Faecal microbiota transplantation: establishment of a clinical application framework.

’Original article Faecal microbiota transplantation: establishment of a clinical application framework Simon M.D. Jørgensena, Mette M. Hansena, Christian Erikstrupb, Jens F. Dahlerupa and Christian L. Hvasa Background Faecal microbiota transplantation (FMT) is currently being established as a second-line treatment for recurrent Clostridium difficile infection. FMT is further being considered for other infectious and inflammatory conditions. Safe and reproducible methods for donor screening, laboratory processing and clinical application of FMT are warranted. Methods Here, we describe the development of a complete clinical application framework for FMT. The framework has been developed to comply with the European Tissue Act, thus considering donor faeces for FMT comparable to a human tissue and not a drug. Results Recruitment and screening of potential faeces donors took place in the public blood donor setting and consisted of questionnaires, blood sampling and faecal sample analysis. Once approved, and following their written informed consent, eligible donors were invited for voluntary faecal donation. Laboratory processing protocols describe the initial handling, cryopreservation and thawing for clinical application. The clinical FMT procedures took place in a gastroenterological setting using a nasojejunal tube or colonoscopy, and follow-ups were performed at 1, 8 and 26 weeks after FMT. Complete traceability of essential equipment, faecal samples and donor–recipient matching data will be maintained and secured for 30 years. Conclusion A clinical FMT service should be consolidated by a complete documentation system that complies with the European Tissue Act. In this paper, we provide a description of such a framework. Eur J Gastroenterol Hepatol 00:000–000 Copyright © 2017 Wolters Kluwer Health, Inc. All rights reserved.

Introduction

Faecal microbiota transplantation (FMT) is a new therapeutic option whereby processed allogenic faeces are transferred from a healthy donor to the gastrointestinal (GI) tract of a recipient with the aim of restoring the composition and functionality of the gut microbiome. The sporadic use of this procedure in clinical medicine has been known for decades [1,2] and has been described in the literature for centuries [3]. Recently, FMT was found to be superior to standard antibiotic therapy for recurrent Clostridium difficile infections (RCDI) [4] and the effect has been confirmed in subsequent clinical trials, with resolution rates ranging from 70 to 94% [4–7]. As a result, FMT is now being recommended as a second-line treatment option for RCDI in the European clinical guidelines [8]. In addition to RCDI, FMT is also being considered as a potential treatment for inflammatory bowel disease, irritable bowel syndrome and other extra-intestinal diseases where a disrupted intestinal microbiome may European Journal of Gastroenterology & Hepatology 2017, 00:000–000 Keywords: Clostridium difficile, faecal microbiota transplantation, faecal microbiota transplantation clinical service, quality improvement Departments of aHepatology and Gastroenterology and bClinical Immunology, Aarhus University Hospital, Aarhus, Denmark Correspondence to Christian L. Hvas, MD, PhD, Department of Hepatology and Gastroenterology, Aarhus University Hospital, Noerrebrogade 44, Building 7, DK-8000 Aarhus C, Denmark Tel: + 45 283 51839; fax: + 45 784 62820; e-mail: [email protected] Received 16 May 2017 Accepted 18 July 2017 Supplemental digital content is available for this article. Direct URL citations appear in the printed text and are provided in the HTML and PDF versions of this article on the journal's website (www.eurojgh.com).

contribute towards pathogenesis, such as obesity, liver diseases and autoimmune disorders [9–14]. The emerging potential for FMT has drawn attention from both the scientific community and the general public in the need for a legislative control system to establish FMT as a safe and sustainable treatment modality. The current legislation leaves national health authorities in a dilemma. As a result, FMT is classified as a drug in some countries, thereby placing it under strict regulations that severely restrict its use [14–17]. In other countries, however, FMT is unregulated. In Denmark, FMT was first filed in 2015 to be regulated by the Danish Tissue Act, derived from the European Tissue and Cells Directive (EUTCD) [18]. In February 2016, the Danish health authorities announced that the legal basis to actively govern FMT was insufficient. Nonetheless, institutions that performed FMT were strongly advised to consider FMT according to the directives of the Danish Tissue Act, albeit without active regulatory oversight. Detailed protocols that describe the administration of FMT have been published [19–21], and consensus reports on overall good conduct exist [22,23]. Yet, standards and methodology for practical implementation, safety and follow-up procedures that specifically comply with current legislation are lacking [24–26]. The EUTCD provides high-quality and safety standards for the handling of tissues and cells of human origin. Utilizing the standards of EUTCD as the regulatory basis when performing FMT provides useful precautionary measures that are equivalent to the guidance of other institutions for handling materials of similar origin [18]. To meet these standards, the adaptation of FMT into a greater framework is necessary.

