Myeloid-derived suppressor cells are associated with disease progression and decreased overall survival in advanced-stage melanoma patients

Myeloid-derived suppressor cells are increased in the peripheral blood of advanced-stage cancer patients; however, no studies have shown a correlation of these immunosuppressive cells with clinical outcomes in melanoma patients. We characterized the frequency and suppressive function of multiple subsets of myeloid-derived suppressor cells in the peripheral blood of 34 patients with Stage IV melanoma, 20 patients with Stage I melanoma, and 15 healthy donors. The frequency of CD14⁺ MDSCs (Lin^? CD11b⁺ HLA-DR^? CD14⁺ CD33⁺) and CD14^? MDSCs (Lin^? CD11b⁺ HLA-DR^? CD14^? CD33⁺) was increased in the peripheral blood of Stage IV melanoma patients relative to healthy donors. The frequency of CD14⁺ and CD14^? MDSCs correlated with each other and with the increased frequency of regulatory T cells, but not with classically defined monocytes. CD14^? MDSCs isolated from the peripheral blood of Stage IV melanoma patients suppressed T cell activation more than those isolated from healthy donors, and the frequency of these cells correlated with disease progression and decreased overall survival. Our study provides the first evidence that the frequency of CD14^? MDSCs negatively correlates with clinical outcomes in advanced-stage melanoma patients. These data indicate that suppressive MDSCs should be considered as targets for future immunotherapies.     

Interleukin-15 rescues tolerant CD8+ T cells for use in adoptive immunotherapy of established tumors

CD8+ T cells can mediate eradication of established tumors, and strategies to amplify tumor-reactive T-cell numbers by immunization or ex vivo expansion followed by adoptive transfer are currently being explored in individuals with cancer. Generating effective CD8+ T cell-mediated responses to tumors is often impeded by T-cell tolerance to relevant tumor antigens, as most of these antigens are also expressed in normal tissues. We examined whether such tolerant T cells could be rescued and functionally restored for use in therapy of established tumors. We used a transgenic T-cell receptor (TCR) mouse model in which peripheral CD8+ T cells specific for a candidate tumor antigen also expressed in liver are tolerant, failing to proliferate or secrete interleukin (IL)-2 in response to antigen. Molecular and cellular analysis showed that these tolerant T cells expressed the IL-15 receptor alpha chain, and could be induced to proliferate in vitro in response to exogenous IL-15. Such proliferation abrogated tolerance and the rescued cells became effective in treating leukemia. Therefore, high-affinity CD8+ T cells are not necessarily deleted by encounter with self-antigen in the periphery, and can potentially be rescued and expanded for use in tumor immunotherapy.     

THE IMPORTANCE OF MOSQUITO BEHAVIOURAL ADAPTATIONS TO MALARIA CONTROL IN AFRICA

Over the past decade the use of long‐lasting insecticidal nets (LLINs), in combination with improved drug therapies, indoor residual spraying (IRS), and better health infrastructure, has helped reduce malaria in many African countries for the first time in a generation. However, insecticide resistance in the vector is an evolving threat to these gains. We review emerging and historical data on behavioral resistance in response to LLINs and IRS. Overall the current literature suggests behavioral and species changes may be emerging, but the data are sparse and, at times unconvincing. However, preliminary modeling has demonstrated that behavioral resistance could have significant impacts on the effectiveness of malaria control. We propose seven recommendations to improve understanding of resistance in malaria vectors. Determining the public health impact of physiological and behavioral insecticide resistance is an urgent priority if we are to maintain the significant gains made in reducing malaria morbidity and mortality.     

Role of PCIF1‐mediated 5′‐cap N6‐methyladeonsine mRNA methylation in colorectal cancer and anti‐PD‐1 immunotherapy

Adenosine N6‐methylation (m6A) and N6,2′‐O‐dimethylation (m6Am) are regulatory modifications of eukaryotic mRNAs. m6Am formation is catalyzed by the methyl transferase phosphorylated CTD‐interacting factor 1 (PCIF1); however, the pathophysiological functions of this RNA modification and PCIF1 in cancers are unclear. Here, we show that PCIF1 expression is upregulated in colorectal cancer (CRC) and negatively correlates with patient survival. CRISPR/Cas9‐mediated depletion of PCIF1 in human CRC cells leads to loss of cell migration, invasion, and colony formation in vitro and loss of tumor growth in athymic mice. Pcif1 knockout in murine CRC cells inhibits tumor growth in immunocompetent mice and enhances the effects of anti‐PD‐1 antibody treatment by decreasing intratumoral TGF‐β levels and increasing intratumoral IFN‐γ, TNF‐α levels, and tumor‐infiltrating natural killer cells. We further show that PCIF1 modulates CRC growth and response to anti‐PD‐1 in a context‐dependent mechanism with PCIF1 directly targeting FOS, IFITM3, and STAT1 via m6Am modifications. PCIF1 stabilizes FOS mRNA, which in turn leads to FOS‐dependent TGF‐β regulation and tumor growth. While during immunotherapy, Pcif1‐Fos‐TGF‐β, as well as Pcif1‐Stat1/Ifitm3‐IFN‐γ axes, contributes to the resistance of anti‐PD‐1 therapy. Collectively, our findings reveal a role of PCIF1 in promoting CRC tumorigenesis and resistance to anti‐PD‐1 therapy, supporting that the combination of PCIF1 inhibition with anti‐PD‐1 treatment is a potential therapeutic strategy to enhance CRC response to immunotherapy. Finally, we developed a lipid nanoparticles (LNPs) and chemically modified small interfering RNAs (CMsiRNAs)‐based strategy to silence PCIF1 in vivo and found that this treatment significantly reduced tumor growth in mice. Our results therefore provide a proof‐of‐concept for tumor growth suppression using LNP‐CMsiRNA to silence target genes in cancer.     

