................................................... 6 
2. 
ELIGIBILITY ASSESSMENT AND ENROLLMENT ......................................................... 9 
2.1 
Eligibility Criteria ............................................................................................................ 9 
2.2. 
Screening Evaluation...................................................................................................... 11 
2.3. 
Registration Procedures.................................................................................................. 12 
3. 
STUDY IMPLEMENTATION ............................................................................................ 12 
3.1 
Study Design .................................................................................................................. 12 
3.2 
Drug Administration ...................................................................................................... 13 
3.3 
On-Study Evaluations: ................................................................................................... 16 
3.4 
Retreatment .................................................................................................................... 16 
3.5 
Post Study Evaluation (Follow-up) ................................................................................ 17 
3.6 
Criteria for Removal from Protocol Therapy and Off Study Criteria ............................ 18 
3.6.1 
Off Treatment Criteria ................................................................................................ 18 
3.6.2 
Off Study Criteria ....................................................................................................... 18 
4. 
CONCOMITANT MEDICATIONS/MEASURES .............................................................. 18 
4.1 
Infection Prophylaxis: .................................................................................................... 18 
4.2 
Blood Product Support ................................................................................................... 19 
4.3 
Empiric antibiotics ......................................................................................................... 19 
4.4 
Other Concomitant medications to control side effects ................................................. 19 
5. 
BIOSPECIMEN COLLECTION .......................................................................................... 20 
5.1 
Correlative Studies for Research .................................................................................... 20 
5.1.1 
Prior to Chemotherapy Administration: ..................................................................... 20 
5.1.2 
Prior To Cell Infusion: ................................................................................................ 20 
5.1.3 
Post Cell Infusion Evaluations: .................................................................................. 20 
5.1.4 
Immunological Testing: .............................................................................................. 20 
5.2. 
Sample Storage, Tracking and Disposition .................................................................... 21 
6. 
DATA COLLECTION AND EVALUATION ..................................................................... 22 
6.1 
Data Collection .................................................................................................................. 22 


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6.2 
Routine Adverse Event Reporting.................................................................................. 22 
6.3 
Response Criteria ........................................................................................................... 23 
6.4 
Toxicity Criteria ............................................................................................................. 25 
7. 
SAFETY REPORTING REQUIREMENTS/DATA AND SAFETY MONITORING PLAN
 
26 
7.1 
Definitions ...................................................................................................................... 26 
7.2 
NCI-IRB Reporting ........................................................................................................ 28 
7.3 
IND Sponsor Reporting Criteria .................................................................................... 29 
7.4 
FDA Reporting Criteria .................................................................................................. 29 
7.4. 
Data and Safety Monitoring Plan ................................................................................... 30 
8. 
STATISTICAL CONSIDERATION .................................................................................... 31 
9. 
COLLABORATIVE AGREEMENTS ................................................................................. 31 
10. 
HUMAN SUBJECTS PROTECTIONS ............................................................................ 31 
10.1. 
Rationale For Subject Selection ................................................................................. 31 
10.2. 
Participation of Children ............................................................................................ 32 
10.3. 
Evaluation of Benefits and Risks/Discomforts ........................................................... 32 
10.4. 
Risks/Benefits Analysis .............................................................................................. 32 
10.5. 
Consent and Assent Process and Documentation ....................................................... 32 
11. 
PHARMACEUTICAL INFORMATION .......................................................................... 33 
11.1. 
Interleukin-2 (Aldesleukin, Proleukin, Recombinant Human Interleukin 2) ............. 33 
11.2. 
Fludarabine: ................................................................................................................ 33 
11.3. 
Cyclophosphamide ..................................................................................................... 34 
11.4. 
YoungTIL Preparation ................................................................................................ 35 
11.5. 
Mesna.......................................................................................................................... 35 
11.6. 
Filgrastim .................................................................................................................... 36 
11.7. 
Trimethoprim and Sulfamethoxazole Double Strength (TMP / SMX DS): ............... 36 
11.8. 
Aerosolized Pentamidine in Place of TMP/SMX DS:................................................ 36 
11.9. 
Herpes Virus Prophylaxis: .......................................................................................... 36 
11.10. 
Support Medications ................................................................................................... 37 
12. 
REFERENCES .................................................................................................................. 39 
13. 
Figures, tables & appendices ............................................................................................. 42 


Abbreviated Title: HPV TIL 
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1. 
INTRODUCTION 
1.1. STUDY OBJECTIVES 
1.1.1. 
Primary Objective: 
• To determine the objective tumor response rate and duration in patients with 
metastatic or recurrent/refractory locally advanced HPV-associated cancers who 
receive autologous Young TIL plus aldesleukin following a lymphodepleting 
preparative regimen.  
 
1.1.2. 
Secondary Objective(s): 
• To study immunologic correlates associated with Young TIL therapy for HPV-
associated cancers. 
• To determine the toxicity of this treatment regimen. 
 
1.2. BACKGROUND AND RATIONALE: 
This is a clinical protocol to treat HPV-associated cancers with adoptive transfer of autologous 
young tumor infiltrating lymphocytes (young TIL). This approach seeks to direct the potent 
immunotherapy of TIL adoptive transfer at the attractive immunologic target of HPV-associated 
cancers. The potential of TIL to mediate complete and durable responses in metastatic melanoma 
has been established by work from the Surgery Branch1. We are now seeking to expand the 
application of young TIL to HPV-positive cancers, which are particularly appealing candidates 
for TIL therapy because i) better treatments are desperately needed, especially for patients that 
have previously received chemotherapy, ii) these cancers express the viral antigens E6 and E7, 
which are good immunologic targets and are important for malignant cell function and survival, 
iii) the presence of known tumor antigens permits preferential transfer of tumor specific T cells 
and enables novel scientific studies that can inform future clinical trials.  
 
1.2.1. HPV-associated malignancies 
HPV-associated cancers, particularly cervical cancer, are a major international health 
concern, but also remain problematic in the United States. In the United states annually, 
there are greater than 19,000 new cases of HPV-associated cancer and approximately 
11,000 deaths from cancer at HPV-associated sites, which include cervix, vagina, vulva, 
anus, penis, and oropharynx (Table 1)2-5. Metastatic or recurrent/refractory locally 
advanced malignancies at these sites are incurable and difficult to palliate. Responses to 
chemotherapy (platinum-based) are variable but generally short-lived with median 
progression-free survival (PFS) around only 3 to 6 months6-8. In a recent Gynecologic 
Oncology Group randomized trial comparing four cisplatin-based doublets as first line 
therapy for cervical cancer the response rates were only 22 to 29 percent9. Median PFS 
was four to six months and median overall survival (OS) was 10 to 13 months9. 
Randomized trials of second line therapy are lacking but response rates for single agents 
are poor at around 3 to 12 percent10. For oropharyngeal cancer, HPV positive tumors 
have improved prognosis in patients receiving chemoradiation as first line treatment of 
the primary tumor; however the effect of HPV status on outcomes of metastatic or 


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7
recurrent/refractory disease is unknown. Furthermore HPV positive tumors have not been 
studied as distinct entities in trials of systemic therapy. The best estimates of the 
chemotherapy responsiveness of these tumors probably comes from looking at the 
oropharyngeal site (an HPV associated site) in subset analyses of broader head and neck 
cancer clinical trials. In a pivotal clinical trial that established platinum, 5-flurouracil, 
plus cetuximab as first line therapy in head and neck cancer, patients with oropharyngeal 
tumors experienced PFS of four to six months and OS of eight to 11 months7. As with 
cervical cancer, randomized trials of second line therapy following platinum treatment 
failure are lacking. A phase II trial of cetuximab, a monoclonal antibody targeting 
epidermal growth factor receptor (EGFR) reported a response rate of 13 percent with 
time to progression of 70 days; cetuximab is now considered to represent the most 
promising second line therapy for this disease11. Thus, second line therapies for HPV-
associated tumors have low response rates and poor response duration, and novel 
treatments must be developed. 
 
1.2.2. 
Adoptive Cell Transfer experience at the Surgery Branch, NCI 
The NCI-SB has pioneered novel T cell based cancer therapies for chemotherapy-
refractory cancers and continues efforts to expand their application. This work has its 
foundation in the successful treatment of metastatic melanoma with adoptive transfer of 
tumor infiltrating lymphocytes (TIL).  We have reported the results of adoptive transfer 
therapy in 93 patients with metastatic melanoma who received TIL following a 
lymphodepleting regimen plus aldesleukin administration, with or without total body 
irradiation (Figure 1)12. Forty-three patients received a non-myeloablative chemotherapy 
consisting of 60 mg/kg cyclophosphamide q daily x 2 and 25mg/m2 fludarabine q daily x 
5 prior to cell transfer and aldesleukin administration. Twenty-five patients each also 
received the same chemotherapy agents in conjunction with either 200 or 1200 cGy total 
body irradiation (TBI) prior to cell infusion and aldesleukin administration. The overall 
objective response rate using RECIST criteria in these 93 patients was 56%. The clinical 
results in these three trials are shown in Table 2, and the toxicities shown in Table 3 for 
the TBI studies (04-C-0288, 06-C-0136) and Table 4 for the initial study without TBI 
(99-C-0158). There was one treatment related death in these 93 patients which occurred 
in a patient who had an undetected diverticular abscess prior to beginning therapy. Of the 
52 responding patients in this trial, 42 had disease that was refractory to aldesleukin 
therapy and 22 had disease that was refractory to prior aldesleukin plus chemotherapy. 
Thus TIL therapy shows promise as an effective treatment for chemotherapy refractory 
metastatic melanoma. 
 
HPV-associated cancers are rational targets for adoptive immunotherapy with TIL 
because i) they express E6 and E7 oncoproteins which are non-self antigens and therefore 
good immunologic targets, ii) E6 and E7 are not only specific to tumor cells but 
important to their survival, iii) better clinical cancer outcomes correlate with T cell 
reactivity against E6 and E7, iv) polyclonal responses against E6 and E7 have been 
identified in TIL, and v) E6 and E7 have been successfully targeted by vaccine therapy in 
premalignant neoplasia. E6 and E7 are small (18 and 13 kDa respectively) oncoproteins 
that cooperate to induce malignant transformation of cells infected with high-risk HPV 


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8
types. These proteins are particularly attractive therapeutic targets because they continue 
to be expressed after transformation and their expression is required for survival of the 
transformed cells. The importance of ongoing E6 and E7 expression for cell survival has 
been demonstrated by studies showing that RNA silencing or expression of HPV E2 
(which suppresses E6 and E7 transcription) causes cellular senescence of HPV-positive 
tumor lines13-16. Expression of E7 is routinely detected by immunohistochemistry for p16 
(CDKN2A, p16Ink4A), which is increased by E7-mediated induction of KDM6B, a 
histone 3 lysine 27-specific demethylase17.The clinical importance of immune responses 
against E6 and E7 is suggested by studies linking immune responses against E6 and E7 to 
improved clinical outcomes 18-23 (Table 5). A number of studies indicated that E6 and E7 
could be targeted with TIL therapy 24,25. In one study, reactivity against the E6 and E7 
antigens could be detected in 23 of 54 TIL and tumor draining lymph node specimens 
from HPV16- and HPV18-positive cervical neoplastic lesions (Table 6)25. TIL could be 
generated from both adenocarcinomas and squamous cell carcionomas, and TIL cultures 
grew to a mean of 68 percent CD3-positive cells at three weeks, with a mix of CD4- and 
CD8-positive cells present (CD3+CD4+ 38% +/- 21 %, CD3+CD8+ 48% +/- 24%). 
Reactivity against HPV was determined by examining T cell proliferation in response to 
autologous monocytes pulsed with full length proteins as well as pools of long 
overlapping peptides spanning E6 and E7 of HPV16 and HPV18. In TIL samples from 19 
of 51 patients HPV E6- and/or E7-specific T cells were detected. The HPV reactive cells 
were both CD4- and CD8-positive and in several cases responded to more than one E6 or 
E7 peptide, a finding consistent with other work showing a polyclonal TIL response 
against multiple epitopes24. MHC restriction analysis revealed frequent use of DP and 
DQ HLA molecules (13 of the 16 class II restricted TIL), although DR and class I 
restricted responses were also present. Thus, TIL from cervical tumors display diverse 
polyclonal reactivity against E6 and E7. While TIL from HPV associated cancers have 
not been tested in clinical trials, E6 and E7 have been targeted with therapeutic vaccines. 
In a trial of long-peptide vaccination for vulvar intraepithelial neoplasia (VIN)26, an 
HPV-induced premalignant condition, 15 of 19 patients had clinical responses to vaccine 
therapy, and 9 of 19 had complete responses all of which were maintained at 24 months 
of follow up. Therapeutic vaccines for advanced HPV-associated cancers, as for 
melanoma, have been universally unsuccessful27. However, TIL therapy has been highly 
successful in treating melanoma, and the existing data indicate that HPV-associated 
cancers may also be excellent targets for TIL therapy. 
In the Surgery Branch, work is underway to extend TIL therapy to non-melanoma 
cancers. In an ongoing protocol we are targeting digestive tract tumors with young TIL. 
We have treated five patients thus far; two patients were taken off study due to 
progressive disease, one patient was treated too recently to evaluate response, and two 
patients died on study and were not evaluable for clinical response. Most of the toxicities 
observed in this study were expected toxicities of the non-myeloablative chemotherapy 
and the IL-2. TIL was grown successfully in 14 of 15 attempts from resected GI tumors 
(Table 7), demonstrating our ability to generate reliably a cell product from a variety of 
cancers. We have also successfully generated TIL from five of five cervical cancers 
(Table 8). These TIL cultures displayed high frequencies of CD3-positive cells, and they 
varied in CD4 and CD8 composition (Table 8). Both CD4- and CD8-predominant 
fragments displayed excellent growth in rapid expansion protocol (REP) expansions 