0954-691X Copyright © 2017 Wolters Kluwer Health, Inc. All rights reserved.

DOI: 10.1097/MEG.0000000000000958

Copyright r 2017 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited.

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Month 2017 • Volume 00 • Number 00

European Journal of Gastroenterology & Hepatology

The aim of this study was to provide a clinicianoriented description of the establishment of an integral clinical FMT framework derived from the standards of the regulatory EUTCD directives. This framework integrates current FMT protocols and describes in detail how to conform to the EUTCD and what to be aware of in clinical practice, thereby easing the overall implementation of an FMT system with sufficient safety and quality standards. A clinical FMT framework

The clinical FMT framework can be divided into three main activity sections: (i) donor recruitment, (ii) laboratory processing of donor material and (iii) clinical application (Fig. 1). This division provides a practical organizational structure and clarifies both the tasks and the necessary personnel to maintain an operational FMT service. Each section is described in detail below. To meet the EUTCD requirements for a tissue establishment, the FMT system should comply with a set of overriding principles and standard requirements (Supplementary Table 1, Supplemental digital content 1, http://links.lww. com/EJGH/A218) [18]. Applied to each element of the FMT process, the standard requirements include the identification and in-depth documentation of critical processes, risk assessment and subsequent development of standard operating procedures (SOPs). When implemented, standardized documentation throughout the entire process will ensure traceability and enables the production of annual activity reports for active, regulatory governing and accrediting inspections by health authorities. Donor recruitment

To sustain FMT as a viable option, the system must rely on a continuous supply of faecal donor material from healthy donors. Donor selection includes the recruitment of potential donors, donor screening and the voluntary delivery of faecal donations for clinical application (Fig. 2). Recruitment

Previous studies of faecal donor recruitment have reported the complexity of the recruitment process in terms of low donor eligibility and low participation and retention rates. In most studies, donors were recruited by advertising to the general community, resulting in overall eligibility rates of 6–10% upon completion of the entire screening process. This makes donor recruitment both expensive and time

Fig. 2. Donor recruitment and screening.

consuming [27–29]. Although patient perceptions have been examined in a small number of studies [30], donor perceptions specific to the donation of faeces are largely unknown. At our institution, we established a collaboration between the Department of Hepatology and Gastroenterology and the Central Danish Blood Centre. During blood donations, blood donors were asked to enter our faecal donor corps. This allowed us to recruit from prescreened and healthy blood donors, thereby increasing the donor eligibility while lowering the cost and time required for donor recruitment. Consequently, the use of the blood bank infrastructure was an efficient means to standardize our recruitment to a confined and adjustable practice, but also proved to be a feasible means to establish a corps of unpaid volunteers. Screening procedures

Fig. 1. A clinical framework for faecal microbiota transplantation (FMT) can be divided into three separate sections: donor screening, laboratory processing and clinical application.

The screening of donors must rely on the analysis of risk related to FMT to avoid transplanting potentially transmittable diseases or contagious traits [14,18]. Previous protocols have established detailed screening standards and we adapted our screening accordingly (Table 1) [19,24,31]. In addition, we consulted the local departments of infectious diseases and clinical microbiology to adjust the screening to the local infectious pressure. Following the suggestions of these departments, we


Faecal microbiota transplantation Jørgensen et al.

