The feasibility of finger prick autologous blood (FAB) as a novel treatment for severe dry eye disease (DED): protocol for a randomised controlled trial

Introduction Patients with severe dry eye disease (DED) often have limited treatment options with standard non-surgical management focused on the use of artificial tears for lubrication and anti-inflammatory drugs. However, artificial tears do not address the extraordinary complexity of human tears. Crudely, human tears with its vast constituents is essentially filtered blood. Blood and several blood-derived products including autologous serum, have been studied as tear substitutes. This study proposes to test the use of whole, fresh, autologous blood obtained from a finger prick for treatment of severe DED. Methods and analysis The research team at the two participating sites will approach patients with severe DED for this study. Recruitment will take place over 12 months and we expect to recruit 60 patients in total. The primary outcome of this feasibility study is to estimate the proportion of eligible patients approached who consent to and comply with study procedures including treatment regimen and completion of required questionnaires. The secondary outcome measures, although not powered for in this feasibility, include corneal inflammation (assessed by the Oxford corneal staining guide), patient pain and symptoms scores (assessed by the Ocular Surface Disease Index Score), and objective signs of DED as indicated by visual acuity (assessed by Schirmer’s test, tear break-up time, lower and/or upper tear meniscus height measurement). Other secondary outcomes include patients’ quality of life (assessed using the validated EQ-5D-5L Questionnaire), cost to the National Health Service (NHS) and patient (assessed via use of NHS services and privately purchased over-the-counter treatment related to DED) and safety measure of pressure within the eye (assessed by the Intraocular Pressure (IOP) Score). Ethics and dissemination This protocol and any subsequent amendments, along with any accompanying material provided to the participant in addition to any advertising material used in this trial have been approved by the East of England - Cambridgeshire and Hertfordshire Research Ethics Committee (REC reference: 17/EE/0508). Written approval from the committee was obtained and subsequently submitted to the respective Trust’s Research and Development (R&D) office with final NHS R&D approval obtained. Data obtained from this study will be published in a suitable peer-review journal and will also presented at international ophthalmic conferences including the American Academy of Ophthalmology, the Royal College of Ophthalmology Annual Congress, the Association for Research and Vision and Ophthalmology, and the European Society of Cataract and Refractive Surgery. Information will be provided to patient groups and charities such as the Sjogren’s Society and the Royal National Institute of Blind People. This will also be shared with the study participants as well as with relevant patient groups and charities. Trial registration number NCT03395431; Pre-results.

Patients with severe dry eye disease often have limited treatment options with standard nonsurgical management focused on the use of artificial tears for lubrication and antiinflammatory drugs. However, artificial tears do not address the extraordinary complexity of human tears. Crudely, human tears with its vast constituents is essentially filtered blood.
Blood and several blood-derived products including autologous serum, have been studied as tears substitutes. This study proposes to test the use of whole, fresh, autologous blood obtained from a fingerprick for treatment of severe dry eye disease.

Methods and Analysis
The research team at the two participating sites will approach patients with severe dry eye disease for this study. Recruitment will take place over 12 months and we expect to recruit 60 patients in total. The primary outcome of this feasibility study is to estimate the proportion of eligible patients approached who consent to and comply with study procedures including treatment regimen and completion of required questionnaires. The secondary outcome measures, although not powered for in this feasibility, include corneal inflammation (assessed by Oxford Corneal Staining Guide), patient pain and symptoms scores (assessed by Ocular Surface Disease Index (OSDI) score), and objective signs of dry eye disease as indicated by visual acuity (assessed by Schirmer's test, tear breakup time, lower and/or upper tear meniscus height measurement). Other secondary outcomes include patients' quality of life (assessed using the validated EQ-5D-5L questionnaire), cost to the NHS and patient (assessed via use of NHS services and privately purchased over the counter treatment related to dry eyes disease) and safety measure of pressure within the eye (assessed by intra ocular pressure (IOP) score).

Ethics and Dissemination
This trial is ongoing and received a favourable research ethics opinion from the East of  (1). If left untreated, DED can lead to severe reduction in the quality of life of the sufferer. It can also cause loss of vision, pain in response to light, painful recurring stabbing sensations, and the feeling of grit in the affected eye(s). Studies show that moderate to severe DED can be as disruptive to patient quality of life as angina (2,3). Despite this, only less than 2% of all United Kingdom (UK) medical research funding is directed at diseases of the eye, leaving eye research critically underfunded (4). No curative agents for DED exist. The treatment of the disorder is essentially symptomatic with standard non-surgical treatment focused on the use of artificial tears for lubrication and anti-inflammatory drugs.
Anti-inflammatory agents used in DED include topical steroids and cyclosporine drops but these can cause side effects that limit their long term use (5). Topical steroids can also cause cataracts and glaucoma, all of which further limit their use. Surgical options for DED include punctal occlusion to reduce tear drainage, punctal cauterization (burning the drainage channel of tears and preventing their outflow) and partial suturing of the eyelids. However, these treatment options are associated with adverse effects and varying levels of patient tolerability.
Overall, available conventional treatment options for DED often only alleviate symptoms, have limited effectiveness, and in most cases patients may fail to respond-although the exact rate of treatment failure is unavailable in the published literature.
The mainstay of non-surgical treatment for DED focuses on the use of artificial tears. These can be purchased over the counter and include brand name products such as Viscotears® and Optrex® (6). Human tears contains an extensive range of growth factors, immunoglobulins, enzymes, cytokines, vitamins and electrolytes and these have been shown in numerous studies to be essential for the maintenance and proliferation of corneal epithelial cells as well as for defence against infection (7). It is well known that artificial tears fail to account for the extraordinarily complex composition of the natural tear film. Also, many artificial tears contain preservatives that have been shown to adversely affect the cornea (8). Crudely, human tears with its vast constituents is essentially filtered blood and as such is an obvious source for a "tear mimic" containing all tear constituents. Blood and several blood-derived products including autologous serum have been studied as tears substitute candidates. Autologous serum (AS) eye drops have been found in uncontrolled trials to be beneficial by improving the ocular surface and reducing symptoms (9)(10)(11). However, a Cochrane review concluded that there is inconsistency in evidence on its benefit (12). Obtaining autologous serum requires frequent drawing of blood from the patient-a feature that excludes patients with anaemia or heart failure from using AS. Furthermore it also appears that 100% autologous serum is more beneficial than 50% serum and requires larger volumes of blood and/or more frequent venesection (10). Patients using AS also require access to a fridge, as the product needs to be stored at low temperatures; a factor that is likely to be inconvenient for patients. In addition, AS is obtained by processing clotted blood with the initial cost to the NHS of £1653.56, and subsequent three-monthly cost of £1131.27 per patient.
The relatively high cost represents the biggest hurdle in the use of AS and is often the reason for delay or inaccessibility in its use as a treatment for DED..We propose that finger prick autologous blood is a simpler, cost-effective and possibly more acceptable method for treating dry eye disease. For this reason, this study proposes to test the use of fingerprick autologous blood (FAB) in which blood obtained from a small finger-prick is applied to the eye. Autologous fresh blood is already used in the sub-conjunctiva to help heal leaking trabeculectomy blebs (13)(14)(15)(16). It is also used to help attach limbal autografts in cases of pterygium (17), and in vitreoretinal macular hole surgery (18)(19)(20)(21) with no adverse effects reported. The FAB method can be used in patients who are awaiting conventional treatment for autologous blood.

Rationale and Risks/Benefits
This study proposes to test the feasibility of the use of whole, fresh, autologous blood as a treatment for severe dry eye disease. The blood can be obtained from a clean finger, pricked by the patient using a diabetic lancet and administered immediately to the eye. This allows the delivery of not only most of the aforementioned beneficial components of tears but additional growth factors and proteins, fresh and unprocessed, which can help heal the ocular surface. If validated, this may replace current autologous serum practice and its ease of use,

Study Design
This is a single blind, two-arm feasibility randomised controlled trial of FAB with conventional treatment versus conventional therapy alone for severe dry eye disease, including a qualitative process evaluation (Figure 1 provides details of the study scheme).
This trial is ongoing and open to recruitment at two participating sites: Bedford Hospital NHS Trust and Moorfields Eye Hospital London. Recruitment into the trial commenced in April 2018 and is due to finish by April 2019.

Sample Size
As this is a feasibility trial, there was no formal sample size calculations (23). The aim of the study is to recruit sufficient patients to evaluate the acceptability and feasibility of the intervention. An outcome of this study will be to estimate parameters such as the standard deviation for a sample size calculation of a subsequent full-scale trial. A sample of 52 patients (26 in each arm) is considered adequate for obtaining reliable sample size estimates (24,25). To allow for a conservative attrition rate of 10%, we aim to recruit 60 patients into the trial.

