A Phase II Study of Metformin, Doxycycline, or a Combination of Both Agents in Head and Neck Squamous Cell Carcinoma
Phase: PHASE2
Conditions: Larynx, LIP, Oral Cavity, Pharynx
Sponsor: Sidney Kimmel Comprehensive Cancer Center at Thomas Jefferson University

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THOMAS JEFFERSON UNIVERSITY 
Sidney Kimmel Cancer Center 
 
A Phase II Study of Metformin, Doxycycline, or a Combination of Both Agents in 
Head and Neck Squamous Cell Carcinoma 
 
Principal Investigator: 
Jennifer M Johnson, MD PhD 
Department of Medical Oncology 
1015 Walnut Street, Suite 700 
Philadelphia, PA 19107 
215-955-8875 
 
Co-Investigator(s): 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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Funding Sponsor: 
Departmentally Funded 
IND/IDE Holder: 
N/A 
IND/IDE Number: 
N/A 
Study Product: 
Metformin, Doxycycline 
Protocol IDs: 
JeffTrial # 9611 
CCRRC # 2016-112 
IRB Control # 16D.703 
 
Version Number: 
Version Date: 
1.0 
7.27.16 
2.0.1 
04.11.17 
3.0 
09.19.17 
4.0 
04/23/2018 
5.0 
08/15/2018 
 
CONFIDENTIAL 
This document is confidential and the property of THOMAS JEFFERSON UNIVERSITY. 
No part of it may be transmitted, reproduced, published, or used by other persons 
without prior written authorization from the study sponsor. 
 

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Table of Contents 
Signature Page .......................................................................................................................... 8 
Statement of Compliance ........................................................................................................... 8 
List of Abbreviations ................................................................................................................... 9 
Study Summary ........................................................................................................................11 
1 
Introduction .........................................................................................................................14 
1.1 
Background Information ..............................................................................................14 
1.2 
Rationale for the Proposed Study ...............................................................................20 
1.3 
Correlative Studies ......................................................................................................22 
1.4 
Potential Risks and Benefits .......................................................................................23 
1.4.1 Potential Risks.........................................................................................................23 
1.4.2 Benefits ...................................................................................................................24 
2 
Study Objectives .................................................................................................................24 
2.1 
Objectives ...................................................................................................................24 
2.1.1 Primary ....................................................................................................................24 
2.1.2 Secondary ...............................................................................................................25 
2.1.3 Exploratory ..............................................................................................................25 
2.2 
Endpoints/Outcome Measures ....................................................................................25 
2.2.1 Primary ....................................................................................................................25 
2.2.2 Secondary ...............................................................................................................25 
2.2.3 Exploratory ..............................................................................................................25 
3 
Study Design ......................................................................................................................26 
3.1 
Characteristics ............................................................................................................26 
3.2 
Number of Subjects ....................................................................................................26 
3.3 
Duration of Therapy ....................................................................................................26 
3.4 
Duration of Follow Up .................................................................................................26 
3.5 
Treatment Assignment Procedures .............................................................................26 

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3.5.1 Randomization Procedures: ....................................................................................26 
3.5.2 Masking Procedures ................................................................................................26 
3.6 
Study Timeline ............................................................................................................26 
3.6.1 Primary Completion .................................................................................................26 
3.6.2 Study Completion ....................................................................................................27 
3.7 
Substudies (if applicable) ............................................................................................27 
4 
Study Enrollment and Withdrawal .......................................................................................27 
4.1 
Eligibility Criteria .........................................................................................................28 
4.1.1 Inclusion Criteria ......................................................................................................28 
4.1.2 Exclusion Criteria ....................................................................................................28 
4.2 
Gender/Minority/Pediatric Inclusion for Research........................................................29 
4.3 
Strategies for Recruitment and Retention ...................................................................29 
4.4 
Subject Withdrawal .....................................................................................................30 
4.4.1 Reasons for Withdrawal ..........................................................................................30 
4.4.2 Handling of Subject Withdrawals or Subject Discontinuation of Study Intervention ..30 
4.5 
Premature Termination or Suspension of Study ..........................................................30 
5 
Study Intervention ...............................................................................................................30 
5.1 
Study Product .............................................................................................................31 
5.2 
Study Product Description ...........................................................................................31 
5.2.1 Acquisition ...............................................................................................................32 
5.2.2 Formulation, Packaging, and Labeling .....................................................................32 
5.2.3 Product Storage and Stability ..................................................................................32 
5.3 
Dosage, Preparation, and Administration ....................................................................33 
5.4 
Dose Modifications and Dosing Delays .......................................................................33 
5.5 
Study Product Accountability ......................................................................................34 
5.6 
Assessing Subject Compliance with Study Product Administration .............................34 
5.7 
Concomitant Medications/Treatments .........................................................................34 

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5.8 
Dietary Restrictions .....................................................................................................35 
6 
Study Schedule ..................................................................................................................35 
6.1 
Pretreatment Period/Screening ...................................................................................40 
6.2 
Enrollment/Baseline ....................................................................................................38 
6.3 
Treatment Period ........................................................................................................38 
6.4 
End of Treatment Study Procedures ...........................................................................39 
6.5 
Post-treatment/Follow-Up ...........................................................................................39 
6.6 
Long Term/Survival Follow-up ....................................................................................39 
6.7 
Withdrawal Visit/Discontinuation of Therapy ...............................................................39 
7 
Study Procedures and Evaluations .....................................................................................40 
7.1 
Study Procedures/Evaluations ....................................................................................40 
7.2 
Laboratory Procedures/Evaluations ............................................................................40 
7.2.1 Clinical Laboratory Evaluations ...............................................................................40 
7.2.2 Special Assays or Procedures .................................................................................40 
7.2.3 Specimen Preparation, Handling, and Storage ........................................................40 
7.2.4 Specimen Shipment ................................................................................................41 
8 
Evaluation of Safety ............................................................................................................41 
8.1 
Specification of Safety Parameters .............................................................................41 
8.1.1 Unanticipated Problems ..........................................................................................41 
8.1.2 Adverse Events .......................................................................................................41 
8.1.3 Serious Adverse Events ..........................................................................................42 
8.2 
Safety Assessment and Follow-Up .............................................................................42 
8.3 
Recording Adverse Events ..........................................................................................42 
8.3.1 Relationship to Study Intervention ...........................................................................42 
8.3.2 Expectedness ..........................................................................................................43 
8.3.3 Severity of Event .....................................................................................................43 
8.4 
Safety Reporting .........................................................................................................43 

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8.4.1 Unanticipated Problem Reporting to IRB .................................................................43 
8.4.1 Adverse Event Reporting to IRB ..............................................................................44 
8.4.2 Serious Adverse Event Reporting to IRB .................................................................44 
8.4.3 AE, SAE, and UAP Reporting to Funding Sponsor ..................................................44 
8.4.4 Reporting of SAEs and AEs to FDA .........................................................................44 
8.4.5 Events of Special Interest (if applicable) ..................................................................45 
8.4.6 Reporting of Pregnancy ...........................................................................................45 
8.5 
Halting Rules ..............................................................................................................45 
9 
Study Oversight ..................................................................................................................45 
10 
Clinical Site Monitoring and Auditing ..............................................................................45 
11 
Statistical Considerations ...............................................................................................45 
11.1 
Study Hypotheses .......................................................................................................46 
11.2 
Analysis Plans ............................................................................................................46 
11.3 
Interim Analyses and Stopping Rules..........................................................................46 
11.3.1 
Safety Review ......................................................................................................46 
11.3.2 
Efficacy Review ...................................................................................................47 
11.4 
Sample Size Considerations .......................................................................................47 
11.4.1 
Replacement Policy .............................................................................................47 
11.4.2 
Accrual Estimates ................................................................................................47 
12 
Source Documents and Access to Source Data/Documents ..........................................47 
13 
Quality Control and Quality Assurance ...........................................................................48 
14 
Ethics/Protection of Human Subjects ..............................................................................48 
14.1 
Ethical Standard .........................................................................................................48 
14.2 
Institutional Review Board ...........................................................................................48 
14.3 
Informed Consent Process ..........................................................................................48 
14.4 
Exclusion of Women, Minorities, and Children (Special Populations) ..........................48 
14.5 
Subject Confidentiality ................................................................................................49 

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14.6 
Future Use of Stored Specimens and Other Identifiable Data .....................................49 
15 
Data Handling and Record Keeping ...............................................................................49 
15.1 
Data Management Responsibilities .............................................................................49 
15.2 
Data Capture Methods ................................................................................................49 
15.3 
Types of Data .............................................................................................................49 
15.4 
Study Records Retention ............................................................................................50 
15.5 
Protocol Deviations .....................................................................................................50 
16 
Study Finances ...............................................................................................................50 
16.1 
Funding Source ..........................................................................................................50 
16.2 
Conflict of Interest .......................................................................................................50 
16.3 
Subject Stipends or Payments ....................................................................................51 
17 
Publication and Data Sharing Policy ...............................................................................51 
18 
Literature References .....................................................................................................51 
SUPPLEMENTAL MATERIALS ................................................................................................55 
Appendices ...............................................................................................................................55 
APPENDIX A: SCHEDULE OF EVENTS ..................................................................................55 
 

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Signature Page 
The signature below constitutes the approval of this protocol and the attachments, and 
provides the necessary assurances that this trial will be conducted according to all 
stipulations of the protocol, including all statements regarding confidentiality, and 
according to local legal and regulatory requirements and applicable US federal 
regulations and ICH guidelines. 
Principal Investigator: 
Signed: 
 
Date:  
Name: 
 
Title:  
 
Statement of Compliance 
This study will be conducted in accordance with the International Conference on Harmonisation 
guidelines for Good Clinical Practice (ICH E6), the Code of Federal Regulations on the 
Protection of Human Subjects (45 CFR Part 46), and Thomas Jefferson University research 
policies 
 

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List of Abbreviations 
AJCC 
American Joint Committee on Cancer 
AE 
Adverse Event/Adverse Experience 
AKT 
Also known as Protein Kinase B 
AMPK 
AMP-activated Protein Kinase 
ATP 
Adenosine Triphosphate 
BGAL 
Beta Galactosidase 
CAF 
Cancer Associated Fibroblast 
CAV1 
Caveolin-1 
CFR 
Code of Federal Regulations 
CIOMS 
Council for International Organizations of Medical Sciences 
CONSORT 
Consolidated Standards of Reporting Trials 
CRF 
Case Report Form 
CRO 
Contract Research Organization 
CSC 
Cancer Stem Cell 
CTCAE 
Common Terminology Criteria for Adverse Events 
DLT 
Dose Limiting Toxicity 
DSMC 
Data and Safety Monitoring Committee 
DSMP 
Data and Safety Monitoring Plan 
ECOG 
Eastern Cooperative Oncology Group  
ERK 
Extracellular Signal-Related Kinases 
ESR 
Erythrocyte Sedimentation Rate 
FDA 
Food and Drug Administration 
FWA 
Federalwide Assurance 
GCP 
Good Clinical Practice 
GPCR 
G Protein Coupled Receptor 
HIPAA 
Health Insurance Portability and Accountability Act 
HOMA 
Homeostatic Model Assessment (of Insulin Resistance) 
HNSCC 
Head and Neck Squamous Cell Carcinoma 
IB 
Investigator’s Brochure 
ICF 
Informed Consent Form 
ICH 
International Conference on Harmonisation 
IDE 
Investigational Device Exemption 
IHC 
Immunohistochemistry 

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IND 
Investigational New Drug Application 
IGF-1 
Insulin-like Growth Factor 1 
IGF-BP3 
Insulin-like Growth Factor Binding Protein 3 
IGFR 
Insulin Growth Factor Receptor 
IRB 
Institutional Review Board 
LKB1 
Liver Kinase B1 
MCT1 
Monocarboxylate Transporter 1 
MCT4 
Monocarboxylate Transporter 4 
MedDRA 
Medical Dictionary for Regulatory Activities 
MOP 
Manual of Procedures 
MSI 
Mass Spectroscopy Imaging 
mTOR 
Mammalian Target of Rapamycin 
mTORC1 
Mammalian Target of Rapamycin Complex 1 
N 
Number (typically refers to subjects) 
NCI 
National Cancer Institute 
OXPHOS 
Oxidative Phosphorylation 
PDE3B 
Phosphodiesterase 3B, CGMP-Inhibited 
PHI 
Protected Health Information 
PI 
Principal Investigator 
PI3K 
Phosphatidylinositol-4,5-bisphosphate 3-kinase 
PTEN 
Phosphatase and tensin homolog 
QA 
Quality Assurance 
QC 
Quality Control 
ROS 
Reactive Oxygen Species 
SAE 
Serious Adverse Event/Serious Adverse Experience 
SKCC 
Sidney Kimmel Cancer Center 
SOP 
Standard Operating Procedure 
TME 
Tumor Microenvironment 
TOMM20 
Transporter of Outer Mitochondrial Membrane 20 
TRAP1 
TNF receptor-associated protein 1 
TSC2 
Tuberous Sclerosis Complex Tumor Suppressor Gene 2 
TUNEL 
Terminal deoxyribonucleotidyl transferase mediated dUTP-digoxigenin nick 
end labeling 
UP 
Unanticipated Problem 

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Study Summary 
Title: 
A Phase II Study of Metformin, Doxycycline, or a Combination of Both 
Agents in Head and Neck Squamous Cell Carcinoma 
Précis: 
 
Objectives: 
 
Primary:  
 
To determine if treatment with metformin, doxycycline, or a 
combination of metformin and doxycycline can increase the 
percentage of cells that express Caveolin-1 in the cancer 
associated fibroblasts or stroma of patients with squamous cell 
carcinoma of the head and neck. 
 
