Single-cell sequencing on CD8+ TILs revealed the nature of exhausted T cells recognizing neoantigen and cancer/testis antigen in non-small cell lung cancer.

BACKGROUND: CD8+tumor infiltrating lymphocytes (TILs) are often observed in non-small cell lung cancers (NSCLC). However, the characteristics of CD8+ TILs, especially T-cell populations specific for tumor antigens, remain poorly understood. METHODS: High throughput single-cell RNA sequencing and single-cell T-cell receptor (TCR) sequencing were performed on CD8+ TILs from three surgically-resected lung cancer specimens. Dimensional reduction for clustering was performed using Uniform Manifold Approximation and Projection. CD8+ TIL TCR specific for the cancer/testis antigen KK-LC-1 and for predicted neoantigens were investigated. Differentially-expressed gene analysis, Gene Set Enrichment Analysis (GSEA) and single sample GSEA was performed to characterize antigen-specific T cells. RESULTS: A total of 6998 CD8+ T cells was analyzed, divided into 10 clusters according to their gene expression profile. An exhausted T-cell (exhausted T (Tex)) cluster characterized by the expression of ENTPD1 (CD39), TOX, PDCD1 (PD1), HAVCR2 (TIM3) and other genes, and by T-cell oligoclonality, was identified. The Tex TCR repertoire (Tex-TCRs) contained nine different TCR clonotypes recognizing five tumor antigens including a KK-LC-1 antigen and four neoantigens. By re-clustering the tumor antigen-specific T cells (n=140), it could be seen that the individual T-cell clonotypes were present on cells at different stages of differentiation and functional states even within the same Tex cluster. Stimulating these T cells with predicted cognate peptide indicated that TCR signal strength and subsequent T-cell proliferation and cytokine production was variable but always higher for neoantigens than KK-LC-1. CONCLUSIONS: Our approach focusing on T cells with an exhausted phenotype among CD8+ TILs may facilitate the identification of tumor antigens and clarify the nature of the antigen-specific T cells to specify the promising immunotherapeutic targets in patients with NSCLC.

New evaluation of the tumor immune microenvironment of non-small cell lung cancer and its association with prognosis.

BACKGROUND: A better understanding of the tumor immune microenvironment (TIME) will facilitate the development of prognostic biomarkers and more effective therapeutic strategies in patients with lung cancer. However, little has been reported on the comprehensive evaluation of complex interactions among cancer cells, immune cells, and local immunosuppressive elements in the TIME. METHODS: Whole-exome sequencing and RNA sequencing were carried out on 113 lung cancers. We performed single sample gene set enrichment analysis on TIME-related gene sets to develop a new scoring system (TIME score), consisting of T-score (tumor proliferation), I-score (antitumor immunity) and S-score (immunosuppression). Lung cancers were classified according to a combination of high or low T-score, I-score, and S-scores (eight groups; G1-8). Clinical and genomic features, and immune landscape were investigated among eight groups. The external data sets of 990 lung cancers from The Cancer Genome Atlas and 76 melanomas treated with immune checkpoint inhibitors (ICI) were utilized to evaluate TIME scoring and explore prognostic and predictive accuracy. RESULTS: The representative histological type including adenocarcinoma and squamous cell carcinoma, and driver mutations such as epidermal growth factor receptor and TP53 mutations were different according to the T-score. The numbers of somatic mutations and predicted neoantigens were higher in Thi (G5-8) than Tlo (G1-4) tumors. Immune selection pressure against neoantigen expression occurred only in Thi and was dampened in Thi/Ilo (G5-6), possibly due to a reduced number of T cells with a high proportion of tumor specific but exhausted cells. Thi/Ilo/Shi (G5) displayed the lowest immune responses by additional immune suppressive mechanisms. The T-score, I-score and S-scores were independent prognostic factors, with survival curves well separated into eight groups with G5 displaying the worst overall survival, while the opposite group Tlo/Ihi/Slo (G4) had the best prognosis. Several oncogenic signaling pathways influenced on T-score and I-scores but not S-score, and PI3K pathway alteration correlated with poor prognosis in accordance with higher T-score and lower I-score. Moreover, the TIME score predicted the efficacy of ICI in patients with melanoma. CONCLUSION: The TIME score capturing complex interactions among tumor proliferation, antitumor immunity and immunosuppression could be useful for prognostic predictions or selection of treatment strategies in patients with lung cancer.