Inclusion Criteria
• Patient age ≥ 18 years • Severe symptomatic dry eye disease diagnosed by: Ocular surface disease index (OSDI) score of greater than 33; OR Oxford Corneal Staining grade 2 or greater; OR Schirmer's without anaesthesia <5mm at 5 minutes • Patients on artificial tears and/or lubricating drops/gel four times a day • Patient able to give consent • Patients able and willing to complete the quality of life (QoL) questionnaires required for the study.

Intervention • Arm A -FAB plus conventional treatment
The patients will use FAB therapy alongside conventional therapy (artificial tears, cyclosporine drops or punctal plugs/cautery) as recommended by their treating ophthalmologist. A fingertip of the hand will be wiped with an alcohol steret and self-pricked using a standard diabetic lancet. The drop of blood is produced as normal and applied to the lower fornix of the affected eye(s) with the lower lid pulled down slightly by the patient. The blood will be applied four times a day. A fresh finger will be used for each eye and blood wiped away with the alcohol steret. Patients will be advised to keep nails short and nail varnish avoided. FAB will be applied at least 15 minutes after any artificial tears and no drops for at least half an hour afterwards.

• Arm B -Conventional treatment only
The patients will use conventional therapy (artificial tears, cyclosporin drops and punctal plugs/cautery) as recommended by their treating ophthalmologist.

Qualitative Approach
A nested qualitative approach using in-depth, semi-structured interviews will help us understand the lived experience of people using FAB and factors relating to the ways that clinical departments and healthcare professionals adapt to working with FAB. Topics, questions and probes in the interviews will emerge from three sources: the relevant literature; our experience of and involvement in clinical ophthalmology; and consultation with patients.
In this feasibility study, we will use a convenience sampling technique for the qualitative work. We will interview all clinicians who administer the intervention in the study and we will aim to interview 10 patients (adjusted for setting, gender and age group representation).
Given the resources, we will aim to draw findings from a larger sample in a future study. This phase will work in parallel to the quantitative data collection so that emerging themes can be investigated in later interviews. Even though the feasibility study sample is reasonably 10 limited, we anticipate some useful insights about the use of FAB to occur because of the triangulation between the RCT and the qualitative findings (for example, reasons for partial adherence).

Economic Evaluation
Good practice recommendations for cost-effectiveness analyses suggest concentrating on the measurement of large cost drivers, with less focus on resources that are not expected to differ between different treatments (26). Estimation of cost-effectiveness is therefore an iterative process and by including a health economic component in a feasibility it is possible to consider how the methods might be refined in any future definitive study. In order to estimate costs, informed by previous data collection instruments (Database of Instruments for Resource Use Measurement: http://www.dirum.org/) and NICE guidance (27), a self-report resource use questionnaire will be devised. This will capture the use of NHS services and privately purchased over the counter treatments such as OPTREX. For benefits, the EQ-5D-5L (28) will be used to measure quality of life, enabling QALY (Quality Adjusted Life Year) scores to be calculated. The main purpose of the economic analysis is to inform the decision regarding how and what cost and effect data would be collected within a more definitive study. In order to inform this decision, we will estimate completion rates and seek to identify big cost drivers.

Outcome Assessment
The primary outcome measure is the proportion of eligible patients approached who consent to and comply with the trial protocol. This will be assessed by: number of eligible patients within the study population and recruitment timeframe, percentage of eligible patients accepted for randomisation, patient compliance with trial protocol measured as per selfreported adherence to intervention, and percentage of patients completing study. The

Data Analysis
A detailed data validation will examine completeness, existence and accuracy of collected data to assess data quality and identify missing and conflicting data. The statistical analysis will be performed on an intention-to-treat basis (i.e. inclusion of patients randomly assigned, regardless of adherence, actual treatment received, and subsequent withdrawal of treatment and/or deviation from the protocol and per-protocol (i.e. inclusion of those who completed the treatment as planned) (29) and will be reported according to 2010 CONSORT guidelines (30). All statistical analyses will be completed using SPSS Statistics v.22.0 (31). A p-value of <0.05 will be accepted as statistical significance. Conforming to recommendations for feasibility studies, data analysis will be primarily descriptive, with means, standard deviations and frequency counts calculated for all variables of interest.
Exploratory efficacy analysis will compare the primary outcome variables derived from the data collected at 8 weeks between the two arms using a marginal general linear model (GLM) with robust standard errors, to allow for clustering by group. Secondary outcomes between the intervention and control groups will be compared at 4 weeks, 8 weeks and 12 weeks (4 weeks post treatment). Safety outcome measures will include IOP rise and any reported infection. Secondary analyses of the primary outcome, controlling for baseline covariates, will be performed using proportional hazards regression models (32). Analysis of treatment discontinuation adjusted for clinical site, baseline status, and whether the patient is randomized to the FAB or control (conventional treatment only) group will be conducted.
Any interaction between baseline status or regimen and treatment will also be assessed.
Interactions between significant covariates and treatment effects will be assessed in the context of the proportional hazards models.
Interviews will be recorded, transcribed and analysed using the framework analysis. We will structure the analysis of collected data to fit two evaluation frameworks relevant to the successful implementation of FAB: • Patient-oriented: Understand the lived experience of patients using FAB with emphasis on intervention acceptability, perceived enablers and barriers for adherence, • Organisation-oriented: Whether and how the use of FAB affected workload and/or workflow; identify process change, adaptation challenges, skills gap; establish treatment fidelity and clinician acceptability.

Study Procedures Informed Consent Procedures
It is the responsibility of the Investigator, or appropriately GCP trained person delegated by the Investigator as documented in the site delegation log, to obtain written informed consent from each participant prior to any participation/study specific procedures. This should follow adequate explanation of the aims, methods, anticipated benefits and potential hazards of the study. The participants will be given ample time to consider giving their consent for the study. For this study a minimum of 24 hours will be given during which the consenting team will be contactable over the phone to answer any questions. The date that the Participant Information Sheet (PIS) is given to the participant must be documented within the patient's notes to confirm that sufficient time was given. The Investigator (or other qualified person) will explain to the potential participant that they are free to refuse any involvement within the study or alternatively withdraw their consent at any point during the study and for any reason.
If there is any further safety information which may result in significant changes in the risk/benefit analysis, the PIS and Informed Consent Form (ICF) will be reviewed and updated accordingly. All participants that are actively enrolled on the study will be informed of the updated information and given a revised copy of the PIS/ICF in order to confirm their wish to continue on the study.

Screening Procedures
Patients will have undergone the below procedures as per standard of care which will in turn feed in to the investigator for their review prior to approaching the potential participant.
-Ocular surface disease index (OSDI) score of greater than 33; OR -Oxford Corneal Staining grade 2 or greater; OR -Schirmer's without anaesthesia <5mm at 5 minutes The results of these which will be reviewed by investigator prior to approaching the potential participant.

Randomisation Procedures
Randomisation will be carried out by the Anglia Ruskin Clinical Trials Unit (ARCTU) using the SEALED ENVELOPE randomisation service. SEALED ENVELOPE is a randomisation and online database service developed for clinical trial services. It is an internet based system and will be set for this study by ARCTU in accordance with the protocol. Enrolled patients will undergo 1:1 block randomisation to Arm A or B. The system stores the pre-determined sequence of randomisation and this list is visible to neither the investigator nor ARCTU staff.
Once a patient has consented to take part in the trial, they will be randomly allocated to either Arm A to receive FAB and convention therapy or Arm B to receive conventional therapy only. The Research Nurse or Fellow or designated staff will log on to a web browser application and enter the patient's eligibility factors into the system. The treatment allocation will be sent to the research team who will make the necessary arrangements for the patient's treatment plan.
The treating ophthalmologist in clinic will prescribe and counsel all patients on the correct technique for conventional treatment. The unblinded research nurses will provide additional training to the patients on the method of delivering FAB, including the use of a training video. Patients will be advised by the research team to inform their ophthalmologist if they have been started on any new treatment during the trial period by other clinicians or themselves.

End of Study Definition
The definition of the end of study will be the point where the last patient recruited had the last follow-up visit.

Participant Withdrawal
Patients will be withdrawn from study based on the following:

PATIENT AND PUBLIC INVOLVEMENT
Patients who participated in a previous pilot (exploratory) study conducted by the study authors (22) were involved in a focus group meeting and provided inputs on research design and study procedures. Two of the patients also participated in an advisory group for this trial provided input on prioritising the research questions and also reviewed the study protocol and participant information resources. Patients in this trial will be invited to request study results from the research team if interested.

