Background & Aims Some patients develop multiple squamous cell carcinomas (SCCs) in the upper aerodigestive tract, attributed to field cancerization; alcohol consumption has been associated with this ...process. We examined the association between multiple areas of dysplastic squamous epithelium with the development of SCC of the esophagus or head and neck cancer, as well as alcohol consumption and smoking. Methods We examined 331 patients with early stage esophageal SCC using Lugol chromoendoscopy to evaluate the dysplastic squamous epithelium in the esophagus. Patients then were assigned to 3 groups, based on the number of Lugol-voiding lesions: A, no lesion; B, 1–9 lesions; or C, 10 or more lesions. Participants completed lifestyle surveys on their history of drinking, smoking, and diet. All participants were evaluated by laryngopharyngoscopy before registration; only those without head and neck cancer were included, except for patients with superficial SCC limited to the subepithelial layer. Lesions detected in the esophagus and head and neck by surveillance were considered to be metachronous. The study end point was the cumulative incidence of metachronous SCCs in the esophagus and head and neck after endoscopic resection of esophageal SCC, according to the grade of Lugol-voiding lesions. At study entry, all patients were instructed to abstain from alcohol and smoking. Results Over the 2-year study period, metachronous SCCs of the esophagus were detected in 4% of patients in group A, in 9.4% of patients in group B, and in 24.7% of patients in group C ( P < .0001 for patients in group A vs B or B vs C). Head and neck SCCs were detected in none of the patients in group A, in 1.7% of the patients in group B, and in 8.6% of the patients in group C ( P = .016 for patients in group A vs C and P = .008 for patients in group B vs C). SCC of the esophagus or head and neck developed in 4.0% of patients in group A, in 10.0% of patients in group B, and in 31.4% of patients in group C ( P < .0001 for group A vs B or A vs C). Alcohol abstinence decreased the risk of multiple SCCs of the esophagus (adjusted hazard ratio, 0.47, 95% confidence interval, 0.25–0.91; P = .025), whereas smoking abstinence did not. Conclusions Multiple dysplastic lesions in the esophagus increase the risk of multiple SCCs. Alcohol abstinence reduces the risk of metachronous SCCs. Clinical Trials registry: UMIN000001676 and UMIN000005466.
CD133 has been reported to be a cancer‐initiating cell marker in colorectal carcinoma. The objective of this study was to evaluate the frequency of CD133 expression in colorectal cancer, the ...distribution of CD133‐positive cancer cells, and their relationship to clinicopathological features, including survival. An immunohistochemical examination of CD133 expression and a clinicopathological analysis were performed in the 189 consecutive colorectal cancer patients. CD133 expression was seen at the luminal surface of cancer glands mainly with cribriform features. Expression was detected in only 29 of the 189 tumors (15.3%). Of these, 21 tumors (11.1%) showed CD133 overexpression. All 21 tumors with CD133 overexpression were diagnosed as well‐ or moderately‐differentiated adenocarcinoma. There was no difference in the distribution of CD133 expressing cells between the invasive area and surface area. Although there was no difference in recurrence‐free survival between patients with CD133 overexpression and without, the patients with CD133 overexpression had significantly poorer overall survival (P = 0.03). CD133 overexpression is a risk factor for poorer overall survival in patients with well‐ and moderately‐differentiated adenocarcinoma. Expression of this cancer‐initiating cell marker may vary with the histological type of the cancer, and further investigation of the relationship between its expression and clinicopathological features may be necessary. (Cancer Sci 2008; 99: 1578–1583)
Background
Cancer progression following chemotherapy is a significant barrier to effective cancer treatment. We aimed to evaluate the role of drug-exposed cancer-associated fibroblasts (CAFs) in the ...growth and progression of drug-exposed gastric cancer (GC) cells and to explore the underlying molecular mechanism.
Methods
The human GC cell line 44As3 and CAFs were treated with 5-fluorouracil and oxaliplatin (5FU + OX). 5FU + OX-pretreated 44As3 cells were then cultured in a conditioned medium (CM) from 5FU + OX-pretreated CAFs, and the growth and migration/invasion ability of the cells were evaluated. We also compared the clinicopathological characteristics of the GC patients treated with S1 + OX in accordance with the properties of their resected specimens, focusing on the number of CAFs. Changes in gene expression in CAFs and 44As3 cells were comprehensively analyzed using RNA-seq analysis.
