The relationship between tumor immunity and the cGAS-STING pathway in breast cancer: An immunohistochemical study.

Breast cancer (BC) is classified into four major histological subtypes, namely luminal A, luminal B, HER2, and basal-like, and its treatment is based on these subtypes. The use of immune checkpoint inhibitors against BC depends on the expression of PD-1/PD-L1. Another tumor immune system-the cGAS-STING pathway-is a potential target for cancer immunotherapy. However, the status of the cGAS-STING pathway in BC has not been fully established. Therefore, we investigated the expression status of the cGAS-STING pathway and immune-related proteins in BC. We classified 111 BCs into six groups-29 hormone receptor-positive carcinomas, 12 HER2+ carcinomas (HER2), 8 luminal-HER2 carcinomas, 26 triple-negative breast carcinomas (TNBCs), 21 lobular carcinomas (LC), and 15 carcinomas with apocrine differentiation (CAD)-and investigated the relationship between BC and tumor immunity via the cGAS-STING pathway using histopathological and immunohistochemical methods. Expression of cGAS was high in CADs (100%) and low in TNBCs (35%); STING-positive lymphocytes were high in TNBC (85%, P = 0.0054). Expression of pSTAT3 was significantly high in patients with TNBC (≥10%, 88%). The proportion of PD-L1-positive tumor cells was higher in TNBCs (54%) than in other BCs (30%). SRGN expression was significantly higher in the TNBC group than in the other BC groups (58%). Tumor immune responses may differ among tumor subtypes. The cGAS-STING pathway may be functional in TNBC and CAD but not in LC. Therefore, targeting the cGAS-STING pathway might be useful in BC, particularly TNBC and CAD.

Clinical characteristics, proteins, and genes related to interstitial pneumonia-associated squamous cell carcinoma of the lungs.

Squamous cell carcinoma (SCC) is a common histological type of lung carcinoma that is associated with interstitial pneumonia (IP). We hypothesized that identifying specific genetic alterations or molecular markers of SCC with IP may aid the development of novel therapeutic strategies for the same. Therefore, in the present study, we aimed to identify tumorigenic genetic alterations and molecular markers in cases of SCC with IP. We included 28 lung SCC cases (14 cases with IP and 14 cases without IP). We performed immunohistochemistry for STAT3, STAT5, and TLE1, and next-generation sequencing was performed using an iSeq 100 system. The panel used in this study targeted 50 cancer-associated genes. Immunohistochemically, the rate of TLE1 positivity was higher in the SCC without IP group (93 %) than in the SCC with IP group (29 %), while that of STAT5 was higher in the SCC with IP group (79 %) than in the SCC without IP group (14 %). STAT3 expression was high in both the groups (SCC with IP, 64 %; SCC without IP, 71 %). Eighteen genes were mutated in more than six samples, and FBXW7 mutation was mainly observed in the SCC with IP group (p < 0.01). Mechanisms underlying tumorigenesis in SCC with IP included STAT5 activation via inflammation, while that in SCC without IP included squamous TLE1-mediated metaplasia. These findings are based on smoking-induced STAT3 activation; therefore, patients with IP who smoke are more likely to have progressive SCC. We also found that FBXW7 mutations may be associated with SCC with IP and keratinization. ERBB4 and KDR mutations were observed in both with or without IP, and these genes may be tumor-related genes in SCC. These molecular markers may help determine the prognoses of patients with SCC with IP and direct the development of treatment approaches.

Severe aortic stenosis during leptin replacement therapy in a patient with generalized lipodystrophy-associated progeroid syndrome due to an LMNA variant: A case report.

Leptin replacement therapy (LRT) has drastically improved the prognosis of patients with lipodystrophy, but pro-inflammatory properties of leptin could become evident in the long term. Here, we report a 30-year-old Japanese woman with generalized lipodystrophy-associated progeroid syndrome due to a heterozygous LMNA variant (c.29C > T; p.T10I), who was diagnosed with severe aortic stenosis (AS) after more than a decade of LRT, which required transcatheter aortic valve implantation. Given her marked hypoadiponectinemia and the LMNA variant, our patient might have been susceptible to progeria-associated disorders, including aortic stenosis, which could have been exaggerated by the prolonged 'imbalanced adipokines' caused by LRT between pro-inflammatory leptin and anti-inflammatory adiponectin. Thus, long-term LRT could be associated with AS in patients with the LMNA variant to cause generalized lipodystrophy-associated progeroid syndrome and hypoadiponectinemia.