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(Table 8). Testing for HPV-specific reactivity was limited by lack of samples of 
peripheral blood mononuclear cells for use in recognition assays; however autologous 
tumor recognition was present in one of two samples that could be tested. As additional 
tissue becomes available through this protocol, we hope to develop tests for E6 and E7 
reactivity. Such testing could be valuable in selecting the optimal TIL to expand for 
therapy. In conclusion, HPV-associated tumors are immunogenic and give rise to TIL 
targeting viral oncoproteins that are critical to malignant cells and that have been 
successfully targeted with cancer vaccine therapy. They are therefore promising targets 
for TIL therapy and a clinical trial of adoptive immunotherapy with TIL is warranted. 
Studies of the first nine patients treated in the cervical cancer cohort of this protocol 
revealed a statistically significant positive association between the HPV reactivity of the 
infusion product and response to therapy. The three patients who received cells with the 
greatest HPV reactivity experienced clinical responses. Five patients with low or 
undetectable reactivity did not respond to treatment. We therefore are introducing 
Amendment I to the protocol, which will require testing for the presence of HPV E6 or 
E7 reactivity in the cell product after the initial TIL outgrowth. If reactivity is low or 
undetectable, a final cell product will not be generated and patients will not be treated. 
This change is intended to spare patients with a low probability of benefiting from the 
treatment the toxicities of the therapy. 
2. 
ELIGIBILITY ASSESSMENT AND ENROLLMENT  
2.1 ELIGIBILITY CRITERIA 
2.1.1 Inclusion Criteria 
a. Measurable metastatic or locally advanced refractory/recurrent malignancies that are 
HPV-16 or HPV-18 HPV positive by in situ hybridization (ISH) or polymerase chain 
reaction (PCR) or any cancer from the uterine cervix.. 
b. All patients must have received at least one standard chemotherapy or 
chemoradiotherapy. 
c. Patients with 3 or fewer brain metastases that are less than 1 cm in diameter and 
asymptomatic are eligible.  Lesions that have been treated with stereotactic 
radiosurgery must be clinically stable for 1 month after treatment for the patient to be 
eligible. 
d. Greater than or equal to 18 years of age and less than or equal to age 70. 
e. Able to understand and sign the Informed Consent Document 
f. Clinical performance status of ECOG 0 or 1. 
g. Life expectancy of greater than three months 
h. Patients of both genders must be willing to practice birth control from the time of 
enrollment on this study and for up to four months after receiving treatment. 
i. Serology:  
• Seronegative for HIV antibody.  (The experimental treatment being evaluated 
in this protocol depends on an intact immune system.  Patients who are HIV 
seropositive can have decreased immune-competence and thus be less 
responsive to the experimental treatment and more susceptible to its 
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• Seronegative for hepatitis B antigen, and seronegative for hepatitis C 
antibody. If hepatitis C antibody test is positive, then patient must be tested 
for the presence of antigen by RT-PCR and be HCV RNA negative.  
j. Women of child bearing potential must have a negative pregnancy test because of the 
potentially dangerous effects of the treatment on the fetus. 
k. Hematology 
• Absolute neutrophil count greater than 1000/mm3 without the support of 
filgrastim 
• WBC ≥ 3000/mm3 
• Platelet count ≥ 100,000/mm3 
• Hemoglobin > 8.0 g/dl 
l. Chemistry: 
• Serum ALT/AST ≤ to 2.5 times the upper limit of normal 
• Serum creatinine ≤ to 1.6 mg/dl 
• Total bilirubin ≤ to 1.5 mg/dl, except in patients with Gilbert’s Syndrome 
who must have a total bilirubin less than 3.0 mg/dl. 
m. More than four weeks must have elapsed since any prior systemic therapy at the time 
the patient receives the preparative regimen, and patients’ toxicities must have 
recovered to a grade 1 or less (except for toxicities such as alopecia or vitiligo). 
Note:  Patients may have undergone minor surgical procedures within the past 3 
weeks, as long as all toxicities have recovered to grade 1 or less or as specified in the 
eligibility criteria in Section 2.1.1. 
n. More than four weeks must have elapsed since any prior radiation therapy. 
 
2.1.2 Exclusion Criteria 
a. Women of child-bearing potential who are pregnant or breastfeeding because of the 
potentially dangerous effects of the treatment on the fetus or infant. 
b. Active systemic infections, coagulation disorders or other active major medical 
illnesses of the cardiovascular, respiratory or immune system, as evidenced by a 
positive stress thallium or comparable test, myocardial infarction, cardiac 
arrhythmias, obstructive or restrictive pulmonary disease.  
c. Any form of primary immunodeficiency (such as Severe Combined 
Immunodeficiency Disease). 
d. Concurrent opportunistic infections (The experimental treatment being evaluated in 
this protocol depends on an intact immune system.  Patients who have decreased 
immune competence may be less responsive to the experimental treatment and more 
susceptible to its toxicities). 
e. Concurrent systemic steroid therapy. 
f. History of severe immediate hypersensitivity reaction to any of the agents used in this 
study. 
g. History of coronary revascularization or ischemic symptoms. 
h. Any patient known to have an LVEF less than or equal to 45%. 
i. Documented LVEF of less than or equal to 45% tested in patients with i) clinically 
significant atrial and/or ventricular arrhythmias including but not limited to: atrial 


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fibrillation, ventricular tachycardia, second or third degree heart block or ii) age ≥ 60 
years old. 
j. Active Bleeding  
2.2.SCREENING EVALUATION 
2.2.1. Within 4 weeks prior to starting the chemotherapy regimen:  
a. Complete history and physical examination, including weight, and vital signs noting 
in detail the exact size and location of any lesions that exist.  (Note: patient history 
may be obtained within 8 weeks.)  
b. Chest x-ray 
c. EKG  
d. Baseline CT of the chest, abdomen and pelvis, and brain MRI to evaluate the 
status of disease. Additional scans and x-rays may be performed if clinically 
indicated based on patients’ signs and symptoms. (Note: brain MRI may be 
obtained within 8 weeks prior to starting the chemotherapy regimen.)  
e. Pulmonary Function testing for patients with a prolonged history of cigarette 
smoking (20 pk/year of smoking within the past 2 years) or symptoms of 
respiratory dysfunction. (Note: may be performed within 8 weeks of treatment).   
f. Cardiac Evaluation (stress thallium, echocardiogram, MUGA etc.) for patients 
who are greater than or equal to age 60, or have a history of ischemic heart 
disease, chest pain, or clinically significant atrial and/or ventricular arrhythmias 
including but not limited to: atrial fibrillation, ventricular tachycardia, or heart 
block.  Patients with a LVEF of less than or equal to 45% will not be eligible. 
Patients under the age of 60 who have cardiac risk factors may also undergo 
cardiac evaluation as noted above (e.g. diabetes, hypertension, obesity). (Note: 
may be performed within 8 weeks of treatment).    
g. HIV antibody titer and HbsAG determination, anti HCV, (Note: may be performed 
within 3 months of chemotherapy start date). 
h. Anti CMV antibody titer, HSV serology, and EBV panel (Note: patients who are 
known to be positive for any of the above do not need to be retested; may be 
performed within 3 months of chemotherapy start date.) 
i. Medical history (may be conducted at any point prior to this time). 
j. Verification that HLA typing is completed or in process. Testing may be conducted at 
any time prior to this point. 
 
2.2.2. Within 14 days prior to starting the chemotherapy regimen: 
a. Baseline blood tests  
• Chem 20 equivalent: (Sodium (Na), Potassium (K), Chloride (Cl), Total CO2 
(bicarbonate), Creatinine, Glucose, Urea nitrogen (BUN), Albumin, Calcium 
total, Magnesium total (Mg), Inorganic Phosphorus, Alkaline Phosphatase, 
ALT/GPT, AST/GOT, Total Bilirubin, Direct Bilirubin, LD, Total Protein, Total 
CK, Uric Acid) 
• Thyroid panel  
• CBC with differential and platelet count 


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• PT/PTT 
b. Urinalysis and culture, if indicated  
 
2.2.3.  Within 7 days prior to starting the chemotherapy regimen: 
a. β-HCG pregnancy test (serum or urine) on all women of child-bearing potential 
b. ECOG performance status of 0 or 1 
c. Calculated creatinine clearance for patients with renal impairment  
Please note: Patients with hydronephrosis or other renal impairment may be required 
to undergo additional testing in order to ensure adequate renal function, (e.g. renal 
ultrasound, renal scan). 
 
2.3.REGISTRATION PROCEDURES 
Patients will be registered on protocol 03-C-0277 (Cell Harvest and Preparation for Surgery 
Branch Adoptive Cell Therapy Protocols) prior to tumor resection for young TIL generation, 
by the clinical fellow or research nurse within 24 hours of the patient signing the consent by 
faxing a completed Eligibility Checklist to the Central Registration Office (CRO) at 301-480-
0757. Once cells exceed the potency requirement and are projected to exceed the minimum 
number specified in the Certificate of Analysis (COA), patients will sign the consent document 
for this protocol. Authorized staff must register an eligible candidate with NCI Central 
Registration Office (CRO) within 24 hours of signing consent.  A registration Eligibility 
Checklist from the web site (http://home.ccr.cancer.gov/intra/eligibility/welcome.htm) must be 
completed and faxed to 301-480-0757.  After confirmation of eligibility at Central Registration 
Office, CRO staff will call pharmacy to advise them of the acceptance of the patient on the 
protocol.  Verification of Registration will be forwarded electronically via e-mail to the 
research team. A recorder is available during non-working hours.  Patients will be stratified 
according to whether they have cervical or non-cervical cancer. 
 
3. 
STUDY IMPLEMENTATION 
3.1 STUDY DESIGN 
The study will be conducted using a Phase II design. Young TIL will be generated from tumor 
fragments and/or digests according to standard operating procedure. Assays for identification 
of E6- and E7-reactive cells will be developed and used to preferentially select wells of TIL for 
rapid expansion and administration. Patients whose TIL cultures display low or undetectable 
HPV E6 and E7 reactivity after initial outgrowth will not be treated.  Patients will receive a 
non-myeloablative, lymphodepleting preparative regimen consisting of cyclophosphamide and 
fludarabine that has been used in prior Surgery Branch trials. The conditioning regimen will be 
followed by intravenous infusion of Young TIL plus aldesleukin.   
3.1.1 Cell Preparation: 
Patients with evaluable metastatic or locally advanced refractory/recurrent HPV-associated 
malignancies who have lesions that can be resected with minimum morbidity will undergo 
resection of tumor.  TIL will be obtained while enrolled on the Surgery Branch protocol 03-C-


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0277, “Cell Harvest and Preparation for Surgery Branch Adoptive Cell Therapy Protocols”.  
Separate tumor biopsies may be performed under protocol 03-C-0277 to obtain TIL if initial 
tumor biopsy could not successfully generate TIL. Young TIL will be grown and expanded for 
this trial according to standard operating procedures submitted in the IND.  After initial 
outgrowth, lymphocyte cultures will be tested for reactivity against HPV E6 and E7 as 
described in TIL Laboratory SOP 4.4, “Screening tumor infiltrating lymphocytes (TIL) from 
patients enrolled on the Human Papillomavirus (HPV) TIL clinical protocol (12-C-0116) for 
reactivity against HPV antigens”.  If three or more cultures demonstrate HPV E6 or E7 
reactivity then production of the cell product will proceed. If two or fewer cultures show HPV 
E6 or E7 reactivity then production of the cell product will be terminated and the patient will 
not be treated.  A positive test for HPV reactivity will be defined as interferon-γ production 
greater than or equal to twice the negative control (gp100) and greater than or equal to 200 
pg/mL.  Young TIL will be assessed for potency by interferon-gamma release as specified in 
the Certificate of Analysis shown in Appendix 4.  Once cells exceed the potency requirement 
and are projected to exceed the minimum number specified in the COA, the patient will be 
registered on this study and receive the lymphocyte depleting preparative regimen consisting of 
fludarabine and cyclophosphamide, followed by infusion of cells and administration of high-
dose aldesleukin.  It is anticipated that Young TIL that meet the COA will not be achievable in 
approximately 20% of patients who undergo resection.  These patients may undergo a second 
resection to grow Young TIL, if another suitable lesion exists.  Cells will be administered at a 
dose of between 1x109 to 2x1011 lymphocytes over 20-30 minutes. All patients will receive one 
course of treatment.  The start date of the course will be the start date of the chemotherapy; the 
end date will be the day of the first post-treatment evaluation. Patients may undergo a second 
treatment as described in Section 3.4. 
 
3.1.2 Protocol Stopping Rules: 
The study will be temporarily halted pending discussions with the FDA and NCI IRB 
regarding safety and the need for protocol revisions if any of the following conditions are 
met: 
• Two or more patients develop a grade 4 or greater toxicity at any point in the study 
only attributable to the cell infusion. 
• If one of the first three patients (or 2 of the first 6 patients, or 3 of the first 9 patients, 
or 4 of the first 12 patients) develop grade 3 autoimmunity, that cannot be resolved to 
less than or equal to a grade 2 autoimmune toxicity within 10 days, or any grade 4 or 
greater autoimmune toxicity. 
• If one or more treatment related deaths occur due to the cell infusion, we will promptly 
discuss this with the NCI IRB and FDA. 
3.2 DRUG ADMINISTRATION 
3.2.1 Preparative Regimen with Cyclophosphamide and Fludarabine: 
(Times are offered as examples and may be changed as long as a similar time 
relationship between administration of the drugs is maintained. Study medication start 
times for drugs given once daily should be given within 2 hours of the scheduled time. All 
other medications should be given +/- one hour of the scheduled time; the length of 


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administration is all +/- 15 minutes. Chemotherapy infusions maybe slowed or delayed 
as medically indicated.  Administration of diuretics, electrolyte replacement, and 
hydration and monitoring of electrolytes should all be performed as clinically indicated – 
the times noted below are offered only as examples.)  
 