Results
The CM from 5FU + OX-pretreated CAFs promoted the migration and invasion of 5FU + OX-pretreated 44As3 cells. Although the number of cases was relatively small (
n
= 21), the frequency of positive cases of lymphovascular invasion and the recurrence rate were significantly higher in those with more residual CAF. RNA-seq analysis revealed 5FU + OX-pretreated CAF-derived glycoprotein 130 (gp130) as a candidate factor contributing to the increased migration of 5FU + OX-pretreated 44As3 cells. Administration of the gp130 inhibitor SC144 prevented the increased migration ability of 5FU + OX-pretreated 44As3 cells owing to drug-treated CAFs.
Conclusions
Our findings provide evidence regarding the interactions between GC cells and CAFs in the tumor microenvironment following chemotherapy, suggesting that ligands for gp130 may be novel therapeutic targets for suppressing or preventing metastasis in GC.
•Podoplanin (PDPN)+ cancer associated fibroblasts (CAFs) promotes cancer progression in non-small cell lung cancer.•PDPN+ CAFs showed higher expression of transforming growth factor-β1 (TGFB1) ...gene.•PDPN+ CAFs were associated with CD204+ tumor-associated macrophage (TAM) infiltration.•PDPN + CAFs might have the connection with the immunosuppressive tumor microenvironment.
Cancer-associated fibroblasts (CAFs) expressing podoplanin (PDPN) harbor a fibrous tumor microenvironment that promotes cancer progression in lung adenocarcinoma. In this study, we investigated whether tumor-promoting PDPN+ CAFs contribute to the immunosuppressive microenvironment in lung squamous cell carcinoma (SqCC).
The gene expression profiles of immunosuppressive cytokines were compared using The Cancer Genome Atlas (TCGA) microarray lung SqCC data (n = 484) between a PDPN-high group and a PDPN-low group. Further, using patient-derived CAFs from surgically resected lung SqCC, the PDPN+ fraction was sorted and gene and protein expressions were analyzed. Finally, immunohistochemical staining was conducted on 131 surgically resected lung SqCC; CD8+ and FOXP3+ tumor infiltrating lymphocytes (TILs), and CD204+ tumor-associated macrophages (TAMs) were evaluated in cases with PDPN+ and PDPN− CAFs.
Analysis of TCGA database revealed that the PDPN-high group exhibited significantly higher expression of interleukin (IL)-1A, IL-1B, IL-6, IL-10, monocyte chemoattractant protein-1 (CCL2), colony stimulating factor 1 (CSF1), fibroblast growth factor 2 (FGF2), galectin 1 (LGALS1), platelet derived growth factor subunit A (PDGFA), PDGFB, and transforming growth factor-β1 (TGFB1) than those in the PDPN-low group. Among them, it was found that TGFB1 expression was higher in patient-derived PDPN+ CAFs. Immunohistochemical analyses revealed that more CD204+ TAMs infiltrated the tumor tissues in cases with PDPN+ CAFs than in cases with PDPN− CAFs (P < 0.03), while CD8+ and FOXP3+ TILs did not. Furthermore, in the same tumor, CD204+ TAMs infiltrated more in PDPN+ CAF-rich areas (P = 0.005).
PDPN+ CAFs showed higher expression of TGFB1 and were associated with CD204+ TAM infiltration in stage-I lung SqCC, suggesting that PDPN+ CAFs were associated with the immunosuppressive tumor microenvironment.
Immunotherapy with anti-PD-1 antibody preliminarily showed promising efficacy for treating esophageal squamous cell carcinoma (ESCC). Herein, we used tissue microarrays and immunohistochemically ...analyzed PD-L1 and various tumor infiltrating immune cells (TIICs) in specimens from 196 ESCC patients who had undergone curative resection without preoperative therapy. PD-L1 expressions in tumor cells (TCs) and TIICs, as well as infiltration of lymphocytes (CD4+, CD8+, FOXP3+, and PD- 1+) and macrophages (CD68+ and CD204+), were evaluated. PD-L1 was expressed in TCs of 18.4% and in TIICs of 83.3% of these patients. PD-L1 expressions in TCs and TIICs were associated with significant infiltration of various TIIC types, especially CD8+ cells. PD-L1 expressions in both TCs and TIICs were significantly associated with favorable overall survival, and combining their levels enhanced prognostic accuracy. Prognostic impacts of PD-L1 expressions in TCs and TIICs, abundant PD-1+ cell infiltration, a high CD8+/FOXP3+ ratio, and the CD8+/CD204+ ratio remained significant after adjusting for clinicopathological factors. In conclusion, PD-L1 expression reflects anti-tumor immunity, and PD-1/PD-L1 expression and the ratio of infiltrating effector to immune suppressor cells have prognostic value. Therapeutic strategies inhibiting the PD-1/PD-L1 signal and immune suppressor cells are anticipated in ESCC patients.
