Researchers designing antitumor treatments have long focused on eliciting tumor-specific CD8 cytotoxic T lymphocytes (CTL) because of their potent killing activity and their ability to reject transplanted organs. The resulting treatments, however, have generally been surprisingly poor at inducing complete tumor rejection, both in experimental models and in the clinic. Although a few scattered studies suggested that CD4 T "helper" cells might also serve as antitumor effectors, they have generally been studied mostly for their ability to enhance the activity of CTL. In this mouse study, we compared monoclonal populations of tumor-specific CD4 and CD8 T cells as effectors against several different tumors, and found that CD4 T cells eliminated tumors that were resistant to CD8-mediated rejection, even in cases where the tumors expressed major histocompatibility complex (MHC) class I molecules but not MHC class II. MHC class II expression on host tissues was critical, suggesting that the CD4 T cells act indirectly. Indeed, the CD4 T cells partnered with NK cells to obtain the maximal antitumor effect. These findings suggest that CD4 T cells can be powerful antitumor effector cells that can, in some cases, outperform CD8 T cells, which are the current "gold standard" effector cell IntroductionResearchers designing antitumor vaccines, or treatments involving transfers of activated antitumor cells, have long focused on methods to elicit tumor-specific CD8 CTLs, envisioning that their potent ability to kill tumor targets in vitro and to reject transplants in vivo would translate into equally potent antitumor activity in vivo. Although many of the resulting treatments have indeed been able to elicit CTLs that recognize tumor cells and/or tumor antigens in vitro, complete tumor regression has been achieved in only a minority of patients. [1][2][3][4][5] Animal models have generated similar results. In a few cases, the transfer of monoclonal T cell receptor transgenic (TCR Tg) CD8 T cells was able to clear small tumors, 6 but in most, the TCR Tg CD8 cells were ineffective without the addition of other aids. In short, though CD8 CTL can clear tumors, they most often do not, unless helped by additional treatments. [6][7][8][9][10][11][12] Over the last 25 years, a few studies have shown that CD4 T cells could also clear tumors completely independently of CD8s. [13][14][15][16][17] Nevertheless, CD4 T cells continue to be studied mainly for their role as helpers for CD8 CTL, 11,18,19 and it has even been suggested that tumor-specific CD4 T regulatory cells could act as suppressors of antitumor responses. 20 Thus, their potential as CD8-independent antitumor effectors has gained only a few proponents, [13][14][15][16][17][21][22][23][24] and only a few of the newly designed cancer vaccines incorporate antigens to stimulate CD4 cells, mostly to enhance their helper activity. 25,26 Most studies using adoptive transfer of tumor-specific T cells continue to focus entirely on CD8 cells. 2,3,[27][28][29][30] We decided to do a direc...

Transglutaminase (TGase) 2 is a ubiquitously expressed enzyme that modifies proteins by cross-linking or polyamination. An aberrant activity of TGase 2 has implicated its possible roles in a variety of diseases including age-related cataracts. However, the molecular mechanism by which TGase 2 is activated has not been elucidated. In this report, we showed that oxidative stress or UV irradiation elevates in situ TGase 2 activity. Neither the expression level nor the in vitro activity of TGase 2 appeared to correlate with the observed elevation of in situ TGase 2 activity. Screening a number of cell lines revealed that the level of TGase 2 activation depends on the cell type and also the environmental stress, suggesting that unrecognized cellular factor(s) may specifically regulate in situ TGase 2 activity. Concomitantly, we observed that human lens epithelial cells (HLE-B3) exhibited about 3-fold increase in in situ TGase 2 activity in response to the stresses. The activated TGase 2 catalyzed the formation of water-insoluble dimers or polymers of ␣B-crystallin, ␤B 2 -crystallin, and vimentin in HLE-B3 cells, providing evidence that TGase 2 may play a role in cataractogenesis. Thus, our findings indicate that in situ TGase 2 activity must be evaluated instead of in vitro activity to study the regulation mechanism and function of TGase 2 in biological and pathological processes.

We established an AQP4-Ab ELISA, which could be a potential monitoring tool of disease activity.

