To evaluate the relationship between the distance from the superior irradiation field border to the aortic bifurcation and non-regional lymph node recurrence (NRLR) in uterine cervical cancer patients treated with definitive radiotherapy. We retrospectively investigated patients with uterine cervical cancer who had received definitive radiotherapy between January 2000 and December 2013 at our hospital. Ninety-four patients were included in this study. According to the International Federation of Obstetrics and Gynecology staging system (FIGO 2008), 14 patients were in stage I, 37 patients were in stage II, 33 patients were in stage III, and 10 patients were in stage IV. Thirty-two patients (34 %) had pelvic lymph node metastasis (N1) at the time of diagnosis. Fifty-three patients (56.4 %) were treated with concurrent chemoradiotherapy. Each external beam radiotherapy field was reviewed and the distance from the superior border of the irradiation field to the aortic bifurcation was measured. The non-regional lymph node recurrence-free survival rate (NRFS) was calculated by the Kaplan-Meier method, and differences in NRFS were evaluated by the log-rank test. All tests were two-sided, and a p value < 0.05 was considered statistically significant. The median follow-up time was 64 months (range 2 – 188 months). There were 35 patients (37.2 %) whose bifurcations were above L4 - L5 interspace. The irradiation field superior border was L4 – L5 interspace in 81 patients (86.2 %). When comparing the superior border to the level of the aortic bifurcation, the border was 1 cm or more caudal in direction to the bifurcation in 28 patients (lower group) (29.8 %) and 1cm or more cranial in direction in 20 patients (21.3 %). NRLR occurred in 19 patients. Of these, 9 patients were initially pelvic node negative (N0), and the rest of the 10 patients were N1. The 5-year NRFS was 83.2 % for the N0 and 64.7 % for the N1, respectively (p = 0.04). With regard to the superior border of the irradiated field to the level of the aortic bifurcation, the 5-year NRFS was 68.5 % in the lower group and 80.9 % in the other group (non-lower group) (p = 0.21). When limited to the N0 group, the 5-year NRFS was significantly worse in the lower group than in the non-lower group (67.5 % versus 91.3 %, p = 0.02). The N1 group had higher NRLR compared to the N0 group. In the N0 group, it could prove prudent to keep a sufficient superior margin of the irradiation field to the aortic bifurcation.

To evaluate the efficacy of nutrition education on changes in daily rectal volume and dose during intensity-modulated radiotherapy (IMRT) treatment in patients with prostate cancer. We retrospectively evaluated changes in rectal volume in patients with prostate cancer who received IMRT as a radical treatment between January 2014 and December 2017 at our hospital. Thirty consecutive patients who underwent a nutrition education program (educated group) and 30 patients who did not undergo the program (non-educated group) were included. The nutrition education program was conducted (1) to ask patients about their dietary and bowel habits, (2) to teach the importance of avoiding constipation and rectal gas filling, (3) to teach habits that should be promoted (for example: consuming a high-fiber diet, drinking 1.5–2 liters of water per day, and engaging in regular exercise), and (4) to teach dietary habits that should be avoided (for example: overconsumption of meat and fast eating). Daily pre-treatment kilovoltage cone beam computed tomography (CBCT) images were acquired for all patients. The treatment-planning CT and 12 CBCT images were used for each patient. The rectum was delineated on the planning CT (planning-rectum) and 12 CBCT (CBCT-rectum) images for each patient. The rectal and overlap volumes between the PTV and rectum were calculated as relative changes between the planning CT and CBCT images (rRectal-V and rOverlap-V, respectively). The changes in relative rectal volumes receiving 40 Gy, 60 Gy, and 70 Gy were defined as rV40, rV60, and rV70, respectively. The volumes of the CBCT-rectum and planning-rectum were compared using Fisher’s exact test, and the rRectal-V, rOverlap-V, rV40, rV60, and rV70 were compared using the Mann–Whitney U test. Seven hundred twenty CBCT images were available for analysis. In the non-educated group, 59.4% of the CBCT-rectums were larger than the planning-rectums, whereas in the educated group, 45.3% of the CBCT-rectums were larger than the planning-rectums (P < 0.001). The median values of rRectum-V and rOverlap-V were significantly lower (97.1% and 102.5%, respectively) in the educated group than those in the non-educated group (103.5% and 131.1%, respectively) (all P < 0.001). The median values of rV40, rV60, and rV70 were significantly lower (97.1%, 98.3%, and 103.6%, respectively) in the educated group than those in the non-educated group (108.4%, 115.2%, and 122.5%, respectively) (all P < 0.001). A nutrition education program decreased daily variations in rectal volume and irradiated dose to the rectum. As a result, the reproducibility of the treatment planning increased. This easy and noninvasive method could reduce late rectal toxicities at low costs.

