Worldwide Regulatory Reliance: Launching A Pilot on A Chemistry, Manufacturing and Control Post Approval Change for A Vaccine.

When an initial marketing authorization of a pharmaceutical product is granted, a substantial number of chemistry, manufacturing and control (CMC) post approval changes (PACs) have to be managed by the manufacturers. Despite efforts undertaken over the years by multiple regulatory jurisdictions, there is still heterogeneity in terms of regulatory requirements and timelines across national regulatory authorities (NRAs). This creates complexity in managing global CMC PACs, putting the supply of medical products at risk. Regulators have developed regulatory mechanisms which aim at accelerating the reviews and approvals of PACs by NRAs. The World Health Organization (WHO) is supporting the concept of 'reliance' amongst NRAs which are encouraged to rely on the assessment completed by a ″high-performing authority″. The objective is to accelerate the overall process for PACs, ultimately fostering more equitable and timely access of medical products to populations who need them. With the support of Health Canada, WHO, Pan American Health Organization, and the Paul-Ehrlich-Institut, Sanofi has launched a pilot using the principles of reliance for a CMC PAC for a vaccine, with 21 NRAs who accepted to participate in the pilot. The objective of this pilot was to apply these principles to reduce the approval timeline to a maximum of 6 months, in all countries after an initial approval is granted by a reference authority. We discussed the opportunities and challenges of implementing reliance principles for CMC PACs. We also described the pilot experience, by sharing initial lessons learned from the Step 1 of this pilot, which consist of engaging the reference authority and the NRAs.

Influence of agricultural amendments on arsenic biogeochemistry and phytotoxicity in a soil polluted by the destruction of arsenic-containing shells.

Agricultural soils can contain high arsenic (As) concentrations due to specific geological contexts or pollution. Fertilizer amendments could influence As speciation and mobility thus increasing its transfer to crops and its toxicity. In the present study, field-relevant amounts of fertilizers were applied to soils from a cultivated field that was a former ammunition-burning site. Potassium phosphate (KP), ammonium sulfate and organic matter (OM) were applied to these soils in laboratory experiments to assess their impact on As leaching, bioavailability to Lactuca sativa and microbial parameters. None of the fertilizers markedly influenced As speciation and mobility, although trends showed an increase of mobility with KP and a decrease of mobility with ammonium sulfate. Moreover, KP induced a small increase of As in Lactuca sativa, and the polluted soil amended with ammonium sulfate was significantly less phytotoxic than the un-amended soil. Most probable numbers of AsIII-oxidizing microbes and AsIII-oxidizing activity were strongly linked to As levels in water and soils. Ammonium sulfate negatively affected AsIII-oxidizing activity in the un-polluted soil. Whereas no significant effect on As speciation in water could be detected, amendments may have an impact in the long term.

Aided-phytostabilization of steel slag dumps: The key-role of pH adjustment in decreasing chromium toxicity and improving manganese, phosphorus and zinc phytoavailability.

Because of their high content in toxic metals, steel slag dumps are potential threats for the environment and public health. Among management methods that could mitigate their hazard, aided-phytostabilization is a relevant, though challenging, option. Indeed, steel slags are very unfavorable for plant growth, due to metal toxicity and very alkaline pH (>10). In this work, we investigated how composted sewage sludge could alleviate slag's toxicity while improving its nutritional status. A pot experiment was performed to study biomass production and leaf ionome composition of five herbaceous species (Achillea millefolium, Bromus erectus, Festuca arundinacea, Melilotus officinalis and Medicago sativa), in relation to soil pore water's pH, concentration of trace and major elements and their chemical speciation. Results showed that pH had a clear-cut effect on plant development. Above pH 8.6, plant biomass was severely affected, due to accumulation of Cr above toxic threshold and deficiencies in Mn, Zn and P. Below pH 8.6, biomass increased significantly, together with a decrease in leaf Cr below toxic level, and an increase in Mn, Zn and P above deficiency levels. Thus, these results bring new insights into the causes of slag phytotoxicity and allow considering aided-pytostabilization as a realistic and efficient approach for the remediation of steel slag dumps, provided soil pH is carefully monitored before seeding.