Table 1. Inclusion, exclusion and screening parameters of potential donors Donor inclusion criteria Currently Age: 25–60 years A general feeling of well-being No medication (except anticonceptives) Within the last 3 months No antibiotics, antiviral or antifungal agents Within the last 6 months No high-risk behaviour (e.g. sexual contact with HIV or AIDS) No tattoos or piercings No history of travel to countries with a high incidence of infectious intestinal diseases No acute bowel disease No recent blood transfusions Donor exclusion criteria Health characteristics Moderate to severe obesity: BMI ≥ 28 kg/m2 Signs of malnutrition: BMI ≤ 18 kg/m2 Atopic disease Depression Sustained need for medication Comorbidities Chronic diseases or autoimmune diseases (e.g. Crohn’s disease, ulcerative colitis, coeliac disease, arthritis, multiple sclerosis, connective tissue disease, psoriasis) History gastrointestinal surgery (except appendectomy) Family history of (siblings and parents) Crohn’s disease, ulcerative colitis and coeliac disease Colorectal cancer Gastrointestinal complaints (within the last 4 weeks) Any moderate to severe complaints to the upper gastrointestinal complaints Diarrhoea Constipation Moderate to severe bloating Moderate to severe nausea Defaecation pain Signs of mucus in faeces Haematochezia or any signs of blood related to the defaecation Blood screening Health functions Pancreas function (pancreatic amylase) Liver function (alanine aminotransferase, bilirubin, alkaline phosphatase, albumin, INR) Electrolytes (sodium, potassium) Kidney function (creatinine, urea) Complete blood count (haemoglobin, leucocytes, platelets, red cell volume) C-reactive protein IgG, IgA, IgM HbA1c Serological screening Cytomegalovirus Epstein–Barr virus Strongyloides stercoralis DNA Entamoeba histolytica DNA Hepatitis Ba Hepatitis Ca HIVa Syphilisa Microbiology Enteropathogenic bacteria Clostridium difficle (toxin A, toxin B, binary toxin, ribotype CD027) Enteropathogenic Escherichia coli Salmonella spp. Shigella spp. Campylobacter jejuni Yersinia enterocolitica Multiresistant bacteria Meropenem-resistant Pseudomonas aeruginosa, Acinetobacter and Enterobacteriaceae Vancomycin-resistant Enterococcus Multiresistant Klebsiella spp. Extended-spectrum β-lactamase Enterobacteriaceae Virus Adenovirus Enterovirus Parechovirus Enteropathogenic parasites Cryptosporidum spp. DNA

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Table 1. (Continued) Giardia duodenalis DNA Entamoeba dispar DNA Entamoeba histolytica DNA Microscopy Worms, eggs and cysts of intestinal parasites (direct microscopy) a

Minimum screening requirements of the EUTCD [17].

extended the faecal screening panel to include resistant bacteria, that is, vancomycin-resistant enterococcus and extended-spectrum β-lactamase enterobacteria. Donors willing to participate first completed an electronic questionnaire, and once completed, eligible donors had their blood drawn for haematological, biochemical and serological screening (Fig. 2). To perform the faeces screening, eligible donors received a faecal collection kit along with sampling instructions. Donors were asked to collect the faecal samples upon their next defaecation at home and ship it immediately thereafter. Faecal samples were received and tested in accordance with third-party agreements for the stool screening protocol at the Department of Clinical Microbiology. Faeces test results are received within 7 days of arrival of the faecal sample. The donors are given 15 days to collect and ship the samples for their screening to remain valid. With an expected low donor eligibility rate, completing the entire screening before entering the donor corps helps decrease the inconvenience of donors, but it also reserves medical resources for qualified donors. This provides an efficient way to screen and include the sufficient number of new donors to maintain the treatment. Inclusion of qualified donors

Once all results from the blood and faecal screenings were available, the results were assessed by a physician to determine eligibility. Only donors with approved screening results were invited for a medical consultation to verify the consistency between the screening results and their overall state of health. During the consultation, the donor first underwent a physical and neurological examination, followed by a medical interview (Supplementary Table 2, Supplemental digital content 1, http://links.lww.com/ EJGH/A218). The practical arrangements of the donations were made at the end of the consultation and the donor was concomitantly instructed on how to collect and transport donor faeces. Once included as an active donor, the donor was assigned a unique identification code to render the donor anonymous. To keep track of the process, a donor record containing all the information mentioned in Supplementary Table 2 (Supplemental digital content 1, http://links.lww. com/EJGH/A218) must be maintained. These records must be stored for a minimum of 30 years to maintain full traceability [18]. To formally include the donor, the health professional undertaking the consultation must obtain and approve all relevant information [18]. In addition, for the donor to legally give consent and enter the faecal donor population, it must be assured and noted that the donor has been informed properly in an understandable manner.


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Donation rounds and quarantine measures