Development of adoptive immunotherapy with KK-LC-1-specific TCR-transduced γδT cells against lung cancer cells.

The present study analyzed the antitumor effect of γδT cells transduced with the TCR of cancer-specific CTLs to establish forceful cancer-specific adoptive immunotherapy. We cloned the TCRαß genes from CTLs showing HLA-B15 restricted recognition of Kita-Kyushu lung cancer antigen-1 (KK-LC-1), a cancer/germline gene antigen, identified in a lung adenocarcinoma case (F1121). The TCRαß and CD8 genes were transduced into γδT cells induced from PBLs of healthy volunteers stimulated with zoledronate and IL-2. The KK-LC-1-specific TCRαß-CD8 γδT cells showed cytotoxic activity against the KK-LC-1 positive lung cancer cell line F1121L and produced IFN-γ against F1121L and KK-LC-1 peptide-pulsed F1121 EBV-B cells. These responses were blocked by HLA class I and HLA-B/C antibodies. An in vivo assay using NOD/SCID mice with xenotransplantation of human lung cancer cells was performed, and the TCRαß-CD8 transduced γδT cells (TCRαß-CD8 γδT cells) were intravenously injected. Growth inhibition of KK-LC-1+ , HLA-B15+ lung cancer cells was confirmed in mice with injection of the TCRαß-CD8 γδT cells from 1 wk after xenotransplantation of cancer cells but not in those treated 2 wk after xenotransplantation. The resected specimens of the tumor, 2 wk after xenotransplantation, highly expressed FasL but not programmed death ligand-1 (PD-L1) by immunohistochemical staining. FasL highly expressed cancer cells xenotransplanted 2 wk ago were resistant to TCRαß-CD8 γδT cells injection. These results suggested that apoptosis of Fas-positive TCRαß-CD8 γδT cells may be induced by a Fas-mediated signal after interacting with FasL-positive cancer cells.

Group B streptococcal empyema necessitatis with pleural fistula after blunt trauma: A case report.

INTRODUCTION: We report the first case of empyema necessitatis (EN) with pleural fistula and septic arthritis caused by Streptococcus agalactiae following blunt trauma. PRESENTATION OF THE CASE: A 46-year-old man with diabetes mellitus and a history of recent right rib fracture and right knee bruising presented with dyspnea and right knee pain. He was diagnosed with EN and underwent chest drainage, followed by open-window thoracotomy. Septic arthritis occurred on day 8 after thoracotomy. The chest wall wound healed after 3 months. DISCUSSION: EN is a rare complication of empyema. In this patient, infection was invasive, causing necrotizing pneumonia with a pleural fistula. To our knowledge, there are no reports of group B streptococcal EN with a pleural fistula resulting from blunt chest trauma. CONCLUSION: Group B streptococcal infection might become invasive in immunocompromised patients, so careful follow-up for those patients is important.

Subxiphoid approach for extracting a giant solitary fibrous tumour of the pleura.

We describe a case of resection of a solitary fibrous tumour of the pleura using video-assisted thoracic surgery and removal of the giant tumour using a subxiphoid incision without the need for minithoracotomy. Use of the subxiphoid approach as a retrieval port is simple and feasible.

Two adult cases of extralobar pulmonary sequestration: A non-complicated case and a necrotic case with torsion.

BACKGROUND: This case report describes two cases of extralobar pulmonary sequestration in adults with and without torsion/necrosis. CASE REPORTS: Non-complicated extralobar pulmonary sequestration was found incidentally in a 50-year-old asymptomatic woman (Case 1), diagnosed with the presence of a branching structure in a mass lesion and blood supply from the right inferior phrenic artery. Another case of a 38-year-old woman presented with a sudden onset of back pain caused by extralobar pulmonary sequestration with torsion/necrosis (Case 2). A 4-cm fusiform mass in the paravertebral region showed enhancement in the peripheral rim only, and no feeding artery. These were the same as it had been reported typical findings in extralobar pulmonary sequestration with necrosis. On magnetic resonance imaging, the masses in both cases showed inhomogeneous low signal and branching high signal on T2-weighted images. That was characteristic for a stroma without dilated alveoli as a solid part and dilated alveoli as fluid regions. CONCLUSIONS: By comparing those two cases, we came to a conclusion that only T2-weighted imaging reflects the native structure, even after infarction. Although differentiation from a cystic tumor with hemorrhage or infection can be problematic, inhomogeneous low signal and branching high signal on T2-weighted images may help us distinguish extralobar pulmonary sequestration from other cystic lesions.