Bacterial diversity on an abandoned, industrial wasteland contaminated by polychlorinated biphenyls, dioxins, furans and trace metals.

Most former industrial sites are contaminated by mixtures of trace elements and organic pollutants. Levels of pollutants do not provide information regarding their biological impact, bioavailability and possible interactions between substances. There is genuine interest in combining chemical analyses with biological investigations. We studied a brownfield where several industrial activities were carried out starting in the 1970s, (incineration of pyralene transformers, recovery of copper by burning cables in the open air). Four representative plots showing different levels of polychlorobiphenyls were selected. Organic and trace metal levels were measured together with soil pedological characteristics. The bacterial community structure and functional diversity were assessed by 16S metagenomics with deep sequencing and community-level physiological profiling. Additionally, a vegetation survey was performed. Polychlorobiphenyls (8 mg.kg-1 to 1500 mg.kg-1) were from 2.4 × 103-fold to 6 × 105-fold higher than the European background level of 2.5 µg.kg-1. Polychlorinated dibenzo-p-dioxins and dibenzofurans ranged from 0.5 to 8.0 µg.kg-1. The soil was also contaminated with trace metals, i.e., Cu > 187, Zn > 217 and Pb > 372 mg.kg-1. Location within the study area, trace metal content and soil humidity were stronger determinants than organic pollutants of bacterial community structures and activities. Thus, the highest biological activity and the greatest bacteriological richness were observed in the plot that was less contaminated with trace metals, despite the high level of organic pollutants in the plot. Moreover, trace element pollution was associated with a relatively low presence of Actinobacteria and Rhizobia. The plot with the highest metal contamination was rich in metal-resistant bacteria such as Sphingomonadales, Geodermatophilaceae and KD4-96 (Chloroflexi phylum). Acidobacteria and Sphingomonadales, capable of resisting trace metals and degrading persistent organic pollutants, were dominant in the plots that had accumulated metal and organic contamination, but bacterial activity was lower in these plots than in the other plots.

Aided phytostabilization of a trace element-contaminated technosol developed on steel mill wastes.

Aided phytostabilization of a barren, alkaline metal(loid)-contaminated technosol developed on steel mill wastes, with high soluble Cr and Mo concentrations, was assessed in a pot experiment using (1) Ni/Cd-tolerant populations of Festuca pratensis Huds., Holcus lanatus L., and Plantago lanceolata L. sowed in mixed stand and (2) six soil treatments: untreated soil (UNT), ramial chipped wood (RCW, 500m3ha-1), composted sewage sludge (CSS, 120t DW ha-1), UNT soil amended with compost (5% w/w) and either vermiculite (5%, VOM) or iron grit (1%, OMZ), and an uncontaminated soil (CTRL). In the CSS soil, pH and soluble Cr decreased whereas soluble Cu, K, Fe, Mn, Mg, Ni and P increased. The RCW treatment enhanced soluble Fe, Mn, and Mg concentrations. After 15 weeks, shoot DW yield and shoot Cd, Cu, Fe, Mn, Mo, Zn, and Mg removals peaked for F. pratensis grown on the CSS soil, with lowest shoot Cr, Ni and Mo concentrations. Holcus lanatus only grew on the CTRL, UNT, and CSS soils and P. lanceolata on the CTRL soil. Best treatment, F. pratensis grown on the CSS soil, led to a dense grass cover but its shoot Mo concentration exceeded the maximum permitted concentration in forage.

Plants increase arsenic in solution but decrease the non-specifically bound fraction in the rhizosphere of an alkaline, naturally rich soil.

We aimed at determining the major physical-chemical processes that drive arsenic (As) dynamic in the rhizosphere of four species (Holcus lanatus, Dittrichia viscosa, Lotus corniculatus, Plantago lanceolata) tested for phytostabilization. Experiments were performed with an alkaline soil naturally rich in As. Composition of the soil solution of planted and unplanted pots was monitored every 15 days for 90 days, with a focus on the evolution of As concentrations in solution and in the non-specifically bound (i.e. easily exchangeable) fraction. The four species similarly increased As concentration in solution, but decreased As concentration in the non-specifically bound fraction. The major part (60%) of As desorbed from the non-specifically bound fraction in planted pots was likely redistributed on the less available fractions of As on the solid phase. A second part (35%) of desorbed As was taken up by plants. The minor part (5%) of desorbed As supplied As increase in solution. To conclude, plants induced a substantial redistribution of As on the less available fractions in the rhizosphere, as expected in phytostabilization strategies. Plants however concomitantly increased As concentration in the rhizosphere solution which may contribute to As transfer through plant uptake and leaching.