All included donors provided donations in a round consisting of five deliveries, scheduled within 1 month. At each delivery, the donor was asked about changes in health and for continued written consent. At the fifth donation, faeces and blood were sampled for a complete rescreening. In the case of any changes in health before the fifth donation, the cycle was stopped at the current donation, and all blood and faecal screenings were performed. Performing the screening both before and after donation helps achieve a documented safe interval of time without changes in the screening criteria [19]. This practice reduces cost and the impracticality of screening the donor at every donation and also limits inconveniences to the donor. In case of any positive postdonation screening test, all donations in the cycle were discarded. All donations from a donation round were held in quarantine and the donor was ineligible to enter a new donation round in this period until all laboratory test results had been approved. Safety was maintained by avoiding potential distribution while awaiting acceptance. Efficient SOPs were in place to define the responsibilities of the involved personnel, thereby confirming the quality and safety of the donor faeces before their certification and release into system. Authorized personnel had to ensure and document that all screening parameters had been met before the release. Currently, no protocols have validated an expiration date of the donor screening when performed in intervals. However, the EUTCD requires laboratory tests to be repeated after an interval of 6 months [18]. Consequently, all approved donors were eligible to enter a new donation round without previous screening if the last screening was performed within the past 6 months. If exceeded, the donor must be tested at the first donation and after the fifth donation. Personnel

The donor-selection process involved coordinated efforts between health professionals and specialists. Most of the initial donor recruitment can be carried out by trained health professionals. However, the EUTCD specifies that the medical consultation must be conducted by a specialist with a medical degree [18]. Thus, in our system, it has proven feasible for trained health professionals to perform all recruitment efforts before the medical consultation, limiting specialist resources only to the medical examination of qualified donors. Laboratory processing

The initial processing of a faecal donation aims to preserve and produce consistent suspensions that are suitable for clinical application. Accordingly, the laboratory processing covers procedures ranging from reception, processing and storage of faecal donations to the final preparation of ready-to-use faecal suspensions (Figs 4 and 5). The EUTCD requires the service to thoroughly review each step of the faecal processing and to identify all processes and procedures that potentially influence quality or safety. These critical processes must be documented and evaluated, and precautionary matters must be validated to


provide the necessary specifications and procedures for SOPs [18]. The validation must be based either on studies carried out by the FMT unit itself, data originating from published studies or well-established processing procedures that have been evaluated retrospectively on the basis of clinical results [18]. No standard laboratory procedures have been established [20], but current protocols [19,24] provide practical guidance on how to process donated faeces. Establishing a frozen faeces biobank

Maintaining an operational service that can handle FMT on a regular basis requires laboratory handling for the processing of donations within a narrow time range. Banking of processed frozen donations that requires only minimal preparation before application considerably eases and simplifies this practice. At our department, we initially used fresh donor faeces from related donors. However, because of both practical and ethical aspects related to the use of family donors [32], family donors were replaced by unrelated, anonymous donors. When using fresh donor faeces, we faced problems such as delays in donations, variations in stool weight and the need for frequent donor screening before the FMT [32,33]. To establish a stable and flexible service, using frozen stool from anonymous donors was an obvious option. This allowed FMTs to be easily scheduled and the donations to be handled separately. Anonymous donors and related donors have been shown to be equal in terms of clinical efficacy [24,33–36]. Similarly, studies have shown that freeze-thawed faeces are equal in terms of efficacy and the frequency of adverse events compared with fresh faeces [6]. Utilizing frozen faeces adds several steps to the laboratory handling. To preserve a viable microbial composition throughout these steps, overall quality control and monitoring of the critical parameters is essential. The critical parameters that influence processing are temperature and time. A previous study examined the effects of these parameters on bacterial viability, examining five different conditions: 8 and 24 h at room temperature, 8 and 24 h at 4°C and immediately snap-frozen in liquid nitrogen (reference measure) [37]. The results of this study included the following findings: (i) decreased temperature and less time before storage are protective for both diversity and total bacterial number and (ii) storing faeces at 4°C for more than 8 h significantly affects the microbial composition [37]. Keeping faecal donations sufficiently cooled within a time range of 8 h from delivery to final processing is therefore essential to ensure minimal impact on viability and quality. The overall quality control of time and temperature must aim to ensure these reference ranges. The EUTCD requires that an expiration date be defined when storing processed material [18], but no studies have specifically examined microbial durability at − 80°C over time. In previous studies, the bacterial viability of faeces dispersed in 10% glycerol and normal saline (NS) stored for up to 6 months at − 80°C appeared unaffected [32]. In a clinical FMT trial, RCDI patients also showed an 88% cure rate when faecal suspensions stored at − 80°C for 2–10 months were used [32]. At our unit, we set the expiration date at − 80°C to 10 months from the date of initial processing.