Tumor invasiveness as defined by the newly proposed IASLC/ATS/ERS classification has prognostic significance for pathologic stage IA lung adenocarcinoma and can be predicted by radiologic parameters.

OBJECTIVES: The International Association for the Study of Lung Cancer, American Thoracic Society, and European Respiratory Society (IASLC/ATS/ERS) have collaborated to propose a new pathologic classification of lung adenocarcinoma. In this classification, noninvasiveness and invasiveness have been newly defined for lung adenocarcinoma. The aims of this study were to validate the prognostic significance of tumor invasiveness as defined by the new IASLC/ATS/ERS classification and to assess the relationship between pathologic invasiveness and radiologic findings in pathologic stage IA lung adenocarcinoma. METHODS: We retrospectively reviewed 123 consecutive patients with pathologic stage IA lung adenocarcinoma. Pathologic data were classified according to the new IASLC/ATS/ERS classification. The following radiologic parameters were assessed using thin-section computed tomography: the ground-glass opacity ratio, tumor disappearance rate, and consolidation diameter. RESULTS: There were 54 noninvasive and 69 invasive adenocarcinomas. Five-year overall survival rates for noninvasive adenocarcinoma and invasive adenocarcinoma were 100% and 78.4%, respectively; this difference was statistically significant (P < .01), indicating the prognostic value of this classification. Receiver operating characteristic curves of the ground-glass opacity ratio, tumor disappearance rate, and consolidation diameter identified the optimal cut-off values for predicting the presence of invasive tumors as 50%, 75%, and 10 mm, respectively. CONCLUSIONS: We found that by using the new IASLC/ATS/ERS classification, histologic subtypes of pathologic stage IA lung adenocarcinoma with prognostic value could be identified. Tumor invasiveness of lung adenocarcinoma as defined by this classification can be predicted by evaluating the ground-glass opacity ratio, tumor disappearance rate, and consolidation diameter on thin-section computed tomography.

Spindle cell component in pleomorphic carcinoma showing polypoid growth in the pulmonary vein.

We describe a rare case of a pleomorphic carcinoma forming polypoid growth in the pulmonary vein. A 69-year-old man was admitted with an abnormal shadow in his right lung. Thoracic computed tomography scans showed a polypoid lesion extending into the right lower pulmonary vein. We performed a right lower lobectomy without any evidence of tumor embolism perioperatively. Pathological examination revealed that the lesion was a pleomorphic carcinoma of the lung and that the polypoid lesion in the pulmonary vein was composed solely of spindle cell components. Early ligation of affected vessel is important to avoid tumor embolism through the pulmonary veins.

Clinical significance of the frequency of regulatory T cells in regional lymph node lymphocytes as a prognostic factor for non-small-cell lung cancer.

BACKGROUND: Regulatory T cells (Tregs) are potent immunosuppressive cells that play a crucial role in tumor immune escape. The purpose of the present study was to evaluate the prognostic significance of the frequency of CD4+CD25+Foxp3+ Tregs in the regional lymph node lymphocytes (RLNL) and peripheral blood lymphocytes (PBL) in patients who underwent surgical resection of non-small cell lung cancer (NSCLC). METHODS: The RLNL and PBL in 158 NSCLC patients who underwent complete surgical resection were collected at the time of surgery. The proportions of CD4+CD25+Foxp3+ cells in the RLNL and PBL were determined by flow cytometry. RESULTS: The average proportions of Tregs in the RLNL and PBL were 1.28% and 0.76%, respectively. The proportion of Tregs in the RLNL was significantly higher than that in the PBL (p < 0.0001). The 5-year overall survival rates of the patients according to the proportion of Tregs in the RLNL were 84.4% and 63.5% in the lower and higher groups, respectively. A significant difference was observed in the survival rate between the higher and lower groups (p = 0.0056). Among the patients with stage I disease, the 5-year survival rate (91.4%) was significantly higher in patients with the lower proportion of Tregs in RLNL that in the higher group (72.1%) (p = 0.0147). CONCLUSIONS: The higher proportion of Tregs in the RLNL was a significant unfavorable prognostic factor, even in patients with node-negative NSCLC. The information about the proportion of Tregs in the RLNL might improve the discriminatory power for assessing the risk of the recurrence of NSCLC.

