Isotype selection for antibody-based cancer therapy.

The clinical application of monoclonal antibodies (mAbs) has revolutionized the field of cancer therapy, as it has enabled the successful treatment of previously untreatable types of cancer. Different mechanisms play a role in the anti-tumour effect of mAbs. These include blocking of tumour-specific growth factor receptors or of immune modulatory molecules as well as complement and cell-mediated tumour cell lysis. Thus, for many mAbs, Fc-mediated effector functions critically contribute to the efficacy of treatment. As immunoglobulin (Ig) isotypes differ in their ability to bind to Fc receptors on immune cells as well as in their ability to activate complement, they differ in the immune responses they activate. Therefore, the choice of antibody isotype for therapeutic mAbs is dictated by its intended mechanism of action. Considering that clinical efficacy of many mAbs is currently achieved only in subsets of patients, optimal isotype selection and Fc optimization during antibody development may represent an important step towards improved patient outcome. Here, we discuss the current knowledge of the therapeutic effector functions of different isotypes and Fc-engineering strategies to improve mAbs application.

Seasonal differences in cytokine expression in the skin of Shetland ponies suffering from insect bite hypersensitivity.

Insect bite hypersensitivity (IBH) in horses is a seasonal, IgE-mediated, pruritic skin disorder primarily caused by Culicoides spp. We hypothesize that a mixed Th2/Th1-type immune status, off season, alters into Th2-dominated immune reactivity in the skin of IBH-affected ponies in the IBH season. To study these immune response patterns Culicoides-specific IgE levels, skin histopathology and cytokine and transcription factor mRNA expression (IL4, IL10, IL13, IFNγ, FoxP3 and CD3(ζ)) in lesional and non-lesional skin of ponies affected by IBH in the IBH season were compared with those of the same animals off season and those in skin of healthy ponies in both seasons. The present study revealed a significantly higher histopathology score in lesional skin of affected ponies than in non-lesional skin and skin of healthy ponies in the IBH season. Culicoides obsoletus-specific IgE serum levels of ponies with IBH were significantly higher than those in healthy ponies in both seasons. Interestingly, C. obsoletus-specific IgE serum levels within each group were the same in the IBH season and off season. The expression of IL4, IL13 and IFNγ mRNA in skin biopsies in the IBH season showed a significant increase compared to off season in both skin derived from healthy control ponies (n=14) as well as in lesional and in non-lesional skin from IBH-affected animals (n=17). This apparently general up-regulation of cytokine expression during the IBH season directly correlated with an increased CD3(ζ) mRNA expression in the skin, indicating an overall increased T cell influx during the summer months. The only significant difference observed between lesional skin from IBH-affected animals as compared to skin from healthy control animals in the IBH season was a lower expression of IL13/CD3(ζ) in the affected animals. FoxP3 and IL10 levels were unaffected, except for a lower expression of FoxP3 in healthy control skin in the IBH season as compared to off season, In addition, the increased level of C. obsoletus-specific IgE did not correlate with higher histological scores in LE skin. In summary, our data indicate a general immune activation in the skin of both healthy and IBH-affected ponies during the IBH season that potentially obscures the Culicoides-specific immune reaction pattern, even in lesional skin of IBH-affected animals.

Considerations in the design of vaccines that induce CD8 T cell mediated immunity.

The protective capacity of many currently used vaccines is based on induction of neutralizing antibodies. Many pathogens, however, have adapted themselves in different ways to escape antibody-based immune protection. In particular, for those infections against which conventional neutralizing antibody-based vaccinations appear challenging, CD8 T-cells are considered to be promising candidates for vaccine targeting. The design of vaccines that induce robust and long-lasting protective CD8 T-cell responses however imposes new challenges, as many factors such as kinetics and efficiency of antigen-processing and presentation by antigen presenting cells, T-cell repertoire and cytokine environment during T cell priming contribute to the specificity and functionality of CD8 T-cell responses. In the following, we review the most prominent aspects that underlie CD8 T-cell induction and discuss how this knowledge may help to improve the design of efficient CD8 T-cell inducing vaccines.

The proteasome inhibitor PI31 competes with PA28 for binding to 20S proteasomes.

PI31 is a previously described inhibitor of 20S proteasomes. Using recombinant PI31 we have analyzed its effect on proteasomal hydrolyzing activity of short fluorogenic substrates and of a synthetic 40-mer polypeptide. In addition, we investigated its influence on the activation of 20S proteasome by the proteasome activator PA28. PI31 inhibits polypeptide degradation already at concentrations which only partially inhibit fluorogenic substrate turnover and immunosubunits do not influence the PI31 binding affinity. Furthermore our data demonstrate that PI31 is a potent competitor of PA28-mediated activation.

A second gene encoding the mouse proteasome activator PA28beta subunit is part of a LINE1 element and is driven by a LINE1 promoter.

The PMSE2 gene encodes the beta-subunit of the proteasome activator PA28 and, as shown by genomic Southern blot analysis, there probably exist four copies sharing sequence homology with PMSE2. Here, we report that in the mouse genome there exist two different chromosomal loci for PA28beta, both of which are transcribed and and which encode a functional PA28beta subunit. One of these represents the previously described PMSE2 gene possessing an intron-exon structure and a gamma interferon (IFNgamma)-inducible promoter. The second one, named PMSE2b, which we describe here, exhibits all the characteristics of an expressed retrotransposon. Our data show that the PA28beta retrotransposon is inserted into a transcriptional active LINE1 element and is driven by a LINE1 F-type monomer promoter as revealed by luciferase assays. The resulting PMSE2b mRNA encodes a protein which is indistinguishable from that encoded by the IFNgamma-inducible PMSE2 gene. Since PA28 plays an important role in major histocompatibility complex class I antigen presentation, the implications for the mouse immune system through a constitutively expressed PA28beta subunit and the biological relevance of this finding are discussed.