Reception of donated faeces

Once a donor entered a donation round, the delivery and collection of the faeces occurred at the donor’s home. Before defaecation, a disposable collection tray covered in a plastic bag was placed in the toilet (Fig. 3). When the defaecation was completed, the faeces were collected in the plastic bag and placed directly in an airtight container. To ensure optimal conditions, the container and freezer packs were placed in a cooler bag validated for transportation of biological substances at a biohazard level 2 (Fig. 3) [18]. The donation was transported to the laboratory within two hours and processed within four hours, thereby utilizing only 6 h of the 8-h viability window [19,24,32,35, 37]. Because the cooling capacity of the freezer packs relies on stool volume, considerable temperature variation must be anticipated [36]. The quantity of stool delivered from our donors in our experience ranges from 30 to 498 g/delivery. At each delivery of donation, the laboratory personnel must verify the donor identity and that the packaging, transportation and labelling requirements have been met. These steps must be documented in the processing report for each donation (Supplementary Table 3, Supplemental digital content 1, http://links.lww.com/EJGH/A218) [18]. If a donation does not meet the requirements, procedures for disposal must be in place. Preparation

Preparation of the donated material proceeded once the donation had been received (Fig. 4). The processing occurred in a clean and sterilized fume cupboard to avoid potential contamination and unpleasant working conditions. Before processing, the stool was weighed and the tubes were labelled with an assigned unique identification code, date of processing and an expiration date. Ideal methods to prepare donor faeces remain to be defined, and no evidence exists on whether anaerobic preparation conditions or the preservation of fibres are crucial for the retention of microbial viability. However, differences in these parameters do not seem to significantly impact clinical efficacy in RCDI trials [6,24,33–35,38].

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NS was added to soften the faeces and to produce homogenous suspensions. There are no specifications on NS volume [6,24,33–35,38]. In our experience, adding a NS volume of 2.5 ml NS per 1 g stool provided sufficient volume to dilute all forms of faeces from a solid to a liquid suspension. The donation was blended using a kitchen hand blender and the blended suspension was further passed through a strainer to diminish the risk of debris blocking the delivery device during the clinical application (Fig. 4). For each donor, we used a new disposable blender and all equipment was disinfected with 70% isopropanol before use. A 10% glycerol solution was added to the blended suspension to improve bacterial viability upon freezing [6, 32,34]. The suspension was then aliquoted into 50 ml Falcon tubes, labelled and immediately stored at − 80°C (Fig. 4). Division of the faeces suspensions into 50 ml Falcon tubes reduces thawing time. Upon completion of the donation processing, it is important to produce and retain a processing report (Supplementary Table 3, Supplemental digital content 1, http://links.lww.com/ EJGH/A218). Thawing and preparation for faecal microbiota transplantation

When a request for FMT is made, the laboratory must receive information on the route of administration as well as the recipient and the clinician performing the FMT to prepare for the treatment modality and to confirm the credentials of the responsible clinician. The thawing procedure (Fig. 5) can be initiated 45 min before the clinical application. Applicable faecal suspensions were consecutively selected and designated to the recipient. If the recipient had previous failed FMT using suspensions from a specific donor, another donor was chosen. The donor and the recipient were linked in a clinical database through the suspension identification number and the identification of the recipient. One transplantation requires the compilation of three 50 ml faecal suspensions, representing a total of 40–50 g undiluted donor faeces. When selected, the suspensions were prepared as shown in Fig. 5. Depending on the route of administration, the compiled suspension was transferred to either an empty NS bottle (endoscopy) or three 60 ml syringes (nasojejunal tube administration). The container or syringes were labelled with a unique identification code, date, additives used, expiration date and the identification of the designated recipient [18]. To ensure microbial viability, the suspensions must be kept cold and used within the next 4 h [19,24,32,35,37]. Facilities and equipment requirements

Fig. 3. Donation kit for one delivery. (a) The transportation freezer bag. (b) Disposable collection tray covered in a plastic bag. (c) The container. (d) Freezing elements.

The facilities and utilized equipment should aim to minimize the risk of contamination and avoid a decrease in quality. At our unit, all equipment should be Conformité Européenne certified and sterilized before use. In addition, the equipment should be validated or certified for the intended purpose [18]. The facilities, all equipment and procedures involved in the critical processes must be identified, and specifications relating to equipment/facility characteristics must be defined in the SOPs [18]. SOPs that detail requirements on working ranges, calibration,


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Fig. 4. Laboratory processing from receipt of donor faeces in the laboratory to cryopreservation at − 80°C.