 
Secondary:   
 
To determine the effect of metformin, doxycycline, or 
metformin and doxycycline treatment on the percentage of 
tumor cells that express MCT4, are TUNEL positive, express 
BGAL, MCT1 and TOMM20 in HNSCC 
 
To assess safety and tolerability of metformin, doxycycline, or 
metformin and doxycycline treatment in subjects with 
squamous cell carcinoma of the head and neck. 
 
Correlatives: 
 
To assess the effect of metformin, doxycycline, or metformin 
and doxycycline therapy on the metabolic profile of cancer 
cells and stroma using mass spectroscopy imaging (MSI) on 
paired samples, comparing metabolite profiles in the pre-
treatment and post-treatment tumor samples when sample is 
available.  
 
To assess the effect of metformin, doxycycline, or metformin 
and doxycycline therapy on oncomiR miR-21 after treatment. 
 
To assess the effect metformin, doxycycline, or metformin and 
doxycycline therapy on the metabolic state of the patient as 
characterized serologically by: adipokines and the IGF-
1/insulin signaling pathways through assessment of serum 
triglycerides, IGF-1, IGF-BP3, erythrocyte sedimentation rate, 
adiponectin, leptin, IGF-1R, exosome evaluation, 
metabolomics profile, and microRNA expression profiles and 
physiologically by performing a nutritional assessment via a 
nutritionist-mediated 3 day dietary recall and comparing a 
patient’s estimated dietary intake against their estimated 
caloric needs.  
 
Population: 
Patients aged 18 years and above. 

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Known or suspected head and neck squamous cell carcinoma  
Have a surgical resection planned as a part of their therapy  
Normal organ function including creatinine less than or equal to 
1.5mg/dL 
 
Phase: 
II 
Number of Sites: 
Thomas Jefferson University Hospital 
Description of 
Intervention: 
Metformin and doxycycline are the therapeutic agents.  The initial 
starting dose of metformin will be 500 mg orally daily for 2 days which 
will then be increased to 500 mg orally twice daily, and if tolerated, 
further increased to 1000 mg twice daily after day 6.  Subjects will 
maintain the maximum tolerated dose (up to 1000 mg twice daily) until 
the day prior to their scheduled definitive surgery. 
For doxycycline the patients will take 100 mg by mouth every twelve 
hours until the day prior to their scheduled definitive surgery. 
 
Study Duration: 
40 months 
Subject Participation 
Duration: 
From the time that patients receive their biopsy and enroll in the trial 
until their definitive surgery is between 1-5 weeks.  The patients will 
be called 30 days after surgical resection to determine if there were 
any further adverse effects of the medications given. 
Estimated Time to 
Complete 
Enrollment: 
36 months 

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Schematic of Study Design: 
 
Prior to  
Enrollment 
 
 
 
 
 
Visit 1 
Day 0 
 
 
 
 
 
 
 
Visit 2 
Anywhere 
between  
Day 7 and  
Day 35 
 
 
 
 
No patient 
visit 
 
 
 
 
 
 
 
No patient 
Visit 
 
 
Immunohistochemistry will be performed for Caveolin-1, TUNEL, BGAL, MCT1, 
MCT4, and TOMM20 on the pre- and post- treatment specimens and evaluated 
for staining distribution and intensity by two blinded pathologists.   
 
The patient will undergo their definitive surgical resection.  Two vials of blood will 
be collected.  The pill diary will be collected.  Any unused pills will be returned to 
the study coordinator.  The surgical pathology information will be recorded. 
Obtain informed consent. Screen potential subjects by inclusion and exclusion 
criteria; obtain history, document. 
Perform baseline assessments. 
Two core biopsies will be taken and two vials of blood collected.  Patients will be 
randomized to receive either metformin alone, doxycycline alone, or a 
combination of metformin and doxycycline.  Patients will be provided with 
metformin and doxycycline information as appropriate by the study coordinator 
and instructed on how to take the medications and fill out the pill diary.  The 
patients will meet with a nutritionist to fill out a dietary recall log when possible. 
Caveolin 1 IHC will be compared 
in pre- and post- treatment 
specimens 

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1 Introduction 
1.1 
Background Information 
Squamous Cell Cancer of the Head and Neck 
 
Head and neck cancers are amongst the most common cancers worldwide accounting 
for more than 550,000 patients with an estimated 46,000 of those patients in the United States.  
An estimated 8,650 of those patients in the US will die.  About 90% of those cancers are 
squamous in origin.  Head and neck squamous cell carcinomas (HNSCC) arise from the 
epithelial lining of the oral cavity, oropharynx, larynx, and hypopharynx. Treatment approaches 
are based upon staging.  For early stage tumors and for those with local involvement of lymph 
nodes, resection is the cornerstone of therapy.  Approximately 50% of head and neck cancer 
patients undergo primary surgery in the treatment of their malignancy.  For patients with 
advanced disease or adverse pathologic features, adjuvant therapy with either radiation therapy 
or concurrent chemoradiotherapy is used to decrease recurrence rates. Even with adjuvant 
treatment, 2-year disease free survival can still be as low as 66%, highlighting a need to better 
understand the biology of these tumors. 
 
The Metabolic Microenvironment as a Potential  
 
Recent studies have contributed significantly to our understanding of the tumor 
microenvironment and the importance of the interaction between tumor epithelial cells and their 
surrounding stromal fibroblasts.  Epithelial cancer cells are capable of converting the 
surrounding stroma in to “cancer associated fibroblasts” which generate lactate and pyruvate via 
aerobic glycolysis and secrete these energy sources.  These nutrients can then be incorporated 
directly into the tricarboxylic acid cycle allowing for high efficiency adenosine triphosphate (ATP) 
generation, and therefore higher proliferative capacity within the cancer cells.  This effectively 
generates a nutrient-rich environment in which the cancer cells can thrive. 
 
Coupled with this evolving understanding of tumor metabolism is an evolving view of 
carcinogenesis and tumor progression in HNSCC.  The high rate of local recurrence in HNSCC 
has led to the notion of “condemned mucosa” or “field cancerization.”  Molecular abnormalities 
are present in epithelial cells beyond the margin of microscopically detectable carcinoma cells 
and this mediates, at least in part, field cancerization. We now also recognize that the field at 
high risk of cancer recurrences is the tumor microenvironment (TME) which contains non-
cancerous epithelium, stromal cells such as fibroblasts, immune cells and their extracellular 
matrix. Our work focuses on the metabolic microenvironment of HNSCC, seeking to better 
characterize the interaction of tumor cells with the surrounding environmental cells as well as 
the overall metabolic state of the patient.   
 
Specifically, we are looking at the effect of two agents - metformin and doxycycline - on 
the metabolic microenvironment of HNSCC. 

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Metformin and Its Potential as an Antineoplastic Agent 
 
Metformin is a biguanide that is best known for its use in the treatment of type II 
diabetes. Extensive preclinical data support the effectiveness of metformin as an antineoplastic 
agent[13].  In head and neck cancer specifically, metformin inhibits proliferation of carcinoma 
cells and induces apoptosis in vitro and in vivo, in addition to reducing colony formation with cell 
cycle arrest in vitro [14].  In vitro and in vivo animal models have also shown that metformin can 
prevent the conversion of premalignant oral lesions to squamous cell carcinoma.  Metformin has 
been shown to reduce the size and numbers of oral tumors in mice models treated with 4NQO 
and halt the progression of potential premalignant lesions[15]. This study, by Vitale-Cross et. al. 
not only showed a decrease in the total number of oral lesions when treated with metformin 
compared to control, but also showed almost complete absence of transformation to squamous 
cell carcinoma.  Retrospective analyses, while limited due to the number of confounders, have 
also shown a potential for decreased in cadence of invasive squamous cell carcinoma of the 
head and neck in patients taking metformin [16, 17].   
 
Mechanism of Action for Metformin: 
 
These observations that metformin may possess antineoplastic properties has 
stimulated interest in its molecular mechanisms.  While the precise mechanisms are still an 
active area of research, overall metformin works by inducing energetic stress.   
 
One proposal for how this is accomplished is by a direct inhibitory effect on 
mitochondrial complex I, thus blocking mitochondrial-dependent production of reactive oxygen 
species (ROS) and adenosine triphosphate (ATP)[18-20].  The decrease in ATP production 
results in the activation of the liver kinase B1 (LKB1) – adenosine monophosphate-activated 
protein kinase (AMPK) signaling pathway[18, 21].  Activation of this pathway usually occurs 
during times of hypoxia and nutrient deprivation and reciprocally it can be suppressed in times 
of over nutrition and hyperglycemia.  AMPK is a key energy sensor that regulates metabolism in 
an attempt to maintain energy homeostasis[22].  The end result of blocking the LKB1-AMPK 
signaling pathway is a down-regulation of energy consuming biosynthetic processes including 
gluconeogenesis, protein and fatty acid synthesis and cholesterol biosynthesis, and promotion 
of catabolic processes such as fatty acid beta oxidation and glycolysis[23].  Metformin may also 
have activity that is independent of LKB1.  In LKB1-deficient cells metformin is still able to affect 
the intracellular energy state [24].   
 
 

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Figure 1.  The proposed mechanism of action of metformin in the cell [25]. 
 
 
Another central signaling molecule impacted by metformin is mammalian target of 
rapamycin (AMPK).  In mouse embryonal fibroblasts and several cancer cell lines, AMPK 
activation by metformin leads to inhibition of mTOR and inhibition of proliferation [24, 26, 27]. 
 
In addition to the effects of metformin and AMPK on metabolic processes, activation of 
AMPK results in rapid inhibition of cellular protein synthesis and growth.  Mechanistically, AMPK 
achieves this by phosphorylation and stabilization of the protein product of the tuberous 
sclerosis complex tumor suppressor gene 2 (TSC2), which serves as an integrator of various 
regulatory inputs implicated in cell growth and transmits them to the master regulator of cellular 
protein synthesis, mTOR.  In addition to mediating the inhibitory effects of AMPK on protein 
synthesis, TSC2 integrates several regulatory inputs that affect cellular protein translation, 
notably signals emanating from the availability of oxygen and growth factor– dependent 
stimulation of the phosphoinositide 3-kinase(PI3K)/phosphatase and tensin homolog(PTEN)/v-
akt murine thymoma viral oncogene homolog (AKT) and the Ras/Raf/extracellular signal-
regulated kinase (TSC2) pathways, two of the most frequently deregulated signaling cascades 
in human cancer.  Metformin has been shown to selectively target stem cells and to have 
synergistic properties with doxorubicin, paclitaxel and carboplatin allowing for conventional 
chemotherapy dose reductions in several carcinoma subtypes[26, 28]. Rozengurt et al [29] 
identified crosstalk between insulin/insulin-like growth factors (IGF-1) receptors and G protein-
coupled receptors (GPCR) signaling systems in pancreatic cancer cells leading to enhanced 
signaling, DNA synthesis, and proliferation.  This crosstalk was dependent on mammalian target 
of rapamycin complex 1 (mTORC1 ). They were able to use metformin, which negatively 
regulates mTORC1, and disrupt crosstalk between insulin/IGF-1 receptors and GPCR signaling 
inhibiting growth of pancreatic cancer cell lines in xenograft models.   Therefore, metformin 
decreases oxidative phosphorylation (OXPHOS) metabolism and generation of reactive oxygen 
species[18] [19, 20].  
 
Epidemiologic Data for Metformin’s Efficacy: 
 
Retrospective studies have shown that diabetics treated with metformin have a cancer 
risk reduction of approximately 40% compared to diabetics not treated with metformin[31, 32].  

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Other studies have also shown a reduction in the frequency of cancer with metformin use [33].  
Evans et al. [34] reported that the risk of subsequent cancer diagnosis was reduced in patients 
with type II diabetes who received metformin (with an odds ratio of 0.85 for any metformin 
exposure versus no metformin exposure).  The protective effect increased with greater 
metformin exposure (measured as total dose prescribed or total duration of use).  
 
Also, there is differential expression of the main transporter of metformin into cells 
between oral premalignant and malignant lesions and head and neck cancer has very high 
expression of the main transporter of metformin[35]. 
 
Current evidence from epidemiologic studies suggests that metformin has clinical activity 
in breast cancer.  Hadad et al. [36] demonstrated biomarker evidence for anti-proliferative 
effects of metformin in women with breast cancer by decreasing Ki67 and messenger RNA 
expression for PDE3B (critical regulator of cAMP levels that affect activation of AMPK).  
Similarly, Niraula et al.[37] showed short-term preoperative metformin with a dosing schedule of 
500mg three times daily was well tolerated and resulted in clinical and cellular changes 
consistent with beneficial anti-cancer effects with increased insulin sensitivity by HOMA in 
subjects and decreased proliferation and increased apoptosis in carcinoma cells. 
 
There are currently multiple completed and on-going clinical trials evaluating the effect of 
metformin in combination with standard treatment of a variety of malignancies including breast, 
colorectal, pancreatic, lung, gynecologic, and prostate cancer[36, 38, 39].  There is one phase II 
study accruing subjects using paclitaxel plus metformin up to 2500 mg a day or placebo in 
recurrent or metastatic head and neck cancer [40].  
 