Effect of fresh and mature organic amendments on the phytoremediation of technosols contaminated with high concentrations of trace elements.

Organic compounds resulting from the decomposition of organic amendments are used in the remediation of trace element (TE) contaminated soils. The mobility, phytoavailability and soil exposure intensity of molybdenum (Mo), chromium (Cr), zinc (Zn), copper (Cu), Cobalt (Co) and Arsenic (As) were evaluated in the phytoremediation of contaminated technosols after the addition of two organic matter types, fresh ramial chipped wood (RCW) and composted sewage sludge (CSS). The experiment consisted of nine main treatment blocks: (A) 3X unamended soil (NE), (B) 3X soil amended with RCW and (C) 3X soil amended with mature CSS. Total dissolved TE concentrations were determined in soil pore water (SPW) sampled by Rhizon samplers. The soil exposure intensity was assessed by standard Chelex 100 DGT (diffusive gradient in thin films) probes. TE phytoavailability was characterized by growing dwarf beans on potted soils and analyzing their foliar TE concentrations. The results of the present study indicate that the addition of fresh RCW and CSS has a positive effect on contaminated technosols. RCW decreased the mobility of all the studied TE in the SPW, whereas CSS reduced the mobility of Mo, Cr and Co, while it increased the mobility of Zn, Cu and As compared with the NE soil. The Zn soil exposure intensity assessed by DGT was not significantly changed by the addition of RCW and CSS, while the Cr soil exposure intensity was significantly decreased after RCW addition compared with the soil treated with CSS and the NE soil. In contrast Cu and Co were non labile in the three soils. Both RCW and CSS decreased the foliar concentration and the mineral mass of Mo, Zn, Cr, As and Co in the bean leaves but increased the foliar Cu concentration.

In situ assessment of phyto and zooavailability of trace elements: A complementary approach to chemical extraction procedures.

For an accurate risk assessment of sites contaminated by trace elements (TE), measurements of bioavailability must be performed. This is routinely achieved using the standardized 0.01M CaCl2 method. However, the suitability of chemical extractions as proxies of bioavailability is questionable. We analyzed the correlations between chemically estimated TE bioavailability and TE actually accumulated by coupling plant and snails bioindicators. Results showed a better correlation between plant TE contents and CaCl2 fraction while total soil concentration better explained snail TE contents. However in both cases chemical measures were not suitable to predict TE accumulation and bioavailability. Considering the soil properties only improve the estimation of Cr, Ni and Pb accumulation by plants while for snails, TE contents in viscera were dependent both on soil and plant contents and soil properties. It highlights the complementarities of biomonitoring methods to assess bioavailability. This dual approach allows a "physiologically defined" evaluation of bioavailability.

Accumulation of Metals and Boron in Phragmites australis Planted in Constructed Wetlands Polishing Real Electroplating Wastewater.

The concentration of metals (Al, Cu, Fe, Mn, Ni, Zn) and B were determined in the above- and belowground biomass of Phragmites australis collected from the microcosm constructed wetland system used for the polishing of real electroplating wastewater. Translocation factor and bioconcentration factor were determined. Pearson correlation test was used to determine correlation between metal concentration in substrate and above- and belowground parts of Phragmites australis. The obtained results suggested that Phragmites australis did not play a major role as an accumulator of metals. It was observed also that the substrate could have exerted an effect on the translocation of Ni, Cu, Zn and Mn. The analysed concentrations of metals and B in biomass were in the range or even below the concentrations reported in the literature with the exception of Ni. The aboveground biomass was found suitable as a composting input in terms of metals concentrations.