Objectives
To determine the feasibility of a definitive randomised controlled trial (RCT) to determine effectiveness of the use of fresh autologous blood (FAB) compared to conventional treatment for patients with dry eye syndrome. Data analysis will be primarily descriptive with means, standard deviations and frequency counts calculated for all variables of interest. Exploratory statistical analysis will compare the primary outcome and secondary outcomes between the intervention and control groups at 4 weeks, 8 weeks and 12 weeks (4 weeks post treatment)   No curative agents for DED exist. The treatment of the disorder is essentially symptomatic with standard non-surgical treatment focused on the use of artificial tears for lubrication and anti-inflammatory drugs. Anti-inflammatory agents include topical steroids and ciclosporin drops but these can cause side effects that limit their long term use (4). Topical steroids can also cause cataracts and glaucoma, all of which further limit its use. Surgical options for DED include punctal occlusion to reduce tear drainage, punctal cauterization (burning the drainage channel of tears, preventing their outflow) and partial suturing of the eyelids; but these are associated with adverse effects and varying levels of patient tolerability. Overall, available conventional treatment options for DED often only alleviate symptoms, have limited effectiveness, and in most cases patients may fail to respond; although the exact rate of treatment failure is unavailable in the published literature.

Number of Participants
The mainstay of non-surgical treatment for DED focuses on the use of artificial tears. These can be purchased over the counter and include brand name products such as Viscotears® and Optrex® (5). Human tears contains an extensive range of growth factors, immunoglobulins, enzymes, cytokines, vitamins and electrolytes and these have been shown in numerous studies to be essential for the maintenance and proliferation of corneal epithelial cells as well as for defence against infection (6). It is well known that artificial tears fails to account for the extraordinarily complex composition of the natural tear film. Also, many artificial tears contain preservatives that have been shown to adversely affect the cornea (7). Crudely, human tears with its vast constituents is essentially filtered blood and as such is an obvious source for a "tear mimic" containing the substances of tears. Blood, and several blood derived products, including autologous serum, have been studied as tear substitute candidates.
Autologous serum (AS) eye drops have been found in uncontrolled trials to be beneficial in DED patient by improving the ocular surface and reducing symptoms (8)(9)(10). However, a Cochrane review concluded that there is inconsistency in evidence on its benefit (11).
Obtaining autologous serum requires frequent drawing of blood from the patient-a feature that excludes patients with anaemia or heart failure from using AS. Furthermore it also appears that 100% autologous serum is more beneficial than 50% serum and requires larger volumes of blood and/or more frequent venesection (9). Patients using AS also require access to a fridge as the product needs to be stored at low temperatures; a factor that is likely to be inconvenient for patients. In addition, AS is obtained by processing clotted blood which is often too expensive for the health service to consistently purchase, given the initial cost of £1653.56 and subsequent three-monthly cost of £1131.27 per patient.
The relatively high cost represents the biggest hurdle in the use of AS and is often the reason for delay or inaccessibility in starting treatment for DED using AS. However, we propose that finger prick autologous blood may be a simpler, cost-effective and possibly more acceptable method for treating dry eye disease. For this reason, this study proposes to test the use of finger prick autologous blood (FAB) technique in which whole blood is applied to the eye from a cleaned finger. Autologous fresh blood is already used subconjunctivally to help heal leaking trabeculectomy blebs (12)(13)(14)(15). It is also used to help attach limbal autografts in cases of pterygium (16) and vitreoretinal macular hole surgery (17)(18)(19)(20), with no adverse effects reported. The FAB method can be used immediately for patients who are awaiting conventional treatment for autologous blood. The objective of this study is therefore to investigate if serum via a drop of fresh blood is an effective treatment for severe dry eye disease which currently require venesected autologous blood and whether this would be particularly useful in the group of patients in whom venesection is contraindicated.

Rationale and Risks/Benefits
This study proposes to test the use of whole, fresh, autologous blood as a treatment for severe dry eye disease. The blood can be obtained from a cleaned finger, pricked by the patient using a diabetic lancet and administered immediately to the dry eye. This allows the delivery of not only most of the aforementioned beneficial components of tears but additional growth factors and proteins, fresh and unprocessed, which can help heal the ocular surface. If validated, this may replace current autologous serum practice and its ease of use, vastly reduced cost with greater convenience may mean that it could be extended to other ocular surface diseases.

Primary Objective and Endpoints
The primary objective is to determine the feasibility of a definitive randomised controlled trial (RCT) to evaluate effectiveness of the use of fresh autologous blood (FAB) compared to conventional treatment for patients with dry eye syndrome. This will involve specifically assessing the following endpoints: i. Number of eligible patients within the study population and recruitment time frame.
ii. Percentage of eligible patients accepted for randomisation.
iii. Patient compliance with trial protocol, measured as per self-reported adherence to intervention.
iv. Percentage of patients completing study.

Secondary Objective and Endpoints
The secondary objectives are:  To determine the effectiveness of the trial intervention and further explore the acceptability of the study design.
 To explore the feasibility of collecting resource use and quality of life data, to inform the design of the health economics component of a future definitive trial.
The secondary end points, which have not been powered for within the feasibility study, include:

Trial Design
Single-blind two-arm feasibility randomised controlled trial of FAB with conventional treatment versus conventional therapy alone for dry eye disease, including a qualitative process evaluation.

 Arm A -FAB plus conventional treatment
The patients will use FAB therapy alongside conventional therapy (artificial tears, ciclosporin drops and punctal plugs/cautery) as recommended by their treating ophthalmologist. A fingertip of the hand will be wiped with an alcohol steret and self-pricked using a standard diabetic lancet. The drop of blood is produced as normal and applied to the lower fornix of the affected eye(s) with the lower lid pulled down slightly by the patient. The blood will be applied four times a day. A fresh finger should be used for each eye. The blood will be wiped away with the alcohol steret. Nails should be kept short and nail varnish should be avoided. FAB should be applied at least 15 minutes after any artificial tears and no drops for at least half an hour afterwards.

 Arm B -Conventional treatment only
The patients will use conventional therapy (artificial tears, cyclosporin drops and punctal plugs/cautery) as recommended by their treating ophthalmologist.

Setting
The study will take place at NHS sites. The main site will be Bedford Hospital NHS Trust and Moorfields Eye Hospital London. Patients will be recruited from corneal and external eye disease clinics at these sites.

Informed Consent Procedures
It is the responsibility of the Investigator, or appropriately GCP trained person delegated by the Investigator as documented in the site delegation log, to obtain written informed consent from each participant prior to any participation/study specific procedures. This should follow adequate explanation of the aims, methods, anticipated benefits and potential hazards of the study. The participants will be given ample time to consider giving their consent for the study.
For this study a minimum of 24 hours will be given during which the consenting team will be contactable over the phone to answer any questions. The date that the Participant Information Sheet (PIS) is given to the participant must be documented within the patient's notes to confirm that sufficient time was given. The Investigator (or other qualified person) will explain to the potential participant that they are free to refuse any involvement within the study or alternatively withdraw their consent at any point during the study and for any reason.
If there is any further safety information which may result in significant changes in the risk/benefit analysis, the PIS and Informed Consent Form (ICF) will be reviewed and updated accordingly. All participants that are actively enrolled on the study will be informed of the updated information and given a revised copy of the PIS/ICF in order to confirm their wish to continue on the study.

Screening Procedures
Patients will have undergone the below procedures as per standard of care which will in turn feed in to the investigator for their review prior to approaching the potential participant.
-Ocular surface disease index (OSDI) score of greater than 33; OR -Oxford Corneal Staining grade 2 or greater; OR -Schirmer's without anaesthesia <5mm at 5 minutes The results of these which will be reviewed by investigator prior to approaching the potential participant.

Randomisation Procedures
Randomisation will be carried out by the Anglia Ruskin Clinical Trials Unit (ARCTU) using the SEALED ENVELOPE randomisation service. SEALED ENVELOPE is a randomisation and online database service developed for clinical trial services. It is an internet based system and will be set for this study by ARCTU in accordance with the protocol. Enrolled patients will undergo 1:1 block randomisation to Arm A or B. The system stores the pre-determined sequence of randomisation and this list is visible to neither the investigator nor ARCTU staff.
Once a patient has consented to take part in the trial, they will be randomly allocated to either Arm A to receive FAB and convention therapy or Arm B to receive conventional therapy only.
The Research Nurse or Fellow or designated staff will log on to a web browser application and enter the patient's eligibility factors into the system. The treatment allocation will be sent to the research team who will make the necessary arrangements for the patient's treatment plan.
The treating ophthalmologist in clinic will prescribe and counsel all patients on the correct technique for conventional treatment. The unblinded research nurses will provide additional training to the patients on the method of delivering FAB, including the use of a training video.

End of Study Definition
The definition of the end of study will be the point where the last patient recruited had the last follow-up visit.

Participant Withdrawal
Patients will be withdrawn from study based on the following: -If finger sore from repeated prick and patient does not want to use another finger.
Any adverse effects will be reported back to the lead investigator at the site and if necessary patients

Adverse Event (AE)
An AE is any untoward medical occurrence in a participant to whom a medicinal product has been administered, including occurrences which are not necessarily caused by or related to that product. An AE can therefore be any unfavourable and unintended sign (including an abnormal laboratory finding), symptom or disease temporarily associated with study activities.