Fig. 5. Thawing procedures before the clinical use of donor faeces.

maintenance and cleaning should be stored in a frequently updated database. A log for each critical process and its parameters must be kept and stored for at least 10 years. All critical equipment, such as freezers, as well as donor material handling instructions must be subject to monitoring, and an alert system must be in place in case of defects [18]. All materials and additives used in the processing must have their required properties defined to ensure quality [18]. In relation to faecal processing, all used materials in contact with the faecal suspension must be food approved to ensure that such materials do not affect the faecal suspension. The packaging of these materials must also be ensured to be durable at the defined low temperatures.

how to handle these deviations and whom to contact if clarification is needed. We collected three safety samples from thawed and used suspensions to ensure full traceability. These safety samples will be stored in a − 80°C freezer for 30 years and are ready to be thawed and screened in case of AE/SAE to investigate the potential contributing causes [18]. Personnel

To maintain good laboratory practice, only laboratorytrained personnel can perform the laboratory-related tasks. Accordingly, they must receive preceding peer-topeer training. In concordance with EUTCD, training procedures are performed every year and kept up to date in case of updates [18].

Distribution and quarantine handling

Faecal suspensions for clinical use should be kept in quarantine until all tests of the final donation have been approved. The storage system should uniquely distinguish between quarantined and released batches. To ensure a clear distinction between the quarantined and the released suspensions, they must be separated physically in distinct freezers [18]. The laboratory must be notified when the materials are released and must reposition the released suspensions to the appropriate − 80°C freezer. If suspensions do not pass postdelivery microbial tests, or if they expire, all suspensions from the donation round must be disposed of according to the SOP. Adverse events and serious adverse events

The EUTCD requires safety procedures and notification systems to be in place in case of any adverse events (AE) or serious adverse events (SAEs) [18]. In case of any AE/SAE related to procedural handling of the faeces in the laboratory, probable causes must be determined and new correcting procedures should be implemented that describe

Clinical application Routes of administration

FMT can be applied through either the upper or the lower GI tract. The most frequent ways to perform FMT are by colonoscopy, nasojejunal tube delivery and enema [20]. When administered by colonoscopy, the suspension was infused in the terminal ileum and right colon. Jejunal delivery required a correctly placed nasojejunal tube through which the donor faeces are applied. Administration by enema was performed using either a suspension-loaded squeeze bottle or a rectal catheter. Both colonoscopy and nasojejunal tube delivery required preprocedure preparation. Colonoscopy required bowel lavage, fasting and sedation. The nasojejunal tube delivery required tube insertion, radiologic imaging verification of intestinal position, fasting and occasional administration of prokinetics to facilitate tube placement. All three modalities have advantages and disadvantages (Table 2) that must be considered. In the majority of studies examining FMT for RCDI, the efficacy is high



irrespective of the application method [39,40]. Colonoscopy may be preferred because it allows for the concomitant diagnosis of colonic disease [9]. However, several patient groups are unable to undergo the required preparation for a colonoscopy, because of either disease severity or comorbidity. In these patients, the gentle approach of nasojejunal tube delivery may be considered favourable. Faecal enema may be preferred in outpatient care because of its ease of use, lack of need for patient preparation and minimal clinical requirements. A disadvantage is that it may require more than one administration to achieve effect rates similar to the other two options [6,41]. Recently, the use of encapsulated suspensions has been reported as a feasible modality [42]. However, this delivery method requires several processing steps and it is unclear whether the encapsulation should be considered a manufacturing step and therefore be subject to drug legislation. Safety issues also remain to be described. Safety

Most of the process of adapting a clinical practice into a greater framework relies on the implementation of

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sufficient standardized safety procedures. This task involves standardized logging protocols, follow-up and handling in case of adverse events [18]. FMT is generally well tolerated, with only a few reports of serious AEs [26] (even in immunocompromised patients) [41]. However, the implemented framework must have a system to follow up all patients and promote the detection of otherwise unknown AEs that may be attributable to the FMT or the processing of the materials. AEs should be registered and documented according to the principles for Good Clinical Practice [43]. The most common AEs associated with FMT are self-limiting nausea, bloating, abdominal cramping and diarrhoea within a couple of hours after the procedure. The most common causes of SAEs related to faeces application include upper GI bleeding, aspiration and mucosal tearing. In terms of SAE linked to the FMT microbial content, cases of peritonitis, bacteraemia and enteritis have been reported [44]. The EUTCD provides standard protocols to be followed in the case of any SAE [18]. In our system, all recipients of FMT were called at week 1 and were required to ship a faecal sample for testing and safekeeping. This procedure was repeated at week 8, but instead of being questioned over the phone, the recipient

Table 2. Advantages and disadvantages of the most commonly used modalities for performing faecal microbiota transplantation Colonoscopy Advantages