Fanconi anemia (FA) is a rare recessive disorder with chromosomal instability, congenital abnormalities, and a high cancer risk. The breast cancer susceptibility gene BRCA2 (FANCD1) is one of the 16 genes involved in this recessive disease. We have identified a novel mutation of the splice donor site of intron 1 in the noncoding region of BRCA2 in a Japanese FA family. This mutation may account for the FA phenotype in a patient originally reported to have biallelic mutations in BRCA2. Subsequent functional studies revealed that one of the mutations, K2729N, was a neutral change. As reported here, a more careful analysis resulted in the identification of a novel splice site mutation. Functional analysis using a mouse embryonic stem cell-based assay revealed that it causes aberrant splicing, reduced transcript levels and hypersensitivity to DNA damaging agents, suggesting that it is likely to be pathogenic. Although similar pathogenic variants in the noncoding region of BRCA1 and 2 were not identified in a cohort of 752 familial breast cancer cases, we still think this finding is relevant for mutation analysis in Hereditary Breast and Ovarian Cancer Syndrome families in a diagnostic setting.

Cytotoxic T lymphocytes (CTLs) play an essential role in immunological responses for tumor rejection. In the past decade, many tumor-associated antigens (TAAs) have been identified predominantly in melanomas. Several clinical trials based on such antigenic peptides with or without adjuvants brought about partially favorable results, suggesting that identification of more immunogenic TAAs is needed. We show here the successful establishment of human leukocyte antigen (HLA)-A24-restricted CTL (TcLHK2 line1) from a pleural effusion of lung cancer patient, using B7.1 (CD80) transduced autologous lung cancer cells as an antigen-presenting cell (APC). TcLHK2 line1 recognized autologous lung adenocarcinoma cell line LHK2 in an HLA-A24-restricted fashion. Moreover, this CTL line also recognized allogeneic HLA-A24-positive lung adenocarcinoma cell line, gastric carcinoma cell line and melanoma cell line. These data raise the possibility that co-stimulatory molecule B7.1 (CD80) plays important role to overcome the immunological tolerance. Furthermore, TcLHK2 line1 is a useful tool for the identification of widely expressed shared antigens restricted by HLA-A24. Further analysis of this CTL and autologous cancer cell line will bring about novel TAAs.

A 47-year-old woman patient who presented with weakness in the proximal parts of the upper and lower limbs and difficulty in swallowing was given a diagnosis of advanced esophageal cancer complicated with polymyositis. Steroid therapy was initiated for the polymyositis, but the CK level remained high. Chemotherapy was then selected as the preferred treatment option for the esophageal cancer, however, the treatment was ineffective and the patient died of respiratory failure. There is currently no consensus on the safety of therapy for malignant carcinoma complicated with collagen diseases, therefore, the selection of treatment modality for the disease must be made with care.

Fanconi anemia (FA) is an inherited disease with congenital abnormalities and an extreme risk of acute myeloid leukemia (AML). Genetic events occurring during malignant transformation in FA and the biology of FA‐associated AML are poorly understood, but are often preceded by the development of chromosomal aberrations involving 3q26‐29 in bone marrow of FA patients. We report here the molecular cytogenetic characterization of FA‐derived AML cell lines SB1685CB and SB1690CB by conventional and array comparative genomic hybridization, fluorescence in situ hybridization, and SKY. We identified gains of a 3.7 MB chromosomal region on 3q26.2‐26.31, which preceded transformation to overt leukemia. This region harbors the oncogenic transcription factor EVI1. A third FA‐derived cell line, FA‐AML1, carried a translocation with ectopic localization of 3q26 including EVI1 . Rearrangements of 3q, which are rare in childhood AML, commonly result in overexpression of EVI1, which determines specific gene expression patterns and confers poor prognosis. We detected overexpression of EVI1 in all three FA‐derived AML. Our results suggest a link between the FA defect, chromosomal aberrations involving 3q and overexpression of EVI1 . We hypothesize that constitutional or acquired FA defects might be a common factor for the development of 3q abnormalities in AML. In addition, cryptic imbalances as detected here might account for overexpression of EVI1 in AML without overt 3q26 rearrangements. © 2007 Wiley‐Liss, Inc.