Serious Adverse Event (SAE)
An SAE fulfils at least one of the following criteria: -Is fatalresults in death (NOTE: death is an outcome, not an event) -Is life-threatening -Requires inpatient hospitalisation or prolongation of existing hospitalisation -Results in persistent or significant disability/incapacity -Is a congenital anomaly/birth defect -Is otherwise considered medically significant by the Investigator

Seriousness
The Chief/Principal Investigator responsible for the care of the participant, or in his absence an authorised medic within the research team, is responsible for assessing whether the event is serious according to the definitions given in section 5.1.

Causality
The Investigator must assess the causality of all serious adverse events in relation to the trial treatment according to the definition given

Expectedness
The investigator must assess the expectedness of all SAEs according to the definition given. If the SAE is unexpected and related, then it needs immediate reporting.

Severity
The Investigator must assess the severity of the event according to the following terms and assessments. The intensity of an event should not be confused with the term "serious" which is a regulatory definition based on participant/event outcome criteria.

Mild: Some discomfort noted but without disruption of daily life
Moderate: Discomfort enough to affect/reduce normal activity Severe: Complete inability to perform daily activities and lead a normal life

Notification and reporting Adverse Events or Reactions
If the AE is not defined as SERIOUS, the AE is recorded in the study file and the participant is followed up by the research team. The AE is documented in the participants' medical notes (where appropriate) and the CRF.

Notification and Reporting of Serious Adverse Events
Serious Adverse Event (SAEs) that are considered to be 'related' and 'unexpected' are to be reported to the sponsor within 24 hours of learning of the event and to the Main REC within 15 days in line with the required timeframe. For further guidance on this matter, please refer to Appendix.

Urgent Safety Measures
The CI may take urgent safety measures to ensure the safety and protection of the clinical trial participants from any immediate hazard to their health and safety, in accordance with Regulation 30. The measures should be taken immediately. In this instance, the approval of the Licensing Authority Approval prior to implementing these safety measures is not required. However, it is the responsibility of the CI to inform the sponsor and Main Research Ethics Committee (via telephone) of this event immediately.
The CI has an obligation to inform both the Main Ethics Committee in writing within 3 days, in the form of a substantial amendment. The sponsor must be sent a copy of the correspondence with regards to this matter. For further guidance on this matter, please refer to Appendix.

5.6
Annual Safety Reporting The CI will send the Annual Progress Report to the main REC using the NRES template (the anniversary date is the date on the MREC "favourable opinion" letter from the MREC) and to the sponsor. Please see appendix for further information.

Overview of the Safety Reporting Process/Pharmacoviligance responsibilities
The CI/PI has the overall pharmacovigilance oversight responsibility. The CI/PI has a duty to ensure that pharmacovigilance monitoring and reporting is conducted in accordance with the sponsor's requirements.  (24)]. The statistical analysis will be performed on an intention-totreat basis and per-protocol and will be reported according to 2010 CONSORT guidelines (25).
All statistical analyses will be completed using SPSS Statistics V.22.0 (26). A p-value of <.05 will be accepted as statistical significance.
Conforming to recommendations for feasibility studies, data analysis will be primarily descriptive, with means, standard deviations and frequency counts calculated for all variables of interest. Exploratory efficacy analysis will compare the primary outcome variables derived from the data collected at 8 weeks between the two arms using a marginal general linear model (GLM) with robust standard errors, to allow for clustering by group. Secondary outcomes between the intervention and control groups will be compared at 4 weeks, 8 weeks and 12 weeks (4 weeks post treatment). Safety outcome measures will include IOP rise and any reported infection.
Secondary analyses of the primary outcome, controlling for baseline covariates, will be performed using proportional hazards regression models (27). Analysis of treatment discontinuation adjusted for clinical site, baseline status, and whether the patient is randomized to the FAB or control (conventional treatment only) group will be conducted. Any interaction between baseline status or regimen and treatment will also be assessed. Interactions between significant covariates and treatment effects will be assessed in the context of the proportional hazards models.

Qualitative approach
A nested qualitative approach using in-depth, semi-structured interviews will help us understand the lived experience of people using FAB and factors relating to the ways that clinical departments and healthcare professionals adapt to working with FAB. Topics, questions and probes in the interviews will emerge from three sources: a) the relevant literature; b) our experience of and involvement in clinical ophthalmology; c) consultation with patients. In this feasibility study, we will use a convenience sampling technique for the qualitative work. We will interview all clinicians who administer the intervention in the study and we will aim to interview 10 patients (adjusted for setting, gender and age group representation). Given the resources, we will aim to draw findings from a larger sample in a future study. Interviews will be recorded, transcribed and analysed using the framework analysis. We will structure the analysis of collected data to fit two evaluation frameworks relevant to the successful implementation of FAB: 1) Patient-oriented: Understand the lived experience of patients using FAB with emphasis on intervention acceptability, perceived enablers and barriers for adherence, perceived clarity of advice and guidance and factors relating to the initial uptake of the intervention.
2) Organisation-oriented: Whether and how the use of FAB affected workload and/or workflow; identify process change, adaptation challenges, skills gap; establish treatment fidelity and clinician acceptability.
This phase will work in parallel to the quantitative data collection so that emerging themes can be investigated in later interviews. Even though the feasibility study sample is reasonably limited, we anticipate some useful insights about the use of FAB to occur because of the triangulation between the RCT and the qualitative findings (e.g. reasons for partial adherence).

Economic Evaluation
Good practice recommendations for cost-effectiveness analyses (28) suggest concentrating on the measurement of large cost drivers, with less focus on resources that are not expected to differ between different treatments. Estimation of cost-effectiveness is therefore an iterative and what cost and effect data would be collected within a more definitive study. in order to inform this decision, we will estimate completion rates and seek to identify big cost drivers.

Confidentiality
The Investigator has a responsibility to ensure that participant anonymity is protected and maintained. They must also ensure that their identities are protected from any unauthorised parties. Information with regards to study participants will be kept confidential and managed in accordance with the Data Protection Act, NHS Caldicott

Case Report Form
Project data collection will be managed by the Clinical Trials Unit Data Manager. The responsibility for data entry rests with the Research Nurse or designated staff who is supported by the Investigator. ARCTU uses an online data management system called MACRO to design and manage eCRFs (Electronic Case Report Forms). ARCTU will work together with the sponsor to design and validate the data collection tools so that they are appropriate for this study. Once a patient is enrolled on the study the research team can access these forms remotely through the Internet portal and study data will be entered and captured for the study. All data will be in anonymised form and patients will be identifiable only by study number. Data will be remotely monitored by ARCTU and discussed at data monitoring committee meetings. Any inconsistencies, validation errors or inaccuracies will be reported to the lead investigator regularly. Once data collection is complete and the data has been validated, a data lock will be performed and analysis will begin.

Record Retention and Archiving
During the course of research, all records are the responsibility of the Chief Investigator and must be kept in secure conditions. When the research trial is complete, it is a requirement of the Research Governance Framework and Trust Policy that the records are kept for a further 20 years.

Compliance
The CI will ensure that the trial is conducted in compliance with the principles of the Declaration of Helsinki (1996), and in accordance with all applicable regulatory requirements including but not limited to the Research Governance Framework, Trust and Research Office policies and procedures and any subsequent amendments.

Ethical Considerations
This protocol and any subsequent amendments, along with any accompanying material provided to the participant in addition to any advertising material will be submitted by the Investigator to an Independent Research Ethics Committee.
Written Approval from the Committee will be obtained and subsequently

Audit and Inspection
Auditing: Definition "A systematic and independent examination of trial related activities and documents to determine whether the evaluated trial related activities were conducted, and the data were recorded, analysed and accurately reported according to the protocol, sponsor's standard operating procedures (SOPs), Good Clinical Practice (GCP), and the applicable regulatory requirement(s)." The FAB in DED study may receive an audit by any of the methods listed below: -A project may be identified via the risk assessment process.
-An individual investigator or department may request an audit.
-A project may be identified via an allegation of research misconduct or fraud or a suspected breach of regulations.
-Projects may be selected at random. The Department of Health states that Trusts should be auditing a minimum of 10% of all research projects.
-Projects may be randomly selected for audit by an external organisation.
Internal audits will be conducted by a sponsor's representative 7.8

Non-Compliance
Non-compliance can be defined as 'a noted systematic lack of both the CI and the study staff adhering to SOPs/protocol/ICH-GCP, which leads to prolonged collection of deviations, breaches or suspected fraud.' These non-compliances may be captured from a variety of different sources including monitoring visits, CRFs, communications and updates. The sponsor will maintain a log of the non-compliances to ascertain if there are any trends developing which to be escalated.
The sponsor will assess the non-compliances and action a timeframe in which they need to be dealt with. Each action will be given a different timeframe dependant on the severity. If the actions are not dealt with accordingly, the sponsor will agree an appropriate action, including an on-site audit. A Trial Management Group (TMG) has been formed comprising the Chief Investigator, other lead investigators (clinical and non-clinical) and members of the Data Centres. The TMG will be responsible for the day-to-day running and management of the trial and will meet at least 3 times a year by teleconference.

