CXCL7 is a predictive marker of sunitinib efficacy in clear cell renal cell carcinomas.

BACKGROUND: Sunitinib is one of the first-line standard treatments for metastatic clear cell renal cell carcinoma (ccRCC) with a median time to progression shorter than 1 year. The objective is to discover predictive markers of response to adapt the treatment at diagnosis. METHODS: Prospective phase 2 multi-centre trials were conducted in ccRCC patients initiating sunitinib (54 patients) or bevacizumab (45 patients) in the first-line metastatic setting (SUVEGIL and TORAVA trials). The plasmatic level of CXCL7 at baseline was correlated with progression-free survival (PFS). RESULTS: The cut-off value of CXCL7 for PFS was 250 ng ml-1. Patients with CXCL7 plasmatic levels above the cut-off at baseline (250 ng ml-1) had a significantly longer PFS (hazard ratio 0.323 (95% confidence interval 0.147-0.707), P=0.001). These results were confirmed in a retrospective validation cohort. The levels of CXCL7 did not influence PFS of the bevacizumab-treated patients. CONCLUSIONS: CXCL7 may be considered as a predictive marker of sunitinib efficacy for ccRCC patients.

The N-terminal domain of the arenavirus L protein is an RNA endonuclease essential in mRNA transcription.

Arenaviridae synthesize viral mRNAs using short capped primers presumably acquired from cellular transcripts by a 'cap-snatching' mechanism. Here, we report the crystal structure and functional characterization of the N-terminal 196 residues (NL1) of the L protein from the prototypic arenavirus: lymphocytic choriomeningitis virus. The NL1 domain is able to bind and cleave RNA. The 2.13 Å resolution crystal structure of NL1 reveals a type II endonuclease α/ß architecture similar to the N-terminal end of the influenza virus PA protein. Superimposition of both structures, mutagenesis and reverse genetics studies reveal a unique spatial arrangement of key active site residues related to the PD…(D/E)XK type II endonuclease signature sequence. We show that this endonuclease domain is conserved and active across the virus families Arenaviridae, Bunyaviridae and Orthomyxoviridae and propose that the arenavirus NL1 domain is the Arenaviridae cap-snatching endonuclease.

Gln151 of HIV-1 reverse transcriptase acts as a steric gate towards clinically relevant acyclic phosphonate nucleotide analogues.

BACKGROUND: In the treatment of HIV, the loose active site of the HIV-1 reverse transcriptase (RT) allows numerous nucleotide analogues to act as proviral DNA 'chain-terminators'. Acyclic nucleotide phosphonate analogues (ANPs) represent a particular class of nucleotide analogue that does not possess a ribose moiety. The structural basis for their substrate efficiency regarding viral DNA polymerases is poorly understood. METHODS: Pre-steady-state kinetics on HIV-1 RT together with molecular modelling, were used to evaluate the relative characteristics of both the initial binding and incorporation into DNA of three different ANP diphosphates with progressively increasing steric demands on the acyclic linker: adefovir-diphosphate (DP), tenofovir-DP, and cidofovir-DP. RESULTS: The increase of steric demand in ANPs induced a proportional loss of the binding affinity to wild-type HIV-1 RT (Kd cidofovir-DP>>Kd tenofovir-DP>Kd adefovir-DP approximately Kd dNTPs), consistent with the lack of HIV-1 inhibitory activity for cidofovir. We show that, starting from adefovir-DP, the steric constraints mainly map to Gln151, as its mutation to alanine provides cidofovir-DP sensitivity. Interactions between the Gln151 residue and the methyl group of tenofovir-DP further increase with the mutation Gln151Met, resulting in a specific discrimination and low-level resistance to tenofovir-DP. This alteration is the result of a dual decrease in the binding affinity (Kd) and the catalytic rate (k(pol)) of incorporation of tenofovir-DP. By contrast, the tenofovir resistance mutation K65R induces a broad 'k(pol)-dependent' nonspecific discrimination towards the three ANPs. CONCLUSIONS: Overall, our results show that the efficiency of ANPs to compete against natural nucleotides as substrates for RT is determined by their close interaction with specific amino acids such as Gln151 within the RT active site. These results should help us to map and predict ANP sensitivity determinants in cellular and viral DNA polymerase active sites for which the understanding of different ANP sensitivity patterns are of medical importance.

In vitro suppression of K65R reverse transcriptase-mediated tenofovir- and adefovir-5'-diphosphate resistance conferred by the boranophosphonate derivatives.

9-[2-(Boranophosphonomethoxy)ethyl]adenine diphosphate (BH(3)-PMEApp) and (R)-9-[2-(boranophosphonomethoxy)propyl]adenine diphosphate (BH(3)-PMPApp), described here, represent the first nucleoside phosphonates modified on their alpha-phosphates that act as efficient substrates for the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT). These analogues were synthesized and evaluated for their in vitro activity against wild-type (WT), K65R, and R72A RTs. BH(3)-PMEApp and BH(3)-PMPApp exhibit the same inhibition properties as their nonborane analogues on WT RT. However, K65R RT was found hypersensitive to BH(3)-PMEApp and as sensitive as WT RT to BH(3)-PMPApp. Moreover, the presence of the borane group restores incorporation of the analogue by R72A HIV RT, the latter being nearly inactive with regular nucleotides. The BH(3)-mediated suppression of HIV-1 RT resistance, formerly described with nucleoside 5'-(alpha-p-borano)-triphosphate analogues, is thus also conserved at the phosphonate level. The present results show that an alpha-phosphate modification is also possible and interesting for phosphonate analogues, a result that might find application in the search for a means to control HIV RT-mediated drug resistance.

Structural and functional basis for ADP-ribose and poly(ADP-ribose) binding by viral macro domains.

Macro domains constitute a protein module family found associated with specific histones and proteins involved in chromatin metabolism. In addition, a small number of animal RNA viruses, such as corona- and toroviruses, alphaviruses, and hepatitis E virus, encode macro domains for which, however, structural and functional information is extremely limited. Here, we characterized the macro domains from hepatitis E virus, Semliki Forest virus, and severe acute respiratory syndrome coronavirus (SARS-CoV). The crystal structure of the SARS-CoV macro domain was determined at 1.8-Angstroms resolution in complex with ADP-ribose. Information derived from structural, mutational, and sequence analyses suggests a close phylogenetic and, most probably, functional relationship between viral and cellular macro domain homologs. The data revealed that viral macro domains have relatively poor ADP-ribose 1"-phosphohydrolase activities (which were previously proposed to be their biologically relevant function) but bind efficiently free and poly(ADP-ribose) polymerase 1-bound poly(ADP-ribose) in vitro. Collectively, these results suggest to further evaluate the role of viral macro domains in host response to viral infection.