Personalized bacteriophage therapy outcomes for 100 consecutive cases: a multicentre, multinational, retrospective observational study.

In contrast to the many reports of successful real-world cases of personalized bacteriophage therapy (BT), randomized controlled trials of non-personalized bacteriophage products have not produced the expected results. Here we present the outcomes of a retrospective observational analysis of the first 100 consecutive cases of personalized BT of difficult-to-treat infections facilitated by a Belgian consortium in 35 hospitals, 29 cities and 12 countries during the period from 1 January 2008 to 30 April 2022. We assessed how often personalized BT produced a positive clinical outcome (general efficacy) and performed a regression analysis to identify functional relationships. The most common indications were lower respiratory tract, skin and soft tissue, and bone infections, and involved combinations of 26 bacteriophages and 6 defined bacteriophage cocktails, individually selected and sometimes pre-adapted to target the causative bacterial pathogens. Clinical improvement and eradication of the targeted bacteria were reported for 77.2% and 61.3% of infections, respectively. In our dataset of 100 cases, eradication was 70% less probable when no concomitant antibiotics were used (odds ratio = 0.3; 95% confidence interval = 0.127-0.749). In vivo selection of bacteriophage resistance and in vitro bacteriophage-antibiotic synergy were documented in 43.8% (7/16 patients) and 90% (9/10) of evaluated patients, respectively. We observed a combination of antibiotic re-sensitization and reduced virulence in bacteriophage-resistant bacterial isolates that emerged during BT. Bacteriophage immune neutralization was observed in 38.5% (5/13) of screened patients. Fifteen adverse events were reported, including seven non-serious adverse drug reactions suspected to be linked to BT. While our analysis is limited by the uncontrolled nature of these data, it indicates that BT can be effective in combination with antibiotics and can inform the design of future controlled clinical trials. BT100 study, ClinicalTrials.gov registration: NCT05498363 .

Recellularizing of human acellular dermal matrices imaged by high-definition optical coherence tomography.

High-definition optical coherence tomography (HD-OCT) permits real-time 3D imaging of the impact of selected agents on human skin allografts. The real-time 3D HD-OCT assessment of (i) the impact on morphological and cellular characteristics of the processing of human acellular dermal matrices (HADMs) and (ii) repopulation of HADMs in vitro by human fibroblasts and remodelling of the extracellular matrix by these cells. Four different skin decellularization methods, Dispase II/Triton X-100, Dispase II/SDS (sodium dodecyl sulphate), NaCl/Triton X-100 and NaCl/SDS, were analysed by HD-OCT. HD-OCT features of epidermal removal, dermo-epidermal junction (DEJ) integrity, cellularity and dermal architecture were correlated with reflectance confocal microscopy (RCM), histopathology and immunohistochemistry. Human adult dermal fibroblasts were in vitro seeded on the NaCl/Triton X-100 processed HADMs, cultured up to 19 days and evaluated by HD-OCT in comparison with MTT proliferation test and histology. Epidermis was effectively removed by all treatments. DEJ was best preserved after NaCl/Triton X-100 treatment. Dispase II/SDS treatment seemed to remove all cellular debris in comparison with NaCl/Triton X-100 but disturbed the DEJ severely. The dermal micro-architectural structure and vascular spaces of (sub)papillary dermis were best preserved with the NaCl/Triton X-100. The impact on the 3D structure and vascular holes was detrimental with Dispase II/SDS. Elastic fibre fragmentation was only observed after Dispase II incubation. HD-OCT showed that NaCl/Triton X-100 processed matrices permitted in vitro repopulation by human dermal fibroblasts (confirmed by MTT test and histology) and underwent remodelling upon increasing incubation time. Care must be taken in choosing the appropriate processing steps to maintain selected properties of the extracellular matrix in HADMs. Processing HADMs with NaCl/Triton X-100 permits in vitro the proliferation and remodelling activity of human dermal fibroblasts. HD-OCT provides unique real-time and non-invasive 3D imaging of tissue-engineered skin constructs and complementary morphological and cytological information.

Real-time three-dimensional imaging of epidermal splitting and removal by high-definition optical coherence tomography.