Trial Steering Committee (TSC)
The Trial Steering Committee (TSC) has membership from TMG plus independent members, including the chair. The role of the TSC is to provide overall supervision for the trial and provide advice through its independent Chairman. The ultimate decision for the continuation of the trial lies with the TSC.

Independent Data Monitoring Committee (IDMC)
The Independent Data Monitoring Committee (IDMC) is the only group who sees the confidential, accumulating data to the trial. Reports to the IDMC will be produced by the trial statistician. The IDMC will meet within 6 months of the trial opening with the frequency of meetings dictated by the IDMC. The IDMC will consider the report for the interim analysis based on the first 30 patients and will decide, keeping in mind the rules described in the statistics section, whether to recommend that the study should continue or be closed. The primary outcome measure for the study addresses toxicity and so it aligns with the IDMC's precautionary role in patient safety. The details of the interim analysis will remain confidential to the IDMC until the study is closed.

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ABSTRACT Introduction
Patients with severe dry eye disease often have limited treatment options with standard nonsurgical management focused on the use of artificial tears for lubrication and antiinflammatory drugs. However, artificial tears do not address the extraordinary complexity of human tears. Crudely, human tears with its vast constituents is essentially filtered blood.
Blood and several blood-derived products including autologous serum, have been studied as tears substitutes. This study proposes to test the use of whole, fresh, autologous blood obtained from a fingerprick for treatment of severe dry eye disease.

Methods and Analysis
The research team at the two participating sites will approach patients with severe dry eye disease for this study. Recruitment will take place over 12 months and we expect to recruit 60 patients in total. The primary outcome of this feasibility study is to estimate the proportion of

Ethics and Dissemination
This trial is ongoing and received a favourable research ethics opinion from the East of England -Cambridgeshire and Hertfordshire Research Ethics Committee (REC reference: 17/EE/0508). The results of this study will be published in a suitable peer-review journal and also presented at international ophthalmic conferences. This will also be shared with the study participants as well as with relevant patient groups and charities.

Trial Registration Number: NCT03395431
Trial Sponsor: Bedford Hospital NHS Trust, Bedford, United Kingdom

Strength and Limitation of Study
• This single-blind randomised control trial addresses a significant problem by investigating the use of whole, fresh, autologous blood obtained from a fingerprick for treatment of severe dry eye disease.
• The study intervention is cheap, simple and with minimal risk of complication.
• The intervention will allow for the delivery of the beneficial components of tears including growth factors and proteins found in whole blood.
• Patients in the intervention arm may experience soreness in fingers that are repeatedly pricked and will be advised to use a different finger for each eye. Anti-inflammatory agents used in DED include topical steroids and cyclosporine drops but these can cause side effects that limit their long term use (5). Topical steroids can also cause cataracts and glaucoma, all of which further limit their use. Surgical options for DED include punctal occlusion to reduce tear drainage, punctal cauterization (burning the drainage channel of tears and preventing their outflow) and partial suturing of the eyelids. However, these treatment options are associated with adverse effects and varying levels of patient tolerability.
Overall, available conventional treatment options for DED often only alleviate symptoms, have limited effectiveness, and in most cases patients may fail to respond-although the exact rate of treatment failure is unavailable in the published literature.
The mainstay of non-surgical treatment for DED focuses on the use of artificial tears. These can be purchased over the counter and include brand name products such as Viscotears® and Optrex® (6). Human tears contains an extensive range of growth factors, immunoglobulins, enzymes, cytokines, vitamins and electrolytes and these have been shown in numerous studies to be essential for the maintenance and proliferation of corneal epithelial cells as well as for defence against infection (7). It is well known that artificial tears fail to account for the extraordinarily complex composition of the natural tear film. Also, many artificial tears contain preservatives that have been shown to adversely affect the cornea (8). Crudely, human tears with its vast constituents is essentially filtered blood and as such is an obvious source for a "tear mimic" containing all tear constituents. Blood and several blood-derived products including autologous serum have been studied as tears substitute candidates. Autologous serum (AS) eye drops have been found in uncontrolled trials to be beneficial by improving the ocular surface and reducing symptoms (9-11). However, a Cochrane review concluded that there is inconsistency in evidence on its benefit (12). Obtaining autologous serum requires frequent drawing of blood from the patient-a feature that excludes patients with anaemia or heart failure from using AS. Furthermore it also appears that 100% autologous serum is more beneficial than 50% serum and requires larger volumes of blood and/or more frequent venesection (10). Patients using AS also require access to a fridge, as the product needs to be stored at low temperatures; a factor that is likely to be inconvenient for patients. In addition, AS is obtained by processing clotted blood with the initial cost to the NHS of £1653.56, and subsequent three-monthly cost of £1131.27 per patient.
The relatively high cost represents the biggest hurdle in the use of AS and is often the reason for delay or inaccessibility in its use as a treatment for DED..We propose that finger prick autologous blood is a simpler, cost-effective and possibly more acceptable method for treating dry eye disease. For this reason, this study proposes to test the use of fingerprick autologous blood (FAB) in which blood obtained from a small finger-prick is applied to the eye. Autologous fresh blood is already used in the sub-conjunctiva to help heal leaking trabeculectomy blebs (13)(14)(15)(16). It is also used to help attach limbal autografts in cases of pterygium (17), and in vitreoretinal macular hole surgery (18)(19)(20)(21) with no adverse effects reported. The FAB method can be used in patients who are awaiting conventional treatment for autologous blood.

Rationale and Risks/Benefits
This study proposes to test the feasibility of the use of whole, fresh, autologous blood as a treatment for severe dry eye disease (detailed study protocol presented as supplementary file).
The blood can be obtained from a clean finger, pricked by the patient using a diabetic lancet and administered immediately to the eye. This allows the delivery of not only most of the aforementioned beneficial components of tears but additional growth factors and proteins, fresh and unprocessed, which can help heal the ocular surface. If validated, this may replace current autologous serum practice and its ease of use, vastly reduced cost with greater convenience may mean that it could be extended to other ocular surface diseases. The proposing team have completed an exploratory study on the use of fingerprick autologous blood (FAB) for persistent epithelial defects and severe dry eye disease and preliminary

PRIMARY AND SECONDARY OBJECTIVES
The primary objective is to determine the feasibility of a definitive randomised controlled

Study Design
This is a single blind, two-arm feasibility randomised controlled trial of FAB with conventional treatment versus conventional therapy alone for severe dry eye disease, including a qualitative process evaluation (Figure 1 provides details of the study scheme).
This trial is ongoing and open to recruitment at two participating sites: Bedford Hospital NHS Trust and Moorfields Eye Hospital London. Recruitment into the trial commenced in April 2018 and is due to finish by April 2019.

Sample Size
As this is a feasibility trial, there was no formal sample size calculations (23). The aim of the study is to recruit sufficient patients to evaluate the acceptability and feasibility of the intervention. An outcome of this study will be to estimate parameters such as the standard deviation for a sample size calculation of a subsequent full-scale trial. A sample of 52 patients (26 in each arm) is considered adequate for obtaining reliable sample size estimates (24,25). To allow for a conservative attrition rate of 10%, we aim to recruit 60 patients into the trial.

Participant Selection
Inclusion Criteria

Intervention • Arm A -FAB plus conventional treatment
The patients will use FAB therapy alongside conventional therapy (artificial tears, cyclosporine drops or punctal plugs/cautery) as recommended by their treating ophthalmologist. A fingertip of the hand will be wiped with an alcohol steret and self-pricked using a standard diabetic lancet. The drop of blood is produced as normal and applied to the lower fornix of the affected eye(s) with the lower lid pulled down slightly by the patient. The blood will be applied four times a day. A fresh finger will be used for each eye and blood wiped away with the alcohol steret. Patients will be advised to keep nails short and nail varnish avoided. FAB will be applied at least 15 minutes after any artificial tears and no drops for at least half an hour afterwards.

• Arm B -Conventional treatment only
The patients will use conventional therapy (artificial tears, cyclosporin drops and punctal plugs/cautery) as recommended by their treating ophthalmologist.

