Identification of Ppy-lineage cells as a novel origin of pancreatic ductal adenocarcinoma.

The Ppy gene encodes pancreatic polypeptide (PP) secreted by PP- or γ-cells, which are a subtype of endocrine cells localised mainly in the islet periphery. For a detailed characterisation of PP cells, we aimed to establish PP cell lines. To this end, we generated a mouse model harbouring the SV40 large T antigen (TAg) in the Rosa26 locus, which is expressed upon Ppy-promoter-mediated Cre-loxP recombination. Whereas Insulin1-CreERT-mediated TAg expression in beta cells resulted in insulinoma, surprisingly, Ppy-Cre-mediated TAg expression resulted in the malignant transformation of Ppy-lineage cells. These mice showed distorted islet structural integrity at 5 days of age compared with normal islets. CK19+ duct-like lesions contiguous with the islets were observed at 2 weeks of age, and mice developed aggressive pancreatic ductal adenocarcinoma (PDAC) at 4 weeks of age, suggesting that PDAC can originate from the islet/endocrine pancreas. This was unexpected as PDAC is believed to originate from the exocrine pancreas. RNA-sequencing analysis of Ppy-lineage islet cells from 7-day-old TAg+ mice showed a downregulation and an upregulation of endocrine and exocrine genes, respectively, in addition to the upregulation of genes and pathways associated with PDAC. These results suggest that the expression of an oncogene in Ppy-lineage cells induces a switch from endocrine cell fate to PDAC. Our findings demonstrate that Ppy-lineage cells may be an origin of PDAC and may provide novel insights into the pathogenesis of pancreatic cancer, as well as possible therapeutic strategies. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Tertiary lymphoid structures in tongue cancer: Association with clinicopathological parameters, preoperative S-1 chemotherapy response, and prognosis.

BACKGROUND: Tertiary lymphoid structures (TLSs) are observed in cancer-invasive sites of various organs, and show evidence of tumor-specific B and/or T cells, suggesting an active humoral antitumor response. The aim of this study was to evaluate the relationship between TLSs and prognosis in patients with tongue squamous cell carcinoma (TSCC) after preoperative S-1 chemotherapy. METHODS: Among 196 TSCC cases, 111 patients who received preoperative S-1 chemotherapy were compared to 85 patients who did not receive chemotherapy. We investigated the incidence of TLSs in both preoperative biopsy and resected specimens. RESULTS: TLSs were present in 24 (12%) biopsy specimens and 31 (16%) resected specimens. TLSs were associated with clinicopathologically advanced cases and positivity for lymphatic invasion. None of the cases with pStage 0 (i.e., noninvasive cancer) showed TLSs. In preoperative S-1 chemotherapy cases, TLSs were significantly more common in those treated with S-1 for more than 21 days and in those with treatment effects 0, Ia, and Ib. TLSs may not be a favorable prognostic factor by themselves but maybe a prognostic factor when combined with preoperative S-1 treatment. CONCLUSION: The presence of TLSs was suggested to be a factor indicating a favorable prognosis when considering the indication for preoperative S-1 chemotherapy. The synergistic effect of S-1 by activating antitumor immunity may be associated with a better prognosis in TSCC patients with TLSs.

Current status and prospects of artificial intelligence in breast cancer pathology: convolutional neural networks to prospective Vision Transformers.

Breast cancer is the most prevalent cancer among women, and its diagnosis requires the accurate identification and classification of histological features for effective patient management. Artificial intelligence, particularly through deep learning, represents the next frontier in cancer diagnosis and management. Notably, the use of convolutional neural networks and emerging Vision Transformers (ViT) has been reported to automate pathologists' tasks, including tumor detection and classification, in addition to improving the efficiency of pathology services. Deep learning applications have also been extended to the prediction of protein expression, molecular subtype, mutation status, therapeutic efficacy, and outcome prediction directly from hematoxylin and eosin-stained slides, bypassing the need for immunohistochemistry or genetic testing. This review explores the current status and prospects of deep learning in breast cancer diagnosis with a focus on whole-slide image analysis. Artificial intelligence applications are increasingly applied to many tasks in breast pathology ranging from disease diagnosis to outcome prediction, thus serving as valuable tools for assisting pathologists and supporting breast cancer management.