DAYS  -7 and -6 
6 AM 
Hydrate: Begin hydration with 0.9% Sodium Chloride Injection containing 10 meq/L of 
potassium chloride at 2.6 ml/kg/hr (starting 11 hours pre-cyclophosphamide and continue 
hydration until 24 hours after last cyclophosphamide infusion). At any time during the 
preparative regimen, if urine output <1.5 ml/kg/hr or if body weight >2 kg over pre-
cyclophosphamide value, furosemide 10-20 mg IV maybe administered. Serum 
potassium should be monitored and treated as indicated following administration of 
furosemide. 
4 PM 
Ondansetron (0.15 mg/kg/dose [rounded to the nearest even mg dose between 8 mg and 
16 mg based on patient weight] IV every 8 hours X 3 days) will be given for nausea.  
5 PM 
Cyclophosphamide 60 mg/kg/day X 2 days IV in 250 ml D5W with Mesna 15 mg/kg/day 
X 2 days over 1 hr. If patient is obese (BMI > 35) drug dosage will be calculated using 
practical weight as described in Table 9.  
6 PM 
Begin mesna infusion at 3 mg/kg/hour intravenously diluted in a suitable diluent (see 
pharmaceutical section) over 23 hours after each cyclophosphamide dose. If patient is 
obese (BMI > 35) drug dosage will be calculated using practical weight as described in 
Table 9.  
DAYS -5 to -1 
Fludarabine 25 mg/m2/day IVPB daily over 30 minutes for 5 days. If patient is obese 
(BMI > 35) drug dosage will be calculated using practical weight as described in Table 9.  
 
3.2.2 Cell Infusion and Aldesleukin Administration: 
The patient’s Young TIL are delivered to the patient care unit by a staff member from the 
Tumor Immunology Cell Processing Laboratory.  Cells will be administered at a dose of 
between 1x109 to 2x1011 lymphocytes.  Prior to infusion, the cell product identity label is 
double-checked by two authorized staff (MD or RN), an identification of the product and 
documentation of administration are entered in the patient’s chart, as is done for blood 
banking protocols.  The cells are to be infused intravenously over 20-30 minutes via non-
filtered tubing, gently agitating the bag during infusion to prevent cell clumping. 
DAY 0 (one to four days after the last dose of fludarabine): 
 


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• Cells will be infused intravenously (i.v.) on the Patient Care Unit over 20 to 30 
minutes (between one and four days after the last dose of fludarabine). 
• Aldeskeukin will be given as described in section 3.2.3. 
DAY 1-4 (Day 0 is the day of cell infusion):  
• Beginning on day 1 or 2, filgrastim may be administered subcutaneously at a dose of 
5 mcg/kg/day (not to exceed 300 mcg/day).  Filgrastim administration will continue 
daily until neutrophil count > 1.0 x109/L X 3 days or > 5.0 x109/L.  
• Aldesleukin will be given as described in section 3.2.3. 
 
Table 3.2 
1One to four days after the last dose of fludarabine 
2Initiate within 24 hours after cell infusion 
3Continue until neutrophils count > 1X109/L for 3 consecutive days or > 5x109/L. 
4The TMP/SMX schedule should be adjusted to QD three times per week (Monday, Wednesday, Friday) and 
continue for at least six months and until CD4 > 200 X 2 
5Continue until ANC > 1000/mm3 
6In patients positive for HSV continue until CD4 > 200 X 2 
 
3.2.3 Aldesleukin Administration 
Aldesleukin will be administered at a dose of 720,000 IU/kg (based on total body weight) 
as an intravenous bolus over a 15 minute period within 24 hours of cell infusion and 
continuing for up to 5 days (maximum 15 doses).  Doses will be preferentially 
administered every eight hours; however, up to 24 hours may elapse between doses 
depending on patient tolerance.  Aldesleukin dosing will be stopped if toxicities are not 
sufficiently recovered by supportive measures within 24 hours of the last dose of 
aldesleukin. Dosing will be delayed or stopped if patients reach Grade 3 or 4 toxicity due 
to aldesleukin, except for the reversible Grade 3 toxicities common to aldesleukin such as 
diarrhea, nausea, vomiting, hypotension, skin changes, anorexia, mucositis, dysphagia, or 
constitutional symptoms and laboratory changes as detailed in Appendix 1.  Toxicities 
will be managed as outlined in Appendix 2.  Dosing may be held or stopped at the 
discretion of the treating investigator (Appendix 3 lists the toxicities seen in patients 
treated with aldesleukin at the NIH Clinical Center).  
Day 
-7 -6 
-5 
-4 
-3 
-2 
-1 
01 
1 
2 
3 
4 
Therapy 
 
 
 
 
 
 
 
 
 
 
 
 
Cyclophosphamide (60 mg/kg) 
X 
X 
 
 
 
 
 
 
 
 
 
 
Fludarabine (25 mg/m2) 
 
 
X 
X 
X 
X 
X 
 
 
 
 
 
Young TIL 
 
 
 
 
 
 
 
X1 
 
 
 
 
Aldesleukin 
 
 
 
 
 
 
 
X2 
X 
X 
X 
X 
Filgrastim3  (5 mcg/kg/day) 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
TMP/SMX4 
160mg/800 mg (example) 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Fluconazole5 (400 mg po) 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
Valacyclovir po or Acyclovir IV6 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 


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Because confusion is a possible side effect of aldesleukin administration, a Durable 
Power of Attorney will be signed by the patient to identify a surrogate to make decisions 
if a patient becomes unable to make decisions. 
3.3 ON-STUDY EVALUATIONS: 
3.3.1 Prior to starting the preparative regimen 
• Apheresis as indicated 
• Within 14 days prior to starting the preparative regimen, patients will have a 
complete blood count, serum chemistries performed including electrolytes, BUN, 
creatinine, and liver function tests.  If any results are beyond the criteria established 
for eligibility, the patient will not proceed until the abnormalities can be resolved.  
3.3.2 During the preparative regimen: DAILY 
• Complete Blood Count 
• Chem 20 equivalent: Sodium (Na), Potassium (K), Chloride (Cl), Total CO2 
(bicarbonate), Creatinine, Glucose, Urea nitrogen (BUN), Albumin, Calcium total, 
Magnesium total (Mg), Inorganic Phosphorus, Alkaline Phosphatase, ALT/GPT, 
AST/GOT, Total Bilirubin, Direct Bilirubin, LD, Total Protein, Total CK, Uric Acid 
• Urinalysis. 
3.3.3 After Cell Infusion: 
• Vital signs will be monitored hourly (+/- 15 minutes) for four hours and then 
routinely (every 4-6 hours) unless otherwise clinically indicated. 
• Once total lymphocyte count is greater than 200/mm3, TBNK for peripheral blood 
CD4 count will be drawn weekly (while the patient is hospitalized).  Please refer to 
section 5 for additional post cell infusion evaluations. 
3.3.4 During Hospitalization:  
Every 1-2 days 
• A review of systems and physical exam 
• CBC 
• Chem 20 equivalent: Sodium (Na), Potassium (K), Chloride (Cl), Total CO2 
(bicarbonate), Creatinine, Glucose, Urea nitrogen (BUN), Albumin, Calcium total, 
Magnesium total (Mg), Inorganic Phosphorus, Alkaline Phosphatase, ALT/GPT, 
AST/GOT, Total Bilirubin, Direct Bilirubin, LD, Total Protein, Total CK, Uric Acid 
• Other tests will be performed as clinically indicated. 
 
3.4 RETREATMENT  
Patients experiencing a partial or complete response may receive a second treatment 
when progression by RECIST criteria is documented after evaluation by the principal 
investigator.  Retreatment will consist of the same regimen that they had been given 
safely previously.  Patients who develop grade 3 or grade 4 toxicity due to cell infusion 


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will not be retreated. Patients must continue to meet the original eligibility criteria to be 
considered for retreatment.  Toxicity related to cyclophosphamide, fludarabine, or 
aldeskeukin should be stable and resolved to less than grade 1 prior to retreatment.  
Retreatment benefits and risks will be carefully explained to the patient.  A maximum of 
1 retreatment course may occur.  
3.5 POST STUDY EVALUATION (FOLLOW-UP) 
3.5.1 All patients will return to the NIH Clinical Center for evaluation 6 weeks (+/- 2 
weeks) following the administration of the cell product.  Please note: Patients 
discharged with grade 3 or greater significant adverse events should be evaluated 
by referring physician within 2 weeks of discharge. 
3.5.2 Patients who experience stable disease, a partial response, or a complete response 
or have unresolved toxicities will be evaluated as noted below:  
• Every month (+/- 2 weeks) x3 
• Every 3 months (+/- 1 month) x3  
• Every 6 months (+/- 1 month) x 2  
• As per PI discretion for subsequent years  
Note: Patients may be seen more frequently as clinically indicated 
3.5.3 At each evaluation patients will undergo:  
• Physical examination  
• Chem 20 equivalent: Sodium (Na), Potassium (K), Chloride (Cl), Total CO2 
(bicarbonate), Creatinine, Glucose, Urea nitrogen (BUN), Albumin, Calcium 
total, Magnesium total (Mg), Inorganic Phosphorus, Alkaline Phosphatase, 
ALT/GPT, AST/GOT, Total Bilirubin, Direct Bilirubin, LD, Total Protein, 
Total CK, Uric Acid  
• Complete blood count  
• Thyroid panel as clinically indicated  
• TBNK, until CD4 > 200 X 2 
• Toxicity assessment, including a review of systems.  
• CT of the chest, abdomen and pelvis. This end of course evaluation will be 
used to determine tumor response. If clinically indicated, other scans or x-rays 
may be performed, e.g. brain MRI, bone scan.  
• Visual symptoms will be evaluated and if changes have occurred from 
baseline, i.e. changes in visual acuity, an ophthalmologic consult will be 
performed.  
• A 5 liter apheresis may be performed.  If the patient is unable to undergo 
aphersis, approximately 96 ml of blood may be obtained at the first follow up 
visit. Subsequently, approximately 60 ml of blood will be obtained at follow 
up visits (approximately monthly) for at least 3 months. Peripheral blood 
mononuclear cells will be cryopreserved so that immunologic testing may be 
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3.5.4 Patients who are unable or unwilling to return for follow up evaluations will be 
followed via phone or email contacts. Patients may be asked to send laboratory, 
imaging and physician exam reports performed by their treating physician.  
3.6 CRITERIA FOR REMOVAL FROM PROTOCOL THERAPY AND OFF STUDY CRITERIA 
3.6.1 OFF TREATMENT CRITERIA 
Patients will be taken off treatment (and followed until progression of disease) for the 
following: 
• Grade 3 autoimmunity that involves vital organs (heart, kidneys, brain, eye, liver, 
colon, adrenal gland, lungs). 
• If a patient experiences a grade 3 or 4 toxicity due to cell infusion (reaction to 
cellular product or infusion reaction) the patient will receive no further treatment. 
• Completion of treatment period 
3.6.2 OFF STUDY CRITERIA 
Patients will be taken off study for the following: 
• The patient voluntarily withdraws 
• There is significant patient noncompliance 
• Radiographic or clinical disease progression, unless the patient is eligible for 
second treatment.  
• General or specific changes in the patient’s condition render the patient 
unacceptable for further treatment on this study in the judgment of the investigator. 
• Death 
Note:  patients who are taken off study for progressive disease or study closure maybe followed 
on Protocol 09-C-0161 Follow up Protocol for Subjects Previously Enrolled in NCI Surgery 
Branch Studies.  
Note:  Patients must be followed until all adverse events have resolved to grade 2 or less with the 
exception of lymphopenia and alopecia.  If an adverse event is not expected to resolve to grade 2 
or less this will be noted in the patient medical record and the patient may be taken off study.  
4. CONCOMITANT MEDICATIONS/MEASURES 
4.1 
INFECTION PROPHYLAXIS: 
Note:  Other anti-infective agents may be substituted at the discretion of the treating physician. 
 
4.1.1 
Pneumocystis Jirovecii Pneumonia 
All patients will receive the fixed combination of trimethoprim and sulfamethoxazole 
(TMP/SMX) as double strength (DS) tab (DS tabs = TMP 160 mg/tab, and SMX 800 
mg/tab) P.O. daily three times a week on non-consecutive days, beginning between days -
5 and -8.  
 


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Pentamidine will be substituted for TMP/SMX-DS in patients with sulfa allergies. It will 
be administered aerosolized at 300 mg per nebulizer within one week of chemotherapy 
start date and monthly thereafter. 
 
 
4.1.2 
Herpes Virus Prophylaxis 
Patients with positive HSV serology will be given valacyclovir orally at a dose of 500 mg 
daily the day after chemotherapy ends, or acyclovir, 250 mg/m2 IV every 12 hrs if the 
patient is not able to take medication by mouth.  Reversible renal insufficiency has been 
reported with IV but not oral acyclovir.  Neurologic toxicity including delirium, tremors, 
coma, acute psychiatric disturbances, and abnormal EEGs have been reported with higher 
doses of acyclovir.  Should this occur, a dosage adjustment will be made or the drug will 
be discontinued.  Acyclovir will not be used concomitantly with other nucleoside analogs 
which interfere with DNA synthesis, e.g. ganciclovir.  In renal disease, the dose is 
adjusted as per product labeling. 
 
Prophylaxis for Pneumocystitis and Herpes will continue for 6 months post 
chemotherapy. If the CD4 count is less that 200 at 6 months post chemotherapy, 
prophylaxis will continue until the CD4 count is greater than 200 x2. 
 
4.1.3 Fungal Prophylaxis 
Patients will start Fluconazole 400 mg p.o. the day after chemotherapy concludes and 
continue until the absolute neutrophil count is greater than 1000/mm3.  The drug may be 
given IV at a dose of 400 mg in 0.9% sodium chloride USP daily in patients unable to 
take it orally. 
4.2 BLOOD PRODUCT SUPPORT 
Using daily CBC’s as a guide, the patient will receive platelets and packed red blood cells 
(PRBC’s) as needed.  Attempts will be made to keep Hb >8.0 gm/dl, and plts >10,000/mm3.  
All blood products will be irradiated.  Leukocyte filters will be utilized for all blood and 
platelet transfusions to decrease sensitization to transfused WBC’s and decrease the risk of 
CMV infection. Empiric antibiotics 
4.3 EMPIRIC ANTIBIOTICS 
Patients will start on broad-spectrum antibiotics, either a 3rd or 4th generation cephalosporin or 
a quinolone for fever of 38.3°C once or two temperatures of 38.0°C or above at least one hour 
apart, AND an ANC <500/mm3. Infectious disease consultation will be obtained for all 
patients with unexplained fever or any infectious complications. 
 