Intron-derived small RNAs for silencing viral RNAs in mosquito cells

Aedes aegypti and Ae. albopictus are the main vectors of mosquito-borne viruses of medical and veterinary significance. Many of these viruses have RNA genomes. Exogenously provided, e.g. transgene encoded, small RNAs could be used to inhibit virus replication, breaking the transmission cycle. We tested, in Ae. aegypti and Ae. albopictus cell lines, reporter-based strategies for assessing the ability of two types of small RNAs to inhibit a chikungunya virus (CHIKV) derived target. Both types of small RNAs use a Drosophila melanogaster pre-miRNA-1 based hairpin for their expression, either with perfect base-pairing in the stem region (shRNA-like) or containing two mismatches (miRNA-like). The pre-miRNA-1 stem loop structure was encoded within an intron; this allows co-expression of one or more proteins, e.g. a fluorescent protein marker tracking the temporal and spatial expression of the small RNAs in vivo. Three reporter-based systems were used to assess the relative silencing efficiency of ten shRNA-like siRNAs and corresponding miRNA-like designs. Two systems used a luciferase reporter RNA with CHIKV RNA inserted either in the coding sequence or within the 3’ UTR. A third reporter used a CHIKV derived split replication system. All three reporters demonstrated that while silencing could be achieved with both miRNA-like and shRNA-like designs, the latter were substantially more effective. Dcr-2 was required for the shRNA-like siRNAs as demonstrated by loss of inhibition of the reporters in Dcr-2 deficient cell lines. These positive results in cell culture are encouraging for the potential use of this pre-miRNA-1-based system in transgenic mosquitoes.     

The DNA damage response pathway regulates the alternative splicing of the apoptotic mediator Bcl-x

Alternative splicing often produces effectors with opposite functions in apoptosis. Splicing decisions must therefore be tightly connected to stresses, stimuli, and pathways that control cell survival and cell growth. We have shown previously that PKC signaling prevents the production of proapoptotic Bcl-x(S) to favor the accumulation of the larger antiapoptotic Bcl-x(L) splice variant in 293 cells. Here we show that the genotoxic stress induced by oxaliplatin elicits an ATM-, CHK2-, and p53-dependent splicing switch that favors the production of the proapoptotic Bcl-x(S) variant. This DNA damage-induced splicing shift requires the activity of protein-tyrosine phosphatases. Interestingly, the ATM/CHK2/p53/tyrosine phosphatases pathway activated by oxaliplatin regulates Bcl-x splicing through the same regulatory sequence element (SB1) that receives signals from the PKC pathway. Convergence of the PKC and DNA damage signaling routes may control the abundance of a key splicing repressor because SB1-mediated repression is lost when protein synthesis is impaired but is rescued by blocking proteasome-mediated protein degradation. The SB1 splicing regulatory module therefore receives antagonistic signals from the PKC and the p53-dependent DNA damage response pathways to control the balance of pro- and antiapoptotic Bcl-x splice variants.     

Adolescent Sexual and Reproductive Health Services and Implications for the Provision of Voluntary Medical Male Circumcision: Results of a Systematic Literature Review

Background

Voluntary medical male circumcision (VMMC) is a critical HIV prevention tool. Since 2007, sub-Saharan African countries with the highest prevalence of HIV have been mobilizing resources to make VMMC available. While implementers initially targeted adult men, demand has been highest for boys under age 18. It is important to understand how male adolescents can best be served by quality VMMC services.

Methods and Findings

A systematic literature review was performed to synthesize the evidence on best practices in adolescent health service delivery specific to males in sub-Saharan Africa. PubMed, Scopus, and JSTOR databases were searched for literature published between January 1990 and March 2014. The review revealed a general absence of health services addressing the specific needs of male adolescents, resulting in knowledge gaps that could diminish the benefits of VMMC programming for this population. Articles focused specifically on VMMC contained little information on the adolescent subgroup. The review revealed barriers to and gaps in sexual and reproductive health and VMMC service provision to adolescents, including structural factors, imposed feelings of shame, endorsement of traditional gender roles, negative interactions with providers, violations of privacy, fear of pain associated with the VMMC procedure, and a desire for elements of traditional non-medical circumcision methods to be integrated into medical procedures. Factors linked to effective adolescent-focused services included the engagement of parents and the community, an adolescent-friendly service environment, and VMMC counseling messages sufficiently understood by young males.

Conclusions

VMMC presents an opportune time for early involvement of male adolescents in HIV prevention and sexual and reproductive health programming. However, more research is needed to determine how to align VMMC services with the unique needs of this population.