Detection of Crimean-Congo haemorrhagic fever virus in Hyalomma marginatum ticks, southern France, May 2022 and April 2023.

Crimean-Congo haemorrhagic fever (CCHF), a potentially severe zoonotic viral disease causing fever and haemorrhagic manifestations in humans. As the Crimean-Congo haemorrhagic fever virus (CCHFV) has been detected in ticks in Spain and antibodies against the virus in ruminant sera in Corsica, it was necessary to know more about the situation in France. In 2022-2023, CCHFV was detected in 155 ticks collected from horses and cattle in southern France.

Innovation in Radionuclide Therapy for the Treatment of Prostate Cancers: Radiochemical Perspective and Recent Therapeutic Practices.

Prostate cancer represents the second cause of death by cancer in males in western countries. While early-stage diseases are accessible to surgery and/or external radiotherapy, advanced metastatic prostate cancers are primarily treated with androgen deprivation therapy, to which new generation androgen receptor antagonists or taxane-based chemotherapies are added in the case of tumor relapse. Nevertheless, patients become invariably resistant to castration with a median survival that rarely exceeds 3 years. This fostered the search for alternative strategies, independent of the androgen receptor signaling pathway. In this line, radionuclide therapies may represent an interesting option as they could target either the microenvironment of sclerotic bone metastases with the use of radiopharmaceuticals containing samarium-153, strontium-89 or radium-223 or tumor cells expressing the prostate-specific membrane antigen (PSMA), a protein found at the surface of prostate cancer cells. This review gives highlights the chemical properties of radioligands targeting prostate cancer cells and recapitulates the clinical trials evaluating the efficacy of radionuclide therapies, alone or in combination with other approved treatments, in patients with castration-resistant prostate tumors. It discusses some of the encouraging results obtained, especially the benefit on overall survival that was reported with [177Lu]-PSMA-617. It also addresses the specific requirements for the use of this particular class of drugs, both in terms of medical staff coordination and adapted infrastructures for efficient radioprotection.

Improving the response to oxaliplatin by targeting chemotherapy-induced CLDN1 in resistant metastatic colorectal cancer cells.

BACKGROUND: Tumor resistance is a frequent cause of therapy failure and remains a major challenge for the long-term management of colorectal cancer (CRC). The aim of this study was to determine the implication of the tight junctional protein claudin 1 (CLDN1) in the acquired resistance to chemotherapy. METHODS: Immunohistochemistry was used to determine CLDN1 expression in post-chemotherapy liver metastases from 58 CRC patients. The effects of oxaliplatin on membrane CLDN1 expression were evaluated by flow cytometry, immunofluorescence and western blotting experiments in vitro and in vivo. Phosphoproteome analyses, proximity ligation and luciferase reporter assays were used to unravel the mechanism of CLDN1 induction. RNAseq experiments were performed on oxaliplatin-resistant cell lines to investigate the role of CLDN1 in chemoresistance. The "one-two punch" sequential combination of oxaliplatin followed by an anti-CLDN1 antibody-drug conjugate (ADC) was tested in both CRC cell lines and murine models. RESULTS: We found a significant correlation between CLDN1 expression level and histologic response to chemotherapy, CLDN1 expression being the highest in resistant metastatic residual cells of patients showing minor responses. Moreover, in both murine xenograft model and CRC cell lines, CLDN1 expression was upregulated after exposure to conventional chemotherapies used in CRC treatment. CLDN1 overexpression was, at least in part, functionally related to the activation of the MAPKp38/GSK3ß/Wnt/ß-catenin pathway. Overexpression of CLDN1 was also observed in oxaliplatin-resistant CRC cell lines and was associated with resistance to apoptosis, suggesting an anti-apoptotic role for CLDN1. Finally, we demonstrated that the sequential treatment with oxaliplatin followed by an anti-CLDN1 ADC displayed a synergistic effect in vitro and in in vivo. CONCLUSION: Our study identifies CLDN1 as a new biomarker of acquired resistance to chemotherapy in CRC patients and suggests that a "one-two punch" approach targeting chemotherapy-induced CLDN1 expression may represent a therapeutic opportunity to circumvent resistance and to improve the outcome of patients with advanced CRC.