4.4 OTHER CONCOMITANT MEDICATIONS TO CONTROL SIDE EFFECTS 
Concomitant medications to control side effects of therapy may be given.  Meperidine (25-50 
mg) will be given intravenously if severe chilling develops.  Other supportive therapy will be 
given as required and may include acetaminophen (650 mg q4h), indomethacin (50-75 mg q6h) 
and ranitidine (150 mg g12h). If patients require steroid therapy they will be taken off 
treatment.  Patients who require transfusions will receive irradiated blood products.  
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Additional antiemetics will be administered as needed for nausea and vomiting uncontrolled by 
ondansetron.  Antibiotic coverage for central venous catheters may be provided at the 
discretion of the investigator. 
5. 
BIOSPECIMEN COLLECTION 
5.1 CORRELATIVE STUDIES FOR RESEARCH 
The amount of blood that may be drawn from adult patients for research purposes shall not 
exceed 10.5 mL/kg or 550 mL, whichever is smaller, over any eight week period.  
5.1.1 Prior to Chemotherapy Administration: 
• 5 CPT tubes (8ml each) 
• 1 SST tube (8ml) 
• 1 SST tube (4ml) daily; starting day of chemotherapy 
 
5.1.2 Prior To Cell Infusion: 
• Blood samples for cytokine analysis (1-8ml tube) 
5.1.3 Post Cell Infusion Evaluations: 
• 
Once total lymphocyte count is greater than 200/mm3, the following samples will 
be drawn and sent to the TIL lab on Monday, Wednesday and Friday x 5 days, 
then weekly (while the patient is hospitalized): 
o 
5 CPT tubes (8 ml each) 
o 
1 SST tube (8 ml) 
5.1.4 Immunological Testing: 
• 
Apheresis may be performed prior to and 4-6 weeks after the treatment.  At other 
time points, patient peripheral blood lymphocytes (PBL) will be obtained from 
whole blood by purification using centrifugation on a Ficoll cushion. Aliquots of 
these PBMC will be cryopreserved for immunological monitoring of cell 
function. 
• 
Lymphocytes will be tested directly and following in vitro culture using some or 
all of the following tests. Direct immunological monitoring will consist of 
quantifying T cells reactive with targets FACS analysis using tetramer staining.  
Ex vivo immunological assays will consist of cytokine release by bulk PBL (+/- 
peptide stimulation) and by other experimental studies such as cytolysis if 
sufficient cells are available. If cell numbers are limiting, preference will be given 
to the direct analysis of immunological activity.  Immunological assays will be 
standardized by the inclusion of 1) pre-infusion PBMC and 2) an aliquot of the 
TIL cryopreserved at the time of infusion. In general, differences of 2 to 3 fold in 
these assays are indicative of true biologic differences.  Foxp3 levels will be 
analyzed by semiquantitative RT-PCR to evaluate for mRNA on PBL samples 
obtained prior to cell infusion and at the follow up time point. 
• 
Blood and tissue specimens collected in the course of this research project may be 
banked and used in the future to investigate new scientific questions related to this 


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study, if the subject provides permission on the optional studies section of the 
consent document. However, this research may only be done if the risks of the 
new questions were covered in the consent document. If new risks are associated 
with the research (e.g., analysis of germ line genetic mutations) the principal 
investigator must amend the protocol and obtain informed consent from all 
research subjects.  Any new use of samples will require prospective IRB review 
and approval. 
• 
Blood and tissue specimens collected in the course of this research project may be 
banked and used in the future to investigate new scientific questions related to this 
study if the subject provides permission on the optional studies section of the 
consent document for protocol 03-C-0277. 
 
Note:  The collection and analysis of research labs will be monitored by the TIL lab and 
not by Harris Technical Services. 
5.2. SAMPLE STORAGE, TRACKING AND DISPOSITION 
Blood and tissue collected during the course of this study will follow the Cell Tracking and 
Labeling System established by the Tumor Immunology Cell Processing Laboratory.  The Cell 
Tracking and Labeling System is designed to unambiguously ensure that patient/data verification 
is consistent.  The patients' cell samples (blood or tissue) are tracked by distinct identification 
labels that include a unique patient identifier and date of specimen collection.  Cryopreserved 
blood and tissue samples also bear the date the sample was frozen.  All cryopreserved samples 
are tracked for freezer location and storage criteria.  All samples are stored in monitored 
freezers/refrigerators in 3NW Surgery Branch Laboratories at specified temperatures with alarm 
systems in place.  Serum samples will be sent to the Clinical Pharmacology Program (CPP) for 
storage.  Samples will be barcoded and stored on site or offsite at NCI Frederick Central 
Repository Services (Fisher Bioservices) in Frederick, MD.  Data is entered and stored securely 
in the Patient Sample Data Management System (PSDMS) utilized by the CPP, and data will be 
updated to the Surgery Branch central computer database weekly. All samples (blood or tissue) 
are entered into a central computer database with identification and storage location, and this 
database is backed up every night. 
At the conclusion of this protocol, if additional studies are to be performed on any samples 
obtained during the conduct of this trial, a Request to Conduct Research for Stored Human 
Samples Specimens, or Data Collected in a Terminated NCI-IRB Protocol will be submitted.  
Otherwise, specimens will be disposed of in accordance with the environmental protection laws, 
regulations, and guidelines of the Federal Government and the State of Maryland. 
Any loss or unintentional destruction of the samples will be reported to the IRB,  the NCI 
Clinical Director, and the office of the CCR, NCI. 


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6. 
DATA COLLECTION AND EVALUATION  
6.1 DATA COLLECTION 
The investigators will be responsible for the collection, maintenance, quality control of the study 
data.  Clinical data will be entered into the NCI CCR C3D database.   
End of study procedures: Data will be stored according to HHS, FDA regulations and NIH 
Intramural Records Retention Schedule as applicable.  
Loss or destruction of data: Should we become aware that a major breech in our plan to protect 
subject confidentiality and trial data has occurred, the IRB will be notified.  
6.2 ROUTINE ADVERSE EVENT REPORTING  
Following registration through 30 days after cell infusion, adverse events will be recorded in the 
source documents, reviewed by the designated data manager or research nurse or principal 
investigator and captured in the C3D database.  All events occurring during the treatment phase 
of the study will be followed until resolution to grade 2 or baseline. During the follow up period, 
only grade 3 and 4 and unexpected  grade 2 events that are related to the treatment will be 
captured in C3D. 
6.2.1 Exclusions to Routine Adverse Event Reporting: 
Patients will be receiving multiple agents which include commercially available agents 
(fludarabine, cyclophosphamide and supportive medications) in combination with the 
investigational agents.  Therefore,  Grade 2 adverse events ‘unrelated’ or ‘unlikely related’ to the 
investigational agent, and ‘possibly’, ‘probably’ or ‘definitely’ related to the commercially 
available agents as specified in the package inserts do not require reporting/recording.  In 
addition all grade 1 events and all expected grade 2 events unrelated to the cell product will not 
be reported/recorded. 
 
6.2.2 Reporting of laboratory events 
Laboratory results will be uploaded in C3D however only those events (including grade 3 and 4 
events) that support the diagnosis of a reportable adverse event or that reflect major organ 
function will be considered adverse events. For example grade 3 and 4, creatinine, liver function 
tests, hemoglobin, ANC, ALC, platelets, and lipase and amylase as indicated will be captured as 
adverse events; electrolytes, BUN, albumin, total protein, uric acid etc, and the remainder of the 
CBC differential will not be captured as adverse events.  
For reportable adverse events: the adverse event start date will be the date the event reaches a 
grade 3; the event will be considered resolved once it reaches grade 2. The highest grade the 
event reaches in that period will be considered the grade of the event. For hematological 
toxicities, the event will not be considered resolved until it reaches grade 2 without the support 
of transfusions or growth factors.  
 


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6.2.3 Reporting of non-laboratory events  
For reportable expected adverse events: the adverse event start date will be the date the event 
reaches a grade 3; the event will be considered resolved once it reaches grade 2. The highest 
grade the event reaches in that period will be considered the grade of the event.  
For unexpected adverse events, the adverse event start date will be the date the event reaches a 
grade 2; the event will be considered resolved once it reaches grade 1 or baseline. 
 
6.2.4 Reporting Infection 
• Febrile neutropenia will be captured as follows:  The start date will be the date the fever 
of 38.5 or greater was first recorded.  The end date will be the date the patient has been 
afebrile greater than 48 hours or the date the patient develops a clinically significant 
infection. 
• If a patient has a positive culture during the period of febrile neutropenia, the event will 
be captured as “infection with neutropenia” with the start date as the date the fever of 
38.5 was first recorded.   
• Infection will only be captured once in any given period regardless of the number of 
organisms cultured or sites involved. 
• Positive cultures seen on routine surveillance cultures with no clinical symptoms will not 
be captured as infections regardless of whether anti-infective agents are given. 
 
6.3 RESPONSE CRITERIA 
Clinical Response will be determined using the Response Evaluation Criteria in Solid 
Tumors (RECIST) guideline (version 1.0).   
6.3.1  Evaluation of target lesions1  
• 
Complete Response (CR):  Disappearance of all target lesions 
• 
Partial Response (PR):  At least a 30% decrease in the sum of the longest 
diameter (LD) of target lesions taking as reference the baseline sum LD. 
• 
Progression (PD):  At least a 20% increase in the sum of LD of target 
lesions taking as reference the smallest sum LD recorded since the 
treatment started or the appearance of one or more new lesions. 
• 
Stable Disease (SD): Neither sufficient shrinkage to qualify for PR nor 
sufficient increase to qualify for PD taking as references the smallest sum 
LD. 
6.3.2  
Evaluation of non-target lesions2  
• 
Complete Response (CR): Disappearance of all non-target lesions and 
normalization of tumor marker level. 
• 
Non-Complete Response:  Persistence of one or more non-target lesions 
• 
Progression (PD):  Appearance of one or more new lesions.  Unequivocal 
progression of existing non-target lesions 
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1 
All measurable lesions up to a maximum of 10 lesions representative of all involved organs should be 
identified as target lesions and recorded and measured at baseline.  Target lesions should be 
selected on the basis of their size (lesions with the longest diameter) and their suitability for accurate 
repetitive measurements (either by imaging techniques or clinically).  A sum of the longest diameter 
(LD) for all target lesions will be calculated and reported as the baseline sum LD.  The baseline sum 
LD will be used as reference to further characterize the objective tumor response of the measurable 
dimension of the disease. 
2 
All other lesions (or sites of disease) should be identified as non-target lesions and should also be 
recorded at baseline.  Measurements are not required, and these lesions should be followed as 
“present” or “absent.” 
6.3.3  
Evaluation of best overall response 
The best overall response is the best response recorded from the start of the treatment 
until disease progression/recurrence (taking as reference for progressive disease the 
smallest measurements recorded since the treatment started).  The patient's best response 
assignment will depend on the achievement of both measurement and confirmation 
criteria. 
 
Target Lesions 
Non-Target Lesions 
New Lesions 
Overall Response 
CR 
CR 
No 
CR 
CR 
Non-CR/Non-PD 
No 
PR 
PR 
Non-PD 
No 
PR 
SD 
Non-PD 
No 
SD 
PD 
Any 
Yes or No 
PD 
Any 
PD 
Yes or No 
PD 
Any 
Any 
Yes 
PD 
 
6.3.4 Confirmatory Measurement/Duration of Response 
6.3.4.1 Confirmation 
To be assigned a status of PR or CR, changes in tumor measurements must be confirmed 
by repeat studies that should be performed at least 4 weeks after the criteria for response 
are first met. In the case of SD, follow-up measurements must have met the SD criteria at 
least once after study entry at a minimum interval of 6-8 weeks. 
6.3.4.2 Duration of Overall Response 
The duration of overall response is measured from the time measurement criteria are met 
for CR/PR (whichever is first recorded) until the first date that recurrent or progressive 


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disease is objectively documented (taking as reference for progressive disease the smallest 
measurements recorded since the treatment started). 
The duration of overall complete response is measured from the time measurement criteria 
are first met for CR until the first date that recurrent disease is objectively documented. 
6.3.4.3 Duration of Stable Disease 
Stable disease is measured from the start of the treatment until the criteria for progression 
are met, taking as reference the smallest measurements recorded since the treatment 
started.  
 
6.4 TOXICITY CRITERIA 
This study will utilize the CTCAE version 3.0 for toxicity and adverse event reporting.  A 
copy of the CTCAE v3.0 can be downloaded from the CTEP home page 
(http://ctep.cancer.gov). All appropriate treatment areas should have access to a copy of the 
CTCAE 3.0.   
 