Collagen type I (Col I) is one of the major extracellular matrix proteins in the cancer tissue. Previously, we have reported that Col I induces epidermal growth factor receptor tyrosine kinase ...inhibitor (EGFR-TKI) resistance by mTOR activation through Akt and ERK1/2 independent pathway. In this study, we aimed to elucidate the molecular mechanism of Col I induced EGFR-TKI resistance. First, we demonstrated the uptake of fluorescently labeled Col I by EGFR-mutated lung cancer cell line PC-9 cells using confocal microscopy and flow cytometry. Metabolome analysis revealed that the metabolic profiles of PC-9 cells was influenced by Col I treatment. Uptake of Col I into PC-9 cells was not inhibited by MMP inhibitor, GM6001, and endocytosis inhibitors, Pitstop2 and Dyngo4a; however, macropinocytosis inhibitor EIPA prevented its uptake. Moreover, the combination of EIPA and EGFR-TKI abrogated Col I-induced EGFR-TKI resistance in PC-9 cells. Inhibition of Rac1, which is essential for micropinocytosis, also decreased the uptake of Col I in PC-9 cells and restored their sensitivity to EGFR-TKI. Thus, EGFR mutated lung cancer cells could develop EGFR-TKI resistance by Col I uptake by macropinocytosis route.
•Collagen type I induced mTOR activation and subsequent drug resistance in EGFR-mutated cancer cells.•Uptake of Collagen type I is via macropinocytosis.•Collagen type I uptake caused metabolic changes in cancer cells.•Combination of macropinocytosis inhibitor and an anticancer drug could be an effective strategy to combat drug resistance induced by collagen type I.
More than 85% of patients with T1 colorectal cancer have no lymph node metastasis and can be cured by endoscopic resection. To avoid unnecessary surgery after complete endoscopic resection, accurate ...histologic methods for evaluating resected specimens are needed to discriminate those at high risk for lymph node metastasis. A retrospective multi-institutional, cross-sectional study of 806 T1 colorectal cancer patients was conducted. A budding/sprouting score was incorporated for predicting lymph node metastasis in addition to other parameters, including the depth of submucosal invasion, histologic grade, and lymphovascular invasion. Lymph node metastasis was detected in 97 patients. Independent predictors of lymph node metastasis by multivariate analysis were depth of submucosal invasion ≥1000 μm (odds ratio (95% confidence interval)=5.56 (2.14-19.10)) and high-grade budding/sprouting (3.14 (1.91-5.21)). Among lesions with a depth of submucosal invasion ≥1000 μm, lymph node metastasis was detected in 59 (29%) of 207 patients with high-grade budding/sprouting, and in 34 (9%) of 396 with low-grade budding/sprouting. Lymph node metastasis was detected in only 4 (2%) of 203 lesions with a depth of submucosal invasion <1000 μm. Of these four tumors, three invaded lymphatic and/or venous vessels. Thus, the risk for lymph node metastasis can be classified into three groups: high risk with a depth of submucosal invasion ≥1000 μm and high-grade budding/sprouting, intermediate-risk with a depth of submucosal invasion ≥1000 μm and low-grade budding/sprouting, and low-risk with a depth of submucosal invasion <1000 μm. These findings revealed that a depth of submucosal invasion ≥1000 μm and high-grade budding/sprouting are powerful predictive parameters for lymph node metastasis in T1 colorectal cancer. This three-tier risk classification system will facilitate the decision for additional major surgery for T1 colorectal cancer patients after successful endoscopic treatment.