Clinical significance of expression of cancer/testis antigen and down-regulation of HLA class-I in patients with stage I non-small cell lung cancer.

AIM: The purpose of this study was to investigate the clinical significance of expression of cancer/testis (CT) antigen and down-regulation of HLA class-I in patients with stage I non-small cell lung cancer (NSCLC), which underwent complete surgical resection. PATIENTS AND METHODS: The expression of HLA class-I molecules was evaluated in 136 resected NSCLC specimens by immunohistochemistry. The results were scored as the percentage of stained tumor cells and categorized into two groups: 0-79%, reduced expression; and >80%, normal expression. The expression of CT antigen was performed by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: The expression of HLA class-I was normal in 49 tumors (36%), and there was reduced expression in 87 tumors (64%). The expression of Melanoma antigen (MAGE)-A3, MAGE-A4, and Kita-Kyushu lung cancer antigen-1 (KK-LC-1) was positive in 34 (25.0%), 22 (16.2%), and 42 (30.9%) patients, respectively. There was no significant difference in the proportion of HLA class-I expression associated with the expression of any of the CT antigens. Among the patients with positive expression of at least one of the CT antigens, the 5-year survival rate of the patients with the normal expression of HLA class-I was 87.5%; however, it was 63.4% in patients with the reduced expression of HLA class-I (p=0.0477). CONCLUSION: Reduced expression of HLA class-I was an unfavorable prognostic factor in patients with positive expression of CT antigen, and represents an important hurdle to antigen-based cancer immunotherapy.

Clinical significance of human leukocyte antigen loss and melanoma-associated antigen 4 expression in smokers of non-small cell lung cancer patients.

BACKGROUND: Melanoma-associated antigen-A4 (MAGE-A4) is one of the candidates for a target of immunotherapy and is expressed in non-small cell lung cancer (NSCLC). However, tumors sometimes lose human leukocyte antigen (HLA) class I expression, and tumor-specific T cells cannot eliminate the tumor with loss of HLA. However, the relationship between MAGE-A4 expression and HLA loss has remained unclear. METHODS: Among 363 NSCLC patients who consecutively underwent curative surgery, 187 cases whose material could be analyzed were reviewed. The expression of HLA class I molecules was assessed by immunohistochemical staining. The expression of MAGE-A4 was analyzed by RT-PCR. RESULTS: Seventy-seven tumors expressed HLA normally; however, 110 tumors lost HLA. The proportion of patients with a smoking habit and expressing the MAGE-A4 gene in patients with HLA loss was higher than those with HLA expression (p = 0.04 and 0.028, respectively). Five-year overall survival (OS) rate in the patients expressing MAGE-A4 but with loss of HLA was 52.4 %, and OS was significantly poorer than their counterparts (74.0 %, p = 0.036). Multivariate analysis indicated that advanced stage or history of smoking and HLA loss was an independently poor prognostic predictor of OS in NSCLC (p < 0.01 and p = 0.04, respectively). CONCLUSION: HLA class I loss in NSCLC was related to smoking history and MAGE-A4 expression of tumors. HLA class I loss in smokers or patients with the MAGE-A4 gene was a prognostic factors in NSCLC.

Prognostic implications of human leukocyte antigen class I expression in patients who underwent surgical resection for non-small-cell lung cancer.

BACKGROUND: The purpose of the present study was to clarify the prognostic significance of human leukocyte antigen (HLA) class I expression in patients with non-small-cell lung cancer who underwent complete surgical resection. PATIENTS AND METHODS: The expression of HLA class I molecules was evaluated in 403 resected NSCLC specimens using immunohistochemistry. The results were scored as the percentage of stained tumor cells and were categorized into three groups: 0%-24% (decreased), 25%-79% (heterogeneous), and 80% or more (normal). RESULTS: The expression of HLA class I was evaluated in 124 tumors in the normal expression group, 181 tumors in the heterogeneous expression group, and 98 tumors in the decreased expression group. The 5-year survival rate of all patients after surgery according to the HLA class I expression in the normal, heterogeneous, and decreased groups was 76.6%, 65.9%, and 76.1%, respectively. The prognosis was significantly better in the normal expression group than in the heterogeneous group. Normal HLA class I expression also correlated with favorable survival in patients with stage I disease. CONCLUSIONS: The normal expression of HLA class I was associated with a favorable prognosis compared with the heterogeneous expression group, but no significant difference was observed between the normal expression and decreased expression groups.

Cancer/testis antigen expression as a predictor for epidermal growth factor receptor mutation and prognosis in lung adenocarcinoma.

OBJECTIVES: Immune therapy targeting cancer/testis (CT) antigens improve the survival in several types of solid tumours. The expression of CT antigens is related to poor survival in non-small-cell lung cancer (NSCLC). The epidermal growth factor receptor (EGFR) mutation is the best predictive factor for the sensitivity to tyrosine kinase inhibitors in lung adenocarcinoma. The aim of this study was to elucidate the correlation between the expression of CT antigens and clinicopathological factors, including the EGFR mutation, and to analyse the prognosis in lung adenocarcinoma. METHODS: Data were collected from a total of 281 lung adenocarcinoma patients who underwent surgery. Among them, 125 cases, whose specimens were too small to extract sufficient DNA and/or RNA, and 2 cases with the coexistence of another histological lung cancer were excluded. A total of 154 patients were reviewed. The expression of CT antigens (melanoma-associated antigen gene [MAGE]-A4 and KK-LC-1) and the EGFR-activating mutation (L858R point mutation in exon 21 and inframe deletion in exon 19) was evaluated by using polymerase chain reaction amplification. RESULTS: The expression of MAGE-A4 and KK-LC-1 was detected in 14 (9%) and 54 patients (35%) with adenocarcinoma. The EGFR-activating mutation was found in 64 patients (42%). Univariate and multivariate analyses demonstrated that tumours expressing at least one CT antigen were associated with no EGFR mutation (odds ratio = 0.3; 95% confidence interval, 0.14-0.71; P < 0.01). A survival analysis was performed in 135 patients who underwent complete resection and the 5-year overall survival rate was 71.1% in those with any expression of CT antigens and 83.2% in those without expression of the genes (P < 0.04). CONCLUSION: Two different therapeutic targets, EGFR-activating mutation and CT antigen, have a negative relationship with each other.

Intrapleural chemotherapy improves the survival of non-small cell lung cancer patients with positive pleural lavage cytology.

PURPOSE: Information regarding the treatment of pleural lavage cytology (PLC)-positive patients is still limited. This study evaluated the efficacy of intrapleural chemotherapy (IPC) in PLC-positive patients. METHODS: Three hundred eighty-six of the 567 lung cancer patients who underwent surgery had undergone PLC after thoracotomy, following by a complete resection were evaluated. IPC was performed after surgery, and cisplatin or adriamycin was injected intrapleurally through the thoracic tube. RESULTS: The pathological diagnosis showed that 17 patients (4.4 %) were positive for (or suspected to have) malignancy in their PLC. The univariate and multivariate analysis showed that only pleural invasion was a significant predictor of a PLC-positive status. The 5-year overall survival in PLC-positive patients was 38 % and that in PLC-negative patients was 84 %. Both the univariate (p < 0.01) and multivariate (p = 0.045) analyses showed that the status of PLC was significantly associated with the overall survival. Eight of the 17 PLC-positive patients underwent IPC. The 2-year OS rate in the patients treated with IPC was 88 % and that of those without IPC was 44 (p = 0.04). CONCLUSION: IPC improved the postoperative survival in PLC-positive NSCLC patients, and a further prospective evaluation regarding this therapy is warranted.

Immunosuppressive effect of regulatory T lymphocytes in lung cancer, with special reference to their effects on the induction of autologous tumor-specific cytotoxic T lymphocytes.

It is not easy to induce cytotoxic T lymphocytes (CTLs) against cancer in in vitro culture. Regulatory T cells (Tregs) are considered to play a pivotal role in tumor immune escape. In this study, we analyzed the distribution of Tregs among tumor-infiltrating lymphocytes (TILs), regional lymph node lymphocytes (RLNLs) and peripheral blood lymphocytes (PBLs) in patients with lung cancer, and analyzed the effect of Tregs on the induction of CTLs in vitro. A total of 84 patients with non-small cell lung cancer underwent surgery between January 2003 and December 2004. The TILs, RLNLs and PBLs from these patients were subjected to a comparison analysis. The proportion of CD4(+)CD25(+)Foxp3(+) cells in these lymphocytes was determined by flow cytometry. The effects of Tregs on the induction of CTLs was analyzed by the depletion of Tregs in mixed lymphocyte-tumor cell culture (MLTC). The average proportions of Tregs in the TILs, RLNLs and PBLs were 10.4±9.5, 4.4±2.4 and 2.8±2.1%, respectively. The proportion of Tregs in the RLNLs was significantly higher than that in the PBLs (P<0.001); furthermore, TILs contained a larger number of Tregs than RLNLs (P=0.034). These Tregs substantially suppressed the induction of CTLs against autologous tumor cells. The depletion of Tregs in the MLTC resulted in the successful induction of CTLs. Tregs were found at a higher frequency in the TILs and RLNLs than in the PBLs in lung cancer patients. Since Tregs inhibited the induction of CTLs, the depletion of Tregs may represent a new therapeutic strategy for lung cancer patients.

An endobronchial lipoma successfully resected by high-frequency electric snare: a report of 2 cases.

Endobronchial lipoma is a rare, benign disease. When it causes chronic cough and pneumonia due to obstruction of the central airway, appropriate treatment is required. We herein report 2 cases of endobronchial lipoma successfully treated with a high-frequency electric snare through a flexible bronchoscope. Case 1, an 83-year-old man, visited a nearby hospital because of dyspnea on exertion. Chest computerized tomography revealed a tumor in the right main bronchus. He was referred to our hospital for further examination and treatment. Bronchoscopy showed a polypoid lesion in the right main bronchus. The tumor was resected by high-frequency electric snare through a flexible bronchoscope. Case 2 was an 83-year-old man who was diagnosed with pneumonia by a primary care physician on the basis of findings on chest computerized tomography. Bronchoscopy showed a polypoid lesion at the orifice of the right B6 bronchus, which caused segmental obstructive pneumonia. The tumor was bronchoscopically resected using a high-frequency electric snare and an neodymium-yttrium-aluminum-garnet laser. In both cases, the pathologic diagnosis was endobronchial lipoma.

Prognostic factors before and after recurrence of resected non-small cell lung cancer.

BACKGROUND: The post-surgical follow-up strategy in non-small cell lung cancer (NSCLC) is still controversial. Data on factors that affect the interval between surgery and recurrence or predict survival after recurrence in NSCLC patients are still limited. METHODS: From a group of 775 NSCLC patients who consecutively underwent curative surgery, 133 patients showing recurrence were retrospectively analyzed. RESULTS: Recurrence was most often seen in smokers and patients with advanced stage disease. In patients experiencing relapse, the 1- and 2-year recurrence rates were 58% and 84%, respectively. A multivariate analysis showed that patients who underwent limited surgery, had non-adenocarcinoma disease, or had metastatic lymph node involvement showed early recurrence (p-values: <0.01, 0.04, and 0.04, respectively). Among all relapsed patients, the 2-year overall survival rate after recurrence was 37%. A multivariate analysis demonstrated that patients with lymph node metastasis at the time of surgery and patients who experienced early recurrence were significantly more likely to have a shorter survival time after recurrence (hazard ratio, 1.73; p=0.03; hazard ratio, 2.56; p<0.001, respectively). CONCLUSIONS: Patients who are node-positive, show non-adenocarcinoma disease, and/or undergo limited surgery should receive careful follow-up during the first year after surgery for NSCLC. The present data provide additional information about postoperative recurrence in NSCLC patients.

Effects of hyaluronic acid and CD44 interaction on the proliferation and invasiveness of malignant pleural mesothelioma.