Over 150 patients have been treated in the Surgery Branch, NCI with tumor infiltrating 
lymphocytes.  Early toxicities related specifically to the infusion of the cells (those which are 
seen immediately following cell infusion and prior to aldesleukin administration) are 
generally mild and include fevers, chills, headache, and malaise.  Toxicities which occur 
following administration of cells can include immune mediated events such as vitiligo, 
transient uveitis, hearing loss and vestibular dysfunction.  The use of the non-myeloablative 
regimen prior to cell administration increases the toxicity of this treatment as profound 
myelosuppression occurs in all patients.  In 93 patients treated with TIL using the non-
myeloablative chemotherapy regimen with or without total body irradiation, there was one 
treatment related death (NMA + 200 cGY TBI) due to an unexpected but preexisting 
diverticular abscess.   
To ensure safety using this treatment, the NCI SB will review safety data on all protocols 
semi-annually at the time of continuing review.  Data will be presented for both the recent 6 
month period and for the entire length of time the protocol has been open.  The toxicity data 
for review will include all toxicities captured on the protocol and will be presented in 
individual tables as follows:  
• all toxicities attributed to the cells,  
• all incidences of intubation including the duration of and reason for intubation, 
• all grade 2 unexpected adverse events, and all grade 3 or greater events regardless of 
attribution.    
Toxicities seen on protocols using this non-myeloablative regimen that occur during the 
follow up period are rare but have included EBV lymphoma following prolonged 
lymphopenia, herpes zoster infection, and sensory neuropathy likely related to fludarabine.   
The major discomforts of the research are those of nausea, mucositis, anorexia, diarrhea, 
fever and malaise.  Side effects of common drugs used in this nonmyeloablative regimen 
include: 
 


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Cyclophosphamide: Marrow suppression, nausea, mucositis, rash, hemorrhagic cystitis, 
myocardial damage, alopecia, infertility, nausea and vomiting, SIADH.   
Fludarabine: Myelosuppression, fever and chills, nausea and vomiting, malaise, fatigue, 
anorexia, weakness, neurologic toxicity including sensory neuropathies and blindness, and 
interstitial pneumonitis.  Serious opportunistic infections have occurred in CLL patients 
treated with fludarabine. 
Antimicrobials in general: Allergic reactions, renal impairment, nausea, vomiting, hepatic 
damage, marrow suppression.  
High-dose aldesleukin: a variety of side effects have been associated with high-dos 
aldesleukin administration. A listing of these side effects in 525 patients treated with high-
dose aldesleukin are listed in Appendix 1. 
7. 
SAFETY REPORTING REQUIREMENTS/DATA AND SAFETY MONITORING 
PLAN 
7.1 DEFINITIONS 
7.1.1 Adverse Event 
An adverse event is defined as any reaction, side effect, or untoward event that occurs 
during the course of the clinical trial associated with the use of a drug in humans, whether 
or not the event is considered related to the treatment or clinically significant. For this 
study, AEs will include events reported by the patient, as well as clinically significant 
abnormal findings on physical examination or laboratory evaluation. A new illness, 
symptom, sign or clinically significant laboratory abnormality or worsening of a pre-
existing condition or abnormality is considered an AE.  All AEs, including clinically 
significant abnormal findings on laboratory evaluations, regardless of severity, will be 
followed until satisfactoryresolution. AEs should be reported up to 30 days following the 
last dose of study drug.  AEs that are considered treatment related, expected, continuing, 
but not resolvable by 30 days after treatment completion (e.g., alopecia) will not be 
followed after the 30-day period.  
7.1.2 Suspected adverse reaction 
Suspected adverse reaction means any adverse event for which there is a reasonable 
possibility that the drug caused the adverse event. For the purposes of IND safety 
reporting, ‘reasonable possibility’ means there is evidence to suggest a causal relationship 
between the drug and the adverse event. A suspected adverse reaction implies a lesser 
degree of certainty about causality than adverse reaction, which means any adverse event 
caused by a drug. 
7.1.3 Unexpected adverse reaction 
An adverse event or suspected adverse reaction is considered “unexpected” if it is not 
listed in the investigator brochure or is not listed at the specificity or severity that has been 
observed; or, if an investigator brochure is not required or available, is not consistent with 
the risk information described in the general investigational plan or elsewhere in the 


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current application. "Unexpected”, also refers to adverse events or suspected adverse 
reactions that are mentioned in the investigator brochure as occurring with a class of drugs 
or as anticipated from the pharmacological properties of the drug, but are not specifically 
mentioned as occurring with the particular drug under investigation. 
7.1.4 Serious 
An Unanticipated Problem or Protocol Deviation is serious if it meets the definition of a 
Serious Adverse Event or if it compromises the safety, welfare or rights of subjects or 
others. 
7.1.5 Serious Adverse Event  
An adverse event or suspected adverse reaction is considered serious if in the view of the 
investigator or the sponsor, it results in any of the following: 
• 
Death, 
• 
A life-threatening adverse drug experience 
• 
Inpatient hospitalization or prolongation of existing hospitalization 
• 
A persistent or significant disability/incapacity 
• 
A congenital anomaly/birth defect. 
• 
Important medical events that may not result in death, be life-threatening, or require 
hospitalization may be considered a serious adverse drug experience when, based 
upon appropriate medical judgment, they may jeopardize the patient or subject and 
may require medical or surgical intervention to prevent one of the outcomes listed in 
this definition. 
7.1.6 Disability 
 A substantial disruption of a person’s ability to conduct normal life functions. 
7.1.7 Life-threatening adverse drug experience 
Any adverse event or suspected adverse reaction that places the patient or subject, in the 
view of the investigator or sponsor, at immediate risk of death from the reaction as it 
occurred, i.e., it does not include a reaction that had it occurred in a more severe form, 
might have caused death. 
7.1.8 Protocol Deviation (NIH Definition) 
Any change, divergence, or departure from the IRB-approved research protocol. 
7.1.9 Non-Compliance (NIH Definition) 
The failure to comply with applicable NIH Human Research Protections Program (HRPP) 
policies, IRB requirements, or regulatory requirements for the protection of human 
research subjects. 


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7.1.10 Unanticipated Problem 
Any incident, experience, or outcome that: 
•  Is unexpected in terms of nature, severity, or frequency in relation to  
(a) the research risks that are described in the IRB-approved research protocol and 
informed consent document; Investigator’s Brochure or other study documents, and 
(b) the characteristics of the subject population being studied; AND 
• Is related or possibly related to participation in the research; AND 
• Places subjects or others at a greater risk of harm (including physical, 
psychological, economic, or social harm) than was previously known or recognized. 
 
7.2 NCI-IRB REPORTING 
7.2.1 NCI-IRB Expedited Reporting of Adverse Events, Unanticipated Problems, and 
Deaths 
The Protocol PI will report to the NCI-IRB: 
• All deaths, except deaths due to progressive disease  
• All Protocol Deviations 
• All Unanticipated Problems 
• All serious non-compliance 
Reports must be received by the NCI-IRB within 7 working days of PI awareness via iRIS. 
7.2.2 NCI-IRB Requirements for PI Reporting at Continuing Review 
The protocol PI will report to the NCI-IRB:  
1. A summary of all protocol deviations in a tabular format to include the date the 
deviation occurred, a brief description of the deviation and any corrective action. 
2. A summary of any instances of non-compliance. 
3. A tabular summary of the following adverse events: 
• 
All Grade 2 unexpected events that are possibly, probably or definitely 
related to the research;  
• 
All Grade 3 and 4 events that are possibly, probably or definitely related to the 
research;  
• 
All Grade 5 events regardless of attribution;  
• 
All Serious Events regardless of attribution.  
NOTE: Grade 1 events are not required to be reported.  


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7.2.3 NCI-IRB Reporting of IND Safety Reports 
Only IND Safety Reports that meet the definition of an unanticipated problem will need to 
be reported to the NCI IRB. 
7.3 IND SPONSOR REPORTING CRITERIA 
An investigator must immediately report to the sponsor, using the mandatory MedWatch 
form 3500a, any serious adverse event, whether or not considered drug related, including 
those listed in the protocol or investigator brochure and must include an assessment of 
whether there is a reasonable possibility that the drug caused the event.  
Study endpoints that are serious adverse events (e.g. all-cause mortality) must be reported in 
accordance with the protocol unless there is evidence suggesting a causal relationship 
between the drug and the event (e.g. death from anaphylaxis).  In that case, the investigator 
must immediately report the death to the sponsor. 
Events will be submitted to Dr. William Dahut, authorized representative for the IND 
Sponsor (CCR) at: 
William Dahut, M.D. 
Bldg 10, Room 3-2571 MSC 1206 
10 Center Drive 
Bethesda, MD 20892 
Telephone: 301-496-4251 
William.Dahut@nih.gov 
Copy all MedWatch forms to: nciprotocolsupportoffice@mail.nih.gov 
7.4 FDA REPORTING CRITERIA 
7.4.1 IND Safety Reports to the FDA (Refer to 21 CFR 312.32) 
The Sponsor will notify the FDA of any unexpected fatal or life-threatening suspected 
adverse reactions as soon as possible but no later than 7 calendar days of initial receipt of 
the information using the MedWatch Form 3500a.  
The Sponsor is also responsible for reporting any:  
• suspected adverse reaction that is both serious and unexpected 
• any findings from clinical, epidemiological, or pooled analysis of multiple 
studies or any findings from animal or in vitro testing that suggest a 
significant risk in humans exposed to the drug 
• clinically important increase in the rate of a serious suspected adverse reaction 
over that listed in the protocol or investigator brochure.  
to the FDA and to all investigators no later than 15 calendar days after determining that the 
information qualifies for reporting using the MedWatch Form 3500a. If FDA requests any 
additional data or information, the sponsor must submit it to the FDA as soon as possible, but 
no later than 15 calendars days after receiving the request. 


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7.4.2 FDA Annual Reports (Refer to 21 CFR 312.33) 
The study Sponsor will submit a brief report annually of the progress of the trial within 
60 days of the anniversary date that the IND went into effect as indicated in 21CFR 
312.33, and any associated FDA correspondences regarding the IND annual report. 
7.4. 
DATA AND SAFETY MONITORING PLAN 
7.4.1 Principal Investigator/Research Team  
The clinical research team will meet on a regular basis when patients are being actively 
treated on the trial to discuss each patient. Decisions about enrollment will be made based 
on the toxicity data from prior patients.  
All data will be collected in a timely manner and reviewed by the principal investigator. 
Adverse events will be reported as required above. Any safety concerns, new information 
that might affect either the ethical and or scientific conduct of the trial, or protocol 
deviations and violations will be reported to the IRB using iRIS. 
The principal investigator will review adverse event and response data on each patient to 
ensure safety and data accuracy. The principal investigator will personally conduct or 
supervise the investigation and provide appropriate delegation of responsibilities to other 
members of the research staff.  
7.4.2 Sponsor Monitoring Plan 
A detailed description of the clinical trial monitoring plan has been included in the initial 
IND submission as required.   
This trial will be monitored by personnel employed by Harris Technical Services on 
contract to the NCI, NIH. Monitors are qualified by training and experience to monitor 
the progress of clinical trials. Personnel monitoring this study will not be affiliated in any 
way with the trial conduct. 
At least 25% of enrolled patients’ will be randomly selected and monitored at least 
quarterly, base on accrual rate. The patients selected will have 100% source document 
verification done. Additional monitoring activities will include: adherence to protocol 
specified study eligibility, treatment plans, data collection for safety and efficacy, 
reporting and time frames of adverse events to the NCI IRB and FDA, and informed 
consent requirements. Written reports will be generated in response to the monitoring 
activities and submitted to the Principal investigator and Clinical Director or Deputy 
Clinical Director, CCR, NCI. 
7.4.3 Safety Monitoring Committee (SMC) 
This protocol will require oversight from the Safety Monitoring Committee (SMC).  
Initial review will occur as soon as possible after the annual NCI-IRB continuing review 
date.   Subsequently, each protocol will be reviewed as close to annually as the quarterly 
meeting schedule permits or more frequently as may be required by the SMC.   For initial 
and subsequent reviews, protocols will not be reviewed if there is no accrual within the 
review period. Written outcome letters will be generated in response to the monitoring 
activities and submitted to the Principal investigator and Clinical Director or Deputy 
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8. 
STATISTICAL CONSIDERATION   
The primary objective of this pilot trial is to determine whether adoptive transfer of Young 
TIL will result in objective clinical responses in patients with either cervical or non-
cervical cancer who are eligible for this trial.   
The study will be conducted using a two-stage optimal design28. Following amendment D, 
patients will be stratified according to whether they have cervical or non-cervical cancer. In 
each cohort, the objective will be to determine if the combination of lymphocyte depleting 
chemotherapy, Young TIL, and high-dose aldesleukin is able to be associated with a 
clinical response rate that can rule out 10% (p0=0.10) in favor of a modest 30% PR + CR 
rate (p1=0.30). The responses determined at 4 month evaluation time point will be the 
definitive results used for this assessment. 
The following design will be used in each of the two cohorts. With alpha=0.05 (5% 
probability of accepting a poor therapy) and beta=0.10 (10% probability of rejecting a good 
therapy), initially 18 evaluable patients will be enrolled in a cohort. If 0 to 2 of the 18 
patients experience a clinical response, then no further patients will be enrolled in that 
cohort. If 3 or more of the first 18 evaluable patients enrolled in a cohort have a clinical 
response, then accrual will continue until a total of 35 evaluable patients have been enrolled 
in that cohort. To reduce the likelihood of transient responses to chemotherapy leading to 
expansion of a cohort, patients must have a PR or CR at least 4 months after cell infusion to 
count toward the clinical responses required for cohort expansion. As it may take several 
weeks to determine if a patient has experienced a clinical response, a temporary pause of 
up to 6 months in the accrual to the trial may be necessary to ensure that enrollment to the 
second stage is warranted. If 3 to 6 of the 35 in a given cohort have a clinical response, then 
this will be considered inadequate for further investigation. If 7 or more of 35 patients have 
a clinical response, then this will indicate that this strategy provides a new approach that 
may be worthy of further consideration for those patients. Under the null hypothesis (10% 
response rate), the probability of early termination is 73%. 
 
To complete the entire study in both cohorts, a total of up to 70 patients may be required.  
In order to allow for a small number of inevaluable patients, the accrual ceiling will be set 
at 73 patients. Provided that about 2 patients per month will be able to be enrolled onto this 
trial, approximately 3-4 years may be needed to accrue the maximum number of required 
patients.   
 
9. 
COLLABORATIVE AGREEMENTS  
We have established a Materials Transfer Agreement (MTA) with Prometheus Laboratories, Inc., 
to supply Proleukin (aldesleukin) for use in this study. 
 
10. HUMAN SUBJECTS PROTECTIONS  
10.1.RATIONALE FOR SUBJECT SELECTION 
The patients to be entered in this protocol have metastatic or recurrent/refractory locally 
advanced HPV-associated cancer which is refractory to standard therapy, and limited life 
expectancies. 