Immunohistochemical Reactivity of Prostate-Specific Membrane Antigen in Salivary Gland Tumors.

Prostate-specific membrane antigen (PSMA) is a transmembrane glycoprotein that is overexpressed in the prostate gland and prostate cancer. PSMA has been recently used in positron emission tomography/computed tomography (PET/CT) imaging and targeted alpha-radiation therapy (TAT) for prostate cancer. Recently, the tubarial gland, a type of minor salivary gland that is described as a new organ situated in the pharynx, is reported to express PMSA. Here, we studied the expression of PSMA in common benign and malignant salivary gland tumors. We performed immunohistochemistry for PSMA in 55 salivary gland tumors comprising 10 pleomorphic adenomas, 10 Warthin tumors, 9 basal cell adenomas, 9 adenoid cystic carcinomas, 9 mucoepidermoid carcinomas, and 8 salivary duct carcinomas. PSMA was expressed in 97% of benign tumors and 77% of malignant tumors. Moreover, PSMA was expressed in 59% of normal salivary glands adjacent to the tumor. PSMA is relatively expressed in salivary gland tumors and salivary glands. Therefore, salivary gland neoplasm, and normal salivary gland, possibly demonstrate the accumulation of PSMA in PET/CT. Thus, we need to monitor the side effects in the salivary glands during TAT for prostate cancer.

Genotype-Structure-Phenotype Correlations of Disease-Associated IGF1R Variants and Similarities to Those of INSR Variants.

We previously reported genotype-phenotype correlations in 12 missense variants causing severe insulin resistance, located in the second and third fibronectin type III (FnIII) domains of the insulin receptor (INSR), containing the α-ß cleavage and part of insulin-binding sites. This study aimed to identify genotype-phenotype correlations in FnIII domain variants of IGF1R, a structurally related homolog of INSR, which may be associated with growth retardation, using the recently reported crystal structures of IGF1R. A structural bioinformatics analysis of five previously reported disease-associated heterozygous missense variants and a likely benign variant in the FnIII domains of IGF1R predicted that the disease-associated variants would severely impair the hydrophobic core formation and stability of the FnIII domains or affect the α-ß cleavage site, while the likely benign variant would not affect the folding of the domains. A functional analysis of these variants in CHO cells showed impaired receptor processing and autophosphorylation in cells expressing the disease-associated variants but not in those expressing the wild-type form or the likely benign variant. These results demonstrated genotype-phenotype correlations in the FnIII domain variants of IGF1R, which are presumably consistent with those of INSR and would help in the early diagnosis of patients with disease-associated IGF1R variants.

Pulmonary wedge aspiration cytology for the rapid diagnosis of pulmonary tumor thrombotic microangiopathy: A case report.

Pulmonary tumor thrombotic microangiopathy (PTTM) is a cancer-related pulmonary complication characterized by rapid progression of dyspnea and pulmonary hypertension, occasionally causing sudden death. Given the condition of patients with dyspnea, lung biopsies are limited because of their invasiveness. A 72-year-old man presented with chronic atrial fibrillation and a high right heart load, as determined using ultrasonography. He had previously undergone resection of the left axillary skin secondary to extramammary Paget's disease (EMPD). Clinically, PTTM was suspected and pulmonary wedge aspiration cytology, collected from the pulmonary artery during catherization, was performed. Cytologically, the tumor demonstrated three-dimensional cell clusters with good cohesion and molding by the blood vessel lumen. Additionally, endothelial-like cells were observed at the periphery of the tumor clusters; fibrin was evident in the clusters. The tumor cells were large, round, and had high nuclear/cytoplasmic ratios. The nuclei demonstrated a variety of sizes and were irregularly shaped, with prominent nucleoli; cells undergoing mitosis were evident. The tumor cells were suspected of being poorly differentiated adenocarcinoma cells, consistent with PTTM. Two days after the aspiration cytology, the patient died and a pathological autopsy was performed. Histologically, the PTTM was determined to have caused the pulmonary hypertension and the primary PTTM site was apparently derived from the EMPD. For rapid diagnoses, an understanding of the tumor's cytological features is important and should contribute to early treatment intervention. Aspiration cytology, using pulmonary artery blood samples, during catherization is a useful tool for diagnosing PTTM.