Utility of human epidermal growth factor 2 heterogeneity as a prognostic factor in triple-negative breast cancer.

In some cases of human epidermal growth factor 2 (HER2)-negative breast cancer, including triple-negative breast cancer, HER2 expression is sporadically and strongly upregulated, a condition known as HER2 heterogeneity. We investigated the clinicopathological features of patients with HER2 heterogeneity in triple-negative breast cancers treated with neoadjuvant chemotherapy. Thirty-nine patients with triple-negative breast cancer who had undergone preoperative chemotherapy participated in this study. To assess for HER2 heterogeneity, we used dual in situ hybridization slides. We evaluated the association between HER2 heterogeneity and clinicopathological factors such as rates of pathologic complete response (pCR) and of recurrence-free survival. Of the 39 patients, 15 (38.5%) had cancers with HER2 heterogeneity. The pCR rates were 13.3% among patients with HER2 heterogeneity and 20.8% among those with HER2 nonheterogeneity, but the difference was not significant. The recurrence-free survival rate was significantly lower in patients with HER2 heterogeneity than in those without (P = 0.025). HER2 heterogeneity is a significant predictor of poor prognosis in patients with triple-negative breast cancer treated with neoadjuvant chemotherapy.

Multiplexed genome editing by in vivo electroporation of Cas9 ribonucleoproteins effectively induces endometrial carcinoma in mice.

Synergistic effects among multiple gene mutations are involved in cancer development and progression. However, developing genetically modified mouse models to analyze various combinations of mutations is extremely labor-intensive and time-consuming. To address these problems, we developed a novel method for in vivo multiplexed genome editing of the murine uterus to model human endometrial carcinoma (EMC). To do this, we injected a CRISPR-Cas9 ribonucleoprotein complex into the uterine cavity of adult female mice, followed by electroporation. Evaluation of reporter mice demonstrated that genome editing occurred specifically in uterine epithelial cells, which are the origin of EMCs. Simultaneous targeting of Pten/Trp53/Lkb1, or targeting of Pten/Lkb1 along with the Ctnnb1ΔEx3 mutation, resulted in efficient generation of invasive tumors in wild-type females within 3 months. This novel method will enable rapid and easy validation of many combinations of gene mutations that lead to endometrial carcinogenesis.

Infiltration of Gastric Cancer Stroma by Tumor-Infiltrating Lymphocytes Correlates with Mechanistic Target of Rapamycin Signaling.

INTRODUCTION: We investigated whether the infiltration of tumor-infiltrating lymphocytes (TILs) in gastric cancer (GC), as evaluated by hematoxylin and eosin (H&E) staining, could be a prognostic marker. We also explored on the relationship between TILs and mechanistic target of rapamycin (mTOR) and how it regulates immune effector responses in GC. METHODS: A total of 183 patients with available data on TIL were included. TIL infiltration was evaluated using H&E staining. We also conducted immunohistochemistry to determine mTOR expression. RESULTS: Positive TIL infiltration was defined as TILs ≥20%. There were 72 (39.3%) and 111 (60.7%) positive and negative cases, respectively. TILs positivity significantly correlated with both absence of lymph node metastasis (p = 0.037) and negative p-mTOR expression (p = 0.040). TIL infiltration correlated with a significantly better overall (p = 0.046) and disease-free (p = 0.020) survival. CONCLUSION: mTOR possibly suppresses TIL infiltration in GC. H&E staining is an effective tool for evaluating the immune status of GC patients. H&E staining may be used in clinical practice to monitor treatment response in GC.

Relationship between histopathological therapeutic effect and prognosis in oral cancer patients after preoperative S-1 chemotherapy followed by surgery.

