How to Better Understand the Influence of Host Genetics on Developing an Effective Immune Response to Thoracic Cancers.

Thoracic cancers pose a significant global health burden. Immune checkpoint blockade therapies have improved treatment outcomes, but durable responses remain limited. Understanding how the host immune system interacts with a developing tumor is essential for the rational development of improved treatments for thoracic malignancies. Recent technical advances have improved our understanding of the mutational burden of cancer cells and changes in cancer-specific gene expression, providing a detailed understanding of the complex biology underpinning tumor-host interactions. While there has been much focus on the genetic alterations associated with cancer cells and how they may impact treatment outcomes, how host genetics affects cancer development is also critical and will greatly determine treatment response. Genome-wide association studies (GWAS) have identified genetic variants associated with cancer predisposition. This approach has successfully identified host genetic risk factors associated with common thoracic cancers like lung cancer, but is less effective for rare cancers like malignant mesothelioma. To assess how host genetics impacts rare thoracic cancers, we used the Collaborative Cross (CC); a powerful murine genetic resource designed to maximize genetic diversity and rapidly identify genes associated with any biological trait. We are using the CC in conjunction with our asbestos-induced MexTAg mouse model, to identify host genes associated with mesothelioma development. Once genes that moderate tumor development and progression are known, human homologues can be identified and human datasets interrogated to validate their association with disease outcome. Furthermore, our CC-MexTAg animal model enables in-depth study of the tumor microenvironment, allowing the correlation of immune cell infiltration and gene expression signatures with disease development. This strategy provides a detailed picture of the underlying biological pathways associated with mesothelioma susceptibility and progression; knowledge that is crucial for the rational development of new diagnostic and therapeutic strategies. Here we discuss the influence of host genetics on developing an effective immune response to thoracic cancers. We highlight current knowledge gaps, and with a focus on mesothelioma, describe the development and application of the CC-MexTAg to overcome limitations and illustrate how the knowledge gained from this unique study will inform the rational design of future treatments of mesothelioma.

Voluntary exercise in mesothelioma: effects on tumour growth and treatment response in a murine model.

OBJECTIVE: There is substantial evidence that exercise can safely reduce the risk of cancer and improve survival in different human cancer populations. Long latency periods associated with carcinogen-induced cancers like asbestos induced mesothelioma provide an opportunity to implement exercise as an intervention to delay or prevent disease development. However, there are limited studies investigating the ability of exercise to prevent or delay cancer, and exercise as a preventive strategy has never been assessed in models with a known carcinogen. We investigated the potential of voluntary exercise (VE) to delay development of asbestos related disease (ARD) in our well-characterised, asbestos induced MexTAg model of mesothelioma. RESULTS: Asbestos exposed MexTAg mice were given continuous or delayed access to VE and ARD assessed over time. We found that the addition of VE did not affect ARD development in asbestos exposed MexTAg mice. However, non-asbestos exposed, aged matched control mice participated in significantly more VE behaviours, suggesting subclinical development of ARD after asbestos exposure had a greater impact on VE participation than age alone. These data highlight the importance of model choice and the potential limitation that some pre-clinical studies may not accurately represent the clinical paradigm, particularly in the context of prevention studies.

A multiplex DNA suspension microarray for simultaneous detection and differentiation of classical swine fever virus and other pestiviruses.

An oligonucleotide suspension microarray (Luminex microsphere system) was developed for detection and differentiation of animal pestiviruses: classical swine fever virus (CSFV), bovine viral diarrhea virus types 1 and 2 (BVDV1 and BVDV2), and border disease virus (BDV). Species-specific and pestivirus-common oligonucleotide probes were designed to the 5' UTR region and conjugated to individual color-coded Luminex carboxy beads (probe beads). Target pestivirus sequences were amplified by asymmetric PCR using a biotinylated reverse primer and a forward and reverse primer ratio of 1:5. The biotinylated products were hybridized to eight probe beads in a multiplex assay and analyzed using streptavidin conjugated to a fluorescent reporter molecule. The assay was able to detect and differentiate all 40 strains of CSFV, BVDV1, BVDV2 and BDV tested. The analytical sensitivity was determined to be 0.2-10 TCID50/ml. The major advantages of the DNA-microsphere suspension microarray, as a low density array, are its ease of handling and ability to simultaneously detect and type multiple infectious agents.

Phylogeny and antigenic relationships of three cervid herpesviruses.

Elk herpesvirus (ElkHV) from North American elk (wapiti, Cervus elaphus nelsoni) is a recently identified alphaherpesvirus related to bovine herpesvirus-1 (BHV-1). In this study, we determined its relationship with European cervid herpesviruses: cervid herpesvirus-1 (CerHV-1) from red deer and rangiferine herpesvirus (RanHV) from reindeer. For phylogenetic analysis, genes for the gC and gD proteins of these viruses were sequenced. These genes demonstrated an extremely high GC content (76-79%). Genetically, ElkHV was found to be closely related to CerHV-1 and both viruses are more closely related to BHV-1 than to RanHV. Antigenically, the same relationships were found. ElkHV shares common neutralizing epitopes with both CerHV-1 and RanHV. A total of 10 epitopes were defined on the gB, gC and gD proteins of these viruses, including a shared neutralizing epitope on gD. The results indicate that ElkHV and CerHV-1 have diverged from a common ancestor virus. Cervid herpesviruses may be useful in determination of evolutionary rates of change for alphaherpesvirus genes.

Mapping of two antigenic domains on the NS3 protein of the pestivirus bovine viral diarrhea virus.

The immunodominant NS3 (p80) protein of the pestivirus bovine viral diarrhea virus (BVDV) functions as a serine protease and a RNA helicase. To identify antigenic domains of the BVDV NS3, a panel of monoclonal antibodies (mAbs) was tested against fragments of the protein expressed in E. coli. Two large overlapping NS3 fragments, A (amino acids [aa] 1-434) and B (aa 368-683) which together contain all NS3 sequences, were used to screen mAbs for reactivity. Two mAbs, 21.5.8 and 1.11.3, were reactive to fragment A (in ELISA only) and one mAb, 20.10.6, was reactive to fragment B (in ELISA and Western blotting). Further mapping demonstrated that the smallest fragment mAbs 21.5.8 and 1.11.3 bound to was comprised of aa 205-369 (domain A). In Western blotting, the smallest fragment reactive with mAb 20.10.6 was comprised of aa 368-549 (domain B). However, in indirect ELISA, mAb 20.10.6 also demonstrated high reactivity to a smaller fragment comprising aa 368-512 (domain B'). This indicated that the epitope of mAb 20.10.6 was conformational and not linear. Blocking ELISAs using these mAbs and type 1 and type 2 BVDV antisera demonstrated that an immunodominant region of the NS3 protein in cattle is defined by aa 205-549.