Early changes in the fatty acid composition of photosynthetic membrane lipids from Populus nigra grown on a metallurgical landfill.

We compared the fatty acid composition of leaves taken from poplars on a metal-contaminated landfill, and on the uncontaminated roadside bordering this site. For the first time, it is shown that the percentage of linolenic acid, which is mainly associated with thylakoid lipids, was significantly lower in tree species within the landfill than within the control area. A correlation study was carried out to investigate relationships between the C18:3/(C18:0 + C18:1 + C18:2) fatty acid ratios and the metal contents in soils and leaves. Lead and chromium leaf contents were significantly negatively correlated to this fatty acid ratio. The impact of each of these metals remains difficult to evaluate, but chromium in leaf likely plays a major role in toxicity. In addition, the decrease in the C18:3/(C18:0 + C18:1 + C18:2) fatty acid ratio occurred at low leaf metal content, and therefore it is shown that this ratio can be used as an early indicator of the effect of metals.

Expression of stress-related genes in tomato plants exposed to arsenic and chromium in nutrient solution.

The molecular responses of hydroponically cultivated tomato plants to As(V) or Cr(VI) were assessed by transcript accumulation analysis of genes coding for products potentially involved in heavy metal tolerance. A quantitative real-time PCR experiment was performed with Hsp90-1, MT2- and GR1-like protein genes using RNA isolated from tomato roots or shoots treated for 24h with As(V) or Cr(VI) at concentrations ranging from 80 to 640 microM. Both transient metallic treatments induced Hsp90-1 transcript accumulation in tomato plants. MT2- and GR1-like transcripts accumulated in tomato roots treated with As(V) but were only slightly affected by Cr(VI) treatment. Tomatoes showed phenotypic symptoms to heavy metal toxicity when plants were exposed to Cr(VI) but not As(V). Plant lethality was observed at 1280 microM Cr(VI), indicating that tomatoes were more tolerant to As than Cr stress under the experimental conditions used here.

Effect of chromium species on phytochemical and physiological parameters in Datura innoxia.

In soil, chromium can be found in two main valence states: hexavalent Cr(VI) and trivalent Cr(III). In this study, we investigated the impact of Cr on photosynthetic gas exchange, photosystem II (PSII) activity, Cr translocation and accumulation, proline content and alkaloids production, i.e. scopolamine and hyoscyamine, in Datura innoxia. Cr uptake was influenced by its oxidation state and its concentration in growth medium. The plant roots were determined as being the main organ of Cr accumulation. Cr(VI) was more toxic than Cr(III) as indicated by reduction in plant biomass and net photosynthesis. The stomatal conductance showed a similar trend to that of photosynthetic capacity. Cr(III) and Cr(VI) had a different impact on substomatal CO(2) concentration then Cr toxicity was related to its oxidation states. In plants stressed with a Cr(VI) excess, a down regulation of PSII activity was observed with an impairment of photochemical activity. Indeed, the maximum quantum yield of PSII (F(v)/F(m)), the quantum yield of PSII (PhiPSII) and the efficiency of excitation capture by open centers (F'(v)/F'(m)) decreased. Cr(III) had little effects on PSII primary photochemistry, whatever its form induces an increase of scopolamine content without changes in hyoscyamine content in leaves of D. innoxia. These results provide that chromium contamination can change the secondary metabolites composition of leaves, thereby, impacting the quality, safety and efficacy of natural plant products synthesized by D. innoxia plants.

A polyepitope DNA vaccine targeted to Her-2/ErbB-2 elicits a broad range of human and murine CTL effectors to protect against tumor challenge.

A cDNA vaccine (pVax1/pet-neu) was designed to encode 12 different Her-2/ErbB-2-derived, HLA-A*0201-restricted dominant and high-affinity heteroclitic cryptic epitopes. Vaccination with pVax1/pet-neu triggered multiple and ErbB-2-specific CTL responses in HLA-A*0201 transgenic HHD mice and in HLA-A*0201 healthy donors in vitro. Human and murine CTL specific for each one of the 12 ErbB-2 peptides recognized in vitro both human and murine tumor cells overexpressing endogenous ErbB-2. Furthermore, vaccination of HHD mice with pVax1/pet-neu significantly delayed the in vivo growth of challenged ErbB-2-expressing tumor (EL4/HHD/neu murine thymoma) more actively when compared with vaccination with the empty vector (pVax1) or vehicle alone. These data indicate that the pVax1/pet-neu cDNA vaccine coding for a poly-ErbB-2 epitope is able to generate simultaneous ErbB-2-specific antitumor responses against dominant and cryptic multiple epitopes.

