Total transcriptome, proteome, and allergome of Johnson grass pollen, which is important for allergic rhinitis in subtropical regions.

BACKGROUND: Genomic data are lacking for many allergen sources. To circumvent this limitation, we implemented a strategy to reveal the repertoire of pollen allergens of a grass with clinical importance in subtropical regions, where an increasing proportion of the world's population resides. OBJECTIVE: We sought to identify and immunologically characterize the allergenic components of the Panicoideae Johnson grass pollen (JGP; Sorghum halepense). METHODS: The total pollen transcriptome, proteome, and allergome of JGP were documented. Serum IgE reactivities with pollen and purified allergens were assessed in 64 patients with grass pollen allergy from a subtropical region. RESULTS: Purified Sor h 1 and Sor h 13 were identified as clinically important allergen components of JGP with serum IgE reactivity in 49 (76%) and 28 (43.8%), respectively, of patients with grass pollen allergy. Within whole JGP, multiple cDNA transcripts and peptide spectra belonging to grass pollen allergen families 1, 2, 4, 7, 11, 12, 13, and 25 were identified. Pollen allergens restricted to subtropical grasses (groups 22-24) were also present within the JGP transcriptome and proteome. Mass spectrometry confirmed the IgE-reactive components of JGP included isoforms of Sor h 1, Sor h 2, Sor h 13, and Sor h 23. CONCLUSION: Our integrated molecular approach revealed qualitative differences between the allergenic components of JGP and temperate grass pollens. Knowledge of these newly identified allergens has the potential to improve specific diagnosis and allergen immunotherapy treatment for patients with grass pollen allergy in subtropical regions and reduce the burden of allergic respiratory disease globally.

Three agonist antibodies in combination with high-dose IL-2 eradicate orthotopic kidney cancer in mice.

BACKGROUND: Combination immunotherapies can be effective against subcutaneous tumors in mice but the effect against orthotopic malignant disease is less well characterized. In particular, a combination of three agonist antibodies, termed Tri-mAb, consisting of anti-DR5, anti-CD40 and anti-CD137 has previously been demonstrated to eradicate a large proportion of subcutaneous renal cell carcinoma (Renca) tumors (75% long-term survival), but the effect against orthotopic disease is not known. PURPOSE: To determine the relative response of orthotopic tumors, we inoculated Renca into the kidney followed by treatment with Tri-mAb. RESULTS: We found that orthotopic tumors responded much less to treatment (approximately 13% survival), but a significant improvement in survival was achieved through the addition of IL-2 to the treatment regimen (55% survival). All three agonist antibodies and high dose IL-2, 100,000 IU for up to six doses, were required. CD8+ T cells were also required for optimal anti-tumor responses. Coadministration of IL-2 led to enhanced T cell activity as demonstrated by an increased frequency of IFN-gamma-producing T cells in tumor-draining lymph nodes, which may have contributed to the observed improvement of therapy against kidney tumors. IMPLICATIONS: Responses of subcutaneous tumors to immunotherapy do not necessarily reflect how orthotopic tumors respond. The use of combination immunotherapy stimulating multiple facets of immunity and including cytokine support for T cells can induce effective anti-tumor responses against orthotopic and metastatic tumors.

A novel ENU-mutation in ankyrin-1 disrupts malaria parasite maturation in red blood cells of mice.

The blood stage of the plasmodium parasite life cycle is responsible for the clinical symptoms of malaria. Epidemiological studies have identified coincidental malarial endemicity and multiple red blood cell (RBC) disorders. Many RBC disorders result from mutations in genes encoding cytoskeletal proteins and these are associated with increased protection against malarial infections. However the mechanisms underpinning these genetic, host responses remain obscure. We have performed an N-ethyl-N-nitrosourea (ENU) mutagenesis screen and have identified a novel dominant (haploinsufficient) mutation in the Ank-1 gene (Ank1(MRI23420)) of mice displaying hereditary spherocytosis (HS). Female mice, heterozygous for the Ank-1 mutation showed increased survival to infection by Plasmodium chabaudi adami DS with a concomitant 30% decrease in parasitemia compared to wild-type, isogenic mice (wt). A comparative in vivo red cell invasion and parasite growth assay showed a RBC-autonomous effect characterised by decreased proportion of infected heterozygous RBCs. Within approximately 6-8 hours post-invasion, TUNEL staining of intraerythrocytic parasites, showed a significant increase in dead parasites in heterozygotes. This was especially notable at the ring and trophozoite stages in the blood of infected heterozygous mutant mice compared to wt (p<0.05). We conclude that increased malaria resistance due to ankyrin-1 deficiency is caused by the intraerythrocytic death of P. chabaudi parasites.

The Lewis-Y carbohydrate antigen is expressed by many human tumors and can serve as a target for genetically redirected T cells despite the presence of soluble antigen in serum.

In this study we aimed to determine the suitability of the Lewis-Y carbohydrate antigen as a target for immunotherapy using genetically redirected T cells. Using the 3S193 monoclonal antibody and immunohistochemistry, Lewis-Y was found to be expressed on a range of tumors including 42% squamous cell lung carcinoma, 80% lung adenocarcinoma, 25% ovarian carcinoma, and 25% colorectal adenocarcinoma. Expression levels varied from low to intense on between 1% and 90% of tumor cells. Lewis- was also found in soluble form in sera from both normal donors and cancer patients using a newly developed enzyme-linked immunosorbent assay. Serum levels in patients was often less than 1 ng/mL, similar to normal donors, but approximately 30% of patients had soluble Lewis-Y levels exceeding 1 ng/mL and up to 9 ng/mL. Lewis-Y-specific human T cells were generated by genetic modification with a chimeric receptor encoding a single-chain humanized antibody linked to the T-cell signaling molecules, T-cell receptor-zeta, and CD28. T cells responded against the Lewis-Y antigen by cytokine secretion and cytolysis in response to tumor cells. Importantly, the T-cell response was not inhibited by patient serum containing soluble Lewis-Y. This study demonstrates that Lewis-Y is expressed on a large number of tumors and Lewis-Y-specific T cells can retain antitumor function in the presence of patient serum, indicating that this antigen is a suitable target for this form of therapy.