Microbiology Society journals contain high-quality research papers and topical review articles. We are a not-for-profit publisher and we support and invest in the microbiology community, to the benefit of everyone. This supports our principal goal to develop, expand and strengthen the networks available to our members so that they can generate new knowledge about microbes and ensure that it is shared with other communities.

Fanconi anemia (FA) is an inherited disease with congenital abnormalities and an extreme risk of acute myeloid leukemia (AML). Genetic events occurring during malignant transformation in FA and the biology of FA-associated AML are poorly understood, but are often preceded by the development of chromosomal aberrations involving 3q26-29 in bone marrow of FA patients. We report here the molecular cytogenetic characterization of FA-derived AML cell lines SB1685CB and SB1690CB by conventional and array comparative genomic hybridization, fluorescence in situ hybridization, and SKY. We identified gains of a 3.7 MB chromosomal region on 3q26.2-26.31, which preceded transformation to overt leukemia. This region harbors the oncogenic transcription factor EVII. A third FA-derived cell line, EA-AMLI, carried a translocation with ectopic localization of 3q26 including EVII. Rearrangements of 3q, which are rare in childhood AML, commonly result in overexpression of EVII, which determines specific gene expression patterns and confers poor prognosis. We detected overexpression of EVII in all three FA-derived AML. Our results suggest a link between the FA defect, chromosomal aberrations involving 3q and overexpression of EVII. We hypothesize that constitutional or acquired FA defects might be a common factor for the development of 3q abnormalities in AML. In addition, cryptic imbalances as detected here might account for overexpression of EVII in AML without overt 3q26 rearrangements. (c) 2007 Wiley-Liss, Inc.

The case of a 20-year-old Japanese man, diagnosed as having autosomal recessive chronic granulomatous disease (CGD), who was being treated with corticosteroids for intractable unclassified colitis, is described. He died from multiple organ failure following disseminated intravascular coagulation secondary to disseminated varicella-zoster virus (VZV) infection. He was diagnosed as an index case of CGD when 2 years old, was inoculated against VZV at the age of 5 years and had had an unremarkable course for 19 years. He was admitted to hospital because of a third episode of recurrent bloody diarrhoea. Clinical remission for each episode was achieved by intravenous corticosteroid therapy. Unclassified colitis associated with CGD was diagnosed based on a colonic biopsy demonstrating characteristic macrophages with lipofuscin deposits. From a treatment viewpoint, idiopathic inflammatory bowel disease (IBD) should be differentiated from secondary IBD occurring in CGD, in which immunosuppressive drugs including corticosteroids, still the mainstay of IBD treatment, should be avoided.

SummaryCytotoxic T lymphocyte (CTL) lines specific for allogeneic antigens were generated by in vitro stimulation of donor‐derived peripheral blood mononuclear cells obtained from patients who received human leucocyte antigen (HLA)‐matched allogeneic haematopoietic stem cell transplantation (HSCT). One of the allogeneic antigen‐specific CD4+ CTL lines, CTL‐A, generated from a patient with T cell acute lymphoblastic leukaemia, recognised HLA‐DPB1*0501‐positive Epstein–Barr virus‐immortalised human B cell line (EBV‐B cells), phytohaemagglutinin blasts and leukaemia cells, but not interferon‐γ (IFN‐γ) treated HLA‐DPB1*0501‐positive fibroblasts, indicating that this CD4+ T‐cell line recognised a minor histocompatibility antigen (mHa) that is preferentially expressed in haematopoietic cells in an HLA‐DPB1*0501‐restricted manner. The other CD4+ CTL line, CTL‐B, generated from a patient with chronic myeloid leukaemia, recognised mismatched HLA‐DQB1*0303 on EBV‐B cells and phytohaemagglutinin (PHA) blasts. Interestingly, this CTL line did not recognise IFN‐γ‐treated recipient's skin fibroblasts, as HLA‐DQ was merely upregulated even after IFN‐γ stimulation in non‐haematopoietic cells including fibroblasts, endothelial cells and hepatocytes. These results suggest that these CD4 positive CTLs, specific for mismatch HLA‐DQ and mHa that are preferentially expressed on haematopoietic cells, may play an important role in induction of selective graft‐versus‐leukaemia effect without development of graft‐versus‐host disease after allogeneic HSCT.