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Subjects from both genders and all racial/ethnic groups are eligible for this study if they meet 
the eligibility criteria.  To date, there is no information that suggests that differences in drug 
metabolism or disease response would be expected in one group compared to another.  
Efforts will be made to extend accrual to a representative population, but in this preliminary 
study, a balance must be struck between patient safety considerations and limitations on the 
number of individuals exposed to potentially toxic and/or ineffective treatments on the one 
hand and the need to explore gender and ethnic aspects of clinical research on the other hand.  
If differences in outcome that correlate to gender or to ethnic identity are noted, accrual may 
be expanded or a follow-up study may be written to investigate those differences more fully. 
10.2.PARTICIPATION OF CHILDREN 
The use of the nonmyeloablative regimen in this protocol is a major procedure which entails 
serious discomforts and hazards for the patient, such that fatal complications are possible.  It 
is therefore only appropriate to carry out this experimental procedure in the context of life 
threatening metastatic cancer.  Since the efficacy of this experimental procedure is unknown, 
it does not seem reasonable to expose children to this risk without further evidence of benefit.  
Should results of this study indicate efficacy in treating metastatic cancer, which is not 
responsive to other standard forms of therapy, future research can be conducted in the 
pediatric population to evaluate potential benefit in that patient population. 
10.3.EVALUATION OF BENEFITS AND RISKS/DISCOMFORTS 
The experimental treatment has a chance to provide clinical benefit though it is not known if 
it will do so. The risks in this treatment are detailed in section 6.4. The success of this effort 
cannot be predicted at this time. Because all patients in this protocol have incurable 
metastatic or recurrent/refractory locally advanced HPV-associated cancers the potential 
benefit is thought to outweigh the risks. 
10.4.RISKS/BENEFITS ANALYSIS 
Because all patients in this protocol have metastatic or recurrent/refractory locally advanced 
HPV-associated cancer and limited life expectancies the potential benefit is thought to 
outweigh the potential risks. 
10.5.CONSENT AND ASSENT PROCESS AND DOCUMENTATION 
Patients initially signs a consent when they agree to have TIL obtained for study and growth 
on protocol 03-C-0277, Cell Harvest and Preparation for Surgery Branch Adoptive Cell 
Therapy Protocols.  If the Young TIL can be generated for infusion and the patient meets the 
thorough screening for eligibility, the patient, with family members or friends at the request 
of the patient, will be presented with a detailed description of the protocol treatment.  The 
specific requirements, objectives, and potential advantages and disadvantages will be 
presented.  The Informed Consent document is given to the patient, who is requested to 
review it and to ask questions prior to agreeing to participate in the treatment portion of this 
protocol.  The patient is reassured that participation on trial is entirely voluntary and that 
he/she can withdraw or decide against treatment at any time without adverse consequences. 
The research nurse, principal investigator, associate investigator, or clinical associate is 
responsible for obtaining written consent from the patient. 


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11. PHARMACEUTICAL INFORMATION   
11.1.INTERLEUKIN-2 (ALDESLEUKIN, PROLEUKIN, RECOMBINANT HUMAN INTERLEUKIN 2) 
How Supplied: Interleukin-2 (aldesleukin) will be provided by the NIH Clinical Pharmacy 
Department from commercial sources.   
Formulation/Reconstitution: Aldesleukin, NSC #373364, is provided as single-use vials 
containing 22 million IU (−1.3 mg) IL-2 as a sterile, white to off-white  lyophilized cake plus 
50 mg mannitol and 0.18 mg sodium dodecyl sulfate, buffered with approximately 0.17 mg 
monobasic and 0.89 mg dibasic sodium phosphate to a pH of 7.5 (range 7.2 to 7.8).  The vial 
is reconstituted with 1.2 mL of Sterile Water for Injection, USP, and the resultant 
concentration is 18 million IU/ml or 1.1 mg/mL.  Diluent should be directed against the side 
of the vial to avoid excess foaming.  Swirl contents gently until completely dissolved.  Do 
not shake.  Since vials contain no preservative, reconstituted solution should be used with 24 
hours. 
Storage: Intact vials are stored in the refrigerator (20 - 80C) protected from light.  Each vial 
bears an expiration date. 
Dilution/Stability: Reconstituted aldesleukin should be further diluted with 50 mL of 5% 
Human Serum Albumin (HSA).  The HSA should be added to the diluent prior to the 
addition of RIL-2.  Dilutions of the reconstituted solution over a 1000-fold range (i.e., 1 
mg/mL to 1 mcg/mL) are acceptable in either glass bottles or polyvinyl chloride bags.  
Aldesleukin is chemically stable for 48 hours at refrigerated and room temperatures, 20 – 
300C. 
Administration:  The dosage will be calculated based on total body weight.  The final dilution 
of aldesleukin will be infused over 15 minutes.  Aldesleukin will be administered as an 
inpatient.   
Toxicities:  Expected toxicities of aldesleukin are listed in the product label and in Appendix 
1 and 2.  Grade 3 toxicities common to aldesleukin include diarrhea, nausea, vomiting, 
hypotension, skin changes, anorexia, mucositis, dysphagia, or constitutional symptoms and 
laboratory changes as detailed in Appendix 1.  Additional grade 3 and 4 toxicities seen with 
aldesleukin are detailed in Appendix 2.   
11.2.FLUDARABINE: 
(Please refer to package insert for complete product information)   
Description: Fludarabine phosphate is a synthetic purine nucleoside that differs from 
physiologic nucleosides in that the sugar moiety is arabinose instead of ribose or 
deoxyribose.  Fludarabine is a purine antagonist antimetabolite.  
How Supplied:  It will be purchased by the NIH Clinical Pharmacy Department from 
commercial sources. Fludarabine is supplied in a 50 mg vial as a fludarabine phosphate 
powder in the form of a white, lyophilized solid cake.   
Stability:  Following reconstitution with 2 mL of sterile water for injection to a concentration 
of 25 mg/ml, the solution has a pH of 7.7.  The fludarabine powder is stable for at least 18 
months at 2-8°C; when reconstituted, fludarabine is stable for at least 16 days at room 
temperature.  Because no preservative is present, reconstituted fludarabine will typically be 


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administered within 8 hours.  Specialized references should be consulted for specific 
compatibility information.  Fludarabine is dephosphorylated in serum, transported 
intracellularly and converted to the nucleotide fludarabine triphosphate; this 2-fluoro-ara-
ATP molecule is thought to be required for the drug’s cytotoxic effects.  Fludarabine inhibits 
DNA polymerase, ribnucleotide reductase, DNA primase, and may interfere with chain 
elongation, and RNA and protein synthesis. 
Storage:  Intact vials should be stored refrigerated (2-8°C). 
Administration:  Fludarabine is administered as an IV infusion in 100 ml 0.9% sodium 
chloride, USP over 15 to 30 minutes.  The doses will be based on body surface area (BSA).  
If patient is obese (BMI > 35) drug dosage will be calculated using practical weight as 
described in Table 9.  
Toxicities:  At doses of 25 mg/m2/day for 5 days, the primary side effect is 
myelosuppression; however, thrombocytopenia is responsible for most cases of severe and 
life-threatening hematologic toxicity.  Serious opportunistic infections have occurred in CLL 
patients treated with fludarabine.  Hemolytic anemia has been reported after one or more 
courses of fludarabine with or without a prior history of a positive Coomb’s test; fatal 
hemolytic anemia has been reported.  In addition, bone marrow fibrosis has been observed 
after fludarabine therapy.  Other common adverse effects include malaise, fever, chills, 
fatigue, anorexia, nausea and vomiting, and weakness.  Irreversible and potentially fatal 
central nervous system toxicity in the form of progressive encephalopathy, blindness, and 
coma is only rarely observed at the currently administered doses of fludarabine.  More 
common neurologic side effects at the current doses of fludarabine include weakness, pain, 
malaise, fatigue, paresthesia, visual or hearing disturbances, and sleep disorders.  Adverse 
respiratory effects of fludarabine include cough, dyspnea, allergic or idiopathic interstitial 
pneumonitis.  Tumor lysis syndrome has been rarely observed in fludarabine treatment of 
CLL.  Treatment on previous adoptive cell therapy protocols in the Surgery Branch have 
caused persistently low (below 200) CD4 counts, and one patient developed polyneuropathy 
manifested by vision blindness, and motor and sensory defects. 
11.3.CYCLOPHOSPHAMIDE 
 (Refer to FDA-approved package insert for complete product information): 
Description:  Cyclophosphamide is a nitrogen mustard-derivative alkylating agent.  
Following conversion to active metabolites in the liver, cyclophosphamide functions as an 
alkyating agent; the drug also possesses potent immunosuppressive activity.  The serum half-
life after IV administration ranges from 3-12 hours; the drug and/or its metabolites can be 
detected in the serum for up to 72 hours after administration.   
How Supplied:  Cyclophosphamide will be obtained from commercially available sources by 
the Clinical Center Pharmacy Department.     
Stability:  Following reconstitution as directed with sterile water for injection, 
cyclophosphamide is stable for 24 hours at room temperature or 6 days when kept at 2-8°C.   
Administration:  It will be diluted in 250 ml D5W and infused over one hour.  The dose will 
be based on the patient’s body weight.  If patient is obese (BMI > 35) drug dosage will be 
calculated using practical weight as described in Table 9. 


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Toxicities:  Hematologic toxicity occurring with cyclophosphamide usually includes 
leukopenia and thrombocytopenia.  Anorexia, nausea and vomiting, rash and alopecia occur, 
especially after high-dose cyclophosphamide; diarrhea, hemorrhagic colitis, infertility, and 
mucosal and oral ulceration have been reported.  Sterile hemorrhagic cystitis occurs in about 
20% of patients; severity can range from microscopic hematuria to extensive cystitis with 
bladder fibrosis.  Although the incidence of hemorrhagic cystitis associated with 
cyclophosphamide appears to be lower than that associated with ifosfamide, mesna (sodium 
2-mercaptoethanesulfonate) has been used prophylactically as a uroprotective agent in 
patients receiving cyclophosphamide.  Prophylactic mesna is not effective in preventing 
hemorrhagic cystitis in all patients.  Patients who receive high dose cyclophosphamide may 
develop interstitial pulmonary fibrosis, which can be fatal.  Hyperuricemia due to rapid 
cellular destruction may occur, particularly in patients with hematologic malignancy.  
Hyperuricemia may be minimized by adequate hydration, alkalinization of the urine, and/or 
administration of allopurinol.  If allopurinol is administered, patients should be watched 
closely for cyclophosphamide toxicity (due to allopurinol induction of hepatic microsomal 
enzymes).  At high doses, cyclophosphamide can result in a syndrome of inappropriate 
antidiuretic hormone secretion; hyponatremia with progressive weight gain without edema 
occurs.  At high doses, cyclophosphamide can result in cardiotoxicity.  Deaths have occurred 
from diffuse hemorrhagic myocardial necrosis and from a syndrome of acute 
myopericarditis; in such cases, congestive heart failure may occur within a few days of the 
first dose.  Other consequences of cyclophosphamide cardiotoxicity include arrhythmias, 
potentially irreversible cardiomyopathy, and pericarditis.  Other reported adverse effects of 
cyclophosphamide include headache, dizziness, and myxedema; faintness, facial flushing, 
and diaphoresis have occurred following IV administration.  Mesna (sodium 2-
mercaptoethanesulphonate; given by IV injection) is a synthetic sulfhydryl compound that 
can chemically interact with urotoxic metabolites of cyclophosphamide (acrolein and 4-
hydroxycyclophosphamide) to decrease the incidence and severity of hemorrhagic cystitis.  
11.4.YOUNGTIL PREPARATION 
The Certificate of Analysis is similar to those approved by the Food and Drug Administration 
and used in other Surgery Branch, NCI TIL clinical studies.  The autologous Young TIL 
product will be provided for investigational use only under a sponsor-investigator IND.  The 
Certificate of Analysis is in Appendix 4 and the Standard Operating Procedures for the 
growth of the Young TIL are included in the IND. For all cohorts, cells will be administered 
at a dose of between 1x109 to 2x1011 lymphocytes in a volume of 100-200 mL.   
 
11.5.MESNA  
(Sodium 2-mercaptoethanesulfonate, Mesnum, Mesnex, NSC-113891):  
(Please refer to the FDA-approved package insert for complete product information) 
Description:  Mesna will be obtained commercially by the Clinical Center Pharmacy 
Department and is supplied as a 100 mg/ml solution.   
Storage:  Intact ampoules are stored at room temperature.   


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Stability:  Diluted solutions (1 to 20 mg/mL) are physically and chemically stable for at least 
24 hours under refrigeration.  Mesna is chemically stable at room temperature for 48-72 
hours in D5W, 48-72 hour in D5W/0.45% NaCl, or 24 hours in 0.9% NaCl. 
Administration:  Dilute to concentrations less than or equal to 20 mg mesna/ml fluid in D5W 
or 0.9% NaCl and to be administered intravenously as a continuous infusion.  If patient is 
obese (BMI > 35) drug dosage will be calculated using practical weight as described in Table 
9.  Toxicities include nausea, vomiting and diarrhea.  
11.6.FILGRASTIM  
(Granulocyte Colony-Stimulating Factor, G-CSF, Filgrastim, Neupogen): 
Filgrastim will be obtained commercially by the Clinical Center Pharmacy Department and is 
supplied in 300 ug/ml and 480 ug/1.6 ml vials.  G-CSF should be refrigerated and not 
allowed to freeze.  The product bears the expiration date.  The product should not be shaken.  
It is generally stable for at least 10 months when refrigerated.  The appropriate dose is drawn 
up into a syringe.  G-CSF will be given as a daily subcutaneous injection.  The side effects of 
G-CSF are skin rash, myalgia and bone pain, an increase of preexisting inflammatory 
conditions, enlarged spleen with occasional associated low platelet counts, alopecia (with 
prolonged use) elevated blood chemistry levels. 
11.7.TRIMETHOPRIM AND SULFAMETHOXAZOLE DOUBLE STRENGTH (TMP / SMX DS): 
TMP/SMX DS will be obtained by the Clinical Center Pharmacy Department from 
commercial sources.  It will be used for the prevention of PCP pneumonia.  The oral dose is 1 
tablet PO daily three times a week (MUST be on non-consecutive days) beginning on day -7 
and continuing for at least 6 months and until the CD4 count is greater than 200 on 2 
consecutive lab studies.  Like other sulfa drugs, TMP/SMX DS can cause allergies, fever, 
photosensitivity, nausea, and vomiting. Allergies typically develop as a widespread itchy red 
rash with fever eight to fourteen days after beginning the standard dose. Neutropenia, a 
reduction in the number of neutrophils, can also occur. 
11.8.AEROSOLIZED PENTAMIDINE IN PLACE OF TMP/SMX DS:   
Patients with sulfa allergies will receive aerosolized Pentamidine 300 mg per nebulizer with  
one week prior to admission and continued monthly until the CD4 count is above 200 on two 
consecutive follow up lab studies and for at least 6 months post chemotherapy.  Pentamidine 
Isethionate will be obtained by the Clinical Center Pharmacy Department from commercial 
sources.  It will be used to prevent the occurrence of PCP infections.  It is supplied in 300 mg 
vials of lyophilized powder and will be administered via nebulizer.  Toxicities reported with 
the use of Pentamidine include metallic taste, coughing, bronchospasm in heavy smokers and 
asthmatics; increased incidence of spontaneous pneumothorax in patients with previous PCP 
infection or pneumatoceles, or hypoglycemia. 
 