[Molecular signatures in castration-resistant prostate cancers: An update]. / Signatures moléculaires dans les cancers de la prostate résistants à la castration : état des lieux.

Castration resistant prostate cancers are highly heterogenous at the molecular level. With the use of new generation sequencing technologies, a series of alterations were identified, opening the way for potential alternative treatments that could be more efficient for specific molecular subtypes. In this brief update, we summarize the new targetable pathways that are currently investigated. In this era of liquid biopsy and of precision medicine, it seems pertinent to be able to screen for these alterations on a more systematic basis before initiating any treatment.

Seroprevalence of Crimean-Congo Hemorrhagic Fever Among Small Ruminants from Southern Romania.

Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne disease that can be contracted by direct contact with viremic animals or humans. Domestic animals are accidental hosts and contribute to the spread and amplification of the virus. The main objective of this study was to provide updated information related to CCHF virus (CCHFV) infection in Southern Romania by assessing the seroprevalence of CCHF in small ruminants (sheep and goats) using a double-antigen sandwich enzyme-linked immunosorbent assay and by detection of CCHFV in engorged ticks and serum samples using real-time RT-PCR. The overall seroprevalence of CCHF in small ruminants was 37.7% (95% CI 31.7 to 43.7). No statistical seroprevalence difference was observed between the two species of ruminants (p = 0.76), but a significant difference was established between the locations (p < 0.01). No CCHFV RNA was detected in tick pools and small ruminant's sera tested by real-time RT-PCR, although the high seroprevalence to CCHFV among ruminants indicates that CCHV or a closely related virus circulates in Southern Romania.

Multiplexed-Based Assessment of DNA Damage Response to Chemotherapies Using Cell Imaging Cytometry.

The current methods for measuring the DNA damage response (DDR) are relatively labor-intensive and usually based on Western blotting, flow cytometry, and/or confocal immunofluorescence analyses. They require many cells and are often limited to the assessment of a single or few proteins. Here, we used the Celigo® image cytometer to evaluate the cell response to DNA-damaging agents based on a panel of biomarkers associated with the main DDR signaling pathways. We investigated the cytostatic or/and the cytotoxic effects of these drugs using simultaneous propidium iodide and calcein-AM staining. We also describe new dedicated multiplexed protocols to investigate the qualitative (phosphorylation) or the quantitative changes of eleven DDR markers (H2AX, DNA-PKcs, ATR, ATM, CHK1, CHK2, 53BP1, NBS1, RAD51, P53, P21). The results of our study clearly show the advantage of using this methodology because the multiplexed-based evaluation of these markers can be performed in a single experiment using the standard 384-well plate format. The analyses of multiple DDR markers together with the cell cycle status provide valuable insights into the mechanism of action of investigational drugs that induce DNA damage in a time- and cost-effective manner due to the low amounts of antibodies and reagents required.

SARS-CoV-2 antibodies seroprevalence in dogs from France using ELISA and an automated western blotting assay.

Dogs are occasionally susceptible to SARS-CoV-2, developing few or no clinical signs. Epidemiological surveillance of SARS-CoV-2 in dogs requires testing to distinguish it from other canine coronaviruses. In the last year, significant advances have been made in the diagnosis of SARS-CoV-2, allowing its surveillance in both human and animal populations. Here, using ELISA and automated western blotting (AWB) assays, we performed a longitudinal study on 809 apparently healthy dogs from different regions of France to investigate anti-SARS-CoV-2 antibodies. There were three main groups: (i) 356 dogs sampled once before the pandemic, (ii) 235 dogs sampled once during the pandemic, and (iii) 218 dogs, including 82 dogs sampled twice (before and during the pandemic), 125 dogs sampled twice during the pandemic and 11 dogs sampled three times (once before and twice during the pandemic). Using ELISA, seroprevalence was significantly higher during the pandemic [5.5% (25/453)] than during the pre-pandemic period [1.1% (5/449)]. Among the 218 dogs sampled twice, at least 8 ELISA-seroconversions were observed. ELISA positive pre-pandemic sera were not confirmed in serial tests by AWB, indicating possible ELISA cross-reactivity, probably with other canine coronaviruses. A significant difference was observed between these two serological tests (Q = 88, p = 0.008). A clear correlation was observed between SARS-CoV-2 seroprevalence in dogs and the incidence of SARS-CoV-2 infection in human population from the same area. AWB could be used as a second line assay to confirm the doubtful and discrepant ELISA results in dogs. Our results confirm the previous experimental models regarding the susceptibility of dogs to SARS-CoV-2, suggesting that viral transmission from and between dogs is weak or absent. However, the new variants with multiple mutations could adapt to dogs; this hypothesis cannot be ruled out in the absence of genomic data on SARS-CoV-2 from dogs.

