Clinical experience with integrase inhibitors in HIV-2-infected individuals in Spain.

BACKGROUND: HIV-2 is a neglected virus despite estimates of 1-2 million people being infected worldwide. The virus is naturally resistant to some antiretrovirals used to treat HIV-1 and therapeutic options are limited for patients with HIV-2. METHODS: In this retrospective observational study, we analysed all HIV-2-infected individuals treated with integrase strand transfer inhibitors (INSTIs) recorded in the Spanish HIV-2 cohort. Demographics, treatment modalities, laboratory values, quantitative HIV-2 RNA and CD4 counts as well as drug resistance were analysed. RESULTS: From a total of 354 HIV-2-infected patients recruited by the Spanish HIV-2 cohort as of December 2017, INSTIs had been given to 44, in 18 as first-line therapy and in 26 after failing other antiretroviral regimens. After a median follow-up of 13 months of INSTI-based therapy, undetectable viraemia for HIV-2 was achieved in 89% of treatment-naive and in 65.4% of treatment-experienced patients. In parallel, CD4 gains were 82 and 126 cells/mm3, respectively. Treatment failure occurred in 15 patients, 2 being treatment-naive and 13 treatment-experienced. INSTI resistance changes were recognized in 12 patients: N155H (5), Q148H/R (3), Y143C/G (3) and R263K (1). CONCLUSIONS: Combinations based on INSTIs are effective and safe treatment options for HIV-2-infected individuals. However, resistance mutations to INSTIs are selected frequently in failing patients, reducing the already limited treatment options.

Overlapping palindromic sequences associated with somatic deletion and meiotic recombination of MHC class I genes.

H-2L-null variants were immunoselected from a transfected murine fibroblast cell line carrying a single copy H-2L gene, and were characterized to determine the basis for the loss of this MHC class I cell surface product. Molecular analysis indicated that inactivation of H-2L expression in nearly every null clone resulted from an apparent deletion or rearrangement of 5'-flanking and 5'-coding H-2L sequences, with breakpoints consistently mapping to within a 550 bp GC-rich region between exon 1 and the middle of intron 2. Notably, this region of the H-2L gene contains a large number of overlapping, inverted repeat sequences as well as potential topoisomerase I cleavage sites. Examination of several in vivo mutant class I genes, believed to have been generated by recombination, has revealed that each of these genes bears similar palindromic structures overlapping or adjacent to the regions of sequence exchange. These findings suggest that inverted repeat sequences may play a role in recombination and deletion within the MHC class I multigene family.

Sentinel surveillance of invasive candidiasis in Spain: epidemiology and antifungal susceptibility.

In order to know the epidemiology and the changes of antifungal resistance in invasive candidiasis (IC) we carried out this prospective study of Candida strains belonging to patients admitted to 26 Spanish hospitals from June 2011 to June 2012 diagnosed with IC. Clinical information and the identity of the Candida species were collected and antifungal susceptibility was tested using broth microdilution in five agents: amphotericin B, fluconazole, voriconazole, caspofungin and anidulafungin. A total of 705 cases-isolates were documented. Most of the patients suffered from candidemia and several underlying diseases and more than half of them were neutropenic or under immunosuppressive therapy, factors associated with higher mortality. Thirty percent of global mortality was documented. C. albicans was the most frequently isolated species, although an increase of non-C. albicans species was observed. Resistance to fluconazole was of 3.4%, lower than in previous years (6.3%). C. parapsilosis presented a higher MIC90 of echinocandins compared to other species.

A demonstration of human beta-2-microglobulin specific association with the surface of teleostean cells.

Purified human Beta-2 microglobulin (beta 2m) and a human beta 2m fluorochrome conjugate were used in exchange reactions to demonstrate that beta 2m associates with a teleostean cell surface protein. beta 2m exchange among brown bullhead, channel catfish, fathead minnow, and rainbow trout cells lines was detected by using either radioimmunoassay or flow cytometry. Evidence that beta 2m binds specifically with the surface of teleostean cells and possibly associates with an expressed class I MHC homologue is provided. Moreover, following exchange on brown bullhead cells, a coprecipitated protein of 45 kDa was observed following subsequent immunoprecipitation with the human beta 2m specific antibody B1.1G6. Given that beta 2m is a peripheral protein which has been shown to exchange with MHC expressing cells from different species, co-precipitation results suggest that the 45 kDa protein may represent a class I MHC homologue.