Qualitative Approach
A nested qualitative approach using in-depth, semi-structured interviews will help us understand the lived experience of people using FAB and factors relating to the ways that clinical departments and healthcare professionals adapt to working with FAB. Topics, questions and probes in the interviews will emerge from three sources: the relevant literature; our experience of and involvement in clinical ophthalmology; and consultation with patients.
In this feasibility study, we will use a convenience sampling technique for the qualitative work. We will interview all clinicians who administer the intervention in the study and we will aim to interview 10 patients (adjusted for setting, gender and age group representation).
Given the resources, we will aim to draw findings from a larger sample in a future study. This phase will work in parallel to the quantitative data collection so that emerging themes can be 10 investigated in later interviews. Even though the feasibility study sample is reasonably limited, we anticipate some useful insights about the use of FAB to occur because of the triangulation between the RCT and the qualitative findings (for example, reasons for partial adherence).

Economic Evaluation
Good practice recommendations for cost-effectiveness analyses suggest concentrating on the measurement of large cost drivers, with less focus on resources that are not expected to differ between different treatments (26). Estimation of cost-effectiveness is therefore an iterative process and by including a health economic component in a feasibility it is possible to consider how the methods might be refined in any future definitive study. In order to estimate costs, informed by previous data collection instruments (Database of Instruments for Resource Use Measurement: http://www.dirum.org/) and NICE guidance (27), a self-report resource use questionnaire will be devised. This will capture the use of NHS services and privately purchased over the counter treatments such as OPTREX. For benefits, the EQ-5D-5L (28) will be used to measure quality of life, enabling QALY (Quality Adjusted Life Year) scores to be calculated. The main purpose of the economic analysis is to inform the decision regarding how and what cost and effect data would be collected within a more definitive study. In order to inform this decision, we will estimate completion rates and seek to identify big cost drivers.

Outcome Assessment
The primary outcome measure is the proportion of eligible patients approached who consent to and comply with the trial protocol. This will be assessed by: number of eligible patients within the study population and recruitment timeframe, percentage of eligible patients accepted for randomisation, patient compliance with trial protocol measured as per selfreported adherence to intervention, and percentage of patients completing study. The  additional NHS services and privately purchased over the counter treatments related to dry eyes disease) and safety measure of pressure within the eye (assessed by Intra ocular pressure (IOP) score).

Data Analysis
A detailed data validation will examine completeness, existence and accuracy of collected data to assess data quality and identify missing and conflicting data. The statistical analysis will be performed on an intention-to-treat basis (i.e. inclusion of patients randomly assigned, regardless of adherence, actual treatment received, and subsequent withdrawal of treatment and/or deviation from the protocol and per-protocol (i.e. inclusion of those who completed the treatment as planned) (29) and will be reported according to 2010 CONSORT guidelines (30). All statistical analyses will be completed using SPSS Statistics v.22.0 (31). A p-value of <0.05 will be accepted as statistical significance. Conforming to recommendations for feasibility studies, data analysis will be primarily descriptive, with means, standard deviations and frequency counts calculated for all variables of interest.
Exploratory efficacy analysis will compare the primary outcome variables derived from the data collected at 8 weeks between the two arms using a marginal general linear model (GLM) with robust standard errors, to allow for clustering by group. Secondary outcomes between the intervention and control groups will be compared at 4 weeks, 8 weeks and 12 weeks (4 weeks post treatment). Safety outcome measures will include IOP rise and any reported infection. Secondary analyses of the primary outcome, controlling for baseline covariates, will be performed using proportional hazards regression models (32). Analysis of treatment discontinuation adjusted for clinical site, baseline status, and whether the patient is randomized to the FAB or control (conventional treatment only) group will be conducted.
Any interaction between baseline status or regimen and treatment will also be assessed.
Interactions between significant covariates and treatment effects will be assessed in the context of the proportional hazards models.
Interviews will be recorded, transcribed and analysed using the framework analysis. We will structure the analysis of collected data to fit two evaluation frameworks relevant to the successful implementation of FAB: • Patient-oriented: Understand the lived experience of patients using FAB with emphasis on intervention acceptability, perceived enablers and barriers for adherence, perceived clarity of advice, and guidance and factors relating to the initial uptake of the intervention.
• Organisation-oriented: Whether and how the use of FAB affected workload and/or workflow; identify process change, adaptation challenges, skills gap; establish treatment fidelity and clinician acceptability.

Study Procedures Informed Consent Procedures
It is the responsibility of the Investigator, or appropriately GCP trained person delegated by the Investigator as documented in the site delegation log, to obtain written informed consent from each participant prior to any participation/study specific procedures. This should follow adequate explanation of the aims, methods, anticipated benefits and potential hazards of the study. The participants will be given ample time to consider giving their consent for the study. For this study a minimum of 24 hours will be given during which the consenting team will be contactable over the phone to answer any questions. The date that the Participant Information Sheet (PIS) is given to the participant must be documented within the patient's notes to confirm that sufficient time was given. The Investigator (or other qualified person) will explain to the potential participant that they are free to refuse any involvement within the study or alternatively withdraw their consent at any point during the study and for any reason.
If there is any further safety information which may result in significant changes in the risk/benefit analysis, the PIS and Informed Consent Form (ICF) will be reviewed and updated accordingly. All participants that are actively enrolled on the study will be informed of the updated information and given a revised copy of the PIS/ICF in order to confirm their wish to continue on the study.

Screening Procedures
Patients will have undergone the below procedures as per standard of care which will in turn feed in to the investigator for their review prior to approaching the potential participant.
-Ocular surface disease index (OSDI) score of greater than 33; OR -Oxford Corneal Staining grade 2 or greater; OR -Schirmer's without anaesthesia <5mm at 5 minutes The results of these which will be reviewed by investigator prior to approaching the potential participant.

Randomisation Procedures
Randomisation will be carried out by the Anglia Ruskin Clinical Trials Unit (ARCTU) using the SEALED ENVELOPE randomisation service. SEALED ENVELOPE is a randomisation and online database service developed for clinical trial services. It is an internet based system and will be set for this study by ARCTU in accordance with the protocol. Enrolled patients will undergo 1:1 block randomisation to Arm A or B. The system stores the pre-determined sequence of randomisation and this list is visible to neither the investigator nor ARCTU staff.
Once a patient has consented to take part in the trial, they will be randomly allocated to either Arm A to receive FAB and convention therapy or Arm B to receive conventional therapy only. The Research Nurse or Fellow or designated staff will log on to a web browser application and enter the patient's eligibility factors into the system. The treatment allocation will be sent to the research team who will make the necessary arrangements for the patient's treatment plan.
The treating ophthalmologist in clinic will prescribe and counsel all patients on the correct technique for conventional treatment. The unblinded research nurses will provide additional training to the patients on the method of delivering FAB, including the use of a training video. Patients will be advised by the research team to inform their ophthalmologist if they have been started on any new treatment during the trial period by other clinicians or themselves.

End of Study Definition
The definition of the end of study will be the point where the last patient recruited had the last follow-up visit.

Participant Withdrawal
Patients will be withdrawn from study based on the following:  Any adverse effects will be reported back to the lead investigator at the site and if necessary patients

Notification and Reporting Adverse Events or Reactions
No serious adverse events (SAEs) or complications associated with use of fingerprick autologous blood in the treatment of ocular disease has been reported by the literature.
Preliminary results of a pilot study conducted by the authors and involving 16 patients with dry eye disease also demonstrated no adverse effects of using FAB in the eye for two months (22). Although, we do not envisage any serious adverse events with the intervention in this trial, if an adverse event (AE) occurs and is not defined as SERIOUS, the AE will be recorded in the study file and the participant followed up by the research team. The AE will also be documented in the participants' medical notes (where appropriate) and the CRF.
Serious Adverse Event (SAEs) that are considered to be 'related' and 'unexpected' will be reported to the sponsor within 24 hours of learning of the event and to the Main REC within 15 days in line with the required timeframe. SAE reporting will commence at the start of the trial and up to 4 weeks after the intervention.

PATIENT AND PUBLIC INVOLVEMENT
Patients who participated in a previous pilot (exploratory) study conducted by the study authors (22) were involved in a focus group meeting and provided inputs on research design and study procedures. Two of the patients also participated in an advisory group for this trial provided input on prioritising the research questions and also reviewed the study protocol and participant information resources. Patients in this trial will be invited to request study results from the research team if interested.

ETHICS AND DISSEMINATION
Participant anonymity will be protected and maintained at all times. Information with regards to study participants will be kept confidential and managed in accordance with the Data Protection Act, NHS Caldicott Guardian, The Research Governance Framework for Health and Social Care and Research Ethics Committee Approval in United Kingdom. Data obtained in the trial will be stored using a unique study code, which is non-identifiable and anonymised. Patient's personal data will be stored separately and confidentially by the  Cataract and Refractive Surgery. Information will be provided to patient groups and charities such as the Sjogren's society and the Royal National Institute of Blind People. Study participants will also be provided with the contact details of the trial team at the respective sites who will be able to answer any additional queries.    1  2  3  4  5  6  7  8  9  10  11  12  13  14  15  16  17  18  19  20  21  22  23  24  25  26  27  28  29  30  31  32  33  34  35  36  37  38  39  40  41  42  43  44  45  46  47  48  49  50  51  52  53  54  55  56  57  58  59

Objectives
To determine the feasibility of a definitive randomised controlled trial (RCT) to determine effectiveness of the use of fresh autologous blood (FAB) compared to conventional treatment for patients with dry eye syndrome.