Doxycycline and Its Potential as an Antineoplastic Agent: 
 
Other agents are also able to affect mitochondria.  In contrast to the mechanism of 
action of metformin, doxycycline inhibits mitochondrial translation and mitochondrial OXPHOS. 
The translation machinery of mitochondria is distinct from that found in the cytosol and in 
bacteria although there are also similarities which make it a promising anticancer drug target. 
Eukaryotic cells originate from a merger of two formerly independent cells—the host cell and the 
α-proteobacteria, which is a precursor of mitochondria. Each one of these cells contributed a 
protein synthesis system and hence eukaryotic cells have cytoplasmic and mitochondrial protein 
translation systems [43]. Host cell translation, which occurs in the cytosol, synthesizes almost all 
cellular proteins, including most mitochondrial proteins. However, 13 key subunits of 
mitochondrial OXPHOS are mitochondrially translated. Up-regulation of mitochondrial 
translation occurs with the development of cancer in a subset of human malignancies [44]. 
Inhibition of mitochondrial protein translation also has anticancer activity. 
 
 
Human mitochondrial ribosomes (mitoribosomes) are composed of a large 39S 
ribosomal subunit and a small 28S subunit, and by contrast, are highly specialized for the 
synthesis of 13 membrane proteins in humans that function in energy production 
[45].  Advances in high-resolution cryo–electron microscopy have proven that the molecular 

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structure of mitoribosomes differs dramatically from the “canonical” cytosolic ribosome of 
bacteria and eukaryotes [44, 46]. Also, the structures of mitoribosomes from different species 
are dramatically different [45]. Human mitoribosomes have increased protein mass compared to 
bacterial ribosomes, which results in a ribosome with a more extensive protein-protein network 
and an rRNA core better shielded from ROS. This has important implications to develop 
anticancer drugs that specifically target mitoribosomes [46]. The human small mitochondrial 28S 
ribosomal subunit shares features with the bacterial 30S subunit. As detailed atomic structures 
of human mitochondrial ribosomes are obtained, this should allow the rational design of 
compounds that specifically block mitochondrial ribosome activity. 
 
Doxycycline, which inhibits mitochondrial ribosome function, eradicates tumors and 
specifically cancer stem cells or CSCs, across many different tumor subtypes including breast 
cancer [47]. Clinical trials are ongoing to determine if tetracyclines can be repurposed as 
anticancer agents.  
 
Mechanism of Action of Doxycycline 
 
 
Doxycycline is a tetracycline antibiotic commonly used to treat non-gonococcal urethritis, 
respiratory tract infections, and acne vulgaris among other bacterial infections [69]. It is 
generally bacteriostatic and affects both gram-positive and gram-negative bacteria by 
transportation into the cell via passive diffusion or through an energy-dependent active transport 
system. Doxycycline is relatively more lipophilic than other tetracyclines, allowing easy passage 
through the bacterial lipid bilayer into the cytosol, where reversible binding to the 30S ribosomal 
subunit occurs; this ultimately inhibits bacterial protein synthesis. High concentrations of 
antibiotic can also interfere with protein synthesis in mammalian cells, but these cells lack the 
active transport systems found in bacteria, making eukaryotic cells less vulnerable targets 
(Clinical Key). 
            Given that mitochondria have a bacterial ancestry and that the 30S bacterial ribosomal 
subunit is homologous to the 28S mitochondrial subunit [70], these antibiotics also target 
mitochondrial translation and impair mitochondrial function. Doxycycline in particular disturbs 
mitochondrial protein synthesis and metabolic activity while altering gene expression [71]. Even 
at low concentrations, it has been shown to increase glycolytic metabolism and markedly reduce 
mitochondrial oxygen consumption [72]. The crystalline structure of mitochondrial proteins also 
plays an important role in the efficacy of various drugs on altering mitochondrial metabolism. 
Lee et al. reported that the target protein in cancer cell mitochondria is TNF receptor-associated 
protein 1 (TRAP1), and TRAP1 inhibitor efficacy depends on the degree of fit between the 
crystalline structure of TRAP1 and the structural conformation of the drug in question. This 
effect of tetracyclines on mitochondrial metabolism is highly relevant for cancer research. 
            Doxycycline, with its long half-life systemically, has recently become an attractive anti-
neoplastic agent. It also encourages the growth of normal stem cells, has anti-inflammatory 
properties, and even increases lifespan, in certain experimental contexts [73, 74]. In tumor 
xenografts and other animal models, doxycycline significantly reduces tumor burden and 

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metastatic cancer cell growth [75]. Further studies are needed to elucidate doxycycline’s anti-
neoplastic properties and support its clinical application in cancer treatment. 
 
Combination of Metformin and Doxycycline for Treatment: 
 
 
Metformin and doxycycline work through independent mechanisms to block 
mitochondrial metabolism.  We hypothesize that these two FDA-approved medications will be 
able to provide an additive effect when used in combination to alter the cellular metabolism of 
cancer cells and their associated stromal fibroblasts. 
 
Preclinical Data in the Martinez Laboratory: 
 
We have shown that head and neck squamous cell carcinomas have high mitochondrial 
OXPHOS metabolism in highly proliferative cells[30].  Also, there is high Monocarboxylate 
Transporter 4 (MCT4) expression in HNSCC cancer associated fibroblasts (CAFs) and in 
carcinoma cells with low proliferation rates[30]. MCT4 expression is a marker of pseudohypoxia, 
oxidative stress and enhanced glycolytic metabolism. We have demonstrated that there is 
metabolic coupling between highly proliferative carcinoma cells with high OXPHOS metabolism 
and low proliferative carcinoma cells and fibroblasts[30]. 
 
We interrogated HNSCC specimens (N=12) to examine if different metabolic and 
oxidative stress compartments co-exist in human tumors. A panel of biomarkers 
(Ki67/TOMM20/COX/MCT1/MCT4) was employed to visualize metabolic compartmentalization.  
Three metabolic compartments were delineated: 1) proliferative and mitochondrial-rich 
carcinoma cells (Ki67+/TOMM20+/COX+), 2) non-proliferative and mitochondrial-poor 
carcinoma cells (Ki67-/TOMM20-/COX-), and 3) non-proliferative and mitochondrial-poor tumor 
stroma (Ki67-/TOMM20-/COX-). High oxidative stress (MCT4+) was specific for cancer tissues. 
 
With this data, we then evaluated its prognostic value in a second cohort (N=40). We 
found that oxidative stress (MCT4+) in non-proliferating carcinoma cells predicted poor clinical 
outcome (p < 0.001), and was functionally associated with PET-avidity (p <0.04).  Similarly, 
oxidative stress (MCT4+) in tumor stromal cells was associated with higher tumor stage (p < 
0.04).  Leading us to propose that oxidative stress is a hallmark of tumor tissues and fuels 
mitochondrial metabolism in proliferating mitochondrial-rich cancer cells, via paracrine energy 
transfer of mitochondrial fuels (such as L-lactate and ketone bodies). 
 
With this, we were able to show for the first time that metabolic and oxidative stress 
compartmentalization exists in HNSCC mucosa with highly proliferative epithelial cancer cells 
having high mitochondrial metabolism with lactate and ketone body uptake while other 
carcinoma cells and CAFs have low mitochondrial metabolism with high lactate and ketone 
body generation and high oxidative stress.  This metabolic and oxidative stress 
compartmentalization of HNSCC mucosa shares similarities with that of normal mucosa and 
likely drives proliferation via OXPHOS metabolism.  MCT4 is an oxidative stress marker and 
may be a marker of CAFs since it is always absent in normal fibroblasts while it is found in the 

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majority of CAFs.  A subgroup of HNSCC has high MCT4 expression in a large carcinoma 
compartment and this subtype is associated with a poor prognosis.  
 
We have studied a cohort of subjects with head and neck cancer who are either human 
papilloma virus positive or negative (HPV+ or HPV-) and have determined that high fibroblast 
MCT4 expression occurs in the majority of cases and TOMM20, which is a biomarker of 
mitochondrial metabolism, is highly expressed in proliferating carcinoma cells. This type of 
metabolic compartmentalization is present irrespective of whether the samples are HPV+ or 
HPV-. 
 
Caveolin-1 or CAV1 is the major structural protein of the caveolae and works as a 
scaffolding molecule for multiple signaling proteins.  Prior work has demonstrated that this 
protein can induce the invasive phenotype in some solid tumor types.  In head and neck cancer, 
it has also already been shown that high expression levels in the primary tumor are associated 
with lymph node metastasis (PMID222133373).  Loss of CAV1 expression in CAFs is mediated 
by oxidative stress, autophagy, and glycolysis. In our pilot trial of metformin treatment in 
HNSCC so far a total of 37 pre- and post-metformin exposure samples have been analyzed 
showing that exposure to metformin alone can increase the percent of samples showing CAV1 
expression in CAFs from an average of 17% to 65% of the samples tested. 
 
Likewise, loss of Beta-galactosidase (BGAL) in tumor stroma is a marker of cancer-
associated fibroblasts. In our pilot trial of metformin in HNSCC, stromal BGAL was significantly 
increased from pre-treatment to post-treatment specimens (p=0.006) as scored by our head and 
neck pathologist. 
 
TUNEL (terminal deoxyribonucleotidyl transferase mediated dUTP-digoxigenin nick end 
labeling) assay is used as a marker of cellular apoptosis which leads to cell death. In our pilot 
trial of metformin treatment in HNSCC, 38 pre- and post-metformin exposure samples have 
shown significantly increased (p<0.001) TUNEL staining after exposure to metformin.  
 
1.2 
Rationale for the Proposed Study 
Metformin, a widely used anti-diabetic drug, which functions as a mitochondrial inhibitor, 
can also be used to selectively target cancer stem cells. Metformin functionally inhibits 
OXPHOS by targeting complex I of the electron transport chain and can even induce lactic 
acidosis, as a lethal side effect [76]. As a result, the use of antibiotics, such as doxycycline, may 
provide a safer and far more effective alternative to anti-cancer therapy with high metformin 
doses. Thus, future clinical trials for testing the efficacy of mitochondrial-targeted antibiotics in 
multiple cancer types are now clearly clinically warranted. 
Doxycycline is a broad-spectrum antibiotic that is commonly used for the treatment of 
many bacterial infections, and functions as an inhibitor of protein synthesis in bacteria. 
Doxycycline also encourages the growth of normal stem cells, has anti-inflammatory properties, 
and even increases lifespan, in certain experimental contexts [77,78]. Thus, the toxic side 
effects of anti-cancer therapy would be minimized. Doxycycline has also been used in human 
tumor xenografts and other animal models to significantly reduce tumor burden and even 

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metastatic cancer cell growth [79-82]. For example, in pancreatic tumor xenografts (with PANC-
1 cells), doxycycline treatment reduced tumor growth by ~80% [83]. In a xenograft model of 
breast cancer bone metastasis (with MDA-MB-231 cells), doxycycline treatment reduced bone 
and bone-associated soft-tissue tumor mass by >60% and ~80%, respectively [84]. 
Overall, we hypothesize that either the use of metformin or doxycycline alone or a 
combination of metformin and doxycycline will target mitochondrial pathways and that 
the combination may possibly be synergistic in impairing mitochondrial function that 
could lead to rescue of CAV1 expression in CAFs and tumor cell death. 
 
Rationale for Dosage/Route of Administration 
Metformin will be administered orally since this is the route of administration currently 
approved by the FDA. The drug will be initiated at a dose of 500mg daily with dose escalation 
every 3 days to a goal of 1000mg twice daily to be continued until the day prior to definitive 
surgery. There are currently multiple studies on-going using doses from between 500 mg twice 
daily up to 2500 mg per day in the treatment arms.  There are also studies using the extended 
release form for a dose of 1500 mg daily. We have chosen our starting dose and escalation 
regimen to minimize side effects.  The chosen standing dose is based on metformin’s 
therapeutic range (minimal therapeutic dose in diabetic patients is 1500-2000 mg a day) [85, 
86]. The minimum time of planned exposure to metformin will be 7 days and the maximum 
planned exposure will be 35 days (average time at TJUH from cancer diagnosis by biopsy to 
definitive surgical treatment is 3 weeks). We will allow a window of 1-5 weeks in the event that 
there are delays in the surgical scheduling but no patients will receive metformin for more than 
35 days. 
The usual dose of oral doxycycline is 200 mg on the first day of treatment (administered 
100 mg every 12 hours) followed by a maintenance dose of 100 mg/day. The maintenance dose 
may be administered as a single dose or as 50 mg every 12 hours. In the management of more 
severe infections (particularly chronic infections of the urinary tract), 100 mg every 12 hours is 
recommended. The study will be administering 200 mg of doxycycline, given as 100 mg every 
12 hours until the day prior to their scheduled definitive surgery.  
Rationale for endpoints 
Primary: We have chosen caveolin-1 (CAV1) expression in tumor stroma as our primary 
endpoint based on studies revealing that metabolic coupling modulates tumor growth in animal 
models, is a prognostic biomarker and preliminary analyses of a pilot study conducted in head 
and neck squamous cell carcinoma.  Immunohistochemistry for CAV1 in the tumor-associated 
stroma will be compared in the pre-treatment specimen and the post-treatment specimens.  All 
stained specimens will be independently viewed and scored by two pathologists who are 
blinded as to the nature of the samples and specifically as to whether the biopsy is pre or post-
metformin/doxycycline treatment as previously described by the investigators (68). The stained 
sections will be scored taking into consideration the intensity of staining and the percentage of 
stained cells within each tissue section.  

