supplement i. study protocol example table of contents

Supplement I.

Study Protocol Example Table of Contents

BMJ Publishing Group Limited (BMJ) disclaims all liability and responsibility arising from any relianceSupplemental material placed on this supplemental material which has been supplied by the author(s) J Immunother Cancer

doi: 10.1136/jitc-2021-003212:e003212. 9 2021;J Immunother Cancer, et al. Jiang CY

Supplement II.

Authorization for Use and Release of Individually Identifiable Health Information



Supplement III.

Informed Consent Sample

INFORMED CONSENT CHECKLIST

*Complete this checklist for each consent obtained and file with the original informed consent

document*

RESEARCH STUDY IDENTIFICATION (REQUIRED)

-Study Title:

-PI Name:

-Name of Study Team Member Obtaining Consent

-Role of Study Team Member Obtaining Consent:

RESEARCH SUBJECT IDENTIFICATION (REQUIRED)

-Last Name, First Name, Middle Initial:

-Date of Birth:

-Todays Date:

A. Date all required signatures (Subject, Witness (if required by IRB), and person

obtaining consent), their printed names and the dates they signed informed

consent document have been checked and verified and appear in proper location.

B. Date and time informed consent was reviewed deemed complete and valid (must

be prior to date/time of Subject’s first study activity). C. Date and time of subject’s first study activity or involvement. Verify and Initial Each Requirement Below

1. Informed consent and HIPPA Authorization, if required by IRB was obtained for

this subject prior to study participation.

2. Appropriate institution paperwork signed by PI and approved by IRB, which

designates me (person obtaining consent) as an authorized agent of the PI and

qualified to obtain consent.

3. This prospective subject was given adequate time necessary to carefully and fully

read the informed consent document and all questions were answered to their

satisfaction.

4. All aspects of this subject’s study involvement, including purpose of study, known

and potential risks, possible benefits, and alternatives to study participation were

explained and discussed prior to subject signed consent document.

5. If required, scanned image of research enrollment form, consent form, and HIPPA

authorization were scanned into the subject’s electronic medical record. 6. A copy of the completed and signed, original informed consent document has

been issued to this subject and the subject was instructed to retain that copy for

reference and to ask any and all questions that might arise throughout their study

involvement.



7. The subject has been informed that participation is entirely voluntary and that

they may withdraw their participation at any time and for any reason.

8. The subject has been shown where in the consent document to locate study team

phone number(s).

9. Original consent documents and all copies are printed and issued as single-sided

documents and that the original signed form must be kept in the investigator’s project files on institution property.

10. Upon completion of the Informed Consent Process, this subject’s name was added to the Master List of All Subjects.

KEY INFORMATION

WHAT IS THE STUDY ABOUT AND WHY ARE WE DOING IT?

Briefly explain in this section what the study is about and the how subject’s participation will change cancer treatment in other patients.

WHAT DOES THE STUDY INVOLVE AND HOW LONG WILL IT LAST?

Discuss what the study will require the participant to do (i.e. provide blood samples). It should

be clearly stated how these samples will be obtained, when, and how often. It should be

discussed what these samples will be used for and what aspects will be measured in the blood.

The option of storing their blood should be discussed, so that future further analysis can be

done. Overall, their direct participation in the research study would be donating a blood

sample. Statement of how many years the study will last should be included and how additional

clinical information may be collected from their medical record.

WHAT ARE KEY REASONS YOU MIGHT CHOOSE TO VOLUNTEER FOR THIS STUDY?

This study is completely voluntary. There is no guarantee that you will benefit from being in this

study. If you do decide to take part in the study, you are free to change your mind at any time

and contact us to stop participating. The following may occur as a result of your participation:

1. List out ways their blood samples will help with future cancer care

2. Example: Identification of changes in genes that result or worsen cancer in Veterans.

3. Example: Pattern may be identified that affect risk of cancer

WHAT ARE KEY REASONS YOU MIGHT CHOOSE NOT TO VOLUNTEER FOR THIS STUDY?

You may not want your tissue to be sequenced (genes studied). You may also not want to

provide any more blood than what is necessary to perform your standard labs ordered by your

doctor.

DO YOU HAVE TO TAKE PART IN THE STUDY?

If you decide to take part in the study, it should be because you really want to volunteer. You

will not lose any services, benefits or rights you would normally have if you choose not to

volunteer.

WHAT IF YOU HAVE QUESTIONS, SUGGESTIONS, OR CONCERNS?



Include the name of the principal investigator along with phone number, address, and any

other additional contact information. Provide details of who to contact if the patient has

questions, suggestions, or wishes to withdraw from the study.

RESEARCH DETAILS

WHAT IS THE PURPOSE OF THIS STUDY?

Provide some background information on why the study is being pursued in order to

adequately explain the purpose and why this specific study is being pursued. Explain what the

study objectives are and how it will be achieved. For example, in lung cancer patients who are

on immunotherapy, there has been an effort to identify markers that could predict response to

therapy. One such way could be measuring specific markers in circulating tumor cells at

different time points of therapy to determine if there are any patterns to predict response. This

would help improve cancer care and outcomes in the future.

WHAT WILL HAPPEN AND WHAT CAN I EXPECT IF I TAKE PART IN THIS STUDY?

Include statement of when subjects can begin the study and how the study team will review

their electronic medical record to collect additional information on their medical history and

oncologic history. Discuss how their blood sample will be collected and where the samples are

sent for additional analysis. Explain what specifically will be tested for and when the blood

draws will occur (i.e. before treatment, every 3 months while on immunotherapy, and at time

of cancer progression). Include how much blood will be drawn each time.

If the patient opts to allow for blood banking for future analysis then include additional

information on this aspect (i.e. where the samples will be stored). Explain that no personal

identifiers will be used when the samples are sent to other places for storage/analysis. Each

participant will be given a unique code that only the study team will know. Only the research

team at the study institution will have access to clinical information. The data obtained from

the blood samples will not be used to make treatment decisions for their cancer. We will not

disclose any information from the results of these studies unless you provide consent to receive

such information. At any time, at your request, we can destroy your specimens as well as any

links to your clinical data. There is no financial gain for any members of this study regardless of

if you choose to participate or not.

WHAT POSSIBLE RISKS OR DISCOMFORTS MIGHT I HAVE IF I TAKE PART IN THIS STUDY?

The risks of simple blood draw commonly include discomfort and/or bruising at the site of the

puncture, and less commonly, the formation of a small blood clot or swelling of the surrounding

area, and/or bleeding from the puncture site. Rarely, fainting or local infection may occur.

Psychological risks include sadness regarding the reminder of cancer diagnosis. There may be

other unforeseeable risks. Despite extreme care to protect personal identifying information,

there is a very small risk of possible loss of confidentiality.

WHAT ARE THE POSSIBLE BENEFITS OF THIS STUDY?



You are not likely to benefit by participating in this study as this study will not affect your

course of treatment for your cancer. However, knowledge gained from this study may help

patients like you in the future. This may lead to new therapies or a combination of treatments

with immunotherapy to better understand the response and improve survival outcomes.

WHAT OTHER CHOICES DO I HAVE IF I DO NOT WANT TO JOINT HIS STUDY?

Participation in this study is entirely voluntary. You do not have to participate in this study. You

may drop out at any time without penalty. By doing so, you will not lose any benefits that you

may be entitled to. There will be no risks to you by not participating in this study. In case you

decide to participate and later withdraw from this study, you will not be harmed in any way.

HOW WILL MY PRIVATE INFORMATION BE PROTECTED?

We will make every effort to protect your privacy and security. All your medical information will

be stored at the research institution and will not be available to our collaborators. Your blood

samples will be coded with a number and only the study team will maintain a link to your

clinical data. Your data (clinical as well as results from your blood analysis) will be stored in an

electronic database behind institution firewall and will be password protected. Access to these

items will be automatically terminated when the study ends or in the unforeseen event of study

staff departure. If results of this study are reported in medical journals or at meetings, you will

not be identified by name, photograph, or any other means without your specific consent.

No information that can identify you will be released or published unless required by the law.

Research results from this study will not change your therapy and will not be made available to

you unless specifically asked for and provided consent.

WHAT ARE THE COSTS TO ME IF I TAKE PART IN THIS STUDY?

There will not be any costs to you for any additional care that you receive as a participant of

this research study. If you currently pay a co-pay you will continue to be responsible for the

same costs associated with having an appointment with your doctor. Every event (research

blood draw) that takes place because of this study will be at no cost to you.

WILL I BE PAID FOR PARTICIPATING IN THE STUDY?

You will not be compensated with money for participating in this study. You should do so

because you really want to participate.

DO I HAVE TO TAKE PART IN THE SUTDY?

No, participation is voluntary. If you do not want to take part in the study, there will be no

penalty or loss of benefits that you are entitled to. Data already collected prior to your

withdrawal, may continue to be reviewed for the study but no more information will be

collected from the time of withdrawal on, except for public records, such as survival data.

Specimens already used cannot be withdrawn. Your health information disclosed pursuant to

this authorization may no longer be protected by Federal laws or regulations and may be

subject to re-disclosure by the recipient. You can revoke this authorization, in writing, at any

time. To revoke your authorization, you must write to the Release of Information Office at this



facility or you can ask a member of the research team to give you a form to revoke the

authorization. Your request will be valid when the Release of Information Office receives it. If

you revoke this authorization, you will not be able to continue to participate in the study. This

will not affect your rights as a patient to treatment or benefit outside of the study.

If you revoke this authorization, the PI and their research team can continue to use information

aboutyou that was collected before receipt of the revocation. The research team will not collect

information about you after you revoke the authorization.

DOES THIS STUDY INVOLVE GENETIC RESEARCH? HOW WILL MY GENETIC INFORMATION BE

PROTECTED?

If using gene sequencing, discuss in this section how that will be done. Explain that the

sequencing done is not related to gene sequencing that discovers issues that could be passed to

future generations. Only the genetic profile of the cancer is being analyzed. The results of these

tests, as mentioned above, will not affect your current cancer treatment. The results of these

gene testing results will not be provided to you unless you specifically provide consent to do so.

In summary, the genes we are sequencing were not considered to be passed on to you from

your parents and are not considered to be mutations which can be passed onto future

generations.

OPTIONAL BLOOD STORAGE

If there is left over blood from your coded samples, we would like to store the leftovers for

blood biobanking. Include location it would be stored. Include discussion on what type of study

the additional blood could be used for. Although, this often can be hard to predict as additional

research questions could arise. Include how long the coded specimen will be stored and that it

will be destroyed after that time period by lab policies.

If you gave consent for the specimen(s) to be used in future research by the institution or its

research partners, an Institutional Review Board (IRB) will review and approve each new study.

The IRB may require that you be contacted for your consent prior to the use of the specimen(s)

in a new study if it decides such consent is required for your protection.

You have the right to withdraw your consent in the future and have your unused specimen

destroyed. You need to notify the investigator of your decision. If you decide to remove

identifiers from your specimen(s), you will not be able to withdraw your specimen later

because it cannot be linked back to you.

AGREEMENT TO PARTICIPATE IN THE RESEARCH STUDY

Dr./Mr./Ms________________________has explained the research study to me. I have been

told of the risks or discomforts and possible benefits of the study. I have been told of other



choices of treatment available to me. I have been given the chance to ask questions and obtain

answers. By signing this document below, I voluntarily consent to participate in this study “and authorize the use and disclosure of my health information for this study”. I also confirm that I have read this consent, or it has been read to me. I will receive a copy of this consent after I

sign it. A copy of this signed consent will also be put in my medical record.

I agree to participate in this research study as has been explained in this document.



Supplement IV.

Standard Operating Procedures for Plasma and Serum Processing

Materials:

• Specimen Collection Kit

• Extra vacutainer tubes

• “RNA Later”

• Refrigerated centrifuge with brake off feature

• Tube racks

• Pipettes: p1000 pipet and p200 pipet

• 9x9 freezer boxes

1. Label vacutainer tubes and transfer vials with patient unique ID, date of collection, and

specimen type.

2. Using a Vacutainer adapter and double ended needle, collect blood into 2 red top

Vacutainer tubes and 3 purple top Vacutainer tubes.

a. Red top Vacutainer tubes (serum): Must sit upright in rack at room temperature

for 30 to 60 minutes in order for clot formation to occur. Must be processed

immediately or stored at 4 degrees Celsius (only up to 4 hours)

b. Purple Vacutainer tubes (plasma): Ensure they are filled to line on label and

invert 4-6 times at 90 degrees angle to mix. Process immediately after obtaining

or store upright in rack at 4 degrees Celsius (up to 4 hours storage).

3. Ensure purple and red tops remain cold at all times while processing (either on ice or in

cold centrifuge)

a. Limit time red top tube sits separated with serum on top of the clot as this could

lead to lysis and hemolysis of serum by red blood cells. Make sure to aliquot

quickly once separated.

4. Place vacutainer tubes in centrifuge at 4 degrees Celsius for 10 minutes at 1300g (brake

off)

5. Place serum, plasma, and buffy coat transfer tubes in separate racks

6. Once centrifuge stops, take the purple Vacutainer tubes out then turn centrifuge brake

on and re-start centrifuge. The red tops will centrifuge for another 10 minutes at 4

degrees Celsius at 1300g.

7. While serum is spinning, remove plasma from 3 purple top vacutainer tubes using a

pipette into 15 ml conical tube. Make sure not to disturb buffy coats (white layer in

tube). Make sure plasma remains cold.

8. Using a pipette place buffy coat into purple transfer vial.

9. With a sterile pipette, place 1.2ml of “RNA Later” to each buffy coat transfer and place

cap on tightly.

10. Buffy coat aliquot tubes are then placed in 9x9 freezer boxes at -80 degrees Celsius

a. Make sure to record temperature and time placed in freezer at top of tube

11. Once centrifuge stops for second time, remove red vacutainer tubes and place conical

tubes in.



12. Centrifuge 15ml conical tube with plasma for 10 minutes at 4 degrees Celsius x 1300g

(brake on).

13. Using a pipette, place serum from red tops into labelled aliquot tubes.

a. 300 ul in the first 20 aliquot tubes

b. Close tubes tightly and store in 9x9 boxes at -80 degrees Celsius

14. When centrifuge stops after third spin (aka second spin for plasma), take the plasma out

and pipette into remaining tubes (blue tops).

a. Aliquot 300 ul in first 20 tubes

b. Close tubes tightly and store in 9x9 boxes at -80 degrees Celsius



Supplement V.

Standard Operating Procedure for Whole Blood Processing

Materials:

• 15 ml centrifuge tubes (sterile)

• Lymphocyte Separation Medium (LSM) (Mediatech 25-072-CV)

• Dulbecco’s PBS w/o Ca2+& Mg2+ (Gibco 14190-144 or equivalent)

• Freezing Media:

• 90% FBS (Gibco 16040-171 or equivalent)

• 10% DMSO (Sigma D-8779 or equivalent)

• 0.5 ml, 1.5 ml, and 2.0 ml cryotubes

• Standard tissue culture supplies (sterile hood, pipettors, sterile pipettes and pipette tips,

70% ethanol, etc.)

1. Make sure that the sample is recorded on the sample log sheet with the patient unique

ID and sample description.

2. For each 10 ml of blood, label one 15 ml tube and add 5 ml of Lymphocyte Separation

Medium (LSM) to the bottom of the tube.

3. Working on one patient sample at a time, spray the tops of the tubes with 70% ethanol,

then remove the caps in the T/C hood. Transfer the blood to the appropriate tubes

dropwise - (do not mix and do not touch the side of the tube with the pipet). Cap the

tube and proceed to the next sample.

4. When all of the samples have been prepared, place the tubes in the Beckman centrifuge

(make sure they are balanced). Set the speed to 1600 rpm (600x g) and the brake to

OFF; centrifuge for 30 minutes.

5. While the samples are spinning, label three 2 ml cryovials, one 0.5 ml cryovial, and two

15 ml centrifuge tubes for each patient. On the cryovials include the following:

a. Date of collection

b. Patient unique identifier

c. Sample description

6. When the centrifuge stops, remove the tubes and see that the separation has worked.

a. Without disturbing the interface, remove the upper layer of fluid into the one of

the 15 ml tubes. Try to leave no more than 1 ml behind.

b. Remove the lymphocytes (interface) to the second 15 ml tube, with minimal

carry-over of LSM. Fill the tube with PBS and cap. Invert to mix.

c. Decant the remaining LSM into the waste container. Discard the tube with the

pelleted RBC (Biohazard waste container).

7. Place the cell samples in the centrifuge and spin @ 1350 rpm for 10 minutes, with the

brake on LOW.

8. Aliquot the isolated plasma into the labelled tubes, then file in the –80 oC freezer. Make

sure to record the amount of plasma isolated on the log sheet and the plasma sample in

the running log for the appropriate freezer.



9. Remove the PBMC from the centrifuge. Decant the supernatant into the waste

container and resuspend the cells. Fill the tubes with PBS and spin down @ 1250 rpm

with the brake on HIGH. After this wash, perform a hypotonic lysis:

a. Decant and resuspend as above.

b. Slowly add 2 ml of sterile water to the resuspended cells while gently mixing.

c. Within 30 seconds, add 10 ml of PBS. If necessary, allow the debris to settle and

remove with a 1 ml pipette.

d. Centrifuge @ 1250 rpm for 10 minutes (brake on high).

10. Resuspend the cells and add 10 ml of PBS, cap and invert the tube a few times. Remove

50 ul to a 96wp. Add 50 ul trypan blue and count. The final cell harvest = (count) 0.2x106

11. Cell storage:

a. If necessary, prepare freezing medium by combining 90% AB serum and 10%

DMSO. Add to the resuspended cells and transfer to the 1.5 ml cryovials (1 ml

per vial). Place the vials in a Nalgene freezer box and place the box in the –80

degrees Celsius freezer overnight.



Supplement VI.

Sample Budget Excel Sheet



supplement i. study protocol example table of contents

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