Main Inclusion Criteria
-Patient age > 18 years -Severe symptomatic dry eye disease diagnosed by: Ocular Surface Disease Index (OSDI) score of greater than 33; OR Oxford Corneal Staining grade 2 or greater; OR Schirmer's without anaesthesia <5mm at 5 minutes. -Patient able to consent -Patients able and willing complete the quality of life (QoL) questionnaires required for the study Statistical Methodology and Analysis Data analysis will be primarily descriptive with means, standard deviations and frequency counts calculated for all variables of interest. Exploratory statistical analysis will compare the primary outcome and secondary outcomes between the intervention and control groups at 4 weeks, 8 weeks and 12 weeks (4 weeks post treatment)   1  2  3  4  5  6  7  8  9  10  11  12  13  14  15  16  17  18  19  20  21  22  23  24  25  26  27  28  29  30  31  32  33  34  35  36  37  38  39  40  41  42  43  44  45  46  47  48  49  50  51  52  53  54  55  56  57  58  59    No curative agents for DED exist. The treatment of the disorder is essentially symptomatic with standard non-surgical treatment focused on the use of artificial tears for lubrication and anti-inflammatory drugs. Anti-inflammatory agents include topical steroids and ciclosporin drops but these can cause side effects that limit their long term use (4). Topical steroids can also cause cataracts and glaucoma, all of which further limit its use. Surgical options for DED include punctal occlusion to reduce tear drainage, punctal cauterization (burning the drainage channel of tears, preventing their outflow) and partial suturing of the eyelids; but these are associated with adverse effects and varying levels of patient tolerability. Overall, available conventional treatment options for DED often only alleviate symptoms, have limited effectiveness, and in most cases patients may fail to respond; although the exact rate of treatment failure is unavailable in the published literature.
The mainstay of non-surgical treatment for DED focuses on the use of artificial tears. These can be purchased over the counter and include brand name products such as Viscotears® and Optrex® (5). Human tears contains an extensive range of growth factors, immunoglobulins, enzymes, cytokines, vitamins and electrolytes and these have been shown in numerous studies to be essential for the maintenance and proliferation of corneal epithelial cells as well as for defence against infection (6). It is well known that artificial tears fails to account for the extraordinarily complex composition of the natural tear film. Also, many artificial tears contain preservatives that have been shown to adversely affect the cornea (7). Crudely, human tears with its vast constituents is essentially filtered blood and as such is an obvious source for a  1  2  3  4  5  6  7  8  9  10  11  12  13  14  15  16  17  18  19  20  21  22  23  24  25  26  27  28  29  30  31  32  33  34  35  36  37  38  39  40  41  42  43  44  45  46  47  48  49  50  51  52  53  54  55  56  57  58  59  "tear mimic" containing the substances of tears. Blood, and several blood derived products, including autologous serum, have been studied as tear substitute candidates.
Autologous serum (AS) eye drops have been found in uncontrolled trials to be beneficial in DED patient by improving the ocular surface and reducing symptoms (8)(9)(10). However, a Cochrane review concluded that there is inconsistency in evidence on its benefit (11).
Obtaining autologous serum requires frequent drawing of blood from the patient-a feature that excludes patients with anaemia or heart failure from using AS. Furthermore it also appears that 100% autologous serum is more beneficial than 50% serum and requires larger volumes of blood and/or more frequent venesection (9). Patients using AS also require access to a fridge as the product needs to be stored at low temperatures; a factor that is likely to be inconvenient for patients. In addition, AS is obtained by processing clotted blood which is often too expensive for the health service to consistently purchase, given the initial cost of £1653.56 and subsequent three-monthly cost of £1131.27 per patient.
The relatively high cost represents the biggest hurdle in the use of AS and is often the reason for delay or inaccessibility in starting treatment for DED using AS. However, we propose that finger prick autologous blood may be a simpler, cost-effective and possibly more acceptable method for treating dry eye disease. For this reason, this study proposes to test the use of finger prick autologous blood (FAB) technique in which whole blood is applied to the eye from a cleaned finger. Autologous fresh blood is already used subconjunctivally to help heal leaking trabeculectomy blebs (12)(13)(14)(15). It is also used to help attach limbal autografts in cases of pterygium (16) and vitreoretinal macular hole surgery (17)(18)(19)(20), with no adverse effects reported. The FAB method can be used immediately for patients who are awaiting conventional treatment for autologous blood. The objective of this study is therefore to investigate if serum via a drop of fresh blood is an effective treatment for severe dry eye disease which currently require venesected autologous blood and whether this would be particularly useful in the group of patients in whom venesection is contraindicated.  1  2  3  4  5  6  7  8  9  10  11  12  13  14  15  16  17  18  19  20  21  22  23  24  25  26  27  28  29  30  31  32  33  34  35  36  37  38  39  40  41  42  43  44  45  46  47  48  49  50  51  52  53  54  55  56  57  58  59

Rationale and Risks/Benefits
This study proposes to test the use of whole, fresh, autologous blood as a treatment for severe dry eye disease. The blood can be obtained from a cleaned finger, pricked by the patient using a diabetic lancet and administered immediately to the dry eye. This allows the delivery of not only most of the aforementioned beneficial components of tears but additional growth factors and proteins, fresh and unprocessed, which can help heal the ocular surface. If validated, this may replace current autologous serum practice and its ease of use, vastly reduced cost with greater convenience may mean that it could be extended to other ocular surface diseases.

Primary Objective and Endpoints
The primary objective is to determine the feasibility of a definitive randomised controlled trial (RCT) to evaluate effectiveness of the use of fresh autologous blood (FAB) compared to conventional treatment for patients with dry eye syndrome. This will involve specifically assessing the following endpoints: i. Number of eligible patients within the study population and recruitment time frame.
ii. Percentage of eligible patients accepted for randomisation.
iii. Patient compliance with trial protocol, measured as per self-reported adherence to intervention.
iv. Percentage of patients completing study.

Secondary Objective and Endpoints
The secondary objectives are:  To determine the effectiveness of the trial intervention and further explore the acceptability of the study design.
 To explore the feasibility of collecting resource use and quality of life data, to inform the design of the health economics component of a future definitive trial.
The secondary end points, which have not been powered for within the feasibility study, include:

Trial Design
Single-blind two-arm feasibility randomised controlled trial of FAB with conventional treatment versus conventional therapy alone for dry eye disease, including a qualitative process evaluation.

 Arm A -FAB plus conventional treatment
The patients will use FAB therapy alongside conventional therapy (artificial tears, ciclosporin drops and punctal plugs/cautery) as recommended by their treating ophthalmologist. A fingertip of the hand will be wiped with an alcohol steret and self-pricked using a standard diabetic lancet. The drop of blood is produced as normal and applied to the lower fornix of the affected eye(s) with the lower lid pulled down slightly by the patient. The blood will be applied four times a day. A fresh finger should be used for each eye. The blood will be wiped away with the alcohol steret. Nails should be kept short and nail varnish should be avoided. FAB should be applied at least 15 minutes after any artificial tears and no drops for at least half an hour afterwards.

 Arm B -Conventional treatment only
The patients will use conventional therapy (artificial tears, cyclosporin drops and punctal plugs/cautery) as recommended by their treating ophthalmologist.

Setting
The study will take place at NHS sites. The main site will be Bedford Hospital NHS Trust and Moorfields Eye Hospital London. Patients will be recruited from corneal and external eye disease clinics at these sites.

Informed Consent Procedures
It is the responsibility of the Investigator, or appropriately GCP trained person delegated by the Investigator as documented in the site delegation log, to obtain written informed consent from each participant prior to any participation/study specific procedures. This should follow adequate explanation of the aims, methods, anticipated benefits and potential hazards of the study. The participants will be given ample time to consider giving their consent for the study.
For this study a minimum of 24 hours will be given during which the consenting team will be contactable over the phone to answer any questions. The date that the Participant Information Sheet (PIS) is given to the participant must be documented within the patient's notes to confirm that sufficient time was given. The Investigator (or other qualified person) will explain to the potential participant that they are free to refuse any involvement within the study or alternatively withdraw their consent at any point during the study and for any reason.
If there is any further safety information which may result in significant changes in the risk/benefit analysis, the PIS and Informed Consent Form (ICF) will be reviewed and updated accordingly. All participants that are actively enrolled on the study will be informed of the updated information and given a revised copy of the PIS/ICF in order to confirm their wish to continue on the study.

