Association between specific human leukocyte antigen alleles and development of thyroid immune-related adverse event.

Aim: Inherent variations in human leukocyte antigen (HLA) alleles have been revealed epidemiologically to influence the development of autoimmune diseases. HLA alleles may thus also be associated with the development of immune-related adverse events (irAEs), such as thyroid irAE. Materials & methods: In this case-control study, 71 cancer patients who received immune checkpoint inhibitors were enrolled and HLA-genotyped and the frequency of HLA alleles was compared. Results: A*26:01, DPA1*01:03 and DPB1*02:01 were significantly more frequent in patients with thyroid irAE than in patients without any irAEs (35.0 vs 3.2% [p = 0.004], 80.0 vs 45.2% [p = 0.020] and 55.0 vs 25.8% [p = 0.044], respectively). Conclusion: A*26:01, DPA1*01:03 and DPB1*02:01 appear to be associated with thyroid irAE.

Everyone has a unique combination of human leukocyte antigens (HLAs) in their body that help the immune system identify threats. HLAs were named from the fact that they were first identified on the surface of human leukocytes. Afterward, HLAs were also found on all human cells. HLAs present antigens to immune cells. These HLAs also influence how the immune system attacks cancer cells. Immune checkpoint inhibitors are drugs that can help the immune system fight cancer, but they sometimes cause severe adverse events. In this study, we investigated whether specific HLA genes are related to the development of an adverse event that affects the thyroid in cancer patients treated with immune checkpoint inhibitors. We found an association between three HLA genes (A*26:01, DPA1*01:03 and DPB1*02:01) and the development of the thyroid adverse event. However, larger studies are needed to confirm and generalize these initial exploratory findings.

Prevalence of pathogenic germline variants in the circulating tumor DNA testing.

BACKGROUND: Somatic and germline variants are not distinguishable by circulating tumor DNA (ctDNA) testing without analyzing non-tumor samples. Although confirmatory germline testing is clinically relevant, the criteria for selecting presumed germline variants have not been established in ctDNA testing. In the present study, we aimed to evaluate the prevalence of pathogenic germline variants in clinical ctDNA testing through their variant allele fractions (VAFs). METHODS: A total of consecutive 106 patients with advanced solid tumors who underwent ctDNA testing (Guardant360®) between January 2018 and March 2020 were eligible for this study. To verify the origin of pathogenic variants reported in ctDNA testing, germline sequencing was performed using peripheral blood DNA samples archived in the Clinical Bioresource Center in Kyoto University Hospital (Kyoto, Japan) under clinical research settings. RESULTS: Among 223 pathogenic variants reported in ctDNA testing, the median VAF was 0.9% (0.02-81.8%), and 88 variants with ≥ 1% VAFs were analyzed in germline sequencing. Among 25 variants with ≥ 30% VAFs, seven were found in peripheral blood DNA (BRCA2: n = 6, JAK2: n = 1). In contrast, among the 63 variants with VAFs ranging from 1 to < 30%, only one variant was found in peripheral blood DNA (TP53: n = 1). Eventually, this variant with 15.6% VAF was defined to be an acquired variant, because its allelic distribution did not completely link to those of neighboring germline polymorphisms. CONCLUSION: Our current study demonstrated that VAFs values are helpful for selecting presumed germline variants in clinical ctDNA testing.

A Case of Squamous Cell Carcinoma of Unknown Primary that Responded to the Multi-Tyrosine Kinase Inhibitor Lenvatinib.