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Secondary: We have chosen TUNEL, BGAL, MCT1, MCT4 and TOMM20 as secondary end-
points of the current clinical trial due to their biologic and prognostic significance as seen in our 
work in head and neck squamous cell carcinoma. CAV1, BGAL, and MCT4 expression 
modulate tumor growth in animal models, they are cancer prognostic biomarkers, and 
preliminary analyses of a pilot study conducted in HNSCC show that metformin modulates 
CAV1, BGAL and TUNEL expression. MCT4 expression in fibroblasts is associated with 
advanced stage disease.  MCT1 and TOMM20 expression in carcinoma cells is associated with 
increased functional mitochondrial mass.  
Correlative: Mass spectroscopy imaging (MSI) will be performed on paired samples comparing 
metabolite profiles including ATP, hexose bisphosphates, lactate and pyruvate in the pre-
treatment and post-treatment tumor sample. MSI will allow spatial characterization of the 
metabolic profile of carcinoma and stromal cells within the tumor. MSI will also allow us to 
determine if distance between carcinoma and stromal cells is associated with different 
metabolite profiles. MSI testing will be performed at Vanderbilt University as has been 
performed in the ongoing metformin HNSCC clinical trial. This testing is performed outside of 
the KCC because we lack the facilities and expertise for MSI. We will also send tumor samples 
from our tumor bank, from patients who have not taken metformin or doxycycline, to use as 
controls. We will also further characterize the baseline metabolic state of each patient by 
performing a nutritional assessment via a nutritionist-mediated 3-day dietary recall and 
comparing a patient’s estimated dietary intake against their estimated caloric needs calculated 
with the Harris-Benedict equation.  This will determine if patients are already calorically 
restricted, a phenotype similar to that of mitochondrial blockade.  Similarly, we will perform a 
serologic assessment at the time of enrollment and at the time of resection using exosomes, 
metabolomics, and microRNA expression profiles. 
1.3 
Correlative Studies 
Mass spectroscopy imaging (MSI) will be performed on paired samples, comparing relative 
proportion of metabolite profiles (specifically ATP, hexose bisphosphates, lactate and pyruvate) 
in the pre-treatment and post-treatment tumor sample. MSI allows assessment of metabolite 
levels in spatially defined tumor regions [87] and hence will allow the evaluation of metabolite 
levels in carcinoma and stromal cell compartments. 
To begin to estimate the patient’s baseline metabolic state, we will also performing a nutritional 
assessment via a nutritionist-mediated 3 day dietary recall and comparing a patient’s estimated 
dietary intake against their estimated caloric needs calculated with the Harris-Benedict equation.  
Patients will be characterized as not meeting, meeting, or exceeding their caloric needs.     
A serologic profile will be built using exosomes and metabolomics, and we will seek to correlate 
this with the patient’s CAV1 expression levels pre- and post-treatment. 
We will also further characterize the baseline metabolic state of the patient by performing a 
nutritional assessment via a nutritionist-mediated 3-day dietary recall and comparing a patient’s 
estimated dietary intake against their estimated caloric needs calculated with the Harris-
Benedict equation.  This will determine if patients are already calorically restricted, a phenotype 
similar to that of mitochondrial blockade.   
 

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1.4 
Potential Risks and Benefits 
1.4.1 
Potential Risks 
 
Metformin- Risks 
Metformin’s most serious toxicity is lactic acidosis, occurring in three of 100,000 patient-
years of use. Risk is significantly reduced when metformin use is avoided in those patients with 
hepatic, cardiac, or renal compromise. However, metformin’s risk of lactic acidosis may be 
overstated since the recent evaluation of metformin associated lactic acidosis cases from 347 
trials showed that the risk of lactic acidosis with metformin was not significantly increased 
compared with other antiglycemic agents [88]. A common side effect of Metformin is diarrhea. 
Minor gastrointestinal upset is the most common toxicity, leading to cessation of therapy in less 
than 5% of individuals. Metformin does not induce hypoglycemia. While metformin alone is 
rarely associated with low blood sugars (hypoglycemia), it has been reported in cases where 
people are fasting or undergoing strenuous physical activity.  It can also cause low blood sugars 
in patients with diabetes who are taking other medications to manage their blood sugars such 
as sulfonylureas 
Doxycycline- Risks 
Due to oral doxycycline’s virtually complete absorption, side effects to the lower bowel, 
particularly diarrhea, have been infrequent. The following adverse reactions have been 
observed in patients receiving tetracyclines:  
Gastrointestinal: Anorexia, nausea, vomiting, diarrhea, glossitis, dysphagia, enterocolitis, and 
inflammatory lesions (with monilial overgrowth) in the anogenital region. Hepatotoxicity has 
been reported rarely. These reactions have been caused by both the oral and parenteral 
administration of tetracyclines. Rare instances of esophagitis and esophageal ulcerations have 
been reported in patients receiving capsule and tablet forms of drugs in the tetracycline class.  
Skin: Maculopapular and erythematous rashes. Exfoliative dermatitis has been reported but is 
uncommon.  
Hypersensitivity reactions: Urticaria, angioneurotic edema, anaphylaxis, anaphylactoid purpura, 
serum sickness, pericarditis, and exacerbation of systemic lupus erythematosus.  
Thyroid Gland Changes: When given over prolonged periods, tetracyclines have been reported 
to produce brown- black microscopic discoloration of thyroid glands. No abnormalities of thyroid 
function are known to occur.  
Combination of these Agents: 
No drug-drug interactions are known to occur between these two agents.  Metformin is 
not metabolized and is excreted unchanged in the urine with a half-life of approximately 5 hours.  
Similarly, doxycycline is not significantly metabolized and no metabolites have been found in 
man.  Some studies have suggested a potential hepatic metabolism pathway.  However, no 

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excess toxicity is expected from the combination of these two agents and thus no additive 
potential harm beyond those associated with the individual agents as described above. 
1.4.2 
Benefits 
Metformin- Benefits 
The possible societal benefits are large since this will allow us to learn about the 
pharmacodynamic effects of metformin and doxycycline in HNSCC.  Information about the 
expression levels of CAV1, TUNEL, BGAL, MCT4, MCT1, TOMM20, and the metabolomics 
described above will also more broadly allow us to gain new information on tumor metabolism.  
The study is designed to study as a primary end-point metformin and doxycycline’s effects on 
expression of CAV1, but not to study if this combination of agents improves outcomes as a 
primary end-point and hence it is unlikely to have significant clinical benefits for subjects.  
Doxycycline- Benefits 
Doxycycline is relatively attractive as a new anti-cancer agent, as it has a long half-life 
systemically and has been used successfully for the long-term treatment of patients with urinary 
tract infections (UTI), prostatitis or acne, for extended periods of time, of up to 4-to-6 months or 
more (200 mg per day). Doxycycline also encourages the growth of normal stem cells, has anti-
inflammatory properties, and even increases lifespan, in certain experimental contexts.  Thus, 
the toxic side effects of anti-cancer therapy would be minimized. 
Other Benefits 
 
Patients in this trial will each meet with a certified oncology nutritionist, whenever 
possible, who will review their dietary choices.   
 
2 Study Objectives 
2.1 
Objectives 
2.1.1 
Primary 
 
To determine if treatment with metformin, doxycycline, or a combination of metformin 
and doxycycline can increase the percentage of stromal cells that express CAV1 in 
patients with squamous cell carcinoma of the head and neck. 
 
2.1.2 
Secondary 
 
To determine the effect of metformin, doxycycline, or metformin and doxycycline 
treatment on the percentage of tumor cells that are apoptotic as determined by the  
TUNEL assay, and express MCT4, MCT1, BGAL, and TOMM20 in squamous carcinoma  
of head and neck region tumor cells. 
 
To assess safety and tolerability of metformin, doxycycline, or metformin and 
doxycycline treatment in subjects with squamous cell carcinoma of the head and neck. 

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2.1.3 
Exploratory 
 
To assess the effect of metformin, doxycycline, or metformin and doxycycline therapy on 
the metabolic profile of cancer cells and stroma using mass spectroscopy imaging (MSI) 
on paired samples, comparing metabolite profiles in the pre-treatment and post-
treatment tumor samples.  
 
To assess the effect of metformin, doxycycline, or metformin and doxycycline therapy on 
the metabolic state of the patient as characterized serologically by: erythrocyte 
sedimentation rate, exosome evaluation, metabolomics profile, and microRNA 
expression profiles and physiologically by performing a nutritional assessment via a 
nutritionist-mediated 3-day dietary recall and comparing a patient’s estimated dietary 
intake against their estimated caloric needs. 
 
2.2 
Endpoints/Outcome Measures 
2.2.1 
Primary 
 
Assess the change in the percent of CAFs expressing CAV1 at an intensity of 1+ or 
greater after treatment with metformin and doxycycline as determined by two blinded 
pathologists. 
2.2.2 
Secondary 
 
Assess safety and tolerability of metformin and doxycycline treatment in subjects with 
cancer. Toxicity will be evaluated using the most recent version (version 4) of the NCI 
toxicity criteria, i.e. the NCI Common Terminology Criteria for Adverse Events (CTCAE) 
version 4.0. A copy of the CTCAE can be found at http://ctep.cancer.gov. Any grade 3 or 
4 SAE will require immediate notification to the DSMB and IRB.  
 
Assess the change in the percent of tumor cells expressing MCT4, that are TUNEL 
positive and that express MCT1, BGAL and TOMM20 in HNSCC tumors using 
semiquantitative scoring and digital pathology using Aperio analyses of expression 
intensity with previously validated algorithms. 
2.2.3 
Exploratory 
 
To assess the effect of combined metformin and doxycycline therapy on the metabolic 
profile of cancer cells and stroma using mass spectroscopy imaging (MSI) on paired 
samples, comparing metabolite profiles in the pre-treatment and post-treatment tumor 
sample.  
 
To assess the impact of a patient’s nutritional status, estimated using 3-day dietary recall 
versus caloric needs as calculated by the Harris-Benedict equation on the baseline and 
net change in CAV1.  Patients will be labeled as below caloric needs, meeting caloric 
needs, and exceeding caloric needs. 
 
To assess the effect of combined metformin and doxycycline therapy on exosome 
evaluation, metabolomics profile, and microRNA expression profile. 
3 Study Design 
 

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This is a Phase II parallel cohort design trial in which patients with known or suspected head 
and neck squamous cell carcinoma who are eligible for resection are randomized to receive 
either metformin (arm A), doxycycline (arm B), or a combination of those two agents (arm C) in 
the 1-5 week window of time between their diagnostic or confirmatory biopsy and their tumor 
resection. All medications should cease at least 12 hours prior to time of resection. 
Immunohistochemical analyses will be performed on the pre-treatment biopsy and the post-
treatment resection specimens, allowing us to more fully characterize the metabolic 
microenvironment of HNSCC and our ability to alter it.  No patients will receive a placebo.  The 
treatment takes place in a period of time during which no other standard therapy exists for these 
patients.  All patients will be treated at TJUH and all samples material analyzed here with the 
exception of MSI performed at Vanderbilt.  Patient participation will last only during this window 
of time between enrollment and surgery, up to 5 weeks, with no additional visits planned.  Data 
collected on the patients will include the IHC and MSI on their specimens, serologic assays on 
their metabolic state, and a characterization of their general nutritional state via the Harris-
Benedict equation.  
3.1 
Characteristics 
Parallel Phase II 
3.2 
Number of Subjects 
33 total, 11 in each arm 
3.3 
Duration of Therapy 
1-5 weeks 
3.4 
Duration of Follow Up 
1 month 
3.5 
Treatment Assignment Procedures 
3.5.1 
Randomization Procedures: The Division of Biostatistics will generate the 
randomization schedule prior to the initiation of the study using the method of 
random permuted blocks.  Randomization assignments will be loaded into a 
REDCap database, and assignments will be accessed by Jennifer Johnson, 
MD, PhD using the REDCap randomization facility. 
3.5.2 
Masking Procedures: Pathologists will not be given information on the 
treatment history of the pathologic samples at the time of interpretation. 
Investigators and clinical coordinators will not be blinded on what treatment 
patients are receiving.  
3.6 
Study Timeline 
3.6.1 
Primary Completion 

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Eligibility Screening: Trial Coordinator will screen the office charts for possible inclusion in the 
trial. Labs for the screening part will be available from pre-admission testing that is standard of 
care for initial biopsy.  
Visit 1, Day 0: Subjects that meet the inclusion criteria after reviewing plan for tissue diagnosis, 
laboratory data and clinical chart will be approached the day of their follow up with a 
subspecialty surgeon. The initial biopsy does not need to take place on Day 0 and previously 
obtained tissue may be used.  The study coordinator will meet with the prospective candidates 
and go over the protocol, answer questions and obtain informed consent. Once the subject’s 
laboratory results are reviewed and eligibility is confirmed, the patient will be randomized and 
metformin and/or doxycycline will be dispensed.  Medication will be dispensed from Thomas 
Jefferson University Investigational Drug Service (IDS) Pharmacy after randomization.  
Instructions on how to take study drug will be given and times for follow up phone calls for 
tolerability and safety will be arranged with participants. Patients will also be asked to meet with 
a nutritionist either in person or by phone within the first 8 days of their enrollment to fill out a 3-
day dietary recall log when possible. A nutritionist will either meet with the patients in person or 
call them within the first 8 days of their enrollment to fill out the 3-day dietary recall log.  The 
nutritionist will do a 2-day guided recall, recording what the patients have eaten in the last 3 
days and how much.  Two 15 mL vials of blood will be drawn for baseline metabolomics and 
exosome profiling. 
Visit 2, Day of Surgery, Week 1-5 based upon surgery: On the day of surgery, two 15 ml vials of 
blood will be drawn for post-treatment metabolomic and exosome profiling. A post-treatment 
tissue sample will be obtained from excess tissue taken from this resection specimen that is not 
needed in the course of clinical care. On either the day of surgery or up until their first post-op 
visit, patients will be approached for bottles and pill reconciliation as well as follow up of side 
effects and adverse events. For patients who, per institutional standard of care, have genetic 
testing performed on their tumor sample, we will review this information in addition to the 
remainder of their pathology report. 
 