Screening Procedures
Patients will have undergone the below procedures as per standard of care which will in turn feed in to the investigator for their review prior to approaching the potential participant.
-Ocular surface disease index (OSDI) score of greater than 33; OR -Oxford Corneal Staining grade 2 or greater; OR -Schirmer's without anaesthesia <5mm at 5 minutes The results of these which will be reviewed by investigator prior to approaching the potential participant.

Randomisation Procedures
Randomisation will be carried out by the Anglia Ruskin Clinical Trials Unit (ARCTU) using the SEALED ENVELOPE randomisation service. SEALED ENVELOPE is a randomisation and online database service developed for clinical trial services. It is an internet based system and will be set for this study by ARCTU in accordance with the protocol. Enrolled patients will undergo 1:1 block randomisation to Arm A or B. The system stores the pre-determined sequence of randomisation and this list is visible to neither the investigator nor ARCTU staff.
Once a patient has consented to take part in the trial, they will be randomly allocated to either Arm A to receive FAB and convention therapy or Arm B to receive conventional therapy only.
The Research Nurse or Fellow or designated staff will log on to a web browser application and enter the patient's eligibility factors into the system. The treatment allocation will be sent to the research team who will make the necessary arrangements for the patient's treatment plan.
The treating ophthalmologist in clinic will prescribe and counsel all patients on the correct technique for conventional treatment. The unblinded research nurses will provide additional training to the patients on the method of delivering FAB, including the use of a training video.

End of Study Definition
The definition of the end of study will be the point where the last patient recruited had the last follow-up visit.

Participant Withdrawal
Patients will be withdrawn from study based on the following: -If finger sore from repeated prick and patient does not want to use another finger.
Any adverse effects will be reported back to the lead investigator at the site and if necessary patients

Adverse Event (AE)
An AE is any untoward medical occurrence in a participant to whom a medicinal product has been administered, including occurrences which are not necessarily caused by or related to that product. An AE can therefore be any unfavourable and unintended sign (including an abnormal laboratory finding), symptom or disease temporarily associated with study activities.

Notification and reporting Adverse Events or Reactions
If the AE is not defined as SERIOUS, the AE is recorded in the study file and the participant is followed up by the research team. The AE is documented in the participants' medical notes (where appropriate) and the CRF.

Notification and Reporting of Serious Adverse Events
Serious Adverse Event (SAEs) that are considered to be 'related' and 'unexpected' are to be reported to the sponsor within 24 hours of learning of the event and to the Main REC within 15 days in line with the required timeframe. For further guidance on this matter, please refer to Appendix.

Urgent Safety Measures
The CI may take urgent safety measures to ensure the safety and protection of the clinical trial participants from any immediate hazard to their health and safety, in accordance with Regulation 30. The measures should be taken immediately. In this instance, the approval of the Licensing Authority Approval prior to implementing these safety measures is not required. However, it is the responsibility of the CI to inform the sponsor and Main Research Ethics Committee (via telephone) of this event immediately.

5.6
Annual Safety Reporting The CI will send the Annual Progress Report to the main REC using the NRES template (the anniversary date is the date on the MREC "favourable opinion" letter from the MREC) and to the sponsor. Please see appendix for further information.
All statistical analyses will be completed using SPSS Statistics V.22.0 (26). A p-value of <.05 will be accepted as statistical significance.
Conforming to recommendations for feasibility studies, data analysis will be primarily descriptive, with means, standard deviations and frequency counts calculated for all variables of interest. Exploratory efficacy analysis will compare the primary outcome variables derived from the data collected at 8 weeks between the two arms using a marginal general linear model (GLM) with robust standard errors, to allow for clustering by group. Secondary outcomes between the intervention and control groups will be compared at 4 weeks, 8 weeks and 12 weeks (4 weeks post treatment). Safety outcome measures will include IOP rise and any reported infection.

Qualitative approach
A nested qualitative approach using in-depth, semi-structured interviews will help us understand the lived experience of people using FAB and factors relating to the ways that clinical departments and healthcare professionals adapt to working with FAB. Topics, questions and probes in the interviews will emerge from three sources: a) the relevant literature; b) our experience of and involvement in clinical ophthalmology; c) consultation with patients. In this feasibility study, we will use a convenience sampling technique for the qualitative work. We will interview all clinicians who administer the intervention in the study and we will aim to interview 10 patients (adjusted for setting, gender and age group representation). Given the resources, we will aim to draw findings from a larger sample in a future study. Interviews will be recorded, transcribed and analysed using the framework analysis. We will structure the analysis of collected data to fit two evaluation frameworks relevant to the successful implementation of FAB: 1) Patient-oriented: Understand the lived experience of patients using FAB with emphasis on intervention acceptability, perceived enablers and barriers for adherence, perceived clarity of advice and guidance and factors relating to the initial uptake of the intervention.
2) Organisation-oriented: Whether and how the use of FAB affected workload and/or workflow; identify process change, adaptation challenges, skills gap; establish treatment fidelity and clinician acceptability.
This phase will work in parallel to the quantitative data collection so that emerging themes can be investigated in later interviews. Even though the feasibility study sample is reasonably limited, we anticipate some useful insights about the use of FAB to occur because of the triangulation between the RCT and the qualitative findings (e.g. reasons for partial adherence).

Case Report Form
Project data collection will be managed by the Clinical Trials Unit Data Manager. The responsibility for data entry rests with the Research Nurse or designated staff who is supported by the Investigator. ARCTU uses an online data management system called MACRO to design and manage eCRFs (Electronic Case Report Forms). ARCTU will work together with the sponsor to design and validate the data collection tools so that they are appropriate for this study. Once a patient is enrolled on the study the research team can access these forms remotely through the Internet portal and study data will be entered and captured for the study. All data will be in anonymised form and patients will be identifiable only by study number. Data will be remotely monitored by ARCTU and discussed at data monitoring committee meetings. Any inconsistencies, validation errors or inaccuracies will be reported to the lead investigator regularly. Once data collection is complete and the data has been validated, a data lock will be performed and analysis will begin.

Record Retention and Archiving
During the course of research, all records are the responsibility of the Chief Investigator and must be kept in secure conditions. When the research trial is complete, it is a requirement of the Research Governance Framework and Trust Policy that the records are kept for a further 20 years.

Ethical Considerations
This protocol and any subsequent amendments, along with any accompanying material provided to the participant in addition to any advertising material will be submitted by the Investigator to an Independent Research Ethics Committee.
Written Approval from the Committee will be obtained and subsequently
-An individual investigator or department may request an audit.
-A project may be identified via an allegation of research misconduct or fraud or a suspected breach of regulations.
-Projects may be selected at random. The Department of Health states that Trusts should be auditing a minimum of 10% of all research projects.
-Projects may be randomly selected for audit by an external organisation.
Internal audits will be conducted by a sponsor's representative 7.8

Non-Compliance
Non-compliance can be defined as 'a noted systematic lack of both the CI and the study staff adhering to SOPs/protocol/ICH-GCP, which leads to prolonged collection of deviations, breaches or suspected fraud.' These non-compliances may be captured from a variety of different sources including monitoring visits, CRFs, communications and updates. The sponsor will maintain a log of the non-compliances to ascertain if there are any trends developing which to be escalated.
The sponsor will assess the non-compliances and action a timeframe in which they need to be dealt with. Each action will be given a different timeframe dependant on the severity. If the actions are not dealt with accordingly, the sponsor will agree an appropriate action, including an on-site audit.  1  2  3  4  5  6  7  8  9  10  11  12  13  14  15  16  17  18  19  20  21  22  23  24  25  26  27  28  29  30  31  32  33  34  35  36  37  38  39  40  41  42  43  44  45  46  47  48  49  50  51  52  53  54  55  56  57  58  59  A Trial Management Group (TMG) has been formed comprising the Chief Investigator, other lead investigators (clinical and non-clinical) and members of the Data Centres. The TMG will be responsible for the day-to-day running and management of the trial and will meet at least 3 times a year by teleconference.

Trial Steering Committee (TSC)
The Trial Steering Committee (TSC) has membership from TMG plus independent members, including the chair. The role of the TSC is to provide overall supervision for the trial and provide advice through its independent Chairman. The ultimate decision for the continuation of the trial lies with the TSC.

Independent Data Monitoring Committee (IDMC)
The Independent Data Monitoring Committee (IDMC) is the only group who sees the confidential, accumulating data to the trial. Reports to the IDMC will be produced by the trial statistician. The IDMC will meet within 6 months of the trial opening with the frequency of meetings dictated by the IDMC. The IDMC will consider the report for the interim analysis based on the first 30 patients and will decide, keeping in mind the rules described in the statistics section, whether to recommend that the study should continue or be closed. The primary outcome measure for the study addresses toxicity and so it aligns with the IDMC's precautionary role in patient safety. The details of the interim analysis will remain confidential to the IDMC until the study is closed.

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