FHR4-based immunoconjugates direct complement-dependent cytotoxicity and phagocytosis towards HER2-positive cancer cells

Directing selective complement activation towards tumour cells is an attractive strategy to promote their elimination. In the present work, we have generated heteromultimeric immunoconjugates that selectively activate the complement alternative pathway (AP) on tumour cells. We used the C4b-binding protein C-terminal-alpha-/beta-chain scaffold for multimerisation to generate heteromultimeric immunoconjugates displaying (a) a multivalent-positive regulator of the AP, the human factor H-related protein 4 (FHR4) with; (b) a multivalent targeting function directed against erbB2 (HER2); and (c) a monovalent enhanced GFP tracking function. Two distinct VHH targeting two different epitopes against HER2 and competing either with trastuzumab or with pertuzumab-recognising epitopes [VHH(T) or VHH(P)], respectively, were used as HER2 anchoring moieties. Optimised high-FHR4 valence heteromultimeric immunoconjugates [FHR4/VHH(T) or FHR4/VHH(P)] were selected by sequential cell cloning and a selective multistep His-Trap purification. Optimised FHR4-heteromultimeric immunoconjugates successfully overcame FH-mediated complement inhibition threshold, causing increased C3b deposition on SK-OV-3, BT474 and SK-BR3 tumour cells, and increased formation of lytic membrane attack complex densities and complement-dependent cytotoxicity (CDC). CDC varies according to the pattern expression and densities of membrane-anchored complement regulatory proteins on tumour cell surfaces. In addition, opsonised BT474 tumour cells were efficiently phagocytosed by macrophages through complement-dependent cell-mediated cytotoxicity. We showed that the degree of FHR4-multivalency within the multimeric immunoconjugates was the key element to efficiently compete and deregulate FH and FH-mediated convertase decay locally on tumour cell surface. FHR4 can thus represent a novel therapeutic molecule, when expressed as a multimeric entity and associated with an anchoring system, to locally shift the complement steady-state towards activation on tumour cell surface

Nutrients limitation of primary productivity in the Southeast Pacific (BIOSOPE cruise)

Revue sans Comité de lectureInternational audienceIron is an essential nutrient involved in a variety of biological processes in the ocean, including photosynthesis, respiration and nitrogen fixation. Atmospheric deposition of aerosols is recognized as the main source of iron for the surface ocean. In high nutrient, low chlorophyll areas, it is now clearly established that iron limits phytoplankton productivity but its biogeochemical role in low nutrient, low chlorophyll environments has been poorly studied. We investigated this question in the unexplored southeast Pacific, arguably the most oligotrophic area of the global ocean. Situated far from any continental aerosol source, the atmospheric iron flux to this province is amongst the lowest of the world ocean. Here we report that, despite low dissolved iron concentrations (~0.1 nmol l-1) measured across the whole gyre (3 stations situated in the center, the western and the eastern edge), photosynthesis and primary productivity are only limited by iron availability at the border of the gyre, but not in the center. The seasonal stability of the gyre has apparently allowed for the development of populations acclimated to these extreme oligotrophic conditions. Moreover, despite clear evidence of nitrogen limitation in the central gyre, we were unable to measure nitrogen fixation in our experiments, even after iron and/or phosphate additions, and cyanobacterial nifH gene abundances were extremely low compared to the North Pacific Gyre. The South Pacific gyre is therefore unique with respect to the physiological status of its phytoplankton populations

R57K Polymorphism in the Human Immunodeficiency Virus Type 1 Protease as Predictor of Early Virological Failure in a Cohort of Antiretroviral-Naive Patients Treated Mostly with a Nelfinavir-Containing Regimen

In 243 antiretroviral-naive human immunodeficiency-infected patients starting a first-line-protease inhibitor (mainly nelfinavir)-containing therapy, the presence of the polymorphism R57K in the protease at the inception of therapy was independently associated with a higher rate of virological failure

Near Full-Length Characterization and Population Dynamics of the Human Immunodeficiency Virus Type I Circulating Recombinant Form 42 (CRF42_BF) in Luxembourg