Lenvatinib is an oral tyrosine kinase inhibitor of vascular endothelial growth factor receptors 1, 2, and 3, fibroblast growth factor receptors 1 through 4, as well as platelet-derived growth factor receptor α, RET, and KIT. At present, lenvatinib is used in the treatment of thyroid cancer and renal cell carcinoma. We herein report a case of a 67-year-old patient with squamous cell carcinoma of unknown primary who was effectively treated with lenvatinib. The patient was initially diagnosed as having undifferentiated thyroid cancer, and after total thyroidectomy and bilateral lymph node dissection, lenvatinib was administered for the treatment of residual lymph node metastasis. A computed tomography scan after 1 month of lenvatinib administration showed marked regression of the lymph nodes, but interstitial pneumonia was also detected. Because the drug lymphocyte stimulation test for lenvatinib was strongly positive, we concluded that the interstitial pneumonia was induced by lenvatinib. The interstitial pneumonia only improved by the withdrawal of lenvatinib. Finally, his thyroid tumor was diagnosed as a metastasis of squamous cell carcinoma; however, we were unable to identify the primary lesion. This is the first reported case of interstitial pneumonia induced by lenvatinib.

How to adjust endoscopic findings to histopathological findings of the stomach: a "histopathology-oriented" correspondence method helps to understand endoscopic findings.

BACKGROUND: Endoscopic findings have now become nearly as detailed as histopathological findings. Thus, one-to-one correspondence and precise feedback of histopathological findings is very desirable but may be very difficult to accomplish. We describe a systematic process called the Kyoto One-to-One Correspondence Method (the KOTO Method) that allows detailed adjustments of endoscopic findings to match histopathological findings. METHODS AND RESULTS: By comparing endoscopic and stereoscopic images of the gastric mucosa, we could obtain one-to-one correspondence between endoscopic images and equivalent histology in 44 of 47 fields. CONCLUSIONS: The histological structure of gastric cancers of the same histological subtype may not be similar. One-to-one correspondence between endoscopic images and gastric mucosal histology (histopathology-oriented correspondence) will improve endoscopic diagnosis and provide more useful information for pathological diagnosis.

Magnifying Endoscopy with Blue Laser Imaging Improves the Microstructure Visualization in Early Gastric Cancer: Comparison of Magnifying Endoscopy with Narrow-Band Imaging.

BACKGROUNDS: Magnifying endoscopy with blue laser imaging (ME-BLI) for diagnosis of early gastric cancer (EGC) is as effective as magnifying endoscopy with narrow-band imaging (ME-NBI). However, there are different EGCs in microstructure visualization between ME-BLI and ME-NBI. This study aimed to clarify the pathological features of the EGCs, in which microstructure visualization was different between ME-NBI and ME-BLI. METHODS: EGCs were classified into groups A (irregular microsurface pattern (MSP) in ME-BLI and absent MSP in ME-NBI), B (irregular MSP in two modalities), or C (absent MSP in two modalities), according to the vessel plus surface classification. We compared the pathological features of EGCs between the three groups. RESULTS: 17, four, and five lesions could be evaluated in detail in groups A, B and C, respectively. Well-differentiated adenocarcinomas with shallow crypts were more frequent in group A than in group B (58.8 and 0%, resp.). The mean crypt depth of group A was significantly shallower than that of group B (56 ± 20, 265 ± 64 µm, resp., P = 0.0002). CONCLUSIONS: ME-BLI could better visualize the microstructures of the EGCs with shallow crypts compared with ME-NBI. Therefore, ME-BLI could enable a more accurate diagnosis of EGC with shallow crypts.

Diagnostic ability of magnifying endoscopy with blue laser imaging for early gastric cancer: a prospective study.

BACKGROUND: Blue laser imaging (BLI) is a new image-enhanced endoscopy technique that utilizes a laser light source developed for narrow-band light observation. The aim of this study was to evaluate the usefulness of BLI for the diagnosis of early gastric cancer. METHODS: This single center prospective study analyzed 530 patients. The patients were examined with both conventional endoscopy with white-light imaging (C-WLI) and magnifying endoscopy with BLI (M-BLI) at Kyoto Prefectural University of Medicine between November 2012 and March 2015. The diagnostic criteria for gastric cancer using M-BLI included an irregular microvascular pattern and/or irregular microsurface pattern, with a demarcation line according to the vessel plus surface classification system. Biopsies of the lesions were taken after C-WLI and M-BLI observation. The primary end point of this study was to compare the diagnostic performance between C-WLI and M-BLI. RESULTS: We analyzed 127 detected lesions (32 cancers and 95 non-cancers). The accuracy, sensitivity, and specificity of M-BLI diagnoses were 92.1, 93.8, and 91.6 %, respectively. On the other hand, the accuracy, sensitivity, and specificity of C-WLI diagnoses were 71.7, 46.9, and 80.0 %, respectively. CONCLUSIONS: M-BLI had improved diagnostic performance for early gastric cancer compared with C-WLI. These results suggested that the diagnostic effectiveness of M-BLI is similar to that of magnifying endoscopy with narrow-band imaging (M-NBI).