Internalization of Foldamer-Based DNA Mimics through a Site-Specific Antibody Conjugate to Target HER2-Positive Cancer Cells.

Inhibition of protein-DNA interactions represents an attractive strategy to modulate essential cellular functions. We reported the synthesis of unique oligoamide-based foldamers that adopt single helical conformations and mimic the negatively charged phosphate moieties of B-DNA. These mimics alter the activity of DNA interacting enzymes used as targets for cancer treatment, such as DNA topoisomerase I, and they are cytotoxic only in the presence of a transfection agent. The aim of our study was to improve internalization and selective delivery of these highly charged molecules to cancer cells. For this purpose, we synthesized an antibody-drug conjugate (ADC) using a DNA mimic as a payload to specifically target cancer cells overexpressing HER2. We report the bioconjugation of a 16-mer DNA mimic with trastuzumab and its functional validation in breast and ovarian cancer cells expressing various levels of HER2. Binding of the ADC to HER2 increased with the expression of the receptor. The ADC was internalized into cells and was more efficient than trastuzumab at inhibiting their growth in vitro. These results provide proof of concept that it is possible to site-specifically graft high molecular weight payloads such as DNA mimics onto monoclonal antibodies to improve their selective internalization and delivery in cancer cells.

PXR Modulates the Prostate Cancer Cell Response to Afatinib by Regulating the Expression of the Monocarboxylate Transporter SLC16A1.

Resistance to castration is a crucial issue in the treatment of metastatic prostate cancer. Kinase inhibitors (KIs) have been tested as potential alternatives, but none of them are approved yet. KIs are subject of extensive metabolism at both the hepatic and the tumor level. Here, we studied the role of PXR (Pregnane X Receptor), a master regulator of metabolism, in the resistance to KIs in a prostate cancer setting. We confirmed that PXR is expressed in prostate tumors and is more frequently detected in advanced forms of the disease. We showed that stable expression of PXR in 22Rv1 prostate cancer cells conferred a resistance to dasatinib and a higher sensitivity to erlotinib, dabrafenib, and afatinib. Higher sensitivity to afatinib was due to a ~ 2-fold increase in its intracellular accumulation and involved the SLC16A1 transporter as its pharmacological inhibition by BAY-8002 suppressed sensitization of 22Rv1 cells to afatinib and was accompanied with reduced intracellular concentration of the drug. We found that PXR could bind to the SLC16A1 promoter and induced its transcription in the presence of PXR agonists. Together, our results suggest that PXR could be a biomarker of response to kinase inhibitors in castration-resistant prostate cancers.

Multiplexed Droplet Digital PCR Assays for the Simultaneous Screening of Major Genetic Alterations in Tumors of the Central Nervous System.