OBJECTIVES: Preoperative S-1 chemotherapy is administered to prevent tumor proliferation before surgery in oral squamous cell carcinoma (OSCC). The aim of this study was to investigate the relationship between the histological therapeutic effect and prognosis in patients with OSCC after pre-operative S-1 chemotherapy. MATERIALS AND METHODS: Among 461 OSCC cases, 281 patients who received preoperative S-1 chemotherapy were compared with 180 patients that did not receive chemotherapy to determine the histological therapeutic effect in the resected specimens and the differences in relapse-free survival. RESULTS: The histological chemotherapeutic effect was well correlated with the subsequent prognosis. In an examination of the combined effect of treatment and ypStage, the groups with good S-1 treatment effects had extremely good prognoses, even if the postoperative resection specimens were within the same ypStage. In a stratified search of patients who received S-1 for more than 7 days and who had a significantly better prognosis than those who did not receive S-1, it was found that the prognosis was significantly better for patients with tongue cancer according to site; furthermore, tongue cancer, age under 70 years of age, male sex, and clinical stage I were factors associated with a significantly better prognosis. CONCLUSIONS: Even if the postoperative resection specimens were within the same ypStage, the groups that responded to S-1 treatment were considered to have extremely good prognoses. CLINICAL RELEVANCE: A good adaptation for S-1 was tongue cancer, and especially tongue cancer with cStage I, male sex, and age less than 70 years old.

Identification of MicroRNAs Associated with Histological Grade in Early-Stage Invasive Breast Cancer.

This study aimed to identify microRNAs associated with histological grade using comprehensive microRNA analysis data obtained by next-generation sequencing from early-stage invasive breast cancer. RNA-seq data from normal breast and breast cancer samples were compared to identify candidate microRNAs with differential expression using bioinformatics. A total of 108 microRNAs were significantly differentially expressed in normal breast and breast cancer tissues. Using clinicopathological information and microRNA sequencing data of 430 patients with breast cancer from The Cancer Genome Atlas (TCGA), the differences in candidate microRNAs between low- and high-grade tumors were identified. Comparing the expression of the 108 microRNAs between low- and high-grade cases, 25 and 18 microRNAs were significantly upregulated and downregulated, respectively, in high-grade cases. Clustering analysis of the TCGA cohort using these 43 microRNAs identified two groups strongly predictive of histological grade. miR-3677 is a microRNA upregulated in high-grade breast cancer. The outcome analysis revealed that patients with high miR-3677 expression had significantly worse prognosis than those with low miR-3677 expression. This study shows that microRNAs are associated with histological grade in early-stage invasive breast cancer. These findings contribute to the elucidation of a new mechanism of breast cancer growth regulated by specific microRNAs.

Dynamic alterations of circulating T lymphocytes and the clinical response in patients with head and neck squamous cell carcinoma treated with nivolumab.

Cancer immunotherapy using immune checkpoint inhibitors (ICIs) has been recognized as a novel therapeutic option for head and neck squamous cell carcinoma (HNSCC). However, only approximately 20-30% of patients with recurrent/metastatic (R/M) HNSCC benefit. Moreover, the mechanisms underlying the response to ICIs remain unclear. We investigated the proportion, activation status, and expression level of immune checkpoint molecules in circulating T cell subsets in R/M HNSCC patients treated with nivolumab using flow cytometry and mass cytometry, and then determined whether treatment response was associated with these values. We also assessed the changes in the frequency of tumor-associated antigens, MAGE-A4 and p53, -specific T cells prior to and after nivolumab treatment using the IFN-γ ELISPOT assay. The proportion of activated CD4+ and CD8+ TEMRA cells significantly increased in the disease-controlled patients but not in disease-progressed patients. As expected, the expression of PD-1 in T cells markedly decreased regardless of the therapeutic response. Meanwhile, T cell immunoglobulin mucin-3 expression on CD8+ T cells was significantly higher in patients with disease progression than in disease-controlled patients after treatment. The frequency of the tumor-associated antigens, MAGE-A4- and p53-specific T cells, was not correlated with clinical responses; however, in the disease-controlled patients, the frequency of MAGE-A4-specific T cells was significantly augmented. We concluded that in R/M HNSCC patients treated with nivolumab, circulating T cells show dynamic alterations depending on treatment efficacy. An analysis of the immunokinetics of circulating T cells could thus provide new insights into rational therapeutic strategies in cancer immunotherapy for HNSCC.

Nuclear morphology in breast lesions: refining its assessment to improve diagnostic concordance.