Clinical usefulness of multigene screening with phenotype-driven bioinformatics analysis for the diagnosis of patients with monogenic diabetes or severe insulin resistance.

AIMS: Monogenic diabetes is clinically heterogeneous and differs from common forms of diabetes (type 1 and 2). We aimed to investigate the clinical usefulness of a comprehensive genetic testing system, comprised of targeted next-generation sequencing (NGS) with phenotype-driven bioinformatics analysis in patients with monogenic diabetes, which uses patient genotypic and phenotypic data to prioritize potentially causal variants. METHODS: We performed targeted NGS of 383 genes associated with monogenic diabetes or common forms of diabetes in 13 Japanese patients with suspected (n = 10) or previously diagnosed (n = 3) monogenic diabetes or severe insulin resistance. We performed in silico structural analysis and phenotype-driven bioinformatics analysis of candidate variants from NGS data. RESULTS: Among the patients suspected having monogenic diabetes or insulin resistance, we diagnosed 3 patients as subtypes of monogenic diabetes due to disease-associated variants of INSR, LMNA, and HNF1B. Additionally, in 3 other patients, we detected rare variants with potential phenotypic effects. Notably, we identified a novel missense variant in TBC1D4 and an MC4R variant, which together may cause a mixed phenotype of severe insulin resistance. CONCLUSIONS: This comprehensive approach could assist in the early diagnosis of patients with monogenic diabetes and facilitate the provision of tailored therapy.

Pulmonary tumor thrombotic microangiopathy associated with extramammary Paget's disease: An autopsy case report.

Pulmonary tumor thrombotic microangiopathy (PTTM) is histologically characterized by micro tumor cell embolism and intimal fibrocellular proliferation of pulmonary arteries or arterioles. We report a secondary case of PTTM associated with extramammary Paget's disease (EMPD). The patient was a 72-year-old man with exertional dyspnea. Clinical examinations found he had pulmonary hypertension and multiple osteolytic lesions of vertebra. Cytological analysis of pulmonary wedge artery sample detected malignant cells and he was dead before treatment was started. Multiple tumor embolisms (>17) were identified in pulmonary arteries or arterioles at autopsy, consistent with PTTM. Metastatic nodules were found in liver and lymph node. Furthermore, disseminated carcinomatosis of the bone marrow (DCBM) was seen. Immunostaining results pointed out that tumor cells possessed mammary gland phenotype. He had 4-years history of EMPD in the left axilla without recurrence, and immunohistochemistry results were the same as the autopsy specimen. Thus, we diagnosed the primary site of PTTM to be EMPD. Our case highlights the usefulness of the recent proposed classification of PTTM, potential association between PTTM and DCBM, and the necessity for long-term follow-up in EMPD. EMPD can rarely cause PTTM to manifest as a paraneoplastic syndrome.

A Case of Dendritic Cell Neurofibroma With Pseudorosettes.

Dendritic cell neurofibroma with pseudorosettes (DCNP) is a rare benign peripheral nerve sheath tumor. Till date, 34 cases of DCNP arising from various sites have been reported. Histopathologically, DCNP is known to present with characteristic pseudorosettes, in which a type II cell is surrounded by type I cells. In the present report, we discuss the rare case of a 63-year-old man diagnosed with DCNP on the left flank (size: approximately 10 mm). On microscopic examination of the resected lesion, we observed the characteristic pseudorosettes with centrally placed type I cells, which exhibited small, dark, slightly irregular oval nuclei with nuclear inclusions, surrounded by type II cells, which showed a large pale nucleus with a constriction, a small nucleolus, and mildly eosinophilic cytoplasm. The type II cells were positive for S-100, CD57, LAMP2, fascin, and factor XIIIa. Although previous reports have suggested that type II cells exhibit a dendritic form, our immunohistochemical analyses revealed that these cells were dermal interstitial dendritic cells.