Blue Laser Imaging-Bright Improves Endoscopic Recognition of Superficial Esophageal Squamous Cell Carcinoma.

Background/Aims. The aim of this study was to evaluate the endoscopic recognition of esophageal squamous cell carcinoma (ESCC) using four different methods (Olympus white light imaging (O-WLI), Fujifilm white light imaging (F-WLI), narrow band imaging (NBI), and blue laser imaging- (BLI-) bright). Methods. We retrospectively analyzed 25 superficial ESCCs that had been examined using the four different methods. Subjective evaluation was provided by three endoscopists as a ranking score (RS) of each image based on the ease of detection of the cancerous area. For the objective evaluation we calculated the color difference scores (CDS) between the cancerous and noncancerous areas with each of the four methods. Results. There was no difference between the mean RS of O-WLI and F-WLI. The mean RS of NBI was significantly higher than that of O-WLI and that of BLI-bright was significantly higher than that of F-WLI. Moreover, the mean RS of BLI-bright was significantly higher than that of NBI. Furthermore, in the objective evaluation, the mean CDS of BLI-bright was significantly higher than that of O-WLI, F-WLI, and NBI. Conclusion. The recognition of superficial ESCC using BLI-bright was more efficacious than the other methods tested both subjectively and objectively.

Linked color imaging improves endoscopic diagnosis of active Helicobacter pylori infection.

BACKGROUND AND STUDY AIMS: Linked color imaging (LCI) is a new image-enhanced endoscopy technique using a laser light source to enhance slight differences in mucosal color. The aim of this study was to compare the usefulness of LCI and conventional white light imaging (WLI) endoscopy for diagnosing Helicobacter pylori (H. pylori). PATIENTS AND METHODS: We retrospectively analyzed images from 60 patients examined with WLI and LCI endoscopy between October 2013 and May 2014. Thirty patients had H. pylori infections, and other thirty patients tested negative for H. pylori after eradication therapy. Four endoscopists evaluated the 2 types of images to determine which was better at facilitating a diagnosis of H. pylori infection. RESULTS: H. pylori infection was identified with LCI by enhancing the red appearance of the fundic gland mucosa. The accuracy, sensitivity, and specificity for diagnosing H. pylori infection using WLI were 74.2 %, 81.7 %, and 66.7 %, respectively, while those for LCI were 85.8 %, 93.3 %, and 78.3 %, respectively. Thus, the accuracy and sensitivity for LCI were significantly higher than those for WLI (P = 0.002 and P = 0.011, respectively). The kappa values for the inter- and intraobserver variability among the 4 endoscopists were higher for LCI than for WLI. CONCLUSIONS: H. pylori infection can be identified by enhancing endoscopic images of the diffuse redness of the fundic gland using LCI. LCI is a novel image-enhanced endoscopy and is more useful for diagnosing H. pylori infection than is WLI.

Incidence of lymphatic involvement in differentiated-type intramucosal gastric cancers as examined by endoscopic resection.