While real-time 3-D evaluation of human skin constructs is needed, only 2-D non-invasive imaging techniques are available. The aim of this paper is to evaluate the potential of high-definition optical coherence tomography (HD-OCT) for real-time 3-D assessment of the epidermal splitting and decellularization. Human skin samples were incubated with four different agents: Dispase II, NaCl 1 M, sodium dodecyl sulphate (SDS) and Triton X-100. Epidermal splitting, dermo-epidermal junction, acellularity and 3-D architecture of dermal matrices were evaluated by High-definition optical coherence tomography before and after incubation. Real-time 3-D HD-OCT assessment was compared with 2-D en face assessment by reflectance confocal microscopy (RCM). (Immuno) histopathology was used as control. HD-OCT imaging allowed real-time 3-D visualization of the impact of selected agents on epidermal splitting, dermo-epidermal junction, dermal architecture, vascular spaces and cellularity. RCM has a better resolution (1 µm) than HD-OCT (3 µm), permitting differentiation of different collagen fibres, but HD-OCT imaging has deeper penetration (570 µm) than RCM imaging (200 µm). Dispase II and NaCl treatments were found to be equally efficient in the removal of the epidermis from human split-thickness skin allografts. However, a different epidermal splitting level at the dermo-epidermal junction could be observed and confirmed by immunolabelling of collagen type IV and type VII. Epidermal splitting occurred at the level of the lamina densa with dispase II and above the lamina densa (in the lamina lucida) with NaCl. The 3-D architecture of dermal papillae and dermis was more affected by Dispase II on HD-OCT which corresponded with histopathologic (orcein staining) fragmentation of elastic fibres. With SDS treatment, the epidermal removal was incomplete as remnants of the epidermal basal cell layer remained attached to the basement membrane on the dermis. With Triton X-100 treatment, the epidermis was not removed. In conclusion, HD-OCT imaging permits real-time 3-D visualization of the impact of selected agents on human skin allografts.

Business oriented EU human cell and tissue product legislation will adversely impact Member States' health care systems.

The transplantation of conventional human cell and tissue grafts, such as heart valve replacements and skin for severely burnt patients, has saved many lives over the last decades. The late eighties saw the emergence of tissue engineering with the focus on the development of biological substitutes that restore or improve tissue function. In the nineties, at the height of the tissue engineering hype, industry incited policymakers to create a European regulatory environment, which would facilitate the emergence of a strong single market for tissue engineered products and their starting materials (human cells and tissues). In this paper we analyze the elaboration process of this new European Union (EU) human cell and tissue product regulatory regime-i.e. the EU Cell and Tissue Directives (EUCTDs) and the Advanced Therapy Medicinal Product (ATMP) Regulation and evaluate its impact on Member States' health care systems. We demonstrate that the successful lobbying on key areas of regulatory and policy processes by industry, in congruence with Europe's risk aversion and urge to promote growth and jobs, led to excessively business oriented legislation. Expensive industry oriented requirements were introduced and contentious social and ethical issues were excluded. We found indications that this new EU safety and health legislation will adversely impact Member States' health care systems; since 30 December 2012 (the end of the ATMP transitional period) there is a clear threat to the sustainability of some lifesaving and established ATMPs that were provided by public health institutions and small and medium-sized enterprises under the frame of the EUCTDs. In the light of the current economic crisis it is not clear how social security systems will cope with the inflation of costs associated with this new regulatory regime and how priorities will be set with regard to reimbursement decisions. We argue that the ATMP Regulation should urgently be revised to focus on delivering affordable therapies to all who are in need of them and this without necessarily going to the market. The most rapid and elegant way to achieve this would be for the European Commission to publish an interpretative document on "placing on the market of ATMPs," which keeps tailor-made and niche ATMPs outside of the scope of the medicinal product regulation.

Beware of the commercialization of human cells and tissues: situation in the European Union.

With this analysis we would like to raise some issues that emerge as a result of recent evolutions in the burgeoning field of human cells, tissues, and cellular and tissue-based product (HCT/P) transplantation, and this in the light of the current EU regulatory framework. This paper is intended as an open letter addressed to the EU policy makers, who will be charged with the review and revision of the current legislation. We propose some urgent corrections or additions to cope with the rapid advances in biomedical science, an extensive commercialization of HCT/Ps, and the growing expectation of the general public regarding the ethical use of altruistically donated cells and tissues. Without a sound wake-up call, the diverging interests of this newly established 'healthcare' industry and the wellbeing of humanity will likely lead to totally unacceptable situations, like some of which we are reporting here.