A new recombinant form representing a mosaic of HIV-1 subtype B and F1 and designated as CRF42_BF was identified in Luxembourg. We confirmed the inedited nature of CRF42_BF by near full-length genome characterization and retrieved a possible ancestor originating from Brazil. The demographic history of CRF42_BF in Luxembourg using Bayesian coalescent-based methods was investigated. The exponential phase of the logistic growth happened in a very short time period of approximately 5 months associated with a high mean rate of population growth of 15.02 new infections per year. However, CRF42_BF was not characterized by either a higher ex vivo replication capacity in peripheral blood mononuclear cells (PBMCs) or a higher ex vivo transmission efficiency from monocyte-derived dendritic cells to PBMCs as compared to B and F1 viruses. These data do not support a high pathogenic potential of CFR42_BF but rather an initial bursting spread of the recombinant probably due to a more favorable transmission route

Factors Associated with Virological Response to Etravirine in Nonnucleoside Reverse Transcriptase Inhibitor-Experienced HIV-1-Infected Patients▿

To identify factors associated with virological response (VR) to an etravirine (ETR)-based regimen, 243 patients previously treated with nonnucleoside reverse transcriptase inhibitors (NNRTIs) were studied. The impact of baseline HIV-1 RNA, CD4 cell count, past NNRTIs used, 57 NNRTI resistance mutations, genotypic sensitivity score (GSS) for nucleoside reverse transcriptase inhibitors (NRTIs) and protease inhibitors (PIs), and the number of new drugs used with ETR for the first time on the VR to an ETR regimen were investigated. Among the 243 patients, the median baseline HIV-1 RNA level was 4.4 log10 copies/ml (interquartile range [IQR], 3.7 to 4.9) and the median CD4 count was 175 cells/mm3 (IQR, 69 to 312). Patients had been previously exposed to a median of 6 NRTIs, 1, NNRTI, and 5 PIs. Overall, 82% of patients achieved a VR at month 2, as defined by a decrease of at least 1.5 log10 copies/ml and/or HIV-1 RNA level of <50 copies/ml. No difference in VR was observed between patients receiving or not a boosted PI in combination with ETR. Factors independently associated with a better VR to ETR were the number of drugs (among enfuvirtide, darunavir, or raltegravir) used for the first time in combination with ETR and the presence of the K103N mutation at baseline. Mutations Y181V and E138A were independently associated with poor VR, whereas no effect of the Y181C on VR was observed. In conclusion, ETR was associated with high response rates in NNRTI-experienced patients in combination with other active drugs regardless of the therapeutic class used

Vertical structure of small eukaryotes in three lakes that differ by their trophic status : a quantitative approach

In lakes, the diversity of eukaryotic picoplankton has been recently studied by the analysis of 18S ribosomal RNA gene sequences; however, quantitative data are rare. In this study, the vertical structure and abundance of the small eukaryotic size fraction (0.2-5 mu m) were investigated in three lakes by tyramide signal amplification-fluorescent in situ hybridization targeting six phylogenetic groups: Chlorophyta, Haptophyta, Cercozoa, LKM11, Perkinsozoa and fungi. The groups targeted in this study are found in all lakes; however, both the abundance and structure of small eukaryotes are dependent on the system's productivity and depth. These data highlighted the presence of Chlorophyta contributing on an average to 19.3%, 14.7% and 41.2% of total small eukaryotes in lakes Bourget, Aydat and Pavin, respectively. This study also revealed the unexpected importance of Haptophyta, reaching 62.8% of eukaryotes in the euphotic zone of Lake Bourget. The high proportions of these pigmented cells highlight the underestimation of these groups by PCR-based methods. The presence of pigmented Chlorophyta in the deepest zones of the lakes suggests a mixotrophic behaviour of these taxa. We also confirmed the presence of putative parasites such as Perkinsozoa (5.1% of small eukaryotes in Lake Pavin and Bourget) and, with lower abundances, fungi (targeted by the MY1574 probe). Cells targeted by LKM11 probes represented the second group of abundance within heterotrophs. Open questions regarding the functional roles of the targeted groups arise from this study, especially regarding parasitism and mixotrophy, which are interactions poorly taken into account in planktonic food web models. The ISME Journal (2010) 4, 1509-1519; doi:10.1038/ismej.2010.83; published online 24 June 201