AIMS: Although evaluation of nuclear morphology is important for the diagnosis and categorisation of breast lesions, the criteria used to assess nuclear atypia rely upon the subjective evaluation of several features that may result in inter- and intraobserver variation. This study aims to refine the definitions of cytonuclear features in various breast lesions. METHODS AND RESULTS: ImageJ was used to assess the nuclear morphological features including nuclear diameter, axis length, perimeter, area, circularity and roundness in 160 breast lesions comprising ductal carcinoma in situ (DCIS), invasive breast carcinoma of no special type (IBC-NST), tubular carcinoma, usual ductal hyperplasia (UDH), columnar cell change (CCC) and flat epithelial atypia (FEA). Reference cells included normal epithelial cells, red blood cells (RBCs) and lymphocytes. Reference cells showed size differences not only between normal epithelial cells and RBCs but also between RBCs in varied-sized blood vessels. Nottingham grade nuclear pleomorphism scores 1 and 3 cut-offs in IBC-NST, compared to normal epithelial cells, were < ×1.2 and > ×1.4 that of mean maximum Feret's diameter and < ×1.6 and > ×2.4 that of mean nuclear area, respectively. Nuclear morphometrics were significantly different in low-grade IBC-NST versus tubular carcinoma, low-grade DCIS versus UDH and CCC versus FEA. No differences in the nuclear features between grade-matched DCIS and IBC-NST were identified. CONCLUSION: This study provides a guide for the assessment of nuclear atypia in breast lesions, refines the comparison with reference cells and highlights the potential diagnostic value of image analysis tools in the era of digital pathology.

The frequency and clinical significance of centromere enumeration probe 17 alterations in human epidermal growth factor receptor 2 immunohistochemistry-equivocal invasive breast cancer.

BACKGROUND AND AIMS: Chromosome 17 alterations affect the assessment of HER2 gene amplification in breast cancer (BC), but its clinical significance remains unclear. This study aimed to identify the prevalence of centromere enumeration probe 17 (CEP17) alterations, and its correlation with response to neoadjuvant therapy (NAT) in BC patients with human epidermal growth factor receptor 2 (HER2) immunohistochemistry-equivocal score. METHODS AND RESULTS: A large BC cohort (n = 6049) with HER2 immunohistochemistry score 2+ and florescent in-situ hybridisation (FISH) results was included to assess the prevalence of CEP17 alterations. Another cohort (n = 885) with available clinicopathological data was used to evaluate the effect of CEP17 in the setting of NAT. HER2-amplified tumours with monosomy 17 (CEP17 copy number < 1.5 per nucleus), normal 17 (CEP17 1.5-< 3.0) and polysomy 17 (CEP17 ≥ 3.0) were observed in 16, 59 and 25%, respectively, compared with 3, 74 and 23%, respectively, in HER2-non-amplified tumours. There was no significant relationship between CEP17 alterations and pathological complete response (pCR) rate in both HER2-amplified and HER2-non-amplified tumours. The independent predictors of pCR were oestrogen (ER) negativity in HER2-amplified tumours [ER negative versus positive; odds ratio (OR) = 11.80; 95% confidence interval (CI) = 1.37-102.00; P = 0.02], and histological grade 3 in HER2 non-amplified tumours (3 versus 1, 2; OR = 5.54; 95% CI = 1.61-19.00; P = 0.007). CONCLUSION: The impacts of CEP17 alterations are not as strong as those of HER2/CEP17 ratio and HER2 copy number. The hormonal receptors status and tumour histological grade are more useful to identify BC patients with a HER2 immunohistochemistry-equivocal score who would benefit from NAT.

PROX1 Was Associated with LGR5 and Wnt Signaling and Contributed to Poor Prognosis in Gastric Cancer.