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Cytotoxic T lymphocyte-associated antigen 4 (CTLA4) has been known to be a strong tolerance-inducing inhibitory receptor on T-cell surface. Systemic blocking of CTLA4 function with blocking antibodies has been regarded as an attractive strategy to enhance antitumor immunity. However, this strategy accompanies systemic autoimmune side effects that are sometimes problematic. Therefore, we developed a novel CTLA4 mutant that could be expressed in tumor antigen-specific T cells to enhance antitumor effect without systemic autoimmunity. This mutant, named CTLA4-CD28 chimera, consists of extracellular and transmembrane domains of CTLA4, linked with cytoplasmic CD28 domain. Overexpression of CTLA4-CD28 chimera in T cells delivered stimulatory signals rather than inhibitory signals of CTLA4 and significantly enhanced T-cell reactivity. Although this effect was observed in both CD4 and CD8 T cells, the effect on CD4 T cells was predominant. CTLA4-CD28 chimera gene IntroductionCytotoxic T lymphocyte-associated antigen 4 (CTLA4) is an inhibitory receptor expressed on T-cell surface that is induced by T-cell activation to negatively regulate excessive activation of T cells. 1 CTLA4-deficient mice develop lethal lymphoproliferative diseases within 3 to 4 weeks of birth. 2 CTLA4 deficiency or anti-CTLA4 antibody treatment ameliorated induction of peripheral T-cell tolerance 3 and exacerbated autoimmune diseases 4,5 in mouse models. Therefore, CTLA4 is thought to play a critical role in maintaining peripheral T-cell tolerance in vivo to prevent autoimmunity. Because tumor cells also were known to induce T-cell tolerance to tumor antigens to resist to antitumor immunity, 6 CTLA4 blockade was believed to be able to break tumor-mediated T-cell tolerance, resulting in enhanced antitumor immunity. Indeed, systemic administration of anti-CTLA4 blocking antibody in preclinical murine tumor models led to efficient regression of several syngenic tumors. [7][8][9] Thus, blocking CTLA4 in human malignancies has been regarded as a promising therapeutic target of antitumor immunotherapy.Therapeutic efficacy of 2 fully human anti-CTLA4 antibodies is currently being tested for several malignancies in clinical trials. 10 For metastatic melanoma, anti-CTLA4 monotherapy or combination therapy with other agents shows an overall response rate of 5% to 15%. A recent study indicated that combination therapy of anti-CTLA4 plus dacarbazine, a chemotherapeutic agent, could improve the overall survival compared with dacarbazine monotherapy in a phase 3 randomized clinical trial. 11 However, a significant proportion of patients who received anti-CTLA4 therapy developed grade III or IV autoimmune side effects, such as enterocolitis, skin rash, hypophysitis, hepatitis, and uveitis. 12 Although most of these side effects could be managed by steroid treatment, several patients experienced the serious conditions such as intestinal perforation. 13 The more intriguing observation was that the degree of autoimmunity was correlated with treatment efficacy. 12 T...

Transglutaminase 2 (TGase 2) is one of a family of enzymes that catalyze protein modi®cation through the incorporation of polyamines into substrates or the formation of protein crosslinks. However, the physiological roles of TGase 2 are largely unknown. To elucidate the functions of TGase 2, we have searched for its interacting proteins. Here we show that TGase 2 interacts with E7 oncoprotein of human papillomavirus type 18 (HPV18) in vitro and in vivo. TGase 2 incorporates polyamines into a conserved glutamine residue in the zinc-binding domain of HPV18 E7 protein. This modi®cation mediates the inhibition of E7's Rb binding ability. In contrast, TGase 2 does not affect HPV16 E7, due to absence of a glutamine residue at this polyamination site. Using E7 mutants, we demonstrate that TGase 2-dependent inhibition of HPV E7 function correlates with the presence of the polyamination site. Our results indicate that TGase 2 is an important cellular interfering factor and de®ne a novel host±virus interaction, suggesting that the inability of TGase 2 to inactivate HPV16 E7 could explain the high prevalence of HPV16 in cervical cancer.

Adenoviruses harboring the herpes simplex virus thymidine kinase (HSVtk) gene under the regulation of a trans-splicing ribozyme targeting human telomerase reverse transcriptase (hTERT-TR) show marked and specific antitumor activity. In addition to inducing tumor cell death by direct cytotoxicity, it is becoming clear that HSVtk also induces antitumor immunity. Programmed death ligand 1 (PD-L1) expressed on tumor cell surfaces mediates tumor-induced immunoresistance by inhibiting PD1-expressing tumor-infiltrating T cells. Here, we explored whether a soluble form of PD1 (sPD1-Ig), which blocks PD-L1, could synergize with TERT-TR-regulated HSVtk to enhance the adenoviral therapeutic efficacy by boosting antitumor immunity. Tumor antigen released by HSVtk-transduced tumors successfully primed tumor antigen-specific CD8 T cells via dendritic cells (DC). Regression of murine tumors was markedly enhanced when sPD1-Ig was incorporated into the adenovirus as compared with a single-module adenovirus expressing only HSVtk. This effect was abolished by CD8 T-cell depletion. Consistent with this, following adoptive transfer of tumor antigen-specific CD8 T cells into tumor-bearing Rag1(-/-) mice, dual-module adenovirus significantly enhanced CD8 T cell-mediated tumor rejection. In addition, secondary tumor challenge at a distal site was completely suppressed in mice treated with a dual-module adenovirus. These results suggest that a dual-targeting strategy to elicit both tumor antigen priming and tumor-induced immunoresistance enhances CD8 T cell-mediated antitumor immunity.

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