Hyaluronic acid (HA) has been proposed as a biochemical marker of malignant pleural mesothelioma (MPM). The present study focused on the implications of HA and CD44 interaction in the proliferation and invasiveness of MPM. The proliferation and invasive activity was evaluated in two human mesothelioma cell lines, ACC-MESO-1 and K921MSO, by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and the transwell chamber model. The knockdown of CD44 gene expression was accomplished by transfection of the cells with small interfering RNA. Flow cytometry revealed that both the ACC-MESO-1 and K921MSO cell lines highly expressed CD44. Treatment with HA enhanced the proliferation in both mesothelioma cell lines in comparison to cells without HA treatment. The treatment with HA (25 µg/ml) also significantly upregulated the invasion of both types of cells. The silencing of CD44 significantly abrogated the effect of HA treatment on the proliferation of ACC-MESO-1 cells and significantly suppressed the proliferation of K921MSO cells. HA-CD44 binding is important for the migration and proliferation of mesothelioma cells. Therefore, the HA-CD44 interaction is a potentially useful therapeutic target in MPM.

Temporal trends in the surgical outcomes of patients with breast cancer.

BACKGROUND: The incidence of breast cancer has been increasing in Japan over the past three decades, and it is the currently the most common malignancy in Japan. This study investigated the temporal trends of the surgical outcomes in patients with breast cancer. METHODS: We evaluated 543 consecutive patients who underwent breast-cancer resection between 1980 and 2009. The temporal trends in the surgical outcome and clinicopathological features were evaluated separately for the periods covering 1980 to 1989, 1990 to 1999, and 2000 to 2009. RESULTS: The number of patients who underwent resection during these three respective periods were 133, 176, and 234, respectively. All patients were women. The percentage of patients at stages 0 or 1 was 63.2%, 58.5%, and 43.6%, respectively, during the three periods. The mean diameter of tumors in each period was 38, 29, and 30 mm, respectively. The percentage of tumors with positive ER expression was 62.5%, 64.3%, and 69.7%, respectively. In terms of surgical procedures, the use of Halsted's radical mastectomy decreased during each period: from 40.6% of cases to 8.5% and then to 0.4%, while the proportion of breast-conserving therapies increased, from 0% to 12.5%, and finally to 35.9%. The postoperative 10-year survival rates during the three periods were 75.9%, 83.5%, and 84.9%, respectively. The 10-year survival rates of patients with stage II disease during the three periods were 66.2%, 75.7%, and 90.7%, respectively. The prognosis of stage III disease in the three periods also showed a tendency toward improvement, increasing from 37.8% to 64.2%, and finally to 84.5%. CONCLUSION: The survival of patients with stage II and III disease has improved during the past 30 years. Along with the recent advances in drug therapy, the surgical treatment has become less invasive, often because of drug therapy-related modifications.

Antitumor activity of human γδ T cells transducted with CD8 and with T-cell receptors of tumor-specific cytotoxic T lymphocytes.

The difficulty in the induction and preparation of a large number of autologous tumor-specific cytotoxic T lymphocytes (CTL) from individual patients is one of major problems in their application to adoptive immunotherapy. The present study tried to establish the useful antitumor effectors by using γδ T cells through tumor-specific TCRαß genes transduction, and evaluated the efficacy of their adoptive transfer in a non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice model. The TCRαß gene was cloned from the HLA-B15-restricted CTL clone specific of the Kita-Kyushu Lung Cancer antigen-1 (KK-LC-1). The cloned TCRαß as well as the CD8 gene were transduced into γδ T cells induced from peripheral blood lymphocytes (PBL). Cytotoxic T lymphocyte activity was examined using a standard 4 h (51) Cr release assay. Mice with a xenotransplanted tumor were treated with an injection of effector cells. Successful transduction of TCRαß was confirmed by the staining of KK-LC-1-specific tetramers. The γδ T cells transduced with TCRαß and CD8 showed CTL activity against the KK-LC-1-positive lung cancer cell line in a HLA B15-restricted manner. Adoptive transfer of the effector cells in a mice model resulted in marked growth suppression of KK-LC-1- and HLA-B15-positive xenotransplanted tumors. Co-transducing TCRαß and CD8 into γδ T cells yielded the same antigen-specific activity as an original CTL in vitro and in vivo. The TCRαß gene transduction into γδ T cells is a promising strategy for developing new adoptive immunotherapy.