Disadvantages

Time consumption (min) Clinician Organization and preparation Clinical application Documentation and follow-up Total Clinical assistant Organization and preparation Clinical application Documentation and follow-up Total Preceding preparation Facilities required

Personnel required

Logging requirements

Covers the entire colon Possibility to take biopsies + disease and severity grading Performed in a controlled environment Preceding bowel lavage and fasting Time consuming Requires more personnel Risk of perforation

5 30 15 50 35 40 20 95 Yes Clinical settings suited for endoscopy Colonoscopy equipment supporting an endowasher Colonoscopy cleaning system Clinical trained gastroenterologist Clinical assistant trained in colonoscopy and the preparation procedure Scope id number Verification of correct scope cleaning Amount of extra saline added Documentation of involved personnel Procedural record The donor suspension code + time of use Suspension transportation and safekeeping

Nasoenteric tube

Enemaa

Easy and fast to perform Gentle application Covers almost the entire length of the GI

Very easy to administer Does not require total bowel lavage Flexible in terms of facilities required

Requires radiological imaging to confirm correct position Wrong placement requires the tube to be replaced

Only reaches the splenic flexure More than one enema therapy may be necessary

20 5 15 40

0 5 15 20

45 5 35 85 Yes Clinical setting or outpatient care Radiograph accessibility

Clinical trained gastroenterologist Clinical assistant trained in the preparation procedure Associated radiologist Tube manufacturer and lot number Amount of extra saline added Documentation of involved personnel Procedural record The donor suspension code + time of use Suspension transportation and safekeeping

15 5 30 50 No Clinical setting or outpatient care

Clinical trained gastroenterologist Clinical assistant trained in the preparation procedure Enema set manufacturer and lot number Documentation of involved personnel Procedural record The donor suspension code + time of use Suspension transportation and safekeeping Suspension transportation and safekeeping

Included is an estimation of the required time, facilities and personnel as well as EUTCD logging requirements [18] for the preparation and FMT on one recipient. FMT, faecal microbiota transplantation; GI, gastrointestinal. a Is based on the described methods in Brandt et al. [9] and our clinical experience in handling the preparation and organization of the FMT process.


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was booked for a clinical consultation. All follow-up data were documented and maintained in a clinical registry. Personnel

Preparative considerations, practical arrangements and the required personnel vary depending on the treatment modality (Table 2). The most time-consuming portion of the transplantation process is the preparation and the preceding organizational tasks. Both a physician and a clinical assistant are needed to perform the transplantation, and the physician must be in possession of a formal degree. From a systematization viewpoint, removing the organizational burden from the physician and assigning it to dedicated personnel allows for the performance of a greater number of FMTs. In our experience, it is also more efficient to prioritize modalities that require the least amount of clinician time. Discussion

The emerging potential of FMT calls for a safe and standardized application system. In this study, we describe the three separate sections of a novel clinical FMT framework: donor recruitment, laboratory processing and clinical application. We further relate the requirements for documentation, traceability and personnel training to published laboratory protocols and the most appropriate legislation. It falls to public health authorities to decide within which jurisdictions to place FMT to ensure sufficient safety. A dedicated public legislation to govern FMT is warranted and a uniform governing policy is mandatory for the future creditability of FMT. The fundamental question in this respect remains whether FMT is considered – or comparable to – a tissue or a drug [16,17, 45–47]. The legislative bases for drug [48] and tissue/cell handling (EUTCD) [18] are similar; both regulations aim to ensure high safety and quality standards. The main differences apply to scope and application. Specifically, whereas drug legislation ensures safety through comprehensive technical and testing requirements at the product level [45], the EUTCD ensures safety by establishing a standardized and precautionary handling system at an organizational level [18]. Regulating FMT as a drug will require the preparative procedures to yield a consistently identical product that is homogenous in terms of its active compounds. This requirement is a restriction that the highly diverse and variable origin of donor stools does not support [16]. Additional manufacturing processes that render it reproducible will therefore be needed. Consequently, the requirement for homogeneity will entail a shift in FMT towards commercialization, promoting the development of predefined therapeutic mixtures of microorganisms. Indeed, such mixtures may be future biologics. Presently, the requirement for standardization and homogeneity severely restricts FMT usage. In contrast, regulating FMT according to the EUTCD enables FMT to be performed continuously if conducted in a safe and standardized system that ensures high-quality standards [18]. In addition, the EUTCD accommodates donor selection and the processing of donated material, thereby securing FMT as the transfer of a heterogonous donor microbiome to manipulate that of the recipient. In doing so, EUTCD also helps delimit the extent of FMT in its definition of