Several types of cancer cells, including colorectal cancer-derived cell lines, show austerity, the resistance to nutrient starvation, but exactly how cancer cells obtain energy sources under ...conditions in which their external nutrient supply is extremely limited remains to be clarified. Because autophagy is a catabolic process by which cells supply amino acids from self-digested organelles, cancer cells are likely to use autophagy to obtain amino acids as alternative energy sources. Amino acid deprivation-induced autophagy was assessed in DLD-1 and other colorectal cancer-derived cell lines. The autophagosome-incorporated LC3-II protein level increased after treatment with a combination of autolysosome inhibitors, which interferes with the consumption of autophagosomes. Autophagosome formation was also morphologically confirmed using ectopically expressed green fluorescent protein-LC3 fusion proteins in DLD-1 and SW480 cells. These data suggest that autophagosomes were actively produced and promptly consumed in colorectal cancer cells under nutrient starvation. Autolysosome inhibitors and 3-methyl adenine, which suppresses autophagosome formation, remarkably enhanced apoptosis under amino acid-deprived and glucose-deprived condition. Similar results were obtained in the cells with decreased ATG7 level by the RNA interference. These data suggest that autophagy is pivotal for the survival of colorectal cancer cells that have acquired austerity. Furthermore, autophagosome formation was seen only in the tumor cells but not in the adjacent noncancerous epithelial cells of colorectal cancer specimens. Taken together, autophagy is activated in colorectal cancers in vitro and in vivo, and autophagy may contribute to the survival of the cancer cells in their microenvironment.
During the metastatic process, cancer cells interact with vascular adventitial fibroblasts (VAF), which are the main components of the outermost connective tissue layer of blood vessels. This ...activity suggests the presence of a specific tumor microenvironment in the perivascular area. The s.c. coinjection of human lung adenocarcinoma cell lines (A549, PC-14, and CRL-5807) and human VAF (hVAF) resulted in a high rate of tumor formation, compared with the coinjection of these cell lines and human lung tissue-derived fibroblasts (hLF). A cDNA microarray analysis revealed a higher expression level of podoplanin in hVAFs than in hLFs (4.7-fold). Flow cytometry analysis also showed a higher expression level of podoplanin in hVAFs (43% ± 17.5%) than in hLFs (16% ± 10.3%). Sorted podoplanin-positive hVAFs displayed enhanced tumor formation, lymph node metastasis, and lung metastasis of A549 compared to sorted podoplanin-negative hVAFs. Knockdown of podoplanin in hVAFs decreased the augmenting effect of tumor formation and in vitro colony formation. The overexpression of podoplanin in hVAFs hastened the tumor formation of A549, compared with control hVAFs. Furthermore, the analysis of small-sized human lung adenocarcinoma (n = 112) revealed that patients with podoplanin-positive cancer-associated fibroblasts had a significantly higher rate of lymph node metastasis and a high risk of recurrence. These results indicate a promotive effect of hVAFs mediated by podoplanin on cancer progression and suggest that the perivascular environment may constitute a specific niche for tumor progression.
Cell line‐derived xenograft (CDX) models created by implanting cancer cell lines into immunodeficient mice have contributed largely to the development of cancer drug therapies. However, cell lines ...often lose their original biological characteristics through many passages and cancer tissues in CDX models have many cancer cells and few cancer stromal cells, therefore CDX models are currently considered not suitable for predicting the results of clinical studies. Conversely, patient‐derived xenograft (PDX) models are gaining importance, as human cancer biological characteristics and microenvironments are recreated by implanting tumor tissue into immunodeficient mice. These highly expected, evidently beneficial PDX models have been used in some basic research and are becoming more generalized. However, quality control and quality assurance criteria have not been established for them, and challenges and problems in the utilization of valuable PDX models in drug development have yet to be clarified. In this report, we conducted a questionnaire survey among researchers in Japanese academic institutions and pharmaceutical companies to understand the current status of PDX models in Japan. Based on the questionnaire results, we summarized the situations surrounding respondent's utilization and quality control in the development of anticancer drugs and proposed several measures to facilitate the utilization of PDX models in the development of anticancer drugs.
We conducted a questionnaire survey among researchers in Japanese academic institutions and pharmaceutical companies to understand the current status of PDX models in Japan. Based on the results of the survey, we proposed several measures to facilitate the utilization of PDX models in the development of anticancer drugs. It is hoped that this report provides information to undertake research using PDX models for the development of anticancer drugs.
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