The increased integration of molecular alterations to define tumor type or grade in central nervous system (CNS) tumor classification brings new challenges for the pathologist to make the best use of a precious limited tissue specimen for molecular studies. Within the different methods available to identify gene alterations, the droplet digital PCR (dPCR) constitutes a rapid, cost-effective, and very sensitive tool. In this study, we describe the development and validation of five multiplexed dPCR assays to detect major CNS biomarkers by using only small amounts of DNA extracted from formalin-fixed paraffin-embedded specimens. When compared to HRM-sequencing, NGS-sequencing, RNA-sequencing, or simplex digital PCR assays used as "gold standard" methods, these multiplexed dPCR assays displayed 100% specificity and sensitivity for the simultaneous detection of: 1/BRAF V600E mutation and KIAA1549:BRAF fusion; 2/FGFR1 N546K and K656E mutations and FGFR1 duplication; 3/H3F3A K27M and G34R/V mutations; 4/IDH1 R132X and IDH2 R172X mutations; and 5/TERT promoter mutations C228T and C250T. In light of the increased integration of molecular alteration, we believe that such strategies might help laboratories to optimize their screening strategies for routine diagnosis of pediatric and adult CNS tumors.

The Anti-Cancer Drug Dabrafenib Is a Potent Activator of the Human Pregnane X Receptor.

The human pregnane X receptor (hPXR) is activated by a large set of endogenous and exogenous compounds and plays a critical role in the control of detoxifying enzymes and transporters regulating liver and gastrointestinal drug metabolism and clearance. hPXR is also involved in both the development of multidrug resistance and enhanced cancer cells aggressiveness. Moreover, its unintentional activation by pharmaceutical drugs can mediate drug-drug interactions and cause severe adverse events. In that context, the potential of the anticancer BRAF inhibitor dabrafenib suspected to activate hPXR and the human constitutive androstane receptor (hCAR) has not been thoroughly investigated yet. Using different reporter cellular assays, we demonstrate that dabrafenib can activate hPXR as efficiently as its reference agonist SR12813, whereas it does not activate mouse or zebrafish PXR nor hCAR. We also showed that dabrafenib binds to recombinant hPXR, induces the expression of hPXR responsive genes in colon LS174T-hPXR cancer cells and human hepatocytes and finally increases the proliferation in LS174T-hPXR cells. Our study reveals that by using a panel of different cellular techniques it is possible to improve the assessment of hPXR agonist activity for new developed drugs.

First detection and genome sequencing of SARS-CoV-2 in an infected cat in France.

After its first description in Wuhan (China), SARS-CoV-2 the agent of coronavirus disease 2019 (COVID-19) rapidly spread worldwide. Previous studies suggested that pets could be susceptible to SARS-CoV-2. Here, we investigated the putative infection by SARS-CoV-2 in 22 cats and 11 dogs from owners previously infected or suspected of being infected by SARS-CoV-2. For each animal, rectal, nasopharyngeal swabs and serum were taken. Swabs were submitted to RT-qPCR assays targeting 2 genes of SARS-CoV-2. All dogs were tested SARS-CoV-2 negative. One cat was tested positive by RT-qPCR on rectal swab. Nasopharyngeal swabs from this animal were tested negative. This cat showed mild respiratory and digestive signs. Serological analysis confirms the presence of antibodies against the SARS-CoV-2 in both serum samples taken 10 days apart. Genome sequence analysis revealed that the cat SARS-CoV-2 belongs to the phylogenetic clade A2a like most of the French human SARS-CoV-2. This study reports for the first time the natural infection of a cat in France (near Paris) probably through their owners. There is currently no evidence that cats can spread COVID-19 and owners should not abandon their pets or compromise their welfare.

High Content Screening Using New U2OS Reporter Cell Models Identifies Harmol Hydrochloride as a Selective and Competitive Antagonist of the Androgen Receptor.