11.9.HERPES VIRUS PROPHYLAXIS: 
11.9.1. Valacyclovir (Valtrex): 


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Valacyclovir will be obtained by the Clinical Center Pharmacy Department from commercial 
sources.  It will be used orally to prevent the occurrence of herpes virus infections in patients 
with positive HSV serology.   It is supplied in 500 mg tablets.  Valcyclovir will be started the 
day after the last dose of fludarabine at a dose of 500 mg orally daily if the patient is able to 
tolerate oral intake.  See package insert for dosing adjustments in patients with renal 
impairment.  Common side effects include headache, upset stomach, nausea, vomiting, 
diarrhea or constipation.  Rare serious side effects include hemolytic uremic syndrome and 
thrombotic thrombocytopenic purpura.       
11.9.2. Acyclovir 
Acyclovir will be obtained by the Clinical Center Pharmacy Department from commercial  
sources.  It will be used to prevent the occurrence of herpes virus infections in patients who 
cannot take oral medications.  It is supplied as powder for injection in 500 mg/vials.  
Reconstitute in 10 mL of sterile water for injection to a concentration of 50 mg/mL.  
Reconstituted solutions should be used within 12 hours.  IV solutions should be diluted to a 
concentration of 7mg/mL or less and infused over 1 hour to avoid renal damage.  Reversible 
renal insufficiency has been reported with IV but not oral acyclovir.  Neurologic toxicity 
including delirium, tremors, coma, acute psychiatric disturbances, and abnormal EEGs have 
been reported with higher doses of acyclovir.  Should this occur, a dosage adjustment will be 
made or the drug will be discontinued. Stomach upset, headache or nausea, rash or hives; 
peripheral edema; pain, elevated liver function tests; and leukopenia, diarrhea, 
lymphadenopathy, myalgias, visual abnormalities and elevated creatinine have been reported. 
Hair loss from prolonged use has been reported. Acyclovir will not be used concomitantly 
with other nucleoside analogs which interfere with DNA synthesis, e.g. ganciclovir. In renal 
disease, the dose is adjusted as per product labeling.  
 
Fluconazole: 
Fluconazole will be obtained by the Clinical Center Pharmacy Department from commercial 
sources.  It will be used to prophylax against fungal infections.  It is available in 200 mg 
tablets.  It can cause headache, nausea, vomiting, diarrhea or abdominal pain, and liver 
damage which may be irreversible.  It can cause rashes and itching, which in rare cases has 
caused Stevens Johnson Syndrome.  It has several significant drug interactions.  The package 
insert should be consulted prior to prescribing.  For IV administration in patients who cannot 
tolerate the oral preparation, Fluconazole comes in 2 MG/ML solution for injection, and 
prepared according to Clinical Center Pharmacy standard procedures.  It should be 
administered at a maximum IV rate of 200 mg/hr. 
11.10. 
SUPPORT MEDICATIONS 
Ondansetron hydrochloride 
Ondansetron hydrochloride will be obtained by the Clinical Center Pharmacy Department 
from commercial sources.  It will be used to control nausea and vomiting during the 
chemotherapy preparative regimen.  It can cause headache, dizziness, myalgias, drowsiness, 
malaise, and weakness.  Less common side effects include chest pain, hypotension, pruritis, 
constipation and urinary retention.  Consult the package insert for specific dosing 
instructions. 
Furosemide 


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Furosemide will be obtained by the Clinical Center Pharmacy Department from commercial 
sources.  It will be used to enhance urine output during the chemotherapy preparative 
regimen with cyclophosphamide.  Adverse effects include dizziness, vertigo, paresthesias, 
weakness, orthostatic hypotension, photosensitivity, rash and pruritis.  Consult the package 
insert for a complete list of all side effects. 
 
 
 


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12. REFERENCES  
1. 
Rosenberg SA, Yang JC, Sherry RM, et al. Durable complete responses in heavily 
pretreated patients with metastatic melanoma using T-cell transfer immunotherapy. Clinical 
Cancer Research 2011;17:4550-7. 
2. 
Chaturvedi AK, Engels EA, Pfeiffer RM, et al. Human Papillomavirus and Rising 
Oropharyngeal Cancer Incidence in the United States. Journal of Clinical Oncology 2011. 
3. 
Cancer Facts & Figures 2010. Atlanta: American Cancer Society; 2010. 
4. 
Gillison ML, Chaturvedi AK, Lowy DR. HPV prophylactic vaccines and the potential 
prevention of noncervical cancers in both men and women. Cancer 2008;113:3036-46. 
5. 
Ryerson AB, Peters ES, Coughlin SS, et al. Burden of potentially human papillomavirus-
associated cancers of the oropharynx and oral cavity in the US, 1998-2003. Cancer 
2008;113:2901-9. 
6. 
Long HJ, 3rd, Bundy BN, Grendys EC, Jr., et al. Randomized phase III trial of cisplatin 
with or without topotecan in carcinoma of the uterine cervix: a Gynecologic Oncology Group 
Study. Journal of Clinical Oncology 2005;23:4626-33. 
7. 
Vermorken JB, Mesia R, Rivera F, et al. Platinum-based chemotherapy plus cetuximab in 
head and neck cancer. The New England Journal of Medicine 2008;359:1116-27. 
8. 
Long HJ, 3rd. Management of metastatic cervical cancer: review of the literature. Journal 
of Clinical Oncology 2007;25:2966-74. 
9. 
Monk BJ, Sill MW, McMeekin DS, et al. Phase III trial of four cisplatin-containing 
doublet combinations in stage IVB, recurrent, or persistent cervical carcinoma: a Gynecologic 
Oncology Group study. Journal of Clinical Oncology 2009;27:4649-55. 
10. 
Monk BJ, Sill MW, Burger RA, Gray HJ, Buekers TE, Roman LD. Phase II trial of 
bevacizumab in the treatment of persistent or recurrent squamous cell carcinoma of the cervix: a 
gynecologic oncology group study. Journal of Clinical Oncology 2009;27:1069-74. 
11. 
Vermorken JB, Trigo J, Hitt R, et al. Open-label, uncontrolled, multicenter phase II study 
to evaluate the efficacy and toxicity of cetuximab as a single agent in patients with recurrent 
and/or metastatic squamous cell carcinoma of the head and neck who failed to respond to 
platinum-based therapy. Journal of Clinical Oncology 2007;25:2171-7. 
12. 
Dudley ME, Yang JC, Sherry R, et al. Adoptive cell therapy for patients with metastatic 
melanoma: evaluation of intensive myeloablative chemoradiation preparative regimens. Journal 
of Clinical Oncology 2008;26:5233-9. 
13. 
Magaldi TG, Almstead LL, Bellone S, Prevatt EG, Santin AD, Dimaio D. Primary human 
cervical carcinoma cells require human papillomavirus E6 and E7 expression for ongoing 
proliferation. Virology 2011. 
14. 
Moody CA, Laimins LA. Human papillomavirus oncoproteins: pathways to 
transformation. Nature Reviews Cancer 2010;10:550-60. 
15. 
Sima N, Wang W, Kong D, et al. RNA interference against HPV16 E7 oncogene leads to 
viral E6 and E7 suppression in cervical cancer cells and apoptosis via upregulation of Rb and 
p53. Apoptosis 2008;13:273-81. 


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16. 
Jiang M, Milner J. Selective silencing of viral gene E6 and E7 expression in HPV-
positive human cervical carcinoma cells using small interfering RNAs. Methods in Molecular 
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17. 
McLaughlin-Drubin ME, Crum CP, Münger K. Human papillomavirus E7 oncoprotein 
induces KDM6A and KDM6B histone demethylase expression and causes epigenetic 
reprogramming. Proceedings of the National Academy of Sciences 2011;108:2130-5. 
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Marur S, D'Souza G, Westra WH, Forastiere AA. HPV-associated head and neck cancer: 
a virus-related cancer epidemic. The lancet oncology 2010;11:781-9. 
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Heusinkveld M, Welters MJ, van Poelgeest MI, et al. The detection of circulating human 
papillomavirus-specific T cells is associated with improved survival of patients with deeply 
infiltrating tumors. International Journal of Cancer 2011;128:379-89. 
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Bell MC, Edwards RP, Partridge EE, et al. CD8+ T lymphocytes are recruited to 
neoplastic cervix. Journal of Clinical Immunology 1995;15:130-6. 
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Piersma SJ, Jordanova ES, van Poelgeest MI, et al. High number of intraepithelial CD8+ 
tumor-infiltrating lymphocytes is associated with the absence of lymph node metastases in 
patients with large early-stage cervical cancer. Cancer Research 2007;67:354-61. 
22. 
Bethwaite PB, Holloway LJ, Thornton A, Delahunt B. Infiltration by immunocompetent 
cells in early stage invasive carcinoma of the uterine cervix: a prognostic study. Pathology 
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23. 
Chao HT, Wang PH, Tseng JY, Lai CR, Chiang SC, Yuan CC. Lymphocyte-infiltrated 
FIGO Stage IIB squamous cell carcinoma of the cervix is a prominent factor for disease-free 
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24. 
de Vos van Steenwijk PJ, Heusinkveld M, Ramwadhdoebe TH, et al. An unexpectedly 
large polyclonal repertoire of HPV-specific T cells is poised for action in patients with cervical 
cancer. Cancer Research 2010;70:2707-17. 
25. 
Piersma SJ, Welters MJ, van der Hulst JM, et al. Human papilloma virus specific T cells 
infiltrating cervical cancer and draining lymph nodes show remarkably frequent use of HLA-DQ 
and -DP as a restriction element. International Journal of Cancer 2008;122:486-94. 
26. 
Kenter GG, Welters MJ, Valentijn AR, et al. Vaccination against HPV-16 oncoproteins 
for vulvar intraepithelial neoplasia. The New England Journal of Medicine 2009;361:1838-47. 
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28. 
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29. 
Ghosh AK, Moore M. Tumour-infiltrating lymphocytes in cervical carcinoma. European 
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30. 
Hilders CG, Ras L, van Eendenburg JD, Nooyen Y, Fleuren GJ. Isolation and 
characterization of tumor-infiltrating lymphocytes from cervical carcinoma. International Journal 
of Cancer 1994;57:805-13. 


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31. 
Badoual C, Hans S, Rodriguez J, et al. Prognostic value of tumor-infiltrating CD4+ T-cell 
subpopulations in head and neck cancers. Clinical Cancer Research 2006;12:465-72. 
32. 
Distel LV, Fickenscher R, Dietel K, et al. Tumour infiltrating lymphocytes in squamous 
cell carcinoma of the oro- and hypopharynx: prognostic impact may depend on type of treatment 
and stage of disease. Oral Oncology 2009;45:e167-74. 
33. 
Rubio CA, Nilsson PJ. Squamous-cell carcinoma of the anus with high intratumoral 
lymphocytosis and its clinical implications. Anticancer Research 2008;28:2469-72. 
 


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13. FIGURES, TABLES & APPENDICES 
 
 
Table 1. Deaths from cancers at HPV-associated sites in the 
United States in 2010. 
Site  
Deaths  
Oral cavity and pharynx 
(not tongue or mouth)  
4,060  
Cervix  
4,210  
Vulva  
920  
Female genital  
780  
Penis  
310  
Anus 
720  
Total  
11,000  
Adapted from: American Cancer Society. Cancer Facts & 
Figures 2010. Atlanta: American Cancer Society; 2010. 


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Table 2. Response rates and duration in patients with melanoma treated with tumor infiltrating 
lymphocytes plus high-dose aldesleukin following three different lymphoconditioning regimens. 


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Table 3. Transfusions and grade 3 and 4 non-hematologic toxicities associated with NMA plus 
TBI lymphodepleting preparative regimens. 
 


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Table 4. Time in hospital and non-hematological grade 3 and 4 toxicities related to 
lymphodepleting chemotherapy and cell transfer. 
 
 


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Table 5. Studies reporting correlations between HPV-associated tumor immunological 
responses and clinical outcomes.  
Site 
Disease extent 
Link to T cell immunity 
Authors 
Cervix 
Large early-
stage 
CD8+ T cell infiltration correlates with absence of 
lymph node metastases 
Piersma, et al21 
Cervix 
Early stage 
Tumor infiltration with T cells associated with 
improved survival 
Bethwaite, et al22 
Cervix 
Resected tumors 
TIL from 23 of 54 patients displayed HPV E6 or E7 
specific responses 
Piersma, et al25 
Cervix 
Deeply 
infiltrating 
HPV-specific immune response correlated with 
improved disease free survival 
Heusinkveld, et 
al19 
Cervix 
Resected tumors 
Tumor-reactive TIL can be generated from cervical 
cancers 
Ghosh, et al29 
Cervix 
Resected tumors 
Tumor-reactive TIL can be generated from cervical 
cancers 
Hilders, et al30 
Head and Neck 
All stages 
Tumor infiltrating activated CD4+ T cells associated 
with improved overall survival 
Badoual, et al31 
Oro- and 
hypopharynx 
“Low-risk” 
High numbers of intraepithelial CD8+ TIL related to 
improved disease-free survival 
Distel, et al32 
Vulva 
Intraepithelial 
neoplasia 
Regression of vulvar intraepithelial neoplasia 
following HPV16 E6/E7 long-peptide vaccination 
Kenter, et al26 
Anal 
All stages 
Improved survival in patients with high number of 
tumoral intraepithelial lymphocytes 
Rubio, et al33 
 
 
 


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Table 6. HPV16 and HPV 18 responses detected in infiltrating lymphocytes25.  