BACKGROUND: Lymphatic involvement may sometimes be detected in differentiated-type intramucosal cancer resected endoscopically during routine examination, but its incidence and clinical significance remain unknown. METHODS: We examined 300 endoscopically resected lesions obtained from 238 patients diagnosed with differentiated-type intramucosal gastric cancer. All sections were subjected to D2-40 monoclonal-based immunohistochemistry. RESULTS: The incidence of lymphatic involvement in differentiated-type intramucosal cancer was 2.0% (6/300). An incidence of 1.8% (5/279) was determined in lesions that were ≤ 3 cm in size and 2.2% (6/276) in those without an ulcer or ulcer scar. Of the six lesions presenting lymphatic involvement, three were determined to be histologically mixed with a predominance of differentiated type, while the other three lesions were pure differentiated type. CONCLUSIONS: We determined that the incidence of lymphatic involvement in differentiated-type intramucosal cancer was 2.0%. To clarify the clinical significance of such lymphatic involvement, it is necessary to detect it with this incidence in mind and to gather and follow up the clinical courses of such cases.

Combination of molecular-targeted drugs with endocrine therapy for hormone-resistant breast cancer.

Overcoming resistance to endocrine therapy is the most intensive research area in estrogen receptor (ER)-positive breast cancer. A strategy to restore endocrine sensitivity using molecular-targeted drugs such as mammalian target of rapamycin inhibitor everolimus along with endocrine therapy has already been used as a treatment option after the progression of previous aromatase inhibitor therapy. Phase II/III clinical trials of several signal pathway inhibitors and cyclin-dependent kinase 4/6 inhibitors are underway. In addition, a randomized phase II trial of the histone deacetylase inhibitor entinostat showed interesting findings. In this review, we summarize the mechanistic principles of combination therapy of molecular-targeted drugs with endocrine therapy by using a hybrid car model.

The inhibitory effect of heat treatment against epithelial-mesenchymal transition (EMT) in human pancreatic adenocarcinoma cell lines.

Epithelial-mesenchymal transition (EMT) plays a crucial role in cancer metastasis. In this study, we evaluated the effect of heat treatment on tumor growth factor-ß1 (TGF-ß1)-induced EMT in pancreatic cancer cells and tried to ascertain the mechanism related to any observed effects. Human pancreatic cancer cell lines (BxPC-3, PANC-1 and MIAPaCa-2) were stimulated by TGF-ß1, and evaluated for morphological changes using immunofluorescence and EMT-related factors (i.e., E-cadherin, Vimentin, Snail or ZEB-1) using RT-PCR. To examine the effect of heat on EMT, the cancer cells were heat-treated at 43°C for 1 h then stimulated with TGF-ß1. We then evaluated whether or not heat treatment changed the expression of EMT-related factors and cell migration and also whether Smad activation was inhibited in TGF-ß signaling. After being treated with TGF-ß1, pancreatic cancer cells resulted in EMT and cell migration was enhanced. Heat treatment inhibited TGF-ß1-induced changes in morphology, inhibited the expression of EMT-related factors, and attenuated TGF-ß1-induced migration in pancreatic cancer cells. Additionally, we observed that heat treatment blocked TGF-ß1-induced phosphorylation of Smad2 in PANC-1 cells. Our results suggest that heat treatment can suppress TGF-ß1-induced EMT and opens the possibility of a new therapeutic use of hyperthermia as a potential treatment for cancer metastasis.

Anoxia/reoxygenation induces epithelial-mesenchymal transition in human colon cancer cell lines.

Epithelial-mesenchymal transition (EMT) is considered to be a crucial event in the development of cancer metastasis. Anoxia/reoxygenation (A/R) is known to occur in cancer tissues due to angiogenesis and changes in tissue pressure that occur during tumor growth. We investigated whether A/R induces EMT in the human colon cancer cell line HT-29. Colon cancer cells were exposed to anoxia (2 h) followed by reoxygenation (4-22 h) and evaluated for EMT changes using immunofluorescence and western blot analyses. We also investigated the expression of EMT-related transcription factors (Snail and ZEB1) using RT-PCR and evaluated the expression of NF-κB using ELISA. To determine whether NF-κB is involved in A/R-induced EMT, HT-29 cells were treated with proteasome inhibitors. Colon cancer cells exposed to A/R underwent EMT morphological changes; the cancer cells acquired a spindle-shaped phenotype. The expression of E-cadherin on the cell surface and the total amount of E-cadherin proteins were reduced after A/R. The expression of EMT-related transcription factors (Snail, ZEB1) was increased after A/R. Pretreatment with proteasome inhibitors significantly attenuated the downregulation of E-cadherin induced by A/R. These results indicate that A/R induces EMT in human colon cancer cells through an NF-κB-dependent transcriptional pathway.