3.6.2 
Study Completion 
 
Follow-up: A follow up phone call will be performed 30 days (± 5 days) after the last dose of 
metformin and/or doxycycline for any other side effects or adverse events. The participant’s 
medical records will be reviewed every 3 months for 12 months to assess final pathologic 
staging.  
4 Study Enrollment and Withdrawal 
Patients must have baseline evaluations performed prior to the first dose of study drug and must 
meet all inclusion and exclusion criteria. Results of all baseline evaluations will be reviewed by 
the Principal Investigator or treating physician prior to enrollment, to verify that all inclusion and 
exclusion criteria have been satisfied. In addition, the patient must be thoroughly informed about 

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all aspects of the study, including the study visit schedule and required evaluations and all 
regulatory requirements for informed consent. The written informed consent must be obtained 
from the patient prior to screening procedures being performed. The following criteria apply to 
all patients enrolled into the study unless otherwise specified. 
 
4.1 
Eligibility Criteria 
4.1.1 
Inclusion Criteria 
Individuals must meet all of the following inclusion criteria in order to be eligible to 
participate in the study: 
1. Diagnosis of head and neck squamous cell carcinoma that is either biopsy proven or 
suspected based on history, physical, and or radiographic findings, and who are planned 
for definitive resection of the tumor without the use of neoadjuvant chemotherapy or 
radiation therapy at TJUH are eligible to participate. 
2. Subjects must be ≥ 18 years of age at time of consent. 
3. Patient must be able to swallow pills.  
4. Patients with serum creatinine levels less than 1.5 mg/dL 
5. Women of childbearing potential must have a negative urine or blood pregnancy test 
within 14 days of study enrollment.  
6. Informed Consent:  All subjects must be able to comprehend and sign a written informed 
consent document.  
7. ECOG Performance status ≤1 
4.1.2 
Exclusion Criteria 
An individual who meets any of the following criteria will be excluded from participation 
in this study: 
1. Subjects that do not have a baseline tumor specimen/biopsy prior to starting study 
medications. 
a. Tumor specimens do not need to be at Jefferson at time of eligibility 
determination. Tumor specimens held at outside institutions should be requested 
for analysis of pre-treatment tumor vs post-treatment tumor.  
2. Subjects who are pregnant or breastfeeding, or may become pregnant during metformin 
and doxycycline administration. 
3. Received prior cancer therapy for the HNSCC that is being resected. 
4. Subjects on metformin or doxycycline for any reason during the preceding 4 weeks. 
5. Diabetic subjects that are managed by taking metformin or insulin  
6. Subjects who have received iodinated contrast dye must wait 12 hours prior to starting 
Metformin. If a CT scan with contrast is scheduled after screening and consent, the 
metformin cannot be taken until after the CT with contrast has been completed and they 
have waited 12 hours. 
7. Patients with serum creatine ≥1.5 mg/dL 
8. Patients with history of lactic or any other metabolic acidosis.  
9. Patients with history of congestive heart failure stage III or greater. 
10. Patients scheduled for definitive cancer surgical resection less than 7 days from 
beginning of study drug administration or greater than 6 weeks from beginning study 

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drug administration.  
11. Patients with history of hepatic dysfunction or hepatic disease and abnormal liver 
function tests defined as AST, ALT, Alk Phos, and or total bilirubin greater than 2.5 times 
the upper limit of normal.  Patients who have a history of hepatic dysfunction or hepatic 
disease and normal liver function tests will be eligible to participate. 
12. Patients with a current history (in the past 30 days) of heavy drinking which is defined in 
accordance with CDC definition as more than 8 drinks per week for women and more 
than 15 drinks per week for men. A standard drink contains .6 ounces of pure alcohol. 
Generally, this amount of pure alcohol is found in 12-ounces of beer, 8-ounces of malt 
liquor, 5-ounces of wine, 1.5-ounces or a “shot” of 80-proof distilled spirits or liquor (e.g., 
gin, rum, vodka, or whiskey). While on study, patients should limit their alcohol 
consumption to no more than 8 drinks per week for women and no more than 15 drinks 
per week for men.  
13.  Patient with prior allergic reaction to metformin, doxycycline, or any other tetracycline 
antibiotic in the past.  
14. Patient is on medications that are contraindicated with metformin or doxycycline under 
current FDA recommendations.  The following is a list of medications identified as class 
D (consider therapy modification) when treatment with metformin or doxycycline is 
considered: 
o Class D: 
 
Bismuth Subsalicylate 
 
Cimetidine 
 
Iodinated contrast agents 
 
Somatropin 
 
4.2 
Gender/Minority/Pediatric Inclusion for Research  
We will not exclude potential subjects from participating in this study based on ethnic origin or 
socioeconomic status.  Every attempt will be made to enter all eligible patients in this protocol 
and therefore address the study objectives in a patient population representative of the entire 
HNSCC cohort at Thomas Jefferson University Hospital.  By the nature of the diseases covered 
in this trial there is a male predominance with 45,330 cancer cases in men and 16,430 cases in 
women projected to be diagnosed in 2016 in the US alone (a 2.8:1 ratio).  Our plan will be to 
monitor accrual in our MDG meetings monthly and to assess whether our accrual ratio matches 
the expected ratio of the disease.  If it does not, we will first query the tumor registry at TJUH to 
assess our recent male to female ratio for head and neck squamous cell cancer, and if there is 
still a discrepancy, we will create an action plan. 
 
4.3 
Strategies for Recruitment and Retention 
33 subjects will be recruited. No advertisement will be conducted. Screening requirements 
include serum measurement of creatinine, aspartate aminotransferase, alanine 
aminotransferase and alkaline phosphatase taken on a non-fasting blood sample. Potential 
research subjects will be identified by a member of the patient’s treatment team, the protocol 
investigator, or research team at participating centers from Medical Oncology, Radiation 
Oncology and Surgical offices.  Investigators will screen the patient’s medical records for 

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suitable research study subjects and discuss the study and their potential for enrolling in the 
research study. Patients will be screened based on pathology, image studies etc. A maximum of 
11 patients will be enrolled onto each arm of the trial (see Statistical Analysis Plan). 
  
4.4 
Subject Withdrawal 
4.4.1 
Reasons for Withdrawal 
Subjects are free to withdraw from participation in the study at any time upon request. 
An investigator may terminate a study subject’s participation in the study if: 
 
Any clinical adverse event (AE), laboratory abnormality, or other medical condition or 
situation occurs such that continued participation in the study would not be in the best 
interest of the subject. 
 
The subject meets an exclusion criterion (either newly developed or not previously 
recognized) that precludes further study participation. 
4.4.2 
Handling of Subject Withdrawals or Subject Discontinuation of Study 
Intervention 
Patients who do not have pre- medication initiation and post-biopsy, or are discontinued from 
the study will not be considered as evaluable for this study and will be replaced. 
Patients will be contacted by phone by a study coordinator 30 days (±5 days) after their planned 
resection to assess for any further adverse events that were not noted at the time of surgical 
resection (Visit 2). 
 
4.5 
Premature Termination or Suspension of Study 
This study may be suspended or prematurely terminated if there is sufficient reasonable cause. 
Written notification, documenting the reason for study suspension or termination, will be 
provided by the suspending or terminating party to investigator, and regulatory authorities. If the 
study is prematurely terminated or suspended, the principal investigator will promptly inform the 
IRB and will provide the reason(s) for the termination or suspension.  
Circumstances that may warrant termination include, but are not limited to: 
 
Determination of unexpected, significant, or unacceptable risk to subjects. 
 
Insufficient adherence to protocol requirements. 
 
Data that is not sufficiently complete and/or evaluable.  
 
Determination of futility. 

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5 Study Intervention 
5.1 
Study Product  
The investigator shall take responsibility for and shall take all steps to maintain appropriate 
records and ensure appropriate supply, storage, handling, distribution and usage of 
investigational product in accordance with the protocol and any applicable laws and regulations. 
 
5.2 
Study Product Description 
 
Metformin: 
Metformin is a biguanide drug currently approved for the treatment of type 2 diabetes mellitus by 
the FDA. It is currently being investigated in multiple cancer treatment trials.  
Metformin is a therapeutic agent in the protocol. The initial starting dose will be 500mg orally 
daily for 3 days which then will be increased to 500 mg twice daily and, if tolerated, further 
increased to 1000mg twice daily after day 6. Patients will maintain 1000 mg twice a day dosing 
until the day prior to their scheduled definitive surgery. This dose schedule has been shown to 
be well tolerated and was able to promote cellular changes consistent with beneficial anti-
cancer effects in breast cancer patients. 
 
The treatments to be used in this trial are outlined in Table 1.  
 
Doxycycline: 
Doxycycline is an antibiotic provided in the formulation doxycycline hyclate.  Doxycycline is 
virtually completely absorbed after oral administration.  Following administration of a single 200 
mg dose to adult volunteers, average peak serum doxycycline levels were 2.6 mcg/mL at 2 
hours, decreasing to 1.45 mcg/mL at 24 hours. The mean Cmax and AUC 0-∞ of doxycycline 
are 24% and 13% lower, respectively, following single dose administration of DORYX tablets, 
100 mg with a high fat meal (including milk) compared to fasted conditions. The mean Cmax of 
doxycycline is 19% lower and the AUC 0-∞ is unchanged following single dose administration 
of DORYX Tablets, 150 mg with a high fat meal (including milk) compared to fasted conditions. 
The clinical significance of these decreases is unknown. 
Tetracyclines are concentrated in bile by the liver and excreted in the urine and feces at high 
concentrations and in a biologically active form. Excretion of doxycycline by the kidney is about 
40%/72 hours in individuals with a creatinine clearance of about 75 mL/min. This percentage 
may fall as low as 1-5%/72 hours in individuals with a creatinine clearance below 10 mL/min.  
Studies have shown no significant difference in the serum half-life of doxycycline (range 18-22 
hours) in individuals with normal and severely impaired renal function. Hemodialysis does not 
alter the serum half-life.  
Doxycycline is an FDA approved product that is indicated for the prophylaxis against and 
treatment of both bacterial and protozoal infections.  For adults the dose is 200 mg on the first 
day of treatment administered as 100 mg every 12 hours followed by a maintenance dose of 
100 mg daily.  The maintenance dose may be administered as a single 100 mg dose of 50 mg 
dose every 12 hours.  Serious infections are treated with a sustained dose of 200 mg per day in 

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divided doses.  Treatment for uncomplicated bacterial infections typically lasts for 7 days total.  
Prophylaxis for malaria can be continued daily for the period of time the patient is in a malarious 
area and continued for 4 weeks thereafter, up to 4 months.  Our planned dosing is to use 100 
mg every twelve hours until the day prior to their scheduled definitive surgery. 
 
Table 1         Trial Treatments 
 
DRUG 
DOSE LEVEL 
DOSE 
ROUTE 
REGIMEN 
METFORMIN 
1 
500 mg daily 
Oral 
Day 1-3 
2 
500 mg every 12 
hours (1000mg 
daily) 
Oral 
Day4-6 
3 
1000 mg every 
12 hours (2000 
mg daily) 
Oral 
Day 7-the day 
prior to surgery 
DOXYCYCLINE 
-1 
100 mg daily 
Oral 
Only if needed 
for inability to 
tolerate level 1 
1 
100 mg every 12 
hours (200 mg 
daily) 
Oral 
Day 1-the day 
prior to surgery 
 
The same dosing and modifications apply either when given alone or together.  
 
5.2.1 
Acquisition 
Drug will be obtained through the hospital drug pharmacy and maintained by the Investigational 
Drug Service Pharmacy. 
 
5.2.2 
Formulation, Packaging, and Labeling 
 
Metformin: 
Metformin will be provided as 500 mg tablets which are round, white to off-white, film-coated 
tablets.  These are stored at 20°–25° C (68°–77° F); excursions permitted to 15°–30° C (59°–
86° F). [See USP Controlled Room Temperature.]  It will be dispensed in light-resistant 
containers labelled as metformin. 
 
Doxycycline: 
Doxycycline will be provided as 100 mg white oval tablets containing yellow pellets.  These are 
stored at 25°C (77°F); excursions permitted to 15°-30°C (59°-86°F) [see USP Controlled Room 
Temperature]; dispense in a tight, light-resistant container (USP) labelled as doxycycline. 
 

