Using Real-World Data in Health Technology Assessment (HTA) Practice:A Comparative Study of Five HTA Agencies

BACKGROUND: Reimbursement decisions are conventionally based on evidence from randomised controlled trials (RCTs), which often have high internal validity but low external validity. Real-world data (RWD) may provide complimentary evidence for relative effectiveness assessments (REAs) and cost-effectiveness assessments (CEAs). This study examines whether RWD is incorporated in health technology assessment (HTA) of melanoma drugs by European HTA agencies, as well as differences in RWD use between agencies and across time. METHODS: HTA reports published between 1 January 2011 and 31 December 2016 were retrieved from websites of agencies representing five jurisdictions: England [National Institute for Health and Care Excellence (NICE)], Scotland [Scottish Medicines Consortium (SMC)], France [Haute Autorité de santé (HAS)], Germany [Institute for Quality and Efficacy in Healthcare (IQWiG)] and The Netherlands [Zorginstituut Nederland (ZIN)]. A standardized data extraction form was used to extract information on RWD inclusion for both REAs and CEAs. RESULTS: Overall, 52 reports were retrieved, all of which contained REAs; CEAs were present in 25 of the reports. RWD was included in 28 of the 52 REAs (54%), mainly to estimate melanoma prevalence, and in 22 of the 25 (88%) CEAs, mainly to extrapolate long-term effectiveness and/or identify drug-related costs. Differences emerged between agencies regarding RWD use in REAs; the ZIN and IQWiG cited RWD for evidence on prevalence, whereas the NICE, SMC and HAS additionally cited RWD use for drug effectiveness. No visible trend for RWD use in REAs and CEAs over time was observed. CONCLUSION: In general, RWD inclusion was higher in CEAs than REAs, and was mostly used to estimate melanoma prevalence in REAs or to predict long-term effectiveness in CEAs. Differences emerged between agencies' use of RWD; however, no visible trends for RWD use over time were observed

Copernicus Marine Service ocean state report, issue 4

This is the final version. Available from Taylor & Francis via the DOI in this record. FCT/MCTE

Mycolactone toxin induces an inflammatory response by targeting the IL-1β pathway: Mechanistic insight into Buruli ulcer pathophysiology

International audienceMycolactone, a lipid-like toxin, is the major virulence factor of Mycobacterium ulcerans, the etiological agent of Buruli ulcer. Its involvement in lesion development has been widely described in early stages of the disease, through its cytotoxic and immunosuppressive activities, but less is known about later stages. Here, we revisit the role of mycolactone in disease outcome and provide the first demonstration of the pro-inflammatory potential of this toxin. We found that the mycolactone-containing mycobacterial extracellular vesicles produced by M. ulcerans induced the production of IL-1β, a potent pro-inflammatory cytokine, in a TLR2dependent manner, targeting NLRP3/1 inflammasomes. We show our data to be relevant in a physiological context. The in vivo injection of these mycolactone-containing vesicles induced a strong local inflammatory response and tissue damage, which were prevented by corticosteroids. Finally, several soluble pro-inflammatory factors, including IL-1β, were detected in infected tissues from mice and Buruli ulcer patients. Our results revisit Buruli ulcer pathophysiology by providing new insight, thus paving the way for the development of new therapeutic strategies taking the pro-inflammatory potential of mycolactone into account

The LipL21 lipoprotein impairs leptospiral peptidoglycan digestion into muropeptides.