Human adipose tissue-derived mesenchymal stem cells secrete functional neprilysin-bound exosomes.

Alzheimer's disease (AD) is characterized by the accumulation of ß-amyloid peptide (Aß) in the brain because of an imbalance between Aß production and clearance. Neprilysin (NEP) is the most important Aß-degrading enzyme in the brain. Thus, researchers have explored virus-mediated NEP gene delivery. However, such strategies may entail unexpected risks, and thus exploration of a new possibility for NEP delivery is also required. Here, we show that human adipose tissue-derived mesenchymal stem cells (ADSCs) secrete exosomes carrying enzymatically active NEP. The NEP-specific activity level of 1 µg protein from ADSC-derived exosomes was equivalent to that of ~ 0.3 ng of recombinant human NEP. Of note, ADSC-derived exosomes were transferred into N2a cells, and were suggested to decrease both secreted and intracellular Aß levels in the N2a cells. Importantly, these characteristics were more pronounced in ADSCs than bone marrow-derived mesenchymal stem cells, suggesting the therapeutic relevance of ADSC-derived exosomes for AD.

Whole blood interferon-γ levels predict the therapeutic effects of adoptive T-cell therapy in patients with advanced pancreatic cancer.

A core challenge in administering immune-based treatments for cancer is the establishment of easily accessible immunological assays that can predict patients' clinical responses to immunotherapy. In this study, our aim was to predict the therapeutic effects of adoptive T-cell therapy in patients with advanced pancreatic cancer. To do this, we evaluated whole blood cytokine levels and peripheral regulatory T cells (Tregs) in 46 patients with unresectable or recurrent pancreatic cancer who received adoptive T-cell therapy at 2-week intervals. To test immune function, venous blood was obtained from patients before the start of therapy and 2 weeks after the 4th treatment. Whole blood interferon (IFN)-α levels (after stimulation with the Sendai virus) were evaluated, as well as the levels of 9 cytokines stimulated with phytohemagglutinin [interleukin (IL)-2, IL-4, IL-5, IL-10, IL-12(p70), IL-13, tumor necrosis factor-α, IFN-γ, and granulocyte-monocyte colony-stimulating factor]. Peripheral Tregs were analyzed by flow cytometry. Using the obtained data, we then observed the relationship between these immunological parameters and clinical outcome of patients. We found that the whole blood production of IFN-γ, IL-2, IL-4, IL-5 and IL-13 significantly increased after adoptive T-cell therapy, whereas the number of peripheral Tregs did not change. Multivariate Cox proportional hazards analyses indicated that the number of peripheral Tregs before receiving adoptive T-cell therapy and the change in IFN-γ levels after adoptive T-cell therapy were independent variables predicting overall survival. The findings of this study indicate that the assay of whole blood IFN-γ production offers promise for evaluating the clinical response of patients to cancer immunotherapy.

Effects of anticancer agents on cell viability, proliferative activity and cytokine production of peripheral blood mononuclear cells.

We investigated the effects of anticancer agents on peripheral blood mononuclear cells for the purpose of providing data to support new translational chemoimmunotherapy regimens. Peripheral-blood mononuclear cells were treated with one of four anticancer agents (5-fluorouracil, irinotecan, cisplatin, and gemcitabine) for 2 h, after which cell viability was determined. For assessment of effects of each drug on proliferation and cytokine production, cells were stimulated with phytohemagglutinin for 48 h. As a result, the anticancer agents did not affect cell viability. Cell proliferation was unaffected by 5-fluorouracil and irinotecan but inhibited by cisplatin and gemcitabine. Treatment with gemcitabine enhanced the production of IFN-γ and decreased the number of regulatory T cells. gemcitabine treatment increased IFN-γ production among CD4 T cells but not among CD8 T cells. The results indicated that GEM had immunoregulatory properties that might support immune response against cancer. This finding has implications for designing chemoimmunotherapy strategies.

