Production of an injectable bacteriophage preparation: identification of the associated critical points

Objective
The trauma department asked to the pharmacy to prepare intra-articular injections of bacteriophages directed against a strain of P. aeruginosa, for compassionate use, under french health authority approval. The objective of this work is to present the critical points of the preparation and to analyse the tests carried out.

Methods
The circuit was defined according to feasibility elements (literacy and feedback).
The aseptic preparation was carried out in a type II laminar flow hood (LFH) in a clean room (B class). Before packaging in syringes, a sterilising filtration through a Poly ether sulphone 0.22µm filter was carried out. The syringe was placed in double aseptic packaging (labelled outer packaging) to maintain asepsis during administration in the operating theatre. To reduce the spread of phages, the preparation was carried out after all those of the week and followed by a double cleaning of the LFH with a disinfectant detergent, then the weekly bio-cleaning of the clean room, associated with the aerosolisation of hydrogen peroxide.
Following an analysis (5M diagrams) identifying the critical points for the preparation, the environment and the manipulator, the following tests were carried out :
1) Sterility test on the finished preparation and environmental samples on agar (contact and sedimentation) under the LFH.
2) Endotoxin test carried out on a sample of the final preparation according to the method described in 2.6.14 and 5.1.10 monographs of the European Pharmacopoeia. The Endotoxin Limit Concentration is 0.034 EU/mL.
3) Objectification of residual contamination by specific swabbing after preparation and indirect detection in microbiology laboratory.

Results
17 critical points were raised (phage conservation, decontamination method, etc.) that could lead to a non-compliant preparation, residual contamination of the LFH and/or cleanroom or of the operator.
The agar plates and sterility tests did not find any CFU.
The vials of bacteriophages (raw materials) were delivered with a certificate of analysis, specifying an endotoxin titre <0.03 EU/mL. The assays on the preparations were 0.016 and 0.010 EU/mL.
Bacteriophages are not class 2 pathogens (french art. R. 4421-2 Code Travail), but the waste was treated via the dedicated infectious disposal route. A meeting with the pharmaceutical staff made it possible to explain the preparation and its risks. Regarding residual contamination, no lysis range was detected from the control swabs.

Discussion/Conclusion
The critical points of the preparation were identified. A risk analysis (i.e. FMEA) or the quantification of the phage titer will be carried out to optimise the performance of this preparation.

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