Background Synovial sarcoma is a high-grade malignant tumor of soft tissue, characterized by the specific chromosomal translocation t(X;18), and its resultant SYT-SSX fusion gene. Despite intensive multimodality therapy, the majority of metastatic or relapsed diseases still remain incurable, thus suggesting a need for new therapeutic options. We previously demonstrated the antigenicity of SYT-SSX gene-derived peptides by in vitro analyses. The present study was designed to evaluate in vivo immunological property of a SYT-SSX junction peptide in selected patients with synovial sarcoma. Methods A 9-mer peptide (SYT-SSX B: GYDQIMPKK) spanning the SYT-SSX fusion region was synthesized. Eligible patients were those (i) who have histologically and genetically confirmed, unresectable synovial sarcoma (SYT-SSX1 or SYT-SSX2 positive), (ii) HLA-A*2402 positive, (iii) between 20 and 70 years old, (iv) ECOG performance status between 0 and 3, and (v) who gave informed consent. Vaccinations with SYT-SSX B peptide (0.1 mg or 1.0 mg) were given subcutaneously six times at 14-day intervals. These patients were evaluated for DTH skin test, adverse events, tumor size, tetramer staining, and peptide-specific CTL induction. Results A total of 16 vaccinations were carried out in six patients. The results were (i) no serious adverse effects or DTH reactions, (ii) suppression of tumor progression in one patient, (iii) increases in the frequency of peptide-specific CTLs in three patients and a decrease in one patient, and (iv) successful induction of peptide-specific CTLs from four patients. Conclusions Our findings indicate the safety of the SYT-SSX junction peptide in the use of vaccination and also give support to the property of the peptide to evoke in vivo immunological responses. Modification of both the peptide itself and the related protocol is required to further improve the therapeutic efficacy.

The identification of tumor antigens recognized by cytotoxic T-lymphocytes and subsequent clinical trials using peptide immunization have shown tumor regression in advanced malignant melanoma patients. However, efficacious therapies for epithelial cancers, and certainly bone and soft tissue sarcomas, seemed to have been significantly delayed. For the development of a peptide-based immunotherapy for epithelial cancer and sarcomas, we have identified novel antigens recognized by CTLs using autologous pairs of tumor cell line-CTLs, or reversed immunological approach. Recently we started the phase I clinical trials of peptide-based immunotherapy. In this review, we described the recent study to establish a peptide-based immunotherapy for epithelial cancers, bone and soft-tissue sarcomas.

We first classified 12 malignant glioma cell lines into three different groups (types 1-3) with respect to major histocompatibility complex (MHC) class II expression and analyzed each group based on the different expression status of the class II transactivator (CIITA) gene. Glioma type 1 (2 of 12) showed constitutive expression of all class II molecules that might be mediated by activation of B cell-specific CIITA promoter III. Glioma type 2 represented the major phenotype (66.7 %) of malignant glioma cell lines, and MHC class II expression was induced by interferon-gamma (IFN-gamma) in this phenotype. Analysis of glioma tissue samples revealed that CIITA promoter IV was detected in 9 of 11 patients (81.8%); however, promoter III was only in two (18.2%). Moreover, cultured glioma cells obtained from a fresh tumor sample upregulated expression of CIITA and class II molecules in the presence of IFN-gamma, strongly suggesting that glioma type 2 might be predominant in glioma tissues. Glioma type 3 (2 of 12) showed CIITA transcripts but loss of MHC class II expression even in the presence of IFN-gamma. In addition, we determined that the constitutive MHC class II expression in the glioma cell lines (type 1) was the result of transcriptional activation of the CIITA gene. This phenomenon was mediated by global histone acetylation over 6 kb upstream from the transcriptional start site of CIITA promoter III. Moreover, stable transfection of CIITA promoter IV as well as promoter III into MHC class II inducible cell lines restored the constitutive expression of all class II molecules. These studies lay the foundation to understand the molecular basis for the expression of class II molecules in gliomas.