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5.2.3 
Product Storage and Stability 
Drug will be stored in a locked cabinet until given to the participants at room temperature in the 
Thomas Jefferson University Hospital Investigational Drug Service (IDS) Pharmacy. Access to 
the locked cabinet will only be granted to the pharmacist.  The investigator must ensure that it is 
stored in accordance with the environmental conditions as defined in the Investigator Brochure. 
Clinical supplies may not be used for any purpose other than that stated in the protocol. 
Each drug is stable at room temperature as above. 
5.3 
Dosage, Preparation, and Administration  
Clinical supplies will be affixed with a clinical label in accordance with regulatory requirements. 
 
5.4 
Dose Modifications and Dosing Delays 
 
Particular attention will be paid in the first three days of treatment. Patients will take 500mg/day 
for 3 days.  From day 4, 500mg twice daily and then in 3 days (day 7) dose escalation to 
1000mg twice daily will be achieved.  This will be taken until the day before surgery after dinner.  
A phone call on the day before (or ± 2 days) each dose escalation will be made in order to 
evaluate the tolerability of the drug and also weekly thereafter.  Patients will be instructed to 
contact the clinical investigators should any toxicity occur during the study (66). 
Metformin will not be started if CT scan with intravenous contrast is administered to decrease 
risk of lactic acidosis. After the scan they will be instructed to start the metformin the following 
day. This approach is more stringent than the most recent recommendation of the American 
College of Radiology which does not recommend holding metformin in the absence of 
comorbidities (renal insufficiency, liver dysfunction, alcohol abuse, cardiac failure, myocardial or 
peripheral muscle ischemia, sepsis or severe infection) which are exclusion criteria for this 
clinical trial (29).  
Toxicity will be evaluated using the most recent version (version 4) of the NCI toxicity criteria, 
i.e. the NCI Common Terminology Criteria for Adverse Events (CTCAE) version 4.0.  
 
Grade 1 toxicity: Patient will be maintained on full dose. 
 
Grade 2 toxicity: Dose will be reduced by 50% until grade 1 or lower. If symptoms are 
not resolved within 3 days the treatment will be discontinued definitively.  
 
Grade 3 toxicity (probably or definitely drug related): Treatment will be interrupted and 
toxicity reassessed daily. If toxicity improves to grade 2, dose will be reduced by 50%. 
 
Grade 4 toxicity (probably or definitely drug related): Treatment will be discontinued 
definitively. 
 
In case of grade 1 or 2 diarrhea (the most frequent side effect) a concomitant administration of 
loperamide will be provided. 
 
Clinical supplies will be affixed with a clinical label in accordance with regulatory requirements. 
If doses of either medication are missed due to toxicity they should not be replaced.  If a dose is 
not taken due to an error, it may be taken up to 6 hours later.  If vomiting occurs within 30 

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minutes of intake, that dose may be repeated.  These events should be recorded by the patient 
on their pill diary. 
 
5.5 
Study Product Accountability 
 
Metformin and doxycycline tablets will be provided by the Department of Medical Oncology and 
will be coordinated through the Thomas Jefferson University Hospital Investigational Drug 
Service (IDS) Pharmacy. Drug will be stored in locked cabinet in the IDS pharmacy until given to 
the participants. An approximately 2-week supply of metformin and/or doxycycline will be 
provided to subjects and the drug will be self-administered by the participants. 
 
Upon receipt of the of the study treatment supplies, an inventory must be performed and a drug 
receipt log filled out and signed by the person accepting the shipment. It is important that the 
designated study staff counts and verifies that the shipment contains all the items noted in the 
shipment inventory. Any damaged or unusable study drug in a given shipment (active drug or 
comparator) will be documented in the study files. The investigator must notify study sponsor of 
any damaged or unusable study treatments that were supplied to the investigator’s site.  
At the completion of the study, there will be a final reconciliation of drug shipped, drug 
consumed, and drug remaining. This reconciliation will be logged on the drug reconciliation 
form, signed and dated. Any discrepancies noted will be investigated, resolved, and 
documented prior to return or destruction of unused study drug. Drug destroyed on site will be 
documented in the study files.  
 
The investigator is responsible for keeping accurate records of the clinical supplies received 
from the Sponsor or designee, the amount dispensed to the subjects and the amount remaining 
at the conclusion of the trial. 
 
Upon completion or termination of the study, all unused and/or partially used investigational 
product will be destroyed at the site per institutional policy. It is the Investigator’s responsibility 
to arrange for disposal of all empty containers, provided that procedures for proper disposal 
have been established according to applicable federal, state, local and institutional guidelines 
and procedures, and provided that appropriate records of disposal are kept. 
5.6 
Assessing Subject Compliance with Study Product Administration 
 
Upon enrollment to the trial, depending on which arm the patient is enrolled to, there will be 
either one or two pill bottles with the appropriate number of metformin and doxycycline tablets 
distributed to the patients.  The pill bottles will be accompanied with detailed instructions on the 
proper dosage/number of tablets to take daily as noted above.  Upon arrival for definitive 
surgical resection, the bottle will be collected by our trial coordinator and the contents will be 
evaluated for compliance.  
5.7 
Concomitant Medications/Treatments 
Medications or vaccinations specifically prohibited in the exclusion criteria are not allowed 
during the ongoing trial. If the patient is on any of the Class C medications as outlined in Section 
5.4.2, they should be monitored closely for any potential interactions or side effects. If there is a 
clinical indication for one of these or other medications or vaccinations specifically prohibited 
during the trial, discontinuation from trial therapy or vaccination may be required. The 
investigator should discuss any questions regarding this with the Sponsor. The final decision on 

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any supportive therapy or vaccination rests with the investigator and/or the subject's primary 
physician. However, the decision to continue the subject on trial therapy or vaccination schedule 
requires the mutual agreement of the Investigator, the Sponsor, and the subject 
All treatments that the investigator considers necessary for a subject’s welfare may be 
administered at the discretion of the investigator in keeping with the community standards of 
medical care. All concomitant medication and therapies will be recorded on the case report form 
(CRF) including all prescription, over-the-counter (OTC), herbal supplements, and IV 
medications and fluids. If changes occur during the trial period, documentation of drug dosage, 
frequency, route, and date should also be included on the CRF. These will be cross referenced 
against the list of drugs in section 4.2 (Exclusion Criteria) to ensure that no agents that are 
contraindicated with metformin or doxycycline are used.   
All concomitant medications received within 28 days before informed consent is signed and 30 
days after the last dose of trial treatment should be recorded. Concomitant medications 
administered to treat SAEs and ECIs should be recorded as defined 
All medications are permitted except those that are contraindicated with metformin and 
doxycycline under current FDA recommendations.  It is important to note that the medications 
that are contraindicated with metformin are contraindicated due to concern for theoretical 
interactions. Class C medications are allowed if they are established at baseline. The treating 
physician should be made aware of any new Class C medications taken during therapy for 
proper monitoring of potential interactions. The following is a list of medications identified as 
class C (monitor therapy) and class D (consider therapy modification) when treatment with 
metformin or doxycycline is considered: 
 
Class C: 
 
 
 
 
 
Class D: 
Carbonic anhydrase inhibitors 
 
 
Bismuth Subsalicylate  
Cephalexin 
 
 
 
 
 
Cimetidine 
 
 
 
Corticosteroids (orally inhaled) 
 
 
Iodinated contrast agents 
Corticosteroids (systemic) 
 
 
 
Somatropin 
Dalfampridine 
Dofetilide 
Glycopyrrolate 
Lamotrigine 
Luteinizing hormone-releasing hormone analogs 
Pegvisomant 
Penicillin 
Trospium 
 
5.8 
Dietary Restrictions 
Subjects should maintain a normal diet unless modifications are required to manage an AE such 
as diarrhea, nausea or vomiting. Patients will be asked to fill a dietary log at the beginning the 
study when they speak with a registered dietician. 
6 Study Schedule  

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All patients will be evaluated by a treating surgeon.  Initial visit will include overall assessment of 
health as well as determination of eligibility. A study calendar details the requisite pretreatment 
evaluations

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Study Procedure 
Screening 
Treatment/Intervention Period D 
End of E 
Treatment 
Follow-up 
G 
Week 1 
Week 2D 
Week 3-5 
 
Day 1 -7 
Day 8-14 
Surgical F 
Week 6  
Week 7 
Resection 
Administrative 
Procedures 
  
Informed consent A and 
research authorization/ 
HIPAA form 
X 
  
  
  
  
  
  
  
  
  
Inclusion/Exclusion 
Criteria 
X 
  
  
  
  
  
  
  
  
  
Demographics, Medical 
history  
X 
 
 
 
 
 
 
 
 
 
Clinical Assessment 
  
  
  
  
Physical examination, 
vitals, weight, blood 
pressure 
X 
  
  
  
  
  
  
XN  
x 
 
Height 
X 
  
  
  
  
  
  
  
  
  
ECOG Performance 
status 
X 
  
  
  
  
  
  
  
X  
 
Pathology Report 
histologic confirmation of 
disease if availableL 
X 
  
  
  
  
  
  
X  
-  
 
Concomitant medsK 
X 
X 
X 
X 
X  
X 
X 
X 
X 
X 
Toxicity and AE 
AssessmentK 
X 
X 
 X 
 X 
X  
X  
 X 
 X 
x 
x 

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Laboratory testsB 
  
  
  
  
Hematology 
X 
  
  
  
  
  
  
  
  
 
Blood Chemistry 
X 
  
  
  
  
  
  
  
  
 
pregnancy testH 
X 
 
 
 
 
 
 
 
 
 
Treatment/ 
Intervention 
  
  
  
MetforminC,D 
  
X 
X 
X 
X 
X 
X 
  
  
  
DoxycyclineC,D 
  
X 
X 
X 
X 
X 
X 
  
  
  
Diagnostic Biopsy or 
tumor sampleI 
X 
 
 
 
 
 
 
X 
 
 
Surgical Resection 
 
 
 
 
 
 
 
X 
 
 
Correlative Studies 
  
  
  
  
Serologic metabolic 
profile (miR21 and 
exosomes)F 
 
X  
  
  
  
  
  
X 
  
  
Nutrition 
Assessment* 
 
XM 
 
 
 
 
 
 
 
 
*Nutrition assessment and Harris Benedict Equation will occur on Day 1 (or up to Day 8) of enrollment when possible, and may be done via a phone interview. 
A. Written informed consent must be obtained before any study-specific screening assessments are performed. Lab results performed as standard-of-care prior to obtaining informed 
consent and within 28 days prior to enrollment may be used; such tests do not need to be repeated for screening. Trial treatment should begin within 5 days of eligibility confirmation.  
B. Blood Chemistry includes: Creatinine, AST, ALT, ALK Phos, T Bili > 2.5 x ULN. Hematology includes: CBC (with differential).  All are non-fasting labs. 
C. Study treatment follow up phone calls will occur on days 3 and 5 ±2 days of dose escalation to evaluate tolerability, document AEs, and concomitant medications. If patient remains on 
study drugs for >7days, study treatment follow-up phone calls will occur a minimum of once a week until surgery to evaluate tolerability, AEs, and con meds. Last doses of metformin 
and/or doxycycline pills should not be taken within 12 hours of scheduled surgery.     
D. Patient must receive a minimum of 7 days of metformin and/or doxycycline before having definitive surgery. Patients will stay on metformin and/or doxycycline until the day prior to 
surgery (up to the maximum of 5 weeks). 
E. End of treatment visit may occur within 14 days ± 7 days of stopping study drug. 
F. Correlative study blood to be collected the morning of surgical resection. Baseline samples are to be drawn after enrollment but prior to start of study treatment.  

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G. Follow up phone call will be performed 30 days after last dose of study medication for any other side effects or toxicity and for pathologic staging information. 
H. serum or urine pregnancy test must be done within 14 days prior to enrollment on study for all women of childbearing potential 
I. If a confirmatory biopsy is performed at TJUH and excess tissue is available tissue will be flash frozen both at the time of biopsy and at the time of resection. A post-treatment tissue 
sample will be obtained from excess tissue taken from this resection specimen that is not needed in the course of clinical care. 
J. Study drug stopped the night prior to surgery.  
K. Concomitant medication reconciliation, and Toxicity and Adverse Event assessments should be done at every patient contact to fully capture any potential dose limiting toxicities or 
AEs that affect the patient’s ability, willingness, and eligibility to continue taking study medications.  
L. Operative reports should be included with pathology reports in source documents when applicable.  
M. To be performed by a registered dietician, when possible. Must meet with patient within five days of enrollment and record the patient’s last 3 days of caloric intake and estimate their 
caloric intake versus their caloric needs per the Harris Benedict Equation. It will be noted in the patient chart if these assessments could not be completed. 
N. Pre-operative nursing or anesthesia exam is acceptable and is to be performed the morning of surgery. 