A case of desmoplastic fibroma of bone with CTNNB1 point mutation.

Desmoplastic fibroma of bone (DFB), a bone tumor, is considered to be an osseous counterpart of desmoid-type fibromatosis (DF). Herein, we report a case of DFB with CTNNB1 point mutation. The 5-year-old male patient had complained of trismus and pain in the jaw. Magnetic resonance imaging revealed a mass in the left mandible. Radical treatment involved surgical resection. Microscopically, the lesion consisted of a bundle-like proliferation of uniform spindle-shaped cells with abundant collagenous stroma, which resembled DF. Immunohistochemical analysis revealed intranuclear accumulation of ß-catenin in the tumor cells. Based on clinical and histologic analysis, we diagnosed the patient as having DFB. We examined the CTNNB1 and APC sequence and found an A-to-G transition at codon 41 of CTNNB1; i.e., Thr was substituted by Ala. Our findings suggest that the dysregulation of Wnt/ß-catenin signaling pathway is related to the tumorigenesis of some cases of DFB. This hypothesis indicates that there are some cases of DFB in which nuclear positive expression of ß-catenin is useful for diagnosis.

Immunohistochemical Reactivity of Prostate-Specific Markers for Salivary Duct Carcinoma.

OBJECTIVES: NKX3.1, a transcription factor related to androgen expression, has recently been introduced as a diagnostic marker of prostate adenocarcinoma. Salivary duct carcinoma (SDC) is typically positive for androgen receptor (AR). Therefore, we hypothesized that NKX3.1 is a new immunohistochemical marker for SDC and aimed to investigate whether NKX3.1 staining in combination with other immunomarkers of prostate carcinoma could have a diagnostic or prognostic value in SDC. METHODS: Materials obtained from 42 resected SDCs were examined by immunohistochemistry using antibodies against AR, NKX3.1, α-methylacyl-CoA racemase (AMACR), prostatic acid phosphatase (PAP), and prostate-specific antigen (PSA). RESULTS: In immunoreactivity among SDC cases, 81.0, 35.7, 58.5, 33.3, and 0% were positive for AR, NKX3.1, AMACR, PAP, and PSA, respectively. AMACR and PAP immunoreactivity rates were higher in recurrence cases than in cases with no recurrence. CONCLUSIONS: NKX3.1 expression is useful for SDC diagnosis, but decreased NKX3.1 expression was not correlated with SDC progression. The immunoreactivity of AMACR and PAP could be useful for assessing prognosis in SDC, but immunohistochemical staining of prostate-specific markers should be interpreted with caution when determining whether a metastatic tumor is of prostate origin, especially when patients have a history of SDC.

Clinical characteristics of insulin resistance syndromes: A nationwide survey in Japan.

AIMS/INTRODUCTION: Insulin resistance syndrome (IRS) of type A or B is triggered by gene abnormalities of or autoantibodies to the insulin receptor, respectively. Rabson-Mendenhall/Donohue syndrome is also caused by defects of the insulin receptor gene (INSR), but is more serious than type A IRS. Here, we carried out a nationwide survey of these syndromes in Japan. MATERIALS AND METHODS: We sent questionnaires to a total of 1,957 academic councilors or responsible individuals at certified facilities of the Japan Diabetes Society, as well as at the department pediatrics or neonatology in medical centers with >300 beds. RESULTS: We received 904 responses with information on 23, 30 and 10 cases of type A or B IRS and Rabson-Mendenhall/Donohue syndrome, respectively. Eight cases with type A IRS-like clinical features, but without an abnormality of INSR, were tentatively designated type X IRS, with five of these cases testing positive for PIK3R1 mutations. Fasting serum insulin levels at diagnosis (mean ± standard deviation) were 132.0 ± 112.4, 1122.1 ± 3292.5, 2895.5 ± 3181.5 and 145.0 ± 141.4 µU/mL for type A IRS, type B IRS, Rabson-Mendenhall/Donohue syndrome and type X IRS, respectively. Type A and type X IRS, as well as Rabson-Mendenhall/Donohue syndrome were associated with low birthweight. Type B IRS was diagnosed most frequently in older individuals, and was often associated with concurrent autoimmune conditions and hypoglycemia. CONCLUSIONS: Information yielded by this first nationwide survey should provide epidemiological insight into these rare conditions and inform better healthcare for affected patients.