[A case of idiopathic omental bleeding treated successfully with transarterial embolization].

A 58-year-old man was brought to our hospital with left upper abdominal pain which suddenly appeared on the previous evening. An abdominal CT scan showed localized retention of ascites, a slightly high density mass under the left upper abdominal wall, with a high density area detected within the mass, which was suggestive of leakage of contrast medium from peripheral branches of the omental artery. From these findings intraperitoneal hemorrhage caused by bleeding from the greater omentum was suspected. Angiographic examination of the abdomen indicated extravasation of contrast medium from blood vessels of the right gastroepiploic artery. Transarterial embolization was carried out and permanent hemostasis was achieved. Injury, anticoagulant, neoplasms, varix, torsion of the omentum, and segmental arterial mediolysis (SAM) etc have been reported as causes of omental bleeding, but none of these were found in our case. We diagnosed the present case as idiopathic omental bleeding.

Aberrant axonal projections from mammillary bodies in Pax6 mutant mice: possible roles of Netrin-1 and Slit 2 in mammillary projections.

Early events in the axonal tract formation from mammillary bodies remain poorly understood. In the present study, we reported an aberrant pattern of axonal projections from mammillary bodies to the dorsal thalamus in mice lacking the transcription factor Pax6. We found that Netrin-1 was ectopically up-regulated and that both Slit1 and Slit2 were down-regulated in the presumptive dorsal thalamus of Pax6 mutant mice. We then examined the effects of Netrin-1 and Slit2 on the mammillary axons by in utero electroporation techniques. Netrin-1 had an attractive action toward the mammillary axons. Moreover, mammillary trajectories were disorganized in Netrin-1-deficient mice. On the other hand, Slit2 had a repulsive effect on the mammillary axons. These findings suggest that the combination of Netrin and Slit may be involved in proper axonal projection from the mammillary bodies and that their misexpression in the diencephalon may cause the misrouting of these axons in Pax6 mutant mice.

Reduced pain sensitivity in mice lacking latexin, an inhibitor of metallocarboxypeptidases.

Latexin, the endogenous protein inhibitor of the A/B subfamily of metallocarboxypeptidases, is expressed in small nociceptive neurons in sensory ganglia and in a subset of neurons in the telencephalon. In this study, we generated latexin-deficient mice that exhibited increased tail-flick latency compared to wild-type animals upon noxious heat stimulation. The reduced pain sensitivity in the mutants was rescued by the systemic administration of a plant carboxypeptidase inhibitor that inhibits the A/B subfamily of metallocarboxypeptidases. These findings suggest that latexin is involved in the transmission of pain.

Correlation of invasion and metastasis of cancer cells, and expression of the RAD21 gene in oral squamous cell carcinoma.

Although RAD21 is involved in the repair of double-strand breaks in DNA and is essential for mitotic growth, its role in cancer has been unclear. In this study, the relevance of RAD21 gene expression to the invasion and metastasis of oral squamous cell carcinoma was clarified using laser microdissection and real-time polymerase chain reaction (PCR). Using two different metastatic potential oral squamous cells [high-metastatic-potential squamous cell carcinoma cells (SAS-Ly) and low-metastatic-potential squamous cell carcinoma cells (SAS)], the relation of RAD21 gene expression to apoptosis, invasion, and metastasis was examined. The results showed that RAD21 gene expression was significantly decreased in oral squamous cell carcinoma when it expressed the INFbeta and INFgamma invasion patterns in comparison with the INFalpha invasion pattern (p<0.01). In addition, in comparison with SAS cells, SAS-Ly cells indicated tolerance to cell death induced by an apoptosis induction reagent, while the expression level of the RAD21 gene in SAS cells was increased by the apoptosis induction reagent. However, in SAS-Ly cells, the reagent induced no significant difference. Our findings indicate that the RAD21 gene was closely related to the invasion and metastasis of cancer cells.