Induction of multiple CD8+ T cell responses against the inducible Hsp70 employing an Hsp70 oligoepitope peptide.

The inducible heat shock protein Hsp70 has been described as a tumour antigen being frequently overexpressed in tumours of various histologic origins, with a role in tumourigenicity, as a critical event in tumour progression. A strategy to enhance the immune response to an antigen is the identification of multiple epitopes and the induction of a polyspecific response. Applied to tumour vaccination, such a polyspecific response should lead to a more robust antitumour efficacy. The long peptide Hsp70380-402 encompasses three nonamer peptides with a high affinity for HLA-A *0201. In a previous paper, we have shown that two of these nonamer peptides, p391 and p393, can raise CTL to recognize tumour cells overexpressing Hsp70. In the present paper, we demonstrate that the third nonamer peptide, p380, is a new epitope efficient in raising an antitumour immune response. The p380-402 polypeptide was able to induce an immune response against each of the three constituent epitopes both in vivo in HLA-A *0201 transgenic mice and in vitro with human PBMC. This polypeptide therefore constitutes an interesting candidate for the induction of multiple HLA-A *0201-restricted anti-Hsp70 antitumour CTL responses.

[Courses of cure: the case of French patients of Samuel and Melanie Hahnemann (1834-1868)]. / Behandlungsverläufe: Die französischen Patienten von Samuel und Mélanie Hahnemann (1834-1868).

Based on the letters sent by French patients to Hahnemann and his wife (or written about them), this essay focuses on the behaviour of patients towards treatment rather than on their representations and interpretations of illness and health. Despite the fact that we don't know exactly who the authors of these letters were, it is clear that they all hope to obtain some of Hahnemann's medicines. The well-known demand for medicine at the beginning of the 19th Century was reinforced by the mystery surrounding homeopathic remedies and the specific way they were made available. Relying on theories which lent an important role to the nerves and the patient's character to explain the origins of sickness, patients hoped that Hahnemann and his new doctrine would be able to change both their life and their psychical characteristics. One can conclude that for the authors of such letters, writing about illness could be a means to construct new behaviour patterns and new attitudes towards health and sickness, rather than an illustration of existing medical, social and literary models.

STEAP, a prostate tumor antigen, is a target of human CD8+ T cells.

STEAP is a recently identified protein shown to be particularly overexpressed in prostate cancer and also present in numerous human cancer cell lines from prostate, pancreas, colon, breast, testicular, cervical, bladder and ovarian carcinoma, acute lymphocytic leukemia and Ewing sarcoma. This expression profile renders STEAP an appealing candidate for broad cancer immunotherapy. In order to investigate if STEAP is a tumor antigen that can be targeted by specific CD8(+) T cells, we identified two high affinity HLA-A*0201 restricted peptides (STEAP(86-94) and STEAP(262-270)). These peptides were immunogenic in vivo in HLA-A*0201 transgenic HHD mice. Peptide specific murine CD8 T cells recognized COS-7 cells co-transfected with HHD (HLA-A*0201) and STEAP cDNA constructs and also HLA-A*0201(+) STEAP(+) human tumor cells. Furthermore, STEAP(86-94) and STEAP(262-270) stimulated specific CD8(+) T cells from HLA-A*0201(+) healthy donors, and these peptide specific CD8(+) T cells recognized STEAP positive human tumor cells in an HLA-A*0201-restricted manner. Importantly, STEAP(86-94)-specific T cells were detected and reactive in the peripheral blood mononuclear cells in NSCLC and prostate cancer patients ex vivo. These results show that STEAP can be a target of anti-tumor CD8(+) T cells and that STEAP peptides can be used for a broad-spectrum-tumor immunotherapy.