Correction to: British Journal of Cancer (2004) 90: 844–852. doi:10.1038/sj.bjc.6601602 Owing to an author error, the author affiliations in the above paper were shown incorrectly. The correct affiliations are given below: Y Morimoto1, M Toyota1,2, H Ikeda3, T Tokino2 and K Imai1 1First Department of Internal Medicine, Cancer Research Institute, Sapporo Medical University, Sapporo 060-8543, Japan; 2Department of Molecular Biology, Cancer Research Institute, Sapporo Medical University, Sapporo 060-8543, Japan; 3Department of Pathology, Sapporo Medical University, Sapporo 060-8543, Japan

The prognosis for patients with osteosarcoma who do not respond to current chemotherapy protocols still remains poor. Toward the goal of establishing efficacious peptide-based immunotherapy for those patients, we previously developed an autologous pair of CTLs and an osteosarcoma cell line. In the current study, we screened the cDNA library of this osteosarcoma cell line using an autologous CTL clone and identified cDNA encoding an antigen. The isolated cDNA was identical to papillomavirus binding factor (PBF), which was recently reported as a DNA binding transcription factor cooperating with RUNX1. Reverse transcription-PCR analysis revealed that PBF was expressed in 16 of 19 cases of bone and soft-tissue sarcoma cell lines (5 of 6 of osteosarcoma lines) and 57 of 76 sarcoma tissue samples (11 of 14 of osteosarcoma tissues). Also, PBF was expressed in 10 of 13 epithelial cancer cell lines and 20 of 34 of cancer tissues. In contrast, PBF was detected in some normal organs including ovary, pancreas, spleen, and liver by reverse transcription-PCR but was restricted in the cytoplasm by immunostaining and undetectable by Western blotting. Furthermore, a 12-mer peptide, CTACRWKKACQR, located at the COOH terminus of PBF, was found to be a minimum requirement for recognition by the CTL clone in the context of the HLA-B*5502 molecule. These findings suggest that PBF is a shared tumor-associated antigen, which may serve as a source of peptides applicable to peptide-based immunotherapy for osteosarcoma and other malignant tumors.

To investigate the effects of anchor substitutions in SYT-SSX junction peptide, an HLA-A24 anchor residue (position 9) of the SYT-SSX B peptide (GYDQIMPKK) was substituted to more favorable residues according to the HLA-A24-binding motif. Among four substitutes constructed, a substitute with isoleucine (termed K9I peptide) most apparently enhanced the affinity for HLA-A24 molecule. Subsequent in vitro CTL induction analysis using PBMCs of 15 HLA-A24(+) synovial sarcoma patients revealed that the original B peptide allowed to induce synovial sarcoma-specific CTLs from 7 patients (47%), whereas such CTLs were inducible from 12 patients (80%) with K9I peptide. Moreover, the extent of cytotoxicity against HLA-A24(+) synovial sarcoma cell lines was higher in K9I peptide-induced CTLs than B peptide-induced CTLs. Influence of anchor substitution on peptide/TCR interaction was evaluated by cytotoxicity assays against autologous cells and tetramer analysis. CTLs induced from a synovial sarcoma patient using K9I peptide did not lyse autologous PHA blasts or EBV-infected B cells. In vitro stimulations of PBMCs from 5 HLA-A24(+) synovial sarcoma patients with K9I peptide increased the frequency of T cells reacting with both HLA-A24/K9I peptide tetramer and HLA-A24/B peptide tetramer. In contrast, the frequency of T cells reacting with HLA/HIV-derived peptide tetramer remained low. These findings support the validity in design of anchor residue substitution in SYT-SSX fusion gene-derived peptide, and provide a potential clue to the current stagnation in vaccination trials of fusion gene-derived natural junction peptides.

Tightly regulated at the level of transcription, expression of MHC class II molecules varies significantly among gastrointestinal cancers. High levels of MHC class II expression are often associated with a better prognosis, which is indicative of the involvement of CD4+ lymphocytes in tumor suppression, but the molecular mechanism by which MHC class II expression is regulated remains unclear. In the present study, we investigated the expression of one inducible MHC class II molecule, HLA-DR, and its coactivators in a panel of colorectal and gastric cancer cell lines. Interferon-γ induced expression of HLA-DR in 14 of 20 cell lines tested; the remaining six cell lines did not express HLA-DR. Analysis of the expression of transcription factors and coactivators associated with HLA-DR revealed that the loss of CIITA expression was closely associated with the absence of HLA-DR induction. Moreover, DNA methylation of the 5′ CpG island of CIITA-PIV was detected in all cancer cells that lacked CIITA. The methylation and resultant silencing of CIITA-PIV depended on the activities of two DNA methyltransferases, DNMT1 and DNMT3B, and their genetic inactivation restored CIITA-PIV expression. It thus appears that CIITA methylation is a key mechanism that enables some gastrointestinal cancer cells to escape immune surveillance.