<p>(A) PG 0.5 h and PG 4 h from <i>L</i>. <i>interrogans</i> Manilae L495 and Fiocruz L1-130 strains, were loaded on coomassie-stained gels (upper panels; protein gels), revealing a 21-kDa protein corresponding to the LipL21. The protein specificity was checked by immunodetection using LipL21 antiserum (lower panels; western blot, (WB)). (B) LipL21 expression checked by immunodetection on bacterial extracts from <i>L</i>. <i>interrogans</i> Manilae L495 (<i>lipl21</i>⁺⁾, the (<i>lipl21</i>^-) M58 mutant and the complemented C5M58 strain (<i>lipl21</i>^-/+). (C) HPLC separation profiles of muropeptides after digestion by mutanolysin of <i>L</i>. <i>interrogans</i> Manilae <i>lipl21</i>⁺, <i>lipl21</i>^- and <i>lipl21</i>^-/+ peptidoglycans, extracted with the 0.5 h protocol. As positive control for the mutanolysin digestion, <i>E</i>. <i>coli</i> PG was extracted with the leptospiral 0.5 h protocol. D) Growth curves of <i>L</i>. <i>interrogans</i> Manilae <i>lipl21</i>⁺, <i>lipl21</i>^- and <i>lipl21</i>^-/+ strains in EMJH medium at 28°C, with agitation. (E) Muropeptides or 6 μg of PGs extracted from the parental Manilae strain (<i>lipl21</i>+), the LipL21 mutant (<i>lipl21</i>^-) and the complemented strain (<i>lipl21</i>^-/+) were co-transfected with the reporters and NOD1 plasmids in HEK293T cells. 24 h after transfection, luciferase activity was measured. MurTriDAP (MTP) (100 nM), the NOD1 agonist was used as positive control and water as negative control (water). Data are expressed as the mean ± SEM of relative light units representing luciferase activity normalized with respect to β-galactosidase activity. This graph is representative of 3 independent experiments. The unpaired <i>t</i> test was used to compare the recognition of each PG by hNOD1 transfected cells to water as a negative control (none). A <i>p</i> value < 0.05 was considered significant. <i>p</i> values: *** <i>p</i> < 0.001. Non significant differences for the + and -/+ PG are not indicated.</p

Expression of LipL21 impairs <i>E</i>. <i>coli</i> peptidoglycan digestion into muropeptides and recognition by NOD receptors.

<p>(A) Color phenotypes of strains expressing alkaline phosphatase (phoA) derivatives and grown as colonies on agar with chloramphenicol and 5-bromo-4-chloro-3-indoyl-phosphate (XP). FL-<i>phoA</i>, Δ(2–22)phoA, FL-<i>lipl21-phoA</i> and ΔN-<i>lipl21-phoA</i> correspond respectively to <i>E</i>. <i>coli</i> (BTH₁₀₁) expressing the full length phoA (positive control), <i>E</i>. <i>coli</i> expressing phoA without signal peptide (negative control), <i>E</i>. <i>coli</i> expressing the full length LipL21 fused with Δ(2–22)<i>phoA</i> and <i>E</i>. <i>coli</i> expressing the LipL21 without signal peptide fused with Δ(2–22)<i>phoA</i>. (B) Growth curves of BL-21 Rosetta-2 <i>E</i>. <i>coli</i> expressing the LipL21 lipoprotein (pLipL21) or not (pEmpty) in the pRSFDuet-1 vector, without IPTG induction. (C) Crude bacterial extracts of BL-21 Rosetta-2 <i>E</i>. <i>coli</i> with empty pASK-IBA6 vector (pEmpty) or LipL21 expressing vector (pLipL21), induced by anhydrous-Tetracyclin or not (NI) and migrated on 12% acrylamide gel. Stain free coloration (left) and immunodetection (right) using a LipL21 antiserum. The Rainbow marker ladder gives an indication for the LipL21size.(D) HPLC separation profiles of muropeptides after digestion with mutanolysin. Each peptidoglycan was extracted with both the 0.5 h and 4 h protocols from BL-21 Rosetta-2 <i>E</i>. <i>coli</i> expressing the LipL21 lipoprotein (pLipL21) or not (pEmpty), after induction with anhydrous-Tetracyclin. (E) and (F) <i>E</i>. <i>coli</i> PGs, extracted from the 0.5 h protocol, were used to stimulate HEK 293T cells expressing human NOD1 (E) or NOD2 (F). HEK 293T cells were co-transfected with 6 μg of PG or 100 nM of muropeptides controls (MTP for NOD1 and MDP for NOD2, along with the reporter constructions and NOD1 or NOD2 expression plasmids. Luciferase activity was measured 24 h after transfection. Cells left untreated with muropeptides were used as negative control (water). Data are expressed as the mean of triplicates ± SEM of relative light units representing luciferase activity normalized with respect to β-galactosidase activity. The graph shown is representative of 3 equivalent experiments. The unpaired <i>t</i> test was used to compare the recognition of PGs prepared from <i>E</i>. <i>coli</i> with the empty vector (pEmpty) to the PG prepared from <i>E</i>. <i>coli</i> expressing LipL21 (pLipL21). A <i>p</i> value < 0.05 was considered significant. <i>p</i> values: *** <i>p</i> < 0.001. For clarity, statistics comparing each PG or muropeptide control to the water treated cells have not been indicated but are all significant with at least p<0,05.</p