A point mutation in beta2-microglobulin results in loss of epitope expression.

A common tool in studying the structure and function of major histocompatibility complex: (MHC) class I is the generation and analysis of beta2-microglobulin (beta2m) mutations. beta2m has been shown to affect proper class I antigen presentation at the level of structural functionality. Many studies characterizing beta2m function in class I presentation have used antibody-based assays. Monitoring the effect of beta2m mutation on antibody epitope expression, therefore, is essential in being able to truly characterize the impact of a mutant interaction between beta2m and class I. Here we describe a mutant beta2m molecule, beta2m #32, that in association with class I loses reactivity with the human beta2m-specific monoclonal antibody, BBM.1. However, the BBM.1 epitope remains intact when beta2m #32 is free from class I association.

The association of H-2Ld with human beta-2 microglobulin induces localized conformational changes in the alpha-1 and -2 superdomain.

We have analyzed changes in the antigenicity of major histocompatibility complex class I molecules resulting from the association of human beta-2 micro-globulin (B2m) with the mouse class I heavy chain. In particular, the H-2Ld molecule exhibited enhanced cross-reactivity for the 34-1-2 monoclonal antibody. In order to assess the nature of this structural alteration induced by human B2m, we utilized H-2 class I hybrid molecules in the mapping of the 34-1-2 determinant to the helical region of the alpha-1 domain. H-2Ld class I hybrid molecules were then used to establish the importance of the alpha-2 and -3 domains in the observed increase of 34-1-2 cross-reactivity following exchange with human B2m. The H-2Ld hybrids suggest that alterations in interdomain contact are responsible for enhanced 34-1-2 cross-reactivity on the H-2Ld molecule. It is likely that this alteration arises through changes in class I conformation at regions of the molecule distant from points of contact between B2m and the class I molecule. This suggests that perturbations induced by association of human B2m with H-2Ld can affect the conformation of the alpha-1 and -2 superdomain. That class I antigenic determinants are altered by the association of human B2m with mouse class I further suggests that the class I molecule is structurally flexible and may reflect the ability of the class I molecule to bind and present a vast array of disparate peptides to the T-cell receptor.

Alteration within a discrete region of the H-2Ld alpha 1 helix upon association with human beta 2 microglobulin.

The utilization of the beta 2-microglobulin (B2m) exchange assay allowed for the association of H-2Ld with human B2m. Upon association with H-2Ld, human B2m induces structural alterations in H-2Ld that appear dependent upon xenogeneic B2m amino acid sequence variability. In this regard, xenogeneic B2m exchange is used as a tool to induce structural alterations in class I as a means of analysing the structural-functional relationship of B2m/class I association. Incorporating H-2Ld site-directed mutants into the experimental approach provided strong evidence that B2m makes indirect contact with discrete class I specific amino acid positions located in the helical region of the alpha 1 domain.

Effects of beta2-microglobulin mutations on the alpha-1 helical region of H2-Ld.

Beta-2 microglobulin (beta2m)has been shown to have an effect on the structural and functional constraints that facilitate proper class I antigen presentation. To date, no evidence has pinpointed the beta2m-specific amino acids that play an integral role in affecting structure in and around the peptide binding region of class I. To delineate beta2m amino acid positions that affect the alpha-1 helical region, we generated a series of mutant beta2m proteins bearing precise amino acid substitutions. The amino acid positions chosen were based upon previous results which demonstrated that human beta2m association with H2-Ld altered the structure of the alpha-1/alpha-2 super-domain. beta2m mutant proteins were used in beta2m exchange assays with cells expressing H2-Ld. Following exchange, cells were assayed to determine whether mutant beta2m association resulted in structural alteration of class I extracellular domains. The alteration in H2-Ld structure was evidenced by an increase in the binding of an antibody (34-1-2), specific for the alpha-1 helical region of H2-Ld. Results demonstrated that amino acid substitutions in beta2m positions 33 and 53 led to a dramatic increase in the reactivity of the alpha-1 domain-specific antibody 34-1-2. Identifying beta2m amino acid positions that influence the structure of the peptide binding region may allow for a better understanding of cellular immune responses that center upon class I/beta2m expression.