 
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6.1 
Pretreatment Period/Screening 
 
Screening Assessment (Day -28 to Day -1) 
 
Before initiating any screening activities, the scope of the study will be explained to each 
patient during informed consent. Patients should be advised of any known risks inherent in the 
planned procedures, any alternative treatment options, their right to withdraw from the study at 
any time for any reason, and their right to privacy. After this explanation, patients should be 
asked to sign and date a Notice of Privacy Practice research authorization/HIPAA form and an 
IRB-approved statement of informed consent that meets the requirements of the Code of 
Federal Regulations (Federal Register Vol. 46, No. 17, January 27, 1981, part 50). During the 
screening period, subject eligibility will be determined according to the inclusion and exclusion 
criteria (Section 4.1, 4.2). The following assessments will be performed during this time:  
 
Obtain informed consent and research authorization 
 
Inclusion/Exclusion Criteria 
 
Record demographics (including age) and medical history  
 
Physical exam 
 
Concomitant meds 
 
ECOG performance status 
 
Discuss concurrent medications  
 
Vital signs [body temperature, blood pressure, pulse, respiratory rate, weight, height] 
 
Obtain histologic and/or (if appropriate) radiologic confirmation of disease 
 
Confirm eligibility according to the inclusion/exclusion criteria 
 
Laboratory tests: 
o Hematology: CBC with differential 
o Blood Chemistry 
 
Creatinine, AST, AT, ALK Phos T Bili > 2.5 x ULN (all are 
Standard Of Care) 
o Serum or urine pregnancy test (within 14 days prior to enrollment for 
women of childbearing potential) 
6.2 
Enrollment/Baseline 
 
In order to be considered for evaluation, the patient must have an evaluable pre-
treatment biopsy specimen. 
 
If a confirmatory diagnostic core biopsy is required and there is excess tissue available 
not needed in the course of clinical care, this tissue will be flash frozen in liquid nitrogen.   
 
Patients who are enrolled in the trial will be randomized to receive metformin, 
doxycycline, or metformin and doxycycline together. Patients will receive a Patient 
Medication Diary to record the time (AM and PM) and number of pills taken for 
metformin and/or doxycycline.  
 
Baseline caloric intake assessment using a dietary log mediated by a nutritionist when 
possible.  This will take place at the patient’s baseline assessment and may be done via 
a phone interview.   

 
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 
Calculation of the patient’s estimated caloric needs via the Harris-Benedict Equation 
when possible. 
*If the nutrition assessment and Harris-Benedict Equation cannot be completed, this will be 
noted in the patient’s chart. 
 
Complete physical examination including ECOG performance status, vital signs 
 
The collection of two serum separator tubes to be drawn after enrollment and prior to the 
start of the study treatment. 
 
Concurrent medications will be recorded 
 
6.3 
Treatment Period 
Subjects receiving metformin and/or doxycycline will return to clinic day 1, week 1. Surgical 
resection will be scheduled a minimum of 7 days after the start of metformin and/or 
doxycycline, and a maximum of 5 weeks. The participant will receive a Patient Medication 
Diary to record the time (AM and PM) and number of pills take for both Metformin and 
Doxycycline as applicable based on their randomization.  
 
The following assessments will be performed at Visit 2, the day of surgery  
 
Physical exam- Pre-operative nursing or anesthesia exam is acceptable and is to be 
performed the morning of surgery. 
 
Concomitant medications 
 
Adverse event assessment 
 
 
Patients will undergo their Surgical Resection at Visit 2. 
o When a pre-treatment sample has been acquired by flash freezing for MSI, and if there 
is excess tissue is available at the time of resection, this tissue samples will also be 
frozen in liquid nitrogen for MSI. A post-treatment tissue sample will be obtained from 
excess tissue taken from this resection specimen that is not needed in the course of 
clinical care. 
 
 
Correlative studies will be drawn at Visit 2: oncomiR miR-21, serum triglycerides, IGF-1, 
IGF-BP3, ESR, adiponectin, leptin, IGF-1R, exosome evaluation, metabolomics profile, and 
microRNA expression profile. 
 
 
Study drug will be taken up until at least 12 hours before surgery.  
 
6.4 
End of Treatment Study Procedures 
 
The following study activities will occur at End of Study Visit, within 14 days ± 7 days of stopping 
the protocol study treatments: 
 
Concomitant medications 
 
Adverse event assessment 
 
6.5 
Post-treatment/Follow-Up 

 
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A follow up phone call will be performed 30 days (± 5 days) after the last dose of metformin and 
doxycycline for any other side effects or adverse events. This is allowed to coincide with the 
End of Treatment visit if it occurs during timing overlap. The participant’s medical records will 
be reviewed every 3 months for 12 months to assess final pathologic staging. 
6.6 
Withdrawal Visit/Discontinuation of Therapy 
Patients can withdraw at any time during the study if they no longer want to participate in the 
trial. If withdrawal occurs no further metformin and or doxycycline administration will occur and 
patients will be required to return remaining tablets which will be logged in to medication 
administration records and destroyed. If a subject withdraws consent to participate in the study, 
permission will be sought to use data pertaining to the subject in the analysis as far as they 
participate and they will be removed from subsequent analyses and replaced.  
 
Description of when a subject’s participation in the trial may be discontinued: 
 
Specific reasons for discontinuing a subject from this study are: 
1. Voluntary discontinuation by the subject who is at any time free to discontinue their 
participation in the study, without prejudice to further treatment. 
2. Safety reasons as judged by the investigator. 
3. Severe non-compliance to the protocol as judged by the investigator. 
4. Incorrect enrollment of the subject. 
5. Subject lost to follow-up 
6. Death 
7 Study Procedures and Evaluations 
Study procedures for this trial include: 
 
Review and immunohistochemical staining of a patient’s diagnostic biopsy, which was 
performed as standard of care. 
 
Randomization to receive metformin alone, doxycycline alone, or metformin and 
doxycycline together. 
 
Administration of the oral medication(s) listed above as per randomization until the day 
before the patient’s planned surgery. 
 
The collection of two serum separator tubes to be drawn after enrollment and prior to the 
start of the study treatment. 
 
Meeting with a registered dietician either in person or by phone to do a guided three day 
dietary recall log within the first 8 days when possible. 
 
Snap freezing a portion of the patient’s biopsy and resection specimen for MSI testing 
may be performed in a subgroup of specimens.  
 
Immunohistochemistry on slides from the patient’s resection specimen. 
 
7.1 
Study Procedures/Evaluations 
 
 
Medical history including intercurrent illnesses and specific attention to diabetes, may be 
obtained from the medical record 
 
Physical exam: height, weight, obtained from the medical record 

 
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 
Concomitant meds: prescription medications taken for the 28 days prior to enrollment 
 
ECOG performance status from the medical record 
 
Obtain histologic confirmation of disease from the medical record 
 
Laboratory tests: Hematology- CBC with differential; Blood Chemistry- Creatinine, AST, 
AT, ALK Phos T Bili to have been performed within 28 days of enrollment and thus may 
be obtained either from the medical record or as a part of the standard of care 
7.2 
Laboratory Procedures/Evaluations 
7.2.1 
Clinical Laboratory Evaluations 
 
Hematology: CBC with differential 
 
Blood Chemistry with Creatinine, AST, AT, ALK Phos T Bili  
 
Serum or urine pregnancy test  
 
7.2.2 
Special Assays or Procedures 
Research Correlates- exosome evaluation, microRNA and metabolomics profile. 
These labs will be obtained from two serum separator tubes which will be provided directly to 
the Martinez laboratory.  After being obtained, the tubes may be kept at room temperature and 
transported to the laboratory at 233 S. 10th Street suite 909. Contact Diana Whitaker Menezes, 
MS, Research Associate, 215-955-9774. 
7.2.3 
Specimen Preparation, Handling, and Storage 
Biopsy specimens both pre-treatment and post-treatment are to be processed as formalin-fixed 
paraffin embedded samples per clinical standards. 
Samples for MSI testing will be flash frozen in liquid nitrogen within 10 minutes of removal of the 
specimen by a member of the Martinez laboratory. Fresh tissue samples will be quick frozen in 
liquid nitrogen, transferred to appropriate storage vials and stored at -80C until shipping to 
Vanderbilt MSI Facility. Each sample storage vial will be labeled with a unique ID number 
without any PHI. 
7.2.4 
Specimen Shipment 
Flash frozen tumor samples will be shipped to the Mass Spectroscopy Imaging facility at 
Vanderbilt University on dry ice by the Martinez laboratory personnel. For shipping, sample vials 
are placed into plastic Ziplock bags that are then placed into a styrofoam container of dry ice 
within a cardboard box. Shipping containers are sent Fed Ex Priority Overnight to the following 
address: 
Mitch Reyzer/Audra Judd, Mass Spectrometry Research Center, 465 21st Avenue South, 9160 
MRB III, Nashville, TN 37240 
8 Evaluation of Safety 
8.1 
Specification of Safety Parameters 
 
 

 
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8.1.1 
Unanticipated Problems 
Unanticipated problems (UAPs) include, in general, any incident, experience, or outcome that 
meets the following criteria: 
 
unexpected in terms of nature, severity, or frequency given (a) the research procedures 
that are described in the protocol-related documents, such as the IRB-approved 
research protocol and informed consent document; and (b) the characteristics of the 
subject population being studied; 
UAPs are considered to pose risk to subjects or others when they suggest that the research 
places subjects or others at a greater risk of harm (including physical, psychological, economic, 
or social harm) than was previously known or recognized. 
8.1.2 
Adverse Events 
An adverse event is any untoward or unfavorable medical occurrence in a human subject, 
including any abnormal sign (for example, abnormal physical exam or laboratory finding), 
symptom, or disease, temporally associated with the subject’s participation in the research, 
whether or not considered related to the subject’s participation in the research. 
 
8.1.3 
Serious Adverse Events 
A serious adverse event (SAE) is one that meets one or more of the following criteria: 
 
Results in death 
 
Is life-threatening (places the subject at immediate risk of death from the event as it 
occurred) 
 
Results in inpatient hospitalization or prolongation of existing hospitalization 
 
Results in a persistent or significant disability or incapacity 
 
Results in a congenital anomaly or birth defect  
 
An important medical event that may not result in death, be life threatening, or require 
hospitalization may be considered an SAE when, based upon appropriate medical 
judgment, the event may jeopardize the subject and may require medical or surgical 
intervention to prevent one of the outcomes listed in this definition. 
 
8.2 
Safety Assessment and Follow-Up 
The PI will follow adverse events with start dates occurring any time after informed consent is 
obtained until 7 (for non-serious AEs) or 30 days (for SAEs) after the last day of study 
participation. At each study visit, the investigator (or designee) will inquire about the occurrence 
of AE/SAEs since the last visit. Events will be followed for outcome information until resolution 
or stabilization. 

 
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8.3 
Recording Adverse Events 
The following subsections detail what information must be documented for each adverse event 
occurring during the time period specified in Section 8.2 Safety Assessment and Follow-Up.  
 
8.3.1 
Relationship to Study Intervention 
The relationship to study intervention or study participation must be assessed and documented 
for all adverse events. Evaluation of relatedness must consider etiologies such as natural 
history of the underlying disease, concurrent illness, concomitant therapy, study-related 
procedures, accidents, and other external factors.  
The following guidelines are used to assess relationship of an event to study intervention: 
1. Related (Possible, Probable, Definite) 
a. The event is known to occur with the study intervention. 
b. There is a temporal relationship between the intervention and event onset. 
c. The event abates when the intervention is discontinued. 
d. The event reappears upon a re-challenge with the intervention. 
2. Not Related (Unlikely, Not Related) 
a. There is no temporal relationship between the intervention and event onset. 
b. An alternate etiology has been established. 
 
8.3.2 
Expectedness 
The PI is responsible for determining whether an AE is expected or unexpected. An AE will be 
considered unexpected if the nature, severity, or frequency of the event is not consistent with 
the risk information previously described for the intervention. Risk information to assess 
expectedness can be obtained from preclinical studies, the investigator’s brochure, published 
medical literature, the protocol, or the informed consent document. 
 
8.3.3 
Severity of Event 
Adverse events will be graded for severity according to the Common Terminology Criteria for 
Adverse Events (CTCAE) version 4.  
 

 
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8.4 
Safety Reporting  
8.4.1 
Unanticipated Problem Reporting to IRB 
All incidents or events that meet criteria for unanticipated problems (UAPs) as defined in 
Section 8.1.1 Unanticipated Problems require the creation and completion of an unanticipated 
problem report form (OHR-20).  
UAPs that pose risk to subjects or others, and that are not AEs, will be submitted to the IRB on 
an OHR-20 form via the eazUP system within 5 working days of the investigator becoming 
aware of the event.  
UAPs that do not pose risk to subjects or others will be submitted to the IRB at the next 
continuing review.  
8.4.1 Adverse Event Reporting to IRB 
Grade 1 AEs are not required to be reported to the IRB.  
Grade 2 AEs will be reported to the IRB at the time of continuing review if, in the opinion of the 
investigator, they represent events that exceed expected frequency or in some other way are 
judged to be unexpected and possibly associated with increased risk.  
 