INTRODUCTION: We investigated whether the expression of prospero homeobox protein-1 (PROX1) in gastric cancer (GC) could be a prognostic marker. We also focused on the relationship between PROX1 and LGR5 and Wnt/ß-catenin activity in GC. METHODS: A total of 196 patients who underwent potentially curative surgery were collected and reviewed retrospectively. Immunohistochemistry was conducted and evaluated the expression PROX1, LGR5, Wnt3a, and ß-catenin expression. And we evaluated the relationship between PROX1 expression and clinicopathological features. RESULTS: The PROX1 low-expression group consisted of 105 patients (53.6%) and the high-expression group consisted of 91 patients (46.4%). For LGR5 expression, 76 patients (38.8%) were classified as low-expression, and 120 patients (61.2%) were classified as high-expression. The PROX1 low-expression group was significantly younger (p = 0.0095), had more intestinal type (p = 0.014), and had smaller tumor size (p = 0.013). The PROX1 high-expression group was significantly correlated with high LGR5 expression (p < 0.0001) and high Wnt3a expression (p = 0.012). In addition, there were significantly more cases of postoperative recurrence in the PROX1 high-expression group (p = 0.013). CONCLUSION: Our findings demonstrate that PROX1 correlated with the cancer stemness markers LGR5 and Wnt3a signaling in GC and had a poor prognosis including postoperative recurrence.

AKT3 is a key regulator of head and neck squamous cell carcinoma.

The phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway plays a vital role in cell proliferation, apoptosis, metabolism, and angiogenesis in various human cancers, including head and neck squamous cell carcinoma (HNSCC). In the present study, we aimed to clarify the role of AKT, which is a major downstream effector of the PI3K-AKT-mTOR pathway, in HNSCC. We first investigated the mRNA expression of AKT isoforms using RNA-sequencing data from The Cancer Genome Atlas database. We observed a specific elevation of AKT3 expression in HNSCC tissues when compared with that in normal tissues. Furthermore, AKT3 expression correlated with genes related to the immunosuppressive microenvironment more than the other AKT isoforms and PIK3CA. Accordingly, we focused on AKT3 and performed a knockdown approach using an HNSCC cell line. AKT3 knockdown cells exhibited impaired proliferation, a shift in the cell cycle from G2/M to G1/G0 phase, an increase in apoptotic cells, and downregulation of gene expression related to immunosuppression, as well as the knockdown of its upstream regulator PIK3CA. We also performed immunohistochemistry for both AKT3 and PIK3CA using surgical specimens from 72 patients with HNSCC. AKT3 expression in tumor cells correlated with immune cell infiltration and unfavorable prognosis when compared with PIK3CA. These findings suggested that AKT3 expression is a potential biomarker for predicting the immunoreactivity and prognosis of HNSCC. Furthermore, the isoform-specific inhibition of AKT3 could be developed as a novel cancer therapy that efficiently suppresses the PI3K-AKT-mTOR pathway.

High membrane expression of CMTM6 in hepatocellular carcinoma is associated with tumor recurrence.

CKLF-like MARVEL transmembrane domain-containing protein 6 (CMTM6) maintains membrane PD-L1 expression by controlling its endosomal recycling. However, in patients with hepatocellular carcinoma (HCC), the correlation among CMTM6, B7 family ligands, and CD8-positive cytotoxic T lymphocytes (CTLs), and the molecular function of CMTM6 in HCC have not been established. We performed immunohistochemistry to evaluate the relationships among CMTM6 expression, clinicopathological factors, B7 family ligands expression, and CTL infiltration in HCC samples. Moreover, we established CMTM6-knockout human HCC cell lines to evaluate the function of human CMTM6 in immune regulation and tumor viability. CMTM6 expression was positively associated with membrane B7 family ligands expression and CTL infiltration in HCC samples. High CMTM6 expression in HCC tissues was associated with the expression of the proliferation marker Ki-67 and shorter recurrence-free survival. In vitro analysis showed the downregulation of membrane B7 family ligands and proliferation potency in the CMTM6-knockout human HCC cell line. High membrane CMTM6 expression was associated with tumor recurrence and proliferation via the regulation of membranous B7 family ligands expression. Thus, CMTM6 might be a biomarker to predict the risk of HCC recurrence and a therapeutic target to suppress tumor growth and increase CTL activity.

Immunological features of circulating monocyte subsets in patients with squamous cell carcinoma of the head and neck.

