No abstract

We have previously described a method for adoptive immunotherapy of cancer based on antigen-specific T(h)1 cells. However, efficient induction of anti-tumor responses using T(h)1 cells remains a formidable challenge, especially for MHC class II-negative tumors. In the present study, we sought to develop a novel strategy to eradicate established tumors of the MHC class II-negative, ovalbumin (OVA)-expressing EG-7 cells. Tumor-bearing mice were intradermally treated with OVA-specific T(h)1 cells, combined with the model tumor antigen (OVA), near the tumor-draining lymph node (DLN). We found that tumor growth was significantly inhibited by this strategy and approximately 50-60% of tumor-bearing mice were completely cured. Tumor eradication was crucially dependent on the generation of OVA/H-2K(b)-specific CTLs in the tumor DLNs and tumor site. The injected T(h)1 cells were mainly distributed in tumor DLNs, where they vigorously proliferated and enhanced the activation of dendritic cells. Strikingly, we also found that the accumulation of CD4(+)CD25(+) regulatory T cells (Tregs) was significantly inhibited in tumor DLNs by T(h)1 cell adjuvant therapy and this abrogation was associated with IFNgamma secreted by T(h)1 cells. These results identify T(h)1 cell adjuvant therapy combined with tumor vaccination as a novel approach to the treatment of human cancer.

The general and efficient method for the site-directed glycosylation of proteins is a key step in order to understand the biological importance of the carbohydrate chains of proteins and to control functional roles of the engineered glycoproteins in terms of the development of improved glycoprotein therapeutics. We have developed a novel method for site-directed glycosylation of proteins based on chemoselective blotting of common reducing sugars by genetically encoded proteins. The oxylamino-functionalized L-homoserine residues, 2-amino-4-O-(N-methylaminooxy) butanoic acid and 2-amino-4-aminooxy butanoic acid, were efficiently incorporated into proteins by using the four-base codon/anticodon pair strategy in Escherichia coli in vitro translation. Direct and chemoselective coupling between unmodified simple sugars and N-methylaminooxy group displayed on the engineered streptavidin allowed for the combinatorial synthesis of novel glycoprotein mimetics.

This paper describes the acid/base-mediated three-state translational isomerization of two [2]rotaxanes, each containing N-alkylaniline and N,N-dialkylamine centers as binding sites for threaded dibenzo[24]crown-8 units. Under neutral conditions, the dialkylamine unit predominantly recognized the crown ether component through cooperative binding of a proton; when both amino units were protonated under acidic conditions, both translational isomers were generated; the addition of a strong base caused aniline-crown ether interactions to dominate. The three states of the [2]rotaxane featuring the 3,5-diphenylaniline terminus in its dumbbell-shaped component were accompanied by distinct absorptive outputs that were detectable using UV spectroscopy.

In unimmunized specific pathogen-free mice, there are unique memory-type CD8(+) T cell populations expressing asialoGM1 (ASGM1). These cells were classified into central memory-type T cells (T(CMT)) judging from their expression profile of CD44, IL-2Rbeta, CD62L and CCR7 cell-surface molecules. Among CD44(high)CD8(+) so-called memory CD8(+) T cell population, ASGM1(+)CD44(high)CD8(+) T(CMT), but not ASGM1(-)CD44(high)CD8(+) memory T cells, produced IFN-gamma by stimulation with anti-CD3 mAb. The physiological significance of ASGM1(+)CD8(+) T(CMT) as early source of IFN-gamma was also demonstrated in vivo. Namely, intravenous injection of anti-CD3 mAb (2 microg) resulted in early activation of IFN-gamma-producing ASGM1(+)CD8(+) T(CMT) cells as well as NKT and NK cells. Unexpectedly, however, few IFN-gamma-producing CD4(+) T cells were detected until 4 h after anti-CD3 mAb administration. Thus, ASGM1(+)CD8(+) T(CMT) were demonstrated to be early IFN-gamma producer, which may be crucial for T(h)1-dependent cellular immunity. Indeed, co-culture of naive CD4(+) T cells with ASGM1(+)CD8(+) T(CMT) but not ASGM1(-)CD8(+) T cells caused a great acceleration of IFN-gamma-producing T(h)1 cells in vitro. Finally, we found that T(h)1-prone C57BL/6 mice possessed higher percentage (10%) of ASGM1(+)CD8(+) T(CMT) in CD8(+) T cells compared with that (3%) of T(h)2-prone BALB/c mice. Moreover, ASGM1(+)CD8(+) T(CMT) derived from C57BL/6 mice produced higher levels of IFN-gamma compared with those from BALB/c mice. Thus, ASGM1(+)CD8(+) T(CMT), whose differentiation in vivo is genetically controlled, appear to play a critical role in the control of type 1 immunity, which is essential for therapy of tumors and infectious diseases.