Colon adenoma and adenocarcinoma with clear cell components - two case reports.

BACKGROUND: Diagnoses reflect clear cell morphologies when tumor cells have clear cytoplasm in many organs, and the nature of such clear cells is typically identified. Colorectal tubular adenoma or adenocarcinoma, conversely, rarely show clear cells, the reason for which remains uncertain. We report 2 colon tumors with clear cell components (Case 1: adenoma; Case 2: adenocarcinoma) and investigate the nature of the clear cells. CASE PRESENTATION: Case 1 was a 75-year-old man with a superficial elevated polyp detected in the rectum for whom endoscopic submucosal dissection was performed. Microscopically, 10% of the tumor showed dysplastic columnar epithelium with clear cytoplasm forming tubular structures accompanied by conventional tubular adenoma. Case 2 was a 58-year-old man with a pedunculated polyp found in his sigmoid colon for which polypectomy was performed. Microscopically, 90% of the tumor showed dysplastic columnar epithelium with clear cytoplasm forming fused glands or cribriform structures adjacent to the ordinal tubular adenocarcinoma. In both cases, clear and ordinary tumor cells were negative for CK7 and positive for CK20 and CDX2, consistent with findings of colorectal origin. Different results were found for CEA and CD10 staining. CEA was positive on the luminal side of the conventional area in contrast diffuse cytoplasmic staining of the clear cell area in both cases. CD10 was only positive for the clear cell component of case 2. The clear cell components were negative for Periodic acid-Schiff (PAS), Alcian blue, and mucicarmine staining and AFP immunohistochemistry. An ultrastructural examination found multiple cytoplasmic lipid-like vacuoles in the clear cell component that were predominantly negative for adipophilin by immunoelectron microscopy. CONCLUSIONS: We investigated tubular adenoma and tubular adenocarcinoma with clear cell components. The accompanying conventional tubular adenoma or adenocarcinoma cells helped us to evaluate the atypia of the clear cells. Diffuse cytoplasmic staining of CEA and CD10 suggested that the clear cell component might harbor malignant potential. We were unable to verify the well-known causes of clear cytoplasm, such as an accumulation of glycogen, lipid, or mucin and enteroblastic differentiation. The causes of clear cells in the colorectal region remain uncertain; however, possible explanations include autolysis and carbohydrate elution.

A Case of Methotrexate-Associated Lymphoproliferative Disorder (Lymphomatoid Granulomatosis) of the Skin.

Iatrogenic lymphoproliferative disorder (LPD) can develop in patients treated with immunosuppressive drugs for autoimmune or other inflammatory diseases. Here, we report a case of lymphomatoid granulomatosis of the skin that occurred as a methotrexate (MTX)-associated LPD. We also review the relevant literature. A 73-year-old woman presented to our department with an approximately 10-year history of MTX therapy for rheumatoid arthritis. Three months earlier, she noticed a small nodule in her right upper arm. It gradually enlarged, and the center began to decay. Grossly, the lesion was 40 × 40 mm in size with ulceration, and the surrounding skin presented dark red erythema. A biopsy specimen was taken for definitive diagnosis. Histologically, infiltrating growth of medium-to-large atypical lymphocytes was observed underneath the ulceration and was accompanied by small reactive lymphocytes. The atypical lymphocytes demonstrated a tendency to infiltrate the vessels, which showed an angiocentric pattