Exploratory Study of Early Biomarkers Allowing Dynamic Assessment of Response to Treatment in Cancers of the Head and Neck
Conditions: Head and Neck Cancer
Sponsor: Royal Marsden NHS Foundation Trust

 
 
Exploratory study of early biomarkers allowing 
Dynamic Assessment of Response to Treatment in 
cancers of the head and neck 
 
Short Title: THE DART STUDY  
 
Protocol version: 
 
 
v1.2 
Dated: 
 
 
 19.08.2022 
 
 
 
RESEARCH REFERENCE NUMBERS 
 
IRAS Number: 
292101 
 
EudraCT Number: 
 
N/A 
ISRCTN Number / Clinical 
trials.gov Number: 
 
N/A 
SPONSORS Number: 
 
 
FUNDERS Number: 
ICR/RMH BRC B106 GPCE theme 
 
 
 
 
 
 
 
 
 
 

 
 
 
The DART Study protocol v1.2 19th August 2022 
 
 
 
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SIGNATURE PAGE 
The undersigned confirm that the following protocol has been agreed and accepted and that the Chief 
Investigator agrees to conduct the trial in compliance with the approved protocol and will adhere to the principles 
outlined in the Medicines for Human Use (Clinical Trials) Regulations 2004 (SI 2004/1031), amended regulations 
(SI 2006/1928) and any subsequent amendments of the clinical trial regulations, GCP guidelines, the Sponsor’s 
SOPs, and other regulatory requirements as amended. 
I agree to ensure that the confidential information contained in this document will not be used for any other 
purpose other than the evaluation or conduct of the clinical investigation without the prior written consent of the 
Sponsor 
I also confirm that I will make the findings of the study publicly available through publication or other 
dissemination tools without any unnecessary delay and that an honest accurate and transparent account of the 
study will be given; and that any discrepancies from the study as planned in this protocol will be explained. 
 
Chief Investigator: 
Signature:  
 
...................................................................................................... 
 
Date: 
  
 19.08.2022 
Name: (please print): 
 
Dr. Ben O’Leary NIHR Academic Clinical Lecturer 
   The Institute of Cancer Research, London 
   The Royal Marsden Hospital, London 
 
 
 
 
 
 
 
 
 

 
 
 
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KEY TRIAL CONTACTS AND PROTOCOL CONTRIBUTORS 
 
Chief Investigator 
Name:     Dr Ben O’Leary  
Email:      ben.oleary@icr.ac.uk  
Phone:     
Trial Manager/ Trial email 
Name:     Jeane Guevara 
Email:     jeane.guevara@rmh.nhs.uk  
Trial Email : dart@rmh.nhs.uk       
Sponsor 
The Royal Marsden NHS Foundation Trust 
For any enquiries, please contact the trial coordinator listed above or email your 
enquiry to: gcpcomplianceteam@rmh.nhs.uk  
Funder(s) 
ICR/RMH BRC B106 GPCE theme 
Key Protocol Contributors 
Name:      Dr,Shree Bhide 
Position:   Consultant Clinical Oncologist, The Royal Marsden  
Email:      Shree.Bhide@rmh.nhs.uk  
Name:      Professor Kevin Harrington 
Position:   Consultant Clinical Oncologist, The Royal Marsden Hospital, London, 
Professor of Biological Cancer Therapies, The Institute of Cancer Research, 
London 
Email:       Kevin.harrington@icr.ac.uk  
 
 
 
TRIAL SUMMARY 
Trial Title 
Exploratory study of early biomarkers allowing Dynamic Assessment of Response to Treatment in 
cancers of the head and neck. 
Summarised Trial Design 
DART is an exploratory molecular analysis study to assess potential early biomarkers 
of treatment response in squamous cell carcinoma of the head and neck (HNSCC). 
Summarised Eligibility Criteria 
• 
Age 18 years or older. 
• 
Patients with histologically confirmed cancer of the head and neck with evidence of 
recurrent or locally advanced cancer not suitable for treatment with curative intent, 
or metastatic disease. 
• 
Ability to give informed consent for biological sample collection 
 
Planned Sample Size 
Up to 50 participants  
Follow up duration 
Up to 5 years clinical follow up for oncological outcomes 
Planned Trial Period 
Prospective identification of patients will take place over 4 years. 
Objectives 

 
 
 
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Primary 
 
Explore circulating tumour DNA dynamics in patients receiving systemic therapies for 
cancers of the head and neck 
Secondary 
 
1. 
To collect longitudinal biological samples, including blood, saliva, stool and tissue, 
for molecular profiling, including extraction of DNA for sequencing, RNA for gene 
expression analysis, expansion of PBMCs, and proteins for proteomic studies. 
2. 
To collect tumour tissue to facilitate molecular analysis of recurrent or metastatic 
cancers of the head and neck. 
3. 
To isolate live tumour and immune cells for studies of therapy resistance and 
biology in cancers of the head and neck. 
4. 
Retrieval and analysis of archival primary tissue blocks for comparison with 
metastatic tumour sites. 
5. 
To correlate assays with clinicopathological data. 
Methods 
• 
This is an exploratory molecular analysis study to provide a platform for clinical 
sample collection and novel biomarker discovery. 
• 
Prospective participants will be identified by screening H&N MDT and clinic lists at 
RMH. 
• 
Following informed consent, blood and saliva will be taken at baseline prior to the 
start of systemic therapy, and then again with each treatment cycle until 
progression. 
• 
Archival samples will be retrieved for longitudinal comparisons. 
• 
Patients undergoing biopsy as part of their usual clinical care will have tissue 
samples taken for research. 
• 
Patients may opt in if willing to an additional research biopsy at the outset of 
treatment and/or at the end of treatment.  
• 
Sequencing of tumour tissue will be used to identify genomic aberrations specific 
to a patient’s cancer 
• 
These personalised genomic changes will be tracked in the longitudinal plasma 
and saliva samples. 
• 
Clinical outcomes such as progression free and overall survival will be compared 
for patients with and without a fall in their circulating tumour DNA in the early part 
of treatment. 
 
FUNDING AND SUPPORT IN KIND 
FUNDER(S) 
FINANCIAL AND NON FINANCIALSUPPORT 
GIVEN 
ICR/RMH BRC B106 GPCE theme 
Sample collection 
The National Institute for Health Research 
Salary support to Chief Investigator 
 
ROLE OF STUDY SPONSOR AND FUNDER 
For this trial some of the duties of the sponsor have been delegated to the Chief Investigator (CI), for example 
the CI has overall responsibility for the design and development of the protocol. The sponsorship agreement 
describes the allocation of such responsibilities, and a summary of this can be provided by the sponsor upon 
request. 
 

 
 
 
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ROLES & RESPONSIBILITIES OF TRIAL MANAGEMENT COMMITTEES, GROUPS AND INDIVIDUALS  
Name 
Ben O’Leary 
Role(s) 
Chief Investigator 
Responsibilities 
Including, not limited to: 
Oversight of project design conduct and reporting 
Liaison with Research Ethics Committee (REC), and other review bodies, 
during the application process, and where necessary during, the conduct 
of the research 
Ensure adherence to protocol 
Molecular analysis 
Analysis and write up 
 
Name 
Kevin Harrington 
Role(s) 
Co-Investigator 
Responsibilities 
Including, not limited to: 
Oversight of project design, conduct and reporting  
Liaison with Research Ethics Committee (REC), and other review bodies, 
during the application process, and where necessary during, the conduct 
of the research 
Ensure adherence to protocol 
Analysis and write up 
Emergency Chief Investigator 
 
Name 
Shreerang Bhide 
Role(s) 
Co-Investigator 
Responsibilities 
Including, not limited to: 
Oversight of project design, conduct and reporting  
Liaison with Research Ethics Committee (REC), and other review bodies, 
during the application process, and where necessary during, the conduct 
of the research 
Ensure adherence to protocol 
Analysis and write up 
 
Name 
Jeane Guevara 
Role(s) 
Trial Manager 
Responsibilities 
Including, not limited to: 
Liaison with Research Ethics Committee (REC), and other review bodies, 
during the application process, and where necessary during, the conduct 
of the research. 
 

 
 
 
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LIST of CONTENTS 
GENERAL INFORMATION 
Page No. 
TITLE PAGE 
 
1 
SIGNATURE PAGE 
2 
TRIAL SUMMARY 
 
4 
FUNDING 
 
5 
ROLES & RESPONSIBILITIES OF TRIAL MANAGEMENT COMMITTEES, GROUPS AND 
INDIVIDUALS   
6 
LIST of CONTENTS 
 
7 
LIST OF ABBREVIATIONS 
8 
SECTION 
1. RATIONALE AND BACKGROUND 
 
9 
2. OBJECTIVES 
 
10 
3. ELIGUBILITY  
11 
4. TRIAL PROCEDURES 
12 
5. STORAGE AND ANALYSIS OF SAMPLES 
14 
6. STATISTICS 
17 
7. DATA HANDLING 
18 
8. TRIAL OVERSIGHT 
19 
9. ETHICAL CONSIDERATIONS 
19 
10. INDEMNITY 
20 
11.AMENDMENTS 
20 
12. DISSEMINATION POLICY 
20 
13. SAFETY REPORTING 
21 
14. REFERENCES 
21 
15. APPENDICES 
22 
 

 
 
 
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LIST OF ABBREVIATIONS 
AE 
Adverse Event 
AR 
Adverse Reaction 
CA 
Competent Authority 
CI 
Chief Investigator 
CRF 
Case Report Form 
CRO 
Contract Research Organisation 
CTA 
Clinical Trial Authorisation 
GCP 
Good Clinical Practice 
ICF 
Informed Consent Form 
ISF 
Investigator Site File 
NHS R&D 
National Health Service Research & Development   
NIMP 
Non-Investigational Medicinal Product 
PIS 
Participant Information Sheet 
PP 
Per protocol 
QA 
Quality Assurance 
QC 
Quality Control 
REC 
Research Ethics Committee 
SDV 
Source Data Verification 
SOP 
Standard Operating Procedure  
TMG 
Trial Management Group 
TMF 
Trial Master File 

 
 
 
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1. RATIONALE AND BACKGROUND INFORMATION 
Recurrent head and neck squamous cell carcinoma (HNSCC) has a very poor prognosis, with only half of 
patients surviving longer than a year after disease recurrence even with optimal treatment [1]. Introduction of 
PD-1 checkpoint inhibitors, an example of immune-oncology (IO) therapy, has improved outcomes for patients 
with advanced HNSCC. Some patients demonstrate prolonged responses, though the biology of this is not 
understood. The evolution of HNSCC on treatment with these agents is unknown and there is an urgent need to 
address this deficit to drive improved treatment strategies. 
PD-1 checkpoint inhibitors have improved the limited survival of patients with advanced HNSCC, with nivolumab 
approved in the UK for patients pre-treated with platinum chemotherapy[2]. Following results from the 
KEYNOTE-048 trial of pembrolizumab this drug is currently available in the first-line setting [1]. Available data in 
other cancers suggests that the factors determining response to IO agents are complex, with a potential role for 
tumour mutational burden[3], the clonal status of tumour neoantigens [4] with a study in HNSCC highlighting a 
role for the tumour microenvironment (TME) and immune milieu [5]. A previous longitudinal study in melanoma 
demonstrated clonal evolution on nivolumab, with evidence of neoantigen-directed immune-editing and changes 
in the T cell receptor repertoire [6], but there are no longitudinal data available for HNSCC. Due to the complexity 
of response to IO, dynamic biomarkers have been proposed as an approach that provides a patient-unique in 
vivo assessment of treatment efficacy [7].  
Circulating tumour DNA (ctDNA) can be identified in patients with a wide variety of cancers, and has been shown 
to allow early prediction of disease relapse after treatment with curative intent in HNSCC [8]. Recent data show 
that early changes in ctDNA can predict which patients will respond better to systemic therapies[9], including 
immunotherapies [10, 11] although there are no data for this yet in HNSCC. 
The DART Study will begin to address the above deficit through a longitudinal study of HNSCC patients 
receiving systemic therapy. Dense sample collection will allow a deep translational analysis of tumour evolution 
and dynamic biomarkers. An initial primary analysis of sub clonal evolution in circulating tumour DNA as an early 
treatment biomarker is planned, with the study samples also providing a sample collection platform allowing 
subsequent analyses, such as mapping the genomic landscape of metastatic HNSCC. Tissue biopsies will be 
collected at the start of systemic therapy with contemporaneous and then longitudinal plasma, and saliva.  

 
 
 
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2. OBJECTIVES  
 
2.1. Goal  
Explore early biomarkers of response to treatment in cancers of the head and neck. 
 
2.2. Primary objective 
Explore circulating tumour DNA dynamics in patients receiving systemic therapies for cancers of the head and 
neck.  
 
2.3. Secondary objectives 
 
1. To collect longitudinal biological samples, including blood, saliva, stool and tissue, for molecular 
profiling, including extraction of DNA for sequencing, RNA for gene expression analysis, expansion 
of PBMCSs, and proteins for proteomic studies. 
2. To collect tumour tissue to facilitate molecular analysis of recurrent or metastatic cancers of the 
head and neck. 
3. To isolate live tumour cells for organoid and co-culture studies of therapy resistance and biology in 
cancers of the head and neck. 
4. Retrieval and analysis of archival primary tissue blocks for comparison with metastatic tumour sites. 
5. To correlate assays with clinicopathological data. 
 
2.4. STUDY SETTING 
Participants will be recruited from the Royal Marsden Hospital (RMH), a tertiary referral H&N cancer unit in 
London, UK.  
 

 
 
 
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3. ELIGIBILITY CRITERIA 
 
3.1.  Inclusion criteria 
I. 
Age 18 years or older. 
II. 
Patients with histologically confirmed cancer of the head and neck with evidence of recurrent or 
locally advanced cancer not suitable for treatment with curative intent, or metastatic disease. 
III. 
Receiving immunotherapy 
IV. 
Ability to give informed consent for biological sample collection. 
 
3.2.  Exclusion criteria 
I. 
Unable to undergo serial sample collection 
II. 
Pregnancy 
 

 
 
 
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4. TRIAL PROCEDURES  
 
 
Study 
entry 
Baseline  
Treatment  Follow 
up 
Informed Consent 
X 
 
 
 
Date of birth 
 
X 
 
 
Demographics 
 
X 
 
 
Cancer treatment history 
 
X 
X 
X 
Height, Weight & BMI 
 
X 
X 
 
Performance status 
 
X 
X 
X 
Intervention for adverse events 
 
X 
X 
X 
Radiological Tumour assessment 
data 
 
X 
X 
X 
Blood Sample & Saliva Sample 
Drawn 
 
X 
X 
 
Next line treatment and survival 
data 
 
 
 
X 
 
 

 
 
4.1. Recruitment 
Patients undergoing systemic therapy for recurrent, metastatic, or locally advanced cancer of the head and neck 
not suitable for treatment with curative intent. This is a biological research study involving the collection of blood, 
tumour tissue, saliva, and other body fluids routinely examined during cancer care (e.g. CSF, ascites, urine, stool 
or pleural fluids). Routine clinicopathological data including treatment details will be collected in linked-
anonymised fashion for correlation and held in a secure database. 
Patients consent to serial collection of blood and saliva with every cycle of systemic therapy. In terms of tissue, 
patients may consent to research biopsies to be taken at the time of routine diagnostic biopsies, or for new 
research biopsies prior to planned treatment. Patients also consent for access to archival tissue and any future 
biopsies. At the time of disease progression, the patient may also be asked whether they consent to have an 
optional new biopsy. 
 
4.2. Patient identification 
The DART study team will screen the weekly H&N MDT meetings and clinic lists at the Royal Marsden Hospital 
(RMH). 
 
4.3.  Consent  
Prospective participants will be informed of the DART study by a member of their usual care team at RMH during 
their routine outpatient appointments for follow up of their H&N cancer. Participants will be provided with the 
Participant Information Sheet and copy of the Informed Consent Form. If they express interest in taking part, 
then consent will be obtained by a member of the DART team, or an appropriately trained delegate, detailed in 
the Delegation Log.  
Consenting procedures will conform to GCP, local and national regulations. Consent will be for collection of new 
research tissue cores, establishment of live tumour cell culture, and sequential blood and saliva collection, with 
additional collection of any excess ascitic, pleural fluid, urine, stool or CSF, in addition to retrieval and analysis of 
archival and future clinical samples. Stool can be collected at any time convenient to the patient. 
The Principal Investigator (or designee) should discuss the study with eligible patients, describing the purpose, 
research objectives and the schedule of data and clinical sample collection. Patients should be provided with the 
ethically approved patient information sheet and consent form for review and given sufficient time to consider 
participation in the study. Written consent should be obtained before any data or clinical samples are collected. 
 
4.4. Withdrawal criteria  
Patients may withdraw from the study at any time at their own request, or they may be discontinued at the 
discretion of the Principal Investigator. This can occur for the following reason: 
Patient decision  
Lost to follow-up 
Pregnancy 
Protocol violation  
Ineligibility  
Patient noncompliance  
PI decision  

 
 
 
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If the patient is withdrawn from the study the primary reason as well as the date of withdrawal will be recorded in 
the in the Excel Data Tool (combined eCRF and database). Should a patient withdraw consent for their samples 
to be used in DART their biological samples will be destroyed and tissue samples returned to the site for 
archiving. Data collected at the point of withdrawal will continue to be used by the study team.  
 
4.5. End of trial 
The sponsor will notify REC end of a clinical trial within 90 days of its completion on the last visit of the last patient 
undergoing the trial. 
 
5. STORAGE AND ANALYSIS OF SAMPLES 
5.1. Blood, saliva and other clinical fluid samples 
A 30 mL blood sample (3x 10 mL blood collection tube, e.g. Streck or other appropriate) for plasma extraction 
and collection of buffy coat and/or immune cells will be taken with each treatment cycle and at disease 
progression. Blood tests will be preferentially taken at the time of venepuncture for routine clinical samples. 
Patients will also be asked to provide a saliva sample. A stool sample will be collected at some point during 
treatment convenient for the participant, ideally before commencing. 
Blood will be centrifuged to separate plasma and buffy coat, or processed for preservation of immune cells as 
appropriate, before being labelled and stored at -80oC. Saliva samples will be stored at -80oC. Ascites, pleural, 
urine, stool or cerebrospinal fluid samples, excess to that required for diagnostic tests, will be collected in a 
sterile fashion for extraction of cell free DNA and isolation of live tumour cells if appropriate. These samples may 
be taken at study entry or in the future. 
5.2. Tumour samples 
This protocol will complement the already established RMH Standard Operating Procedures that are in place for 
the collection of core biopsies for research purposes in patients with suspicious and/or confirmed carcinoma. If 
not all sites of disease have progressed following a prior therapy, a disease site that has progressed must be 
biopsied. A total of 4 research cores should be obtained. The feasibility, appropriateness and safety of additional 
research biopsies will be made on an individual case basis and in consultation with the involved clinicians. 
 2X core biopsy (or resection specimen) will be taken with 14G core biopsy needle for formalin fixation and 
standard paraffin embedding. Sample should be placed in a 20 mL universal container containing 10% neutral 
buffered formalin and sent for embedding in paraffin to the local Histopathology Department. In consultation with 
the treating clinicians fine needle aspiration can be considered for lymph nodes where core biopsy is felt to 
represent an unacceptable risk to safety. 
If the patient has standard diagnostic formalin fixed biopsies taken at the same time as research biopsies then 
NO further research biopsies should be taken for formalin fixation, with all research biopsies taken for fresh 
tissue as below. 
2X core biopsies (or resection specimen) will be taken with 14G core biopsy needle for fresh tissue analysis. 
Samples should be placed on ice in a 1.5 mL Nunc tube. 

 
 
 
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If for technical reasons it is not possible for the patient to have 4 cores taken, then a minimum of two cores, one 
fixed in formalin and one fresh should be taken.  
5.3. Storage, labelling and postage of biological samples  
All the samples will be collected by appropriately trained staff and labelled in accordance with good clinical 
practice and local protocols. They will be stored in a way that it is easily accessible. If the patient consents to 
participate in the study, the name of the study, the study ID and the date of acquisition should also be added to 
the label. Sample storage will be in compliance with the Human Tissue Act. 
The biological samples from each patient will be transferred to the Institute of Cancer Research for further 
analysis, where samples will be stored in secure facilities. Samples labelled in an anonymised fashion with no 
identifiable features may be shared with external institutions and collaborators. Biological samples retained for 
the study will be encoded with a unique identifier and other patient identifiers will be removed prior to storage in 
order to maintain patient confidentiality. 
Laboratory researchers will not have access to any details that identify the patient in almost all circumstances. 
Certain key individuals within the DART team will be able to link the unique identifier with the patient’s 
identification details. This will allow the collection of clinicopathological data, to complement the molecular assay 
data resulting from the study. 
Patients will be asked to provide written informed consent for access to archived tumour sample excess to 
clinical requirements, and for extraction of nucleic acids for analysis. Tumour samples may be from the primary 
cancer, local recurrence, or sites of metastatic disease. Extracted nucleic acids will be stored for future analysis 
in secure facilities at the Institute of Cancer Research. All samples will be held in compliance with the Human 
Tissue Act (HTA). 
All samples should be sent by post to the following address:  
Dr Ben O’Leary 
Molecular Oncology 
Room 2S8 
237 Fulham Road, London SW3 6JB 
The Institute of Cancer Research 
 
5.4. Molecular analyses  
5.4.1. Nucleic acid extraction 
Nucleic acids including DNA and RNA will be extracted from biological samples, with buffy coat used for 
germline DNA. DNA samples will be subjected to sequencing analysis or other molecular techniques to identify 
changes relevant to cancer biology and resistance to treatment. The analysis will focus initially on the 
identification of genomic changes in the tumour which can be identified in the contemporaneous plasma samples 
as ctDNA at the outset of treatment. The levels of ctDNA will then be followed in a longitudinal manner 
throughout treatment in plasma and other biological samples. This will allow exploration of whether non-invasive 
analyses can be used in place of tissue biopsies. 

 
 
 
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5.4.2. Protein analysis 
Biological samples from the study will be analysed by immunohistochemistry, or immunofluorescence, or other 
techniques for analysis of proteins. Tissue samples may be processed for analysis of proteins using alternative 
techniques. For patients receiving immunotherapy this will included tumour infiltrating lymphocytes and PD-L1 
staining. 
 
5.4.3. Live cell analysis 
Fresh tissue sections and other biological samples may be processed in the laboratory to isolate and study live 
tumour and immune cells. This may include isolation of tumour cells, or stromal and immune cells from the 
tumour or blood, for in vitro analysis of cellular properties and response to interventions. Live tumour cells or 
tumour pieces may be introduced into immunocompromised mice in order to study the tumourigenic properties of 
these cells in vivo and study mechanisms of response and resistance of these cells to drugs in vivo. Immune 
cells will be separated from blood samples and stored for potential expansion and co-culture. 
6. STATISTICS AND DATA ANALYSIS 
6.1. Primary endpoint 
The level of circulating tumour DNA pre-treatment will be descriptively compared to the levels detected at 
subsequent time points. 
6.2. Secondary and exploratory endpoints 
This study will facilitate analyses involving cells within the laboratory. For this standard statistical tests for work of 
this type will be employed. 
 
6.3. Sample size 
This is principally a biological sample collection study aiming to assess feasibility of monitoring early changes in 
ctDNA. The initial recruitment cohort size of 50 has been chosen to ensure reasonable representation of the 
most frequently mutated genomic sub groups found in HNSCC. A sample size of 50 allows gives a 75% chance 
of including at least 4 cases of a genomic sub group with a prevalence of 10% in the HNSCC population (e.g. 
CASP8 or NSD1 mutation), using a binomial model of selection.  
For the study of live cells it is anticipated that only a low proportion of samples will grow in culture or in immuno-
compromised mice, with approximately 10%-30% of samples generating growth. This is exploratory work and 
has not been prospectively powered. 
6.4. Study duration 
The study will continue until up to 50 patients have been recruited and either died or completed 5 years of 
clinical follow up. 
 
6.5. Planned recruitment rate 
No limitation will be placed on the recruitment window. 

 
 
 
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6.6. Subject population 
Patients undergoing systemic therapy for recurrent, metastatic, or locally advanced cancer of the head and neck 
not suitable for treatment with curative intent. This is a biological research study involving the collection of blood, 
tumour tissue, saliva, and other body fluids routinely examined during cancer care (e.g. CSF, ascites, urine, stool 
or pleural fluids). Routine clinicopathological data including treatment details will be collected in linked-
anonymised fashion for correlation and held in a secure database. 
Patients consent to serial collection of blood and saliva with every cycle of systemic therapy. In terms of tissue, 
patients may consent to research biopsies to be taken at the time of routine diagnostic biopsies, or for new 
research biopsies prior to planned treatment. Patients also consent for access to archival tissue and any future 
biopsies. At the time of disease progression, the patient may also be asked whether they consent to have an 
optional new biopsy. 
 
6.7. Statistical analysis plan summary 
Descriptive statistics only are planned for the primary endpoint. 
For the secondary and exploratory endpoints standard statistical tests will be employed. Full details of all 
analyses will be described in a separate statistical analysis plan. 
 
 

 
 
 
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7. DATA HANDLING 
Data collection tools and source document identification 
7.1. Clinical data 
7.1.1. We will be collecting identifiable data as part of the DART study. This information will be held by 
the study sponsor on an excel spread sheet stored on a Trust computer in line with local data 
governance policies and GDPR. Each patient enrolled in the study will have a unique DART Study 
ID generated. 
7.1.2. The full dataset is available in an associated Excel Data Tool (combined eCRF and 
database). The dataset is divided into the following areas: 
• 
General patient details: Demographics, smoking/alcohol and co-morbidities 
• 
Cancer diagnosis: Involved sites, staging, procedures and adjuvant treatment 
• 
Previous treatment: Involved sites, staging, procedures, complications and adjuvant treatment 
• 
Outcome: Survival, recurrence, follow-up 
7.2. Source Data 
The electronic patient record will form the principal source for clinical data. Data may also be entered 
directly onto the password protected Excel Data Tool (combined eCRF and database) form where this 
information would not be collected as part of standard care.  As such the Excel Data Tool (combined 
eCRF and database) may be a source document also. 
7.3. Excel Data Tool (combined eCRF and database) 
Clinical data will be collected directly onto the password protected Excel Data Tool (combined eCRF and 
database). This will be stored in the electronic site file on the shared drive for clinical studies conducted by 
the H&N research team. In accordance with local data governance regulations for patient identifiable data.  
7.4. Access to data 
Direct access will be granted to authorised representatives from the Sponsor and the regulatory 
authorities to permit study-related monitoring, audits and inspections, in line with participant consent.  
7.5. Molecular Data 
Biological specimens will be identified only using the DART Study ID. As such, no patient identifiable data 
will be processed outside of RMH or by the ICR laboratories.  
 
7.6. Data handling and record keeping 
Information related to study participants will remain confidential and be managed in accordance with the 
Data Protection Act, NHS Caldecott Principles, The UK Policy Framework for Health and Social Care 
Research’, and the conditions of Research Ethics Committee Approval. 
All case record forms will be held in the site file, which will be kept in a locked drawer in a locked office at 
the study site. They will be destroyed after 5 years. All digital data will be stored on a password-protected 

 
 
 
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NHS computer on site at Royal Marsden Hospital, under the control of the Chief Investigator. Data will be 
erased securely after 5 years 
7.7. Access to Data 
Direct access will be granted to authorised representatives from the Sponsor and the regulatory authorities to 
permit study-related monitoring, audits and inspections, in line with participant consent.  
 
7.8. Archiving 
All study documents will be archived by the Royal Marsden Hospital NHS Foundation Trust following submission 
of the end of study report.   Archiving will be conducted in line with local guidelines. These documents will be 
stored in a location determined by the Sponsor in line with their standard operating procedures.  Any destruction 
of essential documents will require authorisation from the Royal Marsden Hospital NHS Foundation Trust. The 
minimum requirement for document retention is 5 years. 

 
 
 
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8 
TRIAL OVERSIGHT, MONITORING, INSPECTION AND AUDIT 
The Royal Marsden Hospital NHS Foundation Trust is the Sponsor of the study. There are no interventions 
requiring risk assessment. The following study-related responsibilities have been defined: 
 
• Overall responsibility: Chief Investigator (CI) 
• Monitoring study progress: Chief Investigator (CI) 
• Operational oversight: Trial Manger 
• Data collection: Lead Researchers and Co-investigators 
• Consent forms: Lead Researchers and Co-investigators 
• Data analysis & interpretation: Chief Investigator (CI) 
• Data storage: Chief Investigator (CI) / Data Manager 
• Intellectual property: Chief Investigator (CI) 
 
Sponsor and site delegation logs will record which members of staff are appropriately qualified and trained to 
conduct trial-specific activities. 
Trial Management Group  
 
A Trial Management Group (TMG) will be set up and membership will include Chef Investigator, Co-Investigators 
and Senior Trial Manager. Principal Investigators and other key study personnel will be invited to join the TMG 
as appropriate. The TMG has operational responsibility for the conduct of the trial. The TMG is responsible for 
monitoring recruitment, safety and governance of the study as well as collaborating with subsequent 
translational sub-studies. The TMG will also review any safety concerns and can convene a meeting if significant 
concerns exist. The TMG will meet regularly (at least 4 times per year) and will send updates to Co-Investigators 
(via email and/or at Investigator Meetings).  
 
9. ETHICAL CONSIDERATIONS 
Ethical approval will be obtained from the Research Ethics Committee (REC) before commencing recruitment 
and for all trial amendments. All correspondence with the REC will be retained in the Trial Master 
File/Investigator Site File.  The Chief Investigator (or delegate) will produce annual progress reports and notify 
the REC of the end of the study.  If the study is ended prematurely, the Chief Investigator will notify the REC, 
including the reasons for the premature termination. Within one year after the end of the study, the Chief 
Investigator will submit a final report with the results, including any publications/abstracts, to the REC.  The study 

 
 
 
The DART Study protocol v1.2 19th August 2022 
 
 
 
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will be conducted in accordance with the conditions of ethical approval. Local R&D Confirmation of Capacity and 
Capability will be obtained prior to recruitment of patients at collaborating centres.  
 
The protocol will be submitted for ethical review to the Health Research Authority’s ‘Integrated Research 
Application System’ (IRAS).  
The DART Study is sponsored by The Royal Marsden NHS Foundation Trust and will be approved by the 
Sponsor’s Committee for Clinical Research (CCR). The Royal Marsden NHS Foundation Trust will ensure that 
the study has received ethics approval from a research ethics committee (REC) and has received Health 
Research Authority (HRA) approval.  
 
10  
INDEMNITY 
There are no specific compensation arrangements for harmful events which might arise from participation in this 
trial. However, the study is covered for negligent claims occurring with the NHS by Crown indemnity. There is no 
pre-existing arrangement for non-negligent claims arising from the conduct of the study. 
 
 
11 
AMENDMENTS  
Amendments to the protocol or associated documentation will be made when necessary and should be agreed 
by the trial management group or equivalent committee. The CI has responsibility for preparing the amendment 
and deciding whether the amendment is substantial. All amendments will be submitted for sponsorship approval 
prior to making REC and/or HRA applications and will not be implemented until local R&D confirmation of 
capacity and capability at the research site has been granted, where appropriate. 
Amendments to the protocol will be summarised in the relevant appendix. 
 
12  
DISSEMINATION POLICY 
12.1  
Dissemination policy 
Data arising from the study are owned by the Sponsor.  Findings will be submitted for publication in relevant 
H&N or Cancer peer reviewed journals.  Funders will be acknowledged in any subsequent reports.  Authorship 
will be granted in line with criteria defined by The International Committee of Medical Journal Editors.  

 
 
 
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12.2  
Authorship eligibility guidelines and any intended use of professional writers 
The trial results will be submitted for publication in a relevant medical journal with authorship according to the 
criteria defined by the ICMJE (http://www.icmje.org). These state that: Authorship credit should be based: 
i. 
On substantial contributions to the conception or design of the work; or the acquisition, analysis, 
or interpretation of data for the work; AND  
ii. 
Drafting the work or revising it critically for important intellectual content; AND  
iii. 
Final approval of the version to be published; AND  
iv. 
Agreement to be accountable for all aspects of the work in ensuring that questions related to the 
accuracy or integrity of any part of the work are appropriately investigated and resolved.  
 
13 
SAFETY REPORTING 
All adverse events potentially related a research procedure will be reviewed by two clinicians in accordance with 
the Royal Marsden’s standard operating procedures (RM-CTU/SOP-18). Adverse events will be recorded using 
the R&D/TEM-21 Non-CTIMP SAE Report Form (other than 
Medical Devices). Adverse events should be reported as soon as possible but within a maximum of 14 calendar 
days. 
 
14 
REFERENCES 
1. 
Burtness B, Harrington KJ, Greil R, et al. Pembrolizumab alone or with chemotherapy versus cetuximab 
with chemotherapy for recurrent or metastatic squamous cell carcinoma of the head and neck (KEYNOTE-048): 
a randomised, open-label, phase 3 study. Lancet 2019;394(10212):1915-1928. 
2. 
Ferris RL, Blumenschein GJ, Fayette J, et al. Nivolumab for Recurrent Squamous-Cell Carcinoma of the 
Head and Neck. New England Journal of Medicine 2016;0(0):null. 
3. 
Hugo W, Zaretsky JM, Sun L, et al. Genomic and Transcriptomic Features of Response to Anti-PD-1 
Therapy in Metastatic Melanoma. Cell 2016;165(1):35-44. 
4. 
McGranahan N, Furness AJ, Rosenthal R, et al. Clonal neoantigens elicit T cell immunoreactivity and 
sensitivity to immune checkpoint blockade. Science 2016;351(6280):1463-9. 
5. 
Hanna GJ, Lizotte P, Cavanaugh M, et al. Frameshift events predict anti–PD-1/L1 response in head and 
neck cancer. JCI Insight 2018;3(4). 
6. 
Riaz N, Havel JJ, Makarov V, et al. Tumor and Microenvironment Evolution during Immunotherapy with 
Nivolumab. Cell 2017;171(4):934-949.e16. 
7. 
Havel JJ, Chowell D, Chan TA. The evolving landscape of biomarkers for checkpoint inhibitor 
immunotherapy. Nat Rev Cancer 2019;19(3):133-150. 
8. 
Chera BS, Kumar S, Beaty BT, et al. Rapid Clearance Profile of Plasma Circulating Tumor HPV Type 16 
DNA during Chemoradiotherapy Correlates with Disease Control in HPV-Associated Oropharyngeal Cancer. 
Clinical Cancer Research 2019; 10.1158/1078-0432.CCR-19-0211:clincanres.0211.2019. 
9. 
O’Leary B, Hrebien S, Morden JP, et al. Early circulating tumor DNA dynamics and clonal selection with 
palbociclib and fulvestrant for breast cancer. Nature Communications 2018;9(1):896. 
10. 
Forschner A, Battke F, Hadaschik D, et al. Tumor mutation burden and circulating tumor DNA in 
combined CTLA-4 and PD-1 antibody therapy in metastatic melanoma - results of a prospective biomarker 
study. J Immunother Cancer 2019;7(1):180. 

 
 
 
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11. 
Anagnostou V, Forde PM, White JR, et al. Dynamics of Tumor and Immune Responses during Immune 
Checkpoint Blockade in Non–Small Cell Lung Cancer. Cancer Research 2019;79(6):1214. 
 
 
15 
Appendix 1 – Amendment History 
Amendment 
No. 
Protocol 
version no. 
Date issued 
Author(s) of 
changes 
Details of changes made 
NSA1 
1.1 
19/08/2022 
Orla Batchelor