Prostate cancer is the most commonly diagnosed malignancy in men. Its growth mainly relies on the activity of the androgen receptor (AR), justifying the use of androgen deprivation therapy as a gold standard treatment for the metastatic disease. Inhibition of the androgen axis using second generation antagonists has improved patients' survival, but is systematically confronted to resistance mechanisms, leading to a median survival that does not exceed 5 years. Counteracting this resistance has been the object of a large number of investigations, with a particular emphasis towards the identification of new AR inhibitors, whether they antagonize the receptor by a competitive or a non-competitive binding. To this end, many high content screens have been performed, to identify new non-steroidal AR antagonists, using a variety of approaches, but reported somewhat controversial results, depending on the approach and on the cell model that was used for screening. In our study, we used the U2OS osteosarcoma cells stably transfected with AR or ARv7 and a luciferase reporter as a previously validated model to screen the Prestwick Phytochemical library. The results of our screen identified ellipticine, harmol, and harmine hydrochloride as confirmed hits. Surprisingly, we could demonstrate that harmol hydrochloride, previously identified as a non-competitive inhibitor of AR or a weak inhibitor of androgen signaling, was actually a competitive antagonist of AR, which inhibits the growth of VCaP prostate cancer line, at concentrations for which it did not affect the growth of the AR negative DU145 and PC3 cells. Interestingly, we also report for the first time that harmol hydrochloride was selective for AR, as it could not alter the activity of other nuclear receptors, such as the glucocorticoid receptor (GR), the progesterone receptor (PR), or the mineralocorticoid receptor (MR). Additionally, we demonstrate that, conversely to enzalutamide, harmol hydrochloride did not show any agonistic activity towards the pregnane X receptor (PXR), a master regulator of drug metabolism. Together, our results shed light on the importance of the cellular context for the screening of new AR antagonists. They further indicate that some of the potential hits that were previously identified may have been overlooked.

Evaluation of a commercial ELISA for detection of epizootic haemorrhagic disease antibodies in domestic and wild ruminant sera.

Bluetongue (BT) and epizootic haemorrhagic disease (EHD) are vector-borne viral diseases affecting domestic and wild ruminants. Both are notifiable under OIE rules. BT and EHD viruses (BTV and EHDV) are closely related Orbiviruses with structural, antigenic and molecular similarities. Both viruses can produce analogous clinical signs in susceptible animals. Serological tests are commonly used for BT and EHD diagnosis and surveillance. Competitive ELISA (c-ELISA) is the most widely used serological test for the specific detection of BTV or EHDV viral protein 7 (VP7) antibodies (Abs). The specificity and sensitivity of the BTV c-ELISA kits available on the market are recognized for the detection of BTV Abs. Concerning EHD, a single commercial EHDV c-ELISA kit (ELISA A kit) commonly used for diagnosis in Europe and Africa was available between 2011 and 2018 but is now no longer on the market. In this study, we evaluated a new commercial c-ELISA to detect ruminant EHDV VP7 Abs in 2,199 serum samples from cattle, sheep, goats, wild deer and zoo animals. The results showed that this ELISA kit is specific and can detect the presence of IgG anti-EHDV VP7 with a very good diagnostic specificity and a satisfactory sensitivity in domestic ruminants, zoo animals and wild deer. Therefore, the evaluated c-ELISA can detect the introduction of EHDV into an area where BTV-seropositive domestic animals are present. The performance of this kit is similar to that of the c-ELISA A kit and can thus be used for diagnosis.

Carboxylate-functionalized foldamer inhibitors of HIV-1 integrase and Topoisomerase 1: artificial analogues of DNA mimic proteins.

Inspired by DNA mimic proteins, we have introduced aromatic foldamers bearing phosphonate groups as synthetic mimics of the charge surface of B-DNA and competitive inhibitors of some therapeutically relevant DNA-binding enzymes: the human DNA Topoisomerase 1 (Top1) and the human HIV-1 integrase (HIV-1 IN). We now report on variants of these anionic foldamers bearing carboxylates instead of phosphonates. Several new monomers have been synthesized with protecting groups suitable for solid phase synthesis (SPS). Six hexadecaamides have been prepared using SPS. Proof of their resemblance to B-DNA was brought by the first crystal structure of one of these DNA-mimic foldamers in its polyanionic form. While some of the foldamers were found to be as active as, or even more active than, the original phosphonate oligomers, others had no activity at all or could even stimulate enzyme activity in vitro. Some foldamers were found to have differential inhibitory effects on the two enzymes. These results demonstrate a strong dependence of inhibitory activity on foldamer structure and charge distribution. They open broad avenues for the development of new classes of derivatives that could inhibit the interaction of specific proteins with their DNA target thereby influencing the cellular pathways in which they are involved.