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Table 7. Patient characteristics and success at generating tumor infiltrating lymphocytes from digestive 
tract cancers. 
Patients 
Age, 
gender 
Prior 
chemo-
therapy 
regimens 
Metastatic adenocarcinoma    
(site of TIL harvest) 
Technique for 
growing TIL 
Successful 
first 
outgrowth 
Total cell 
count, end of 
REP (x e9) 
Preclinical studies 
 
 
 
 
 
1) 
R. W. 
63, M 
2 
Colon (liver) 
GMACS,     O/N digest 
yes 
47.5 
2) 
A. S. 
44, F 
3 
Gastric (liver) 
GMACS,     O/N digest 
yes 
38.3 
3) 
T. H. 
45, M 
1 
Colon (liver) 
GMACS,     O/N digest 
yes 
25.1 
Clinical studies 
 
 
 
 
 
4) 
M. L. 
45, F 
3 
Colon (liver) 
GMACS,     O/N digest 
yes 
18.5 
5) 
D. A. 
57, M 
3 
Biliary tract (omentum) 
GMACS,     O/N digest 
yes 
no REP 
6) 
J.F. L. 
42, M 
1 
Colon (retro-peritoneum) 
Fragments 
yes 
32.1 
7) 
J. M. 
51, M 
4 
Rectal (lung) 
Fragments, GMACS 
yes 
20.0 
8) 
M. M. 
51, M 
2 
Colon (lung) 
Fragments, GMACS 
yes 
no REP 
9) 
M. S. 
53, F 
2 
Colon (abdominal wall) 
Fragments, GMACS 
no 
- 
11) 
S.M. 
57, F 
3 
Colon (liver) 
Fragments, GMACS 
yes 
no REP 
12) 
J. R. 
51, F 
1 
Colon (liver, omental nodule) 
Fragments, GMACS 
yes 
30.3 
13) 
A. H. 
52, F 
6 
Colon (lung) 
Fragments, GMACS 
yes 
69.5 
14) 
B. D. 
41, M 
5 
Colon (Axillary node) 
Fragments, GMACS 
yes 
no REP 
15) 
M. W. 
37, F 
2 
Colon (liver) 
Fragments, GMACS 
yes 
75.0 


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Table 8. Characteristics of tumor infiltrating lymphocytes grown from five cervical cancer 
specimens. 
 
 
 
 
 
 
Frequency of cells in the 
lymphocyte gate 
 
Patient 
Prior 
therapy 
Growth positive 
wells/total wells1 
Autologous 
tumor 
reactivity2 
Fragments 
for REP 
CD3+ 
CD4+ CD8+ 
CD3-
CD56+ 
REP fold 
expansion 
1 
none 
8/24 
n/a 
F1 
97 
73 
24 
3 
1216 
 
 
 
 
F2 
96 
9 
85 
2 
1110 
 
 
 
 
F3 
79 
35 
38 
18 
1326 
 
 
 
 
F4 
62 
9 
93 
6 
1890 
2 
chemo/RT 
5/24 
positive 
F1 
80 
10 
42 
19 
344 
 
 
 
 
F2 
78 
31 
36 
19 
1752 
 
 
 
 
F3 
86 
70 
10 
11 
1224 
 
 
 
 
F4 
78 
8 
71 
21 
816 
3 
none 
17/26 
negative 
F1 
99 
75 
24 
0 
2226 
 
 
 
 
F2 
99 
15 
84 
1 
1632 
 
 
 
 
F3 
93 
35 
32 
3 
2240 
 
 
 
 
F4 
99 
90 
7 
0 
3 
4 
none 
2/19 
n/a 
F1 
95 
69 
8 
3 
2737 
 
 
 
 
F2 
95 
11 
84 
5 
1968 
5 
none 
13/13 
n/a 
n/a 
n/a 
n/a 
n/a 
n/a 
n/a 
1 Growth positive defined as lymphocyte confluence in 4 wells of a 24-well plate, approximately 10e6 cells. 
2Autologous tumor reactivity was determined by IFN-γ production in overnight coculture with cryopreserved 
autologous tumor targets. Positive recognition was defined as two times the negative control and greater than 200 
pg/mL of IFN-γ. 
3Pooled with F3 for the REP. 
n/a = not tested 
 
 
 


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Table 9: 
Modification of Dose Calculations* in patients whose BMI is greater than 35 
Unless otherwise specified in this  protocol, actual body weight is used for dose 
calculations of treatment agents.   In patients who are determined to be obese (BMI > 
35), the practical weight (see 3 below) will be used. 
 
 
1. BMI Determination: 
 
BMI = weight (kg) / [height (m)]2 
 
 
2. Calculation of ideal body weight 
 
Male = 50 kg + 2.3 (number of inches over 60 inches) 
Example:  ideal body weight of 5’10’’ male 
50 + 2.3 (10) = 73 kg 
 
Female = 45.5 kg + 2.3 (number of inches over 60 inches) 
Example:  ideal body weight of 5’3’’ female 
45.5 + 2.3 (3) = 57 kg 
 
3. Calculation of “practical weight” 
 
Calculate the average of the actual and the ideal body weights.  This is the 
practical weight to be used in calculating the doses of chemotherapy and 
associated agents designated in the protocol. 


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Figure 1. Survival of patients with metastatic melanoma treated with autologous tumor infiltration 
lymphocytes and IL-2 following three different lymphoconditioning regimens. 


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Appendix 1: 
ADVERSE EVENTS OCCURRING IN ≥10% OF PATIENTS TREATED WITH ALDESLEUKIN 
(n=525)1 
Body System  
% Patients  
 
Body System  
   % Patients 
Body as a Whole  
 
 
 
Metabolic and Nutritional Disorders 
Chills   
 
 
52  
 
Bilirubinemia   
 
40 
Fever   
 
 
29  
 
Creatinine increase   
33 
Malaise  
 
 
27  
 
Peripheral edema  
 
28 
Asthenia  
 
 
23 
 
 SGOT increase  
 
23 
Infection  
 
 
13  
 
Weight gain   
 
16 
Pain   
 
 
12  
 
Edema  
 
 
15 
Abdominal pain  
 
11  
 
Acidosis  
 
 
12 
Abdomen enlarged  
 
10  
 
Hypomagnesemia  
 
12 
Cardiovascular  
 
 
 
Hypocalcemia  
 
11 
Hypotension   
 
71  
 
Alkaline phosphatase incr 
10 
Tachycardia   
 
23 
 
Nervous 
Vasodilation   
 
13  
 
Confusion  
 
 
34 
Supraventricular tachycardia  12  
 
Somnolence   
 
22 
Cardiovascular disordera  
11  
 
Anxiety  
 
 
12 
Arrhythmia  
 
 
10  
 
Dizziness  
 
 
11 
Digestive  
 
 
 
 
Respiratory 
Diarrhea  
 
 
67  
 
Dyspnea  
 
 
43 
Vomiting  
 
 
50  
 
Lung disorderb  
 
24 
Nausea  
 
 
35  
 
Respiratory disorderc   
11 
Stomatitis  
 
 
22  
 
Cough increase  
 
11 
Anorexia  
 
 
20  
 
Rhinitis  
 
 
10 
Nausea and vomiting   
19  
 
Skin and Appendages 
Hemic and Lymphatic  
 
 
Rash   
 
 
42 
Thrombocytopenia  
 
37 
 
 Pruritus  
 
 
24 
Anemia  
 
 
29  
 
Exfoliative dermatitis   
18 
Leukopenia 
 
 
16  
 
Urogenital 
Oliguria  
 
 
63 
a Cardiovascular disorder: fluctuations in blood pressure, asymptomatic ECG changes, CHF. 
b Lung disorder: physical findings associated with pulmonary congestion, rales, rhonchi. 
c Respiratory disorder: ARDS, CXR infiltrates, unspecified pulmonary changes. 
1Source: Proleukin® Prescribing Information – June 2007 


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Appendix 2 
Expected IL-2 Toxicities and their Management 
 
Expected 
toxicity 
Expected 
grade 
Supportive 
Measures 
Stop Cycle* 
 
Stop Treatment ** 
Chills 
3 
IV Meperidine 25-50 
mg, IV q1h, prn, 
No 
No 
Fever 
3 
Acetaminophen 650 
mg, po, q4h; 
Indomethicin 50-75 
mg, po, q8h 
No 
No 
Pruritis 
3 
Hydroxyzine HCL 
10-20 mg po q6h, 
prn; 
Diphenhydramine 
HCL25-50 mg, po, 
q4h, prn 
No 
No 
Nausea/ Vomiting/ 
Anorexia 
3 
Ondansetron 10 mg, 
IV, q8h, prn; 
Granisetron 0.01 
mg/kg IV daily prn; 
Droperidol 1 mg, IV 
q4-6h, prn; 
Prochlorperazine 25 
mg pr, prn or 10 mg 
IV q6h prn 
No 
No 
Diarrhea 
3 
Loperamide 2mg, po, 
q3h, prn; 
Diphenoxylate HCl 
2.5 mg and atropine 
sulfate 25 mcg, po, 
q3h, prn; codeine 
sulfate 30-60 mg, po, 
q4h, prn 
If uncontrolled after 
24 hours despite all 
supportive measures 
No 
Malaise 
3 or 4 
Bedrest 
If other toxicities 
occur 
simultaneously 
No 
Hyperbilirubinemia 
3 or 4 
Observation 
If other toxicities 
occur 
simultaneously 
No 
Anemia 
3 or 4 
Transfusion with 
PRBCs 
If uncontrolled 
despite all 
supportive measures 
No 


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Thrombocytopenia 
3 or 4 
Transfusion with 
platelets 
If uncontrolled 
despite all 
supportive measures 
No 
Edema/Weight gain 
3 
Diuretics prn 
No 
No 
Hypotension 
3 
Fluid resuscitation 
Vasopressor support 
If uncontrolled 
despite all 
supportive measures 
No 
Dyspnea 
3 or 4 
Oxygen or 
ventilatory support 
If requires 
ventilatory support 
No 
Oliguria 
3 or 4 
Fluid boluses or 
dopamine at renal 
doses 
If uncontrolled 
despite all 
supportive measures 
No 
Increased creatinine 
3 or 4 
Observation 
Yes (grade 4) 
No 
Renal failure 
3 or 4 
Dialysis 
Yes 
Yes 
Pleural effusion 
3 
Thoracentesis 
If uncontrolled 
despite all 
supportive measures 
No 
Bowel perforation 
3 
Surgical intervention 
Yes 
Yes 
Confusion 
3 
Observation 
Yes 
No 
Somnolence 
3 or 4 
Intubation for airway 
protection 
Yes 
Yes 
Arrhythmia 
3 
Correction of fluid 
and electrolyte 
imbalances; chemical 
conversion or 
electrical conversion 
therapy 
If uncontrolled 
despite all 
supportive measures 
No 
Elevated Troponin 
levels 
3 or 4 
Observation 
Yes 
If changes in LV function 
have not improved to 
baseline by next dose 
Myocardial 
Infarction 
4 
Supportive care 
Yes 
Yes 
Elevated 
transaminases 
3 or 4 
Observation 
For grade 4 without 
liver metastases 
If changes have not 
improved to baseline by 
next dose 
Hyperbilirubinemia 
3 or 4 
Observation 
For grade 4 without 
liver metastases 
If changes have not 
improved to baseline by 
next dose 
Electrolyte 
imbalances 
3 or 4 
Electrolyte 
replacement 
If uncontrolled 
despite all 
supportive measures 
No 
Neutropenia 
4 
Observation 
No 
No 
*Unless the toxicity is not reversed within 12 hours 
** Unless the toxicity is not reversed to grade 2 or less by next treatment. 
 


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Appendix 3: Interleukin-2 toxicities observed in patients treated at the NIH Clinical Center 
 


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Appendix 4:  
 
 
Certificate of Analysis:                     
Young TIL from HPV+ Tumors 
Patient: 
Date of preparation of final product: 
Unique TIL identifier (tumor and culture number): 
 
Tests performed on final product: 
Test 
Method 
Limits 
Result 
Initials/Date 
Cell viability1 
trypan blue exclusion 
>70% 
 
 
Total viable cell 
number1 
visual microscopic 
count 
between 109 and  
2 X 1011 
 
 
Identity 
FACs  
> 80 % CD3+ on REP 
cells 
 
 
TIL potency2 
OKT3-stimulated IFN 
release  
>200 pg/ml per 105 
cells and > 2 times 
background 
 
 
Microbiological 
studies  
aerobic culture5  
no growth 
 
 
 
anaerobic culture5 
no growth 
 
 
 
gram stain1,3 
no micro-organisms 
seen 
 
 
 
aerobic culture3,4 
no growth 
 
 
 
fungal culture3,4 
no growth 
 
 
 
anaerobic culture3,4 
no growth 
 
 
 
mycoplasma test2 
negative 
 
 
Endotoxin1  
limulus assay 
#5 E.U./kg 
 
 
Presence of 
tumor cells2 
Cytopathology 
No tumor cells per 
200 cells examined 
 
 
1 Performed on the final product prior to infusion.  Results are available at the time of infusion. 
2 Performed 2 - 10 days prior to infusion (test performed prior to final manipulation).  Results are available at the 
time of infusion. 
3 Performed 2-4 days prior to infusion.  Results are available at the time of infusion but may not be definitive. 
4 Sample for test collected on the final product prior to infusion.  Results will not be available before cells are 
infused into the patient. 
5 Sample for test collected on the in process cells prior to the REP.  Results will be available before cells are infused 
into the patient. 
 
Prepared by: 
_________________________________________ 
Date:  ____________________ 
 
QC sign-off: 
__________________________________________ 
Date:  ____________________ 
Qualified laboratory or Clinical Supervisor