8.4.2 
Serious Adverse Event Reporting to IRB 
SAEs will be reported to the IRB on OHR-10 forms via the electronic reporting system (eSAEy) 
according to the required time frames described below. 
Grade 3-4 AEs that are unexpected and deemed to be at least possibly or definitely related to 
the study will be reported to the IRB within 2 working days of knowledge of the event. 
Grade 3-4 AEs that are deemed unrelated to the study will be reported to the IRB within 5 
working days.  
Grade 5 AEs will be reported to the IRB within one working day of knowledge of the event. 
8.4.3 
Reporting to SKCC DSMC 
All AEs and SAEs, safety and toxicity data, and any corrective actions will be submitted to the 
DSMC per the frequency described in the SKCC DSMP. The report to the SKCC DSMC will 
also include any unanticipated problems that in the opinion of the PI should be reported to the 
DSMC. 
For expedited reporting requirements, see table below:                                             
DSMC AE/SAE Reporting Requirements 

 
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8.4.4 
Reporting of Pregnancy 
Pregnant women are excluded from this trial.  Should a woman of child-bearing potential 
become pregnant during the course of this trial she will be discontinued from treatment and 
replaced in the study. 
8.5 
Halting Rules 
 
This study will be monitored by the SKCC DSMC.  
Acute toxicity will be monitored using CTCAE version 4.0 criteria during therapy and up to 30-
days post completion of surgical resection.  If the lowest dose of metformin and doxycycline 
develop toxicity ≥4, the study may be terminated after discussion with PI and sponsor.  
9 Study Oversight 
In addition to the PI’s responsibility for oversight, study oversight will be under the direction of 
the SKCC’s Data and Safety Monitoring Committee (DSMC). The DSMC operates in 

 
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compliance with a Data and Safety Monitoring Plan (DSMP) that is approved by the Clinical 
Trials Oversight Committee (CTOC).  
10 Clinical Site Monitoring and Auditing 
Clinical site monitoring and auditing is conducted to ensure that the rights of human subjects are 
protected, that the study is implemented in accordance with the protocol and/or other operating 
procedures, and that the quality and integrity of study data and data collection methods are 
maintained. Monitoring and auditing for this study will be performed in accordance with the 
SKCC’s Data and Safety Monitoring Plan (DSMP) developed by the SKCC Data and Safety 
Monitoring Committee (DSMC). The DSMP specifies the frequency of monitoring, monitoring 
procedures, the level of clinical site monitoring activities (e.g., the percentage of subject data to 
be reviewed), and the distribution of monitoring reports. Some monitoring activities may be 
performed remotely, while others will take place at the study site(s). Appropriate staff will 
conduct monitoring activities and provide reports of the findings and associated action items in 
accordance with the details described in the DSMP. 
11 Statistical Considerations  
11.1 Study Hypotheses 
 
Primary: Exposure to metformin and/or doxycycline administered together will alter the 
metabolic profile of squamous cell carcinoma of the head and neck, specifically as measured by 
the expression level of CAV1 in tumor-associated stromal cells. 
 
Secondary:   Exposure to metformin and/or doxycycline administered together will alter the 
metabolic profile of squamous cell carcinoma of the head and neck, as measured by the 
expression levels of BGAL and MCT4 in tumor associated stroma, and TUNEL, MCT1 and 
TOMM20 in epithelial tumor cells.  We hypothesize that these drugs, given either as 
monotherapy or in combination will be safe and tolerable for patients in the pre-surgical period. 
 
11.2 Analysis Plans 
 
Analysis for primary outcomes: The primary objective of the study is to assess the impact of 
metformin and or doxycycline on CAV1 expression by immunohistochemistry (IHC) in tumor-
associated stromal cells.  All patients from whom samples are obtained both pre- and post-
treatment will be included in the primary analyses. Within-patient change in IHC scores will be 
analyzed using the Wilcoxon signed-rank test. Comparisons will be made between pre-
treatment and post-treatment within each of the cohorts. The study is not powered to make 
comparisons between the three different cohort groups. 
 
Analysis for secondary outcomes:  Analysis of change in TUNEL, BGAL, TOMM20, MCT1, 
and MCT4 will be performed using the Wilcoxon signed-rank test.  Safety will be provided in 
descriptive tables. 
 
11.2.1 Safety Review 

 
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Incidence of DLTs will be monitored continuously with potential stopping after 10 patients have 
been enrolled based on the Bayesian method of Thall and Simon.  We are comparing our 
experimental treatment to a standard of no treatment and assume no DLTs under standard of 
care.  Both agents are extremely safe and DLTs should be exceedingly rare.  In 900 patients 
treated with metformin alone, the discontinuation rate was 0.6%.  In 31 patients treated with IV 
doxycycline, 4 SAEs were observed, but none were related to the study drug.  Thus, it is 
reasonable to assume a prior distribution for the DLT rate in the combination of Beta 
(0.04,1.96). That is, we assume a 2% event rate for the combination with information equivalent 
to data from 2 patients.  We will stop the study for safety if the posterior probability that the rate 
of DLTs in the combination treatment is 5% or greater is greater than 90%.  Stopping rules are 
outlined in the table below. 
 
Number of patients enrolled 
Stop if number of patients with Grade 3 or 
higher toxicities is ≥  
10 
2 
11-22 
3 
23-35 
4 
36 
5 
Thall C.P., Simon R. Practical Bayesian guidelines for phase IIB clinical trials. Biometrics. 
1994;50:337–349. 
 
11.3 Sample Size Considerations 
 
The sample in each cohort is based on having 85% power to detect an average increase in 
CAV1 staining of 40% using a two-sided Wilcoxon signed-rank test with alpha=0.05.  Based on 
pilot data in other cancers, we assume the standard deviation of change scores to be 32%.  A 
sample size of 9 per cohort is required under these assumptions.  Allowing for approximately 
15% of patients to have missing pre- and or post-treatment data, we will recruit 11 patients per 
cohort for a total of 33 patients. 
 
11.3.1 Replacement Policy 
 
Patients who do not have sufficient pre-treatment biopsy material obtained in the course of their 
medical care to be evaluated for CAV1 by IHC, or are discontinued from the study will not be 
considered as evaluable for this study and will be replaced. 
 
11.3.2 Accrual Estimates 
Our total goal is 33 patients.  We estimate this will be possible in a 3-year period with 11 per 
year. 

 
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12 Source Documents and Access to Source Data/Documents 
Study staff will maintain appropriate medical and research records for this study, in compliance 
with ICH E6, and regulatory and institutional requirements for the protection of confidentiality of 
subject information. Study staff will permit authorized representatives of SKCC and regulatory 
agencies to examine (and when required by applicable law, to copy) research records for the 
purposes of quality assurance reviews, audits, and evaluation of the study safety, progress and 
data validity.  

 
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13 Quality Control and Quality Assurance 
The investigator will allocate adequate time for monitoring activities. The Investigator will also 
ensure that the medical monitor or other compliance or quality assurance reviewer is given 
access to all of the  above  noted  study-related  documents  and  study  related  facilities  (e.g. 
pharmacy, diagnostic laboratory, etc.), and has adequate space to conduct the monitoring visit.  
14 Ethics/Protection of Human Subjects  
14.1 Ethical Standard 
The investigator will ensure that this study is conducted in full conformity with the principles set 
forth in The Belmont Report: Ethical Principles and Guidelines for the Protection of Human 
Subjects of Research, as drafted by the US National Commission for the Protection of Human 
Subjects of Biomedical and Behavioral Research (April 18, 1979) and codified in 45 CFR Part 
46 and/or the ICH E6.  
 
14.2 Institutional Review Board 
The protocol, informed consent form(s), recruitment materials, and all subject materials will be 
submitted to the IRB for review and approval. Approval of both the protocol and the consent 
form must be obtained before any subject is enrolled. Any amendment to the protocol will 
require review and approval by the IRB before the changes are implemented in the study.  
14.3 Informed Consent Process 
Informed consent is a process that is initiated prior to the individual agreeing to participate in the 
study and continues throughout study participation. Extensive discussion of risks and possible 
benefits of study participation will be provided to subjects and their families, if applicable. A 
consent form describing in detail the study procedures and risks will be given to the subject. 
Consent forms will be IRB-approved, and the subject is required to read and review the 
document or have the document read to him or her. The investigator or designee will explain the 
research study to the subject and answer any questions that may arise. The subject will sign the 
informed consent document prior to any study-related assessments or procedures. Subjects will 
be given the opportunity to discuss the study with their surrogates or think about it prior to 
agreeing to participate. They may withdraw consent at any time throughout the course of the 
study. A copy of the signed informed consent document will be given to subjects for their 
records. The rights and welfare of the subjects will be protected by emphasizing to them that the 
quality of their clinical care will not be adversely affected if they decline to participate in this 
study. The consent process will be documented in the clinical or research record.  
14.4 Exclusion of Women, Minorities, and Children (Special Populations) 
Pregnant and breastfeeding women are excluded from the trial due to the potential for the use 
of doxycycline.  Otherwise, we will attempt to accrue women and men equally on this study, 
representative of the epidemiologic distribution of the disease. 
14.5 Subject Confidentiality 

 
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Subject confidentiality is strictly held in trust by the investigators, study staff, and the sponsor(s) 
and their agents. This confidentiality is extended to cover testing of biological samples and 
genetic tests in addition to any study information relating to subjects. 
The study protocol, documentation, data, and all other information generated will be held in 
strict confidence. No information concerning the study or the data will be released to any 
unauthorized third party without prior written approval of the sponsor. 
The study monitor or other authorized representatives of the sponsor may inspect all study 
documents and records required to be maintained by the investigator, including but not limited 
to, medical records (office, clinic, or hospital) for the study subjects. The clinical study site will 
permit access to such records. 
 
14.6 Future Use of Stored Specimens and Other Identifiable Data  
No identifiable data will be retained. 
15 Data Handling and Record Keeping 
The investigators are responsible for ensuring the accuracy, completeness, legibility, and 
timeliness of the data reported. All source documents should be completed in a neat, legible 
manner to ensure accurate interpretation of data. The investigators will maintain adequate case 
histories of study subjects, including accurate case report forms (CRFs), and source 
documentation. 
 
15.1 Data Management Responsibilities 
Data collection and accurate documentation are the responsibility of the study staff under the 
supervision of the investigator. All source documents and laboratory reports must be reviewed 
by the study team and data entry staff, who will ensure that they are accurate and complete. 
Unanticipated problems and adverse events must be reviewed by the investigator or designee.  
 
15.2 Data Capture Methods 
Data will be captured on paper case report forms and transferred to an electronic database 
which will be kept behind a password protected firewall- RedCap. 
15.3 Types of Data 
 
Data that will be collected include patient history and physical examination data, safety, 
laboratory studies, pathology staging studies, and immunohistochemistry as determined in the 
laboratory. 
15.4 Study Records Retention 

 
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Study records will be maintained for at least three years from the date that the grant federal 
financial report (FFR) is submitted to the NIH. 
Study documents should be retained for a minimum of 2 years after the last approval of a 
marketing application in an ICH region and until there are no pending or contemplated 
marketing applications in an ICH region or until at least 2 years have elapsed since the formal 
discontinuation of clinical development of the investigational product. These documents should 
be retained for a longer period, however, if required by local regulations. No records will be 
destroyed without the written consent of the sponsor, if applicable. It is the responsibility of the 
sponsor to inform the investigator when these documents no longer need to be retained. 
 
15.5 Protocol Deviations 
A protocol deviation is any noncompliance with the clinical study protocol, Good Clinical 
Practice, or Manual of Procedures requirements. The noncompliance may be on the part of the 
subject, the investigator, or study staff. As a result of deviations, corrective actions are to be 
developed by the study staff and implemented promptly. 
All deviations from the protocol must be addressed in study subject source documents and 
promptly reported to the IRB and other regulatory bodies according to their requirements.  
16 Study Finances 
16.1 Funding Source 
The department of medical Oncology at Thomas Jefferson University is funding this trial. 
16.2 Conflict of Interest 
Any investigator who has a conflict of interest with this study (patent ownership, royalties, or 
financial gain greater than the minimum allowable by their institution, etc.) must have the conflict 
reviewed by a properly constituted Conflict of Interest Committee with a Committee-sanctioned 
conflict management plan that has been reviewed and approved by the study sponsor prior to 
participation in this study. All Jefferson University Investigators will follow the TJU Conflicts of 
Interest Policy for Employees (107.03). 
16.3 Subject Stipends or Payments 
Subjects will not receive payment for participation in the study. 
17 Publication and Data Sharing Policy 
This study will comply with the NIH Public Access Policy, which ensures that the public has 
access to the published results of NIH funded research. It requires scientists to submit final 
peer-reviewed journal manuscripts that arise from NIH funds to the digital archive PubMed 
Central upon acceptance for publication. 
The International Committee of Medical Journal Editors (ICMJE) member journals have adopted 
a clinical trials registration policy as a condition for publication. The ICMJE defines a clinical trial 
as any research project that prospectively assigns human subjects to intervention or concurrent 

 
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comparison or control groups to study the cause-and-effect relationship between a medical 
intervention and a health outcome. Medical interventions include drugs, surgical procedures, 
devices, behavioral treatments, process-of-care changes, and the like. Health outcomes include 
any biomedical or health-related measures obtained in patients or participants, including 
pharmacokinetic measures and adverse events. The ICMJE policy requires that all clinical trials 
be registered in a public trials registry such as ClinicalTrials.gov, which is sponsored by the 
National Library of Medicine. Other biomedical journals are considering adopting similar 
policies. The ICMJE does not review specific studies to determine whether registration is 
necessary; instead, the committee recommends that researchers who have questions about the 
need to register err on the side of registration or consult the editorial office of the journal in 
which they wish to publish. 
U.S. Public Law 110-85 (Food and Drug Administration Amendments Act of 2007 or FDAAA), 
Title VIII, Section 801 mandates that a "responsible party" (i.e., the sponsor or designated 
principal investigator) register and report results of certain "applicable clinical trials:"  
Trials of Drugs and Biologics: Controlled, clinical investigations, other than Phase I 
investigations, of a product subject to FDA regulation;  
Trials of Devices: Controlled trials with health outcomes of a product subject to FDA regulation 
(other than small feasibility studies) and pediatric postmarket surveillance studies. 
NIH grantees must take specific steps to ensure compliance with NIH implementation of 
FDAAA. 
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