BACKGROUND: Circulating monocytes are classified into three subsets according to their CD14 and CD16 expressions. Here we investigated all three subsets in patients with squamous cell carcinoma of the head and neck (SCCHN). METHODS: Peripheral blood from 54 patients with SCCHN and 24 healthy donors (HDs) was tested for flowcytometry. Immunohistochemical staining of the primary tumor was performed. SCCHN cells were co-cultured with human monocytes in vitro. RESULTS: The level of intermediate monocytes was significantly lower in SCCHN than in HDs. The expression levels of HLA-G, PD-L1, and CD51 on intermediate monocytes was evidently greater in patients with SCCHN. In vitro co-culturing of SCCHN cells with monocytes revealed a significant increase in CD51 expression levels on monocytes. The decrease in expression levels of the maturation markers CX3CR1 and CD68 was significantly correlated to poor clinical outcomes. CONCLUSION: The level of intermediate monocytes was decreased in cancer patients in favor of immature and expressed immunosuppressive molecules.

Predictors of pathological complete response to neoadjuvant treatment and changes to post-neoadjuvant HER2 status in HER2-positive invasive breast cancer.

The response of human epidermal growth factor receptor2 (HER2)- positive breast cancer (BC) patients to anti-HER2 targeted therapy is significant. However, the response is not uniform and a proportion of HER2-positive patients do not respond. This study aims to identify predictors of response in the neoadjuvant treatment and to assess the discordance rate of HER2 status between pre- and post-treatment specimens in HER2-positive BC patients. The study group comprised 500 BC patients treated with neoadjuvant chemotherapy (NACT) and/or neoadjuvant anti-HER2 therapy and surgery who had tumours that were 3+ or 2+ with HER2 immunohistochemistry (IHC). HER2 IHC 2+ tumours were classified into five groups by fluorescence in situ hybridisation (FISH) according to the 2018 ASCO/CAP guidelines of which Groups 1, 2 and 3 were considered HER2 amplified. Pathological complete response (pCR) was more frequent in HER2 IHC 3+ tumours than in HER2 IHC 2+/HER2 amplified tumours, when either in receipt of NACT alone (38% versus 13%; p = 0.22) or neoadjuvant anti-HER2 therapy (52% versus 20%; p < 0.001). Multivariate logistic regression analysis showed that HER2 IHC 3+ and histological grade 3 were independent predictors of pCR following neoadjuvant anti-HER2 therapy. In the HER2 IHC 2+/HER2 amplified tumours or ASCO/CAP FISH Group 1 alone, ER-negativity was an independent predictor of pCR following NACT and/or neoadjuvant anti-HER2 therapy. In the current study, 22% of HER2-positive tumours became HER2-negative by IHC and FISH following neoadjuvant treatment, the majority (74%) HER2 IHC 2+/HER2 amplified tumours. Repeat HER2 testing after neoadjuvant treatment should therefore be considered.

High L-Type Amino Acid Transporter 1 Levels Are Associated with Chemotherapeutic Resistance in Gastric Cancer Patients.

INTRODUCTION: We investigated whether the expression of L-type amino acid transporter 1 (LAT-1) in clinical gastric cancer (GC) patients could predict patient therapeutic response to postoperative adjuvant chemotherapy. METHODS: Immunohistochemistry was used to investigate LAT-1, CD98, and phosphorylated-mammalian target of rapamycin (p-mTOR) expression in 111 GC patients. To clarify whether LAT-1 influences the therapeutic effects of chemotherapy, the correlation between disease-free survival rates and LAT-1 was determined in 2 groups: 59 patients who did not undergo postoperative adjuvant chemotherapy and 52 patients who did undergo postoperative adjuvant chemotherapy. RESULTS: LAT-1 was significantly correlated with CD98 and p-mTOR expressions. We did not find any statistically significant correlation between LAT-1 and recurrence in the nontreated group. In contrast, a significant association was found between LAT-1 expression and disease-free survival in the chemotherapy group. Moreover, multivariate regression analysis demonstrated that LAT-1 was an independent predictor of disease-free survival in the postoperative adjuvant chemotherapy group (p = 0.012). CONCLUSION: Our findings demonstrate that LAT-1 is a useful predictive marker for a successful postoperative adjuvant chemotherapy treatment.

Role of PD-L1 Expression during the Progression of Submucosal Gastric Cancer.