what is to be considered transplantation and what is considered a drug. On this basis, we believe that considering FMT within the EUTCD provides the best regulatory basis for FMT as a transplantation procedure. Introducing an FMT framework helps facilitate all aspects related to FMT, but both the implementation of this framework and its additional safety procedures are time consuming. Although the addition of standardized protocols that ensure logging and safety may seem excessive and troublesome to the clinical application, recent studies indicate that AEs are under-reported because of inadequate standardization and lack of long-term followup [26]. At our institution, we have performed a total of 122 FMTs since the beginning of 2014, primarily on RCDI patients. Ninety-three of these have been performed after the introduction of the FMT framework. Since this introduction, we have experienced a similar trend towards an increase in documented AEs, further suggesting that standards should be developed. This experience also highlights the need for clinical trials that use similar follow-up strategies to sufficiently address any AEs of FMT. One of the main challenges in establishing an FMT service is the documentation of parameters and processes that have the potential to critically influence the microbial composition and to adapt the protocols accordingly. Even though the EUTCD permits the validation of protocols to be based on retrospective follow-up [18], it is largely unknown how processing and storage precisely affect microbial viability. Hence, estimation of accurate safety intervals of variables, such as temperature and expiration, becomes difficult. Moreover, precise functional knowledge is lacking, and most FMT protocols currently rely on results from RCDI trials. In the case of RCDI, however, FMT appears to be effective irrespective of application and preparation methods [23,40]. Consequently, RCDI may not be the ideal model to validate FMT for other diseases that are characterized by more complex pathogenesis and mechanisms of action [13]. Extending indications beyond RCDI thus requires FMT to be performed in a standardized framework to ensure that potential hazardous risks are limited. The recruitment of faecal donors among blood donors has obvious advantages and could secure a continuous supply of faeces. In our experience, recruitment from a prescreened cohort of healthy individuals eases access to potential donors while also increasing eligibility. An important aspect to bear in mind is the management and information of unqualified donors who are rejected during the screening process. Even with an increase in the expected eligibility rates of 6–10% [27–29], the management and information of unqualified donors add a timeconsuming task that requires the availability of a specialist in case of influential findings. The practical handling of different donation volumes and storage space become urgent as an FMT service becomes functional. Although all donors are booked for the same number of donations, faecal donation volumes between donors vary markedly. The time of delivery and pressure for delivery are heavily influencing factors; hence, offering flexible opening hours and using experienced donors can make donations more reliable. Different donation volumes still make estimation of the yield from one donation difficult and raise the concern of when to



cancel processing because of small donation sizes. Therefore, banking more suspensions is required and further storage of both suspensions currently in quarantine and safety samples necessitates considerable amounts of freezer capacity. The FMT service presented here has limitations. First, it is based on a single-centre experience and does not account for the costs of implementation. Although it may be beneficial to establish such a system from common European law, direct applicability outside Europe would be limited. Thus, the framework represents an attempt to link regulatory measures with the clinical application rather than dictating protocols. Further incorporation of these protocols into existing institutions, such as the blood bank system, may be needed for them to function adequately on a greater multicentre scale.

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Conclusion

FMT constitutes an effective treatment option with emerging potential. With its increased use in clinical settings, the need for regulatory oversight becomes urgent, requiring the adaptation of current research protocols into regulatory frameworks. The principles for a clinical FMT service outlined here define an application framework that complies with the EUTCD and provide institutions looking to establish FMT as a clinical treatment modality with a pragmatic and sustainable structural basis. Acknowledgements

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Conflicts of interest

There are no conflicts of interest.

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Editorial: a simple faecal preparation protocol for faecal microbiota transplantation.

Gastroenterologist perceptions of faecal microbiota transplantation.

Faecal microbiota transplantation: Key points to consider.

Faecal microbiota transplantation--the Austrian approach.

Faecal microbiota transplantation for Clostridium difficile infection.

Faecal microbiota transplantation in Clostridium difficile infections.

Clinical and immunologic effects of faecal microbiota transplantation in a patient with collagenous colitis.

Editorial: a simple faecal preparation for faecal microbiota transplantation--authors' reply.

Toward Rational Donor Selection in Faecal Microbiota Transplantation for IBD.

Faecal microbiota transplantation: applications and limitations in treating gastrointestinal disorders.

Faecal microbiota transplantation protects against radiation-induced toxicity.

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