INTRODUCTION: Programmed death-ligand 1 (PD-L1) expression is a prognostic marker for gastric cancer that correlates with tumor diameter and depth of penetration. But the role of PD-L1 and mechanism(s) employed in the initial phase of invasion in early gastric cancer is yet to be understood. OBJECTIVE: This study aims to elucidate the role of PD-L1 during the progression of gastric cancer, specifically invading the submucosa beyond the lamina muscularis mucosa. METHODS: Using 107 patients with pathological submucosal gastric cancer, we determined the expression of PD-L1 based on the staining of the cell membrane or cytoplasm of tumor cells in the central and invasive front of the tumor. Samples were categorized into 3 groups based on the intensity of PD-L1 expression. CD8+ lymphocytes expressing PD-1 and CD163+ macrophages were used to determine the number of cell nuclei at the invasive front, similar to PD-L1. CMTM6 levels were determined and used to stratify samples into 3 groups. RESULTS: PD-L1 expression was higher in the invasive front (26.2%) than in the central portion of the tumors (7.4%; p < 0.001). Moreover, lymphatic and vascular invasion were more frequently observed in samples with high levels of PD-L1 (lymphatic invasion: 60.7 vs. 35.4%, p = 0.0026, and vascular invasion: 39.3 vs. 16.5%, p = 0.0018). There was no correlation between PD-L1 expression and the levels of PD-1, CD8, CD163, and CMTM6. CONCLUSIONS: PD-L1-expressing cancer cells at the invasive front of gastric cancer influence the initial stages of tumor invasion and lymphovascular permeation in early-stage gastric cancers. Immune checkpoint signaling may be the driving force in the invasive front during the invasion of the submucosa beyond the lamina muscularis mucosa.

In vitro assessment of antitumor immune responses using tumor antigen proteins produced by transgenic silkworms.

The evaluation of antitumor immune responses is essential for immune monitoring to predict clinical outcomes as well as treatment efficacies in cancer patients. In this study, we produced two tumor antigen (TA) proteins, melanoma antigen family A4 and wild type p53, using TG silkworm systems and evaluated anti-TA-specific immune responses by enzyme-linked immunosorbent spot assays in patients with head and neck cancer. Eleven (61.1%) of 18 patients showed significant IFN-γ production in response to at least one TA; however, the presence of TA-specific immune responses did not significantly contribute to better prognosis (overall survival, p = 0.1768; progression-free survival, p = 0.4507). Further studies will need to be performed on a larger scale to better assess the clinical significance of these systems. The production of multiple TA proteins may provide new avenues for the development of immunotherapeutic strategies to stimulate a potent and specific immune response against tumor cells as well as precise assessment of antitumor immune responses in cancer patients.

Diagnostic value of 18F-FDG-PET to predict the tumour immune status defined by tumoural PD-L1 and CD8+tumour-infiltrating lymphocytes in oral squamous cell carcinoma.

BACKGROUND: Lately, immune checkpoint proteins, such as programmed death 1 (PD-1) and its ligand-1 (PD-L1), have garnered attention as a new target in oral squamous cell carcinoma (OSCC). Reportedly, fluoro-D-glucose (FDG)-uptake alteration by anti-PD-1 antibody treatment depicts the response in patients with lung cancer. This study aims to elucidate the correlations between tumour immune status, clinicopathological factors, 18F-FDG-uptake and cold tumour phenotypes as low PD-L1 expression/low CD8+tumour-infiltrating lymphocytes (TILs) in OSCC. METHODS: We performed immunohistochemical analysis of PD-L1, hypoxia-inducible factor 1 A (HIF-1A), glucose transporter type 1 (GLUT1), CD8, E-cadherin and Ki-67 on 59 operable OSCC samples. We assessed the correlations between these factors and preoperative 18F-FDG-uptake, clinicopathological characteristics and prognosis. RESULTS: Low expression of PD-L1 in OSCC correlated with cancer aggressiveness, poor prognosis, high 18F-FDG-uptake with HIF-1A/GLUT1 and low E-cadherin expression and low CD8. Cold tumour phenotypes as low PD-L1 tumour cells and low stromal CD8 correlated with the poor prognosis, high 18F-FDG-uptake and E-cadherin suppression. Furthermore, the high level of preoperative 18F-FDG-uptake in OSCC was an independent predictor of the cold tumour immune status. CONCLUSIONS: 18F-FDG-uptake is an independent predictor of cold tumour in OSCC. 18F-FDG-PET imaging could be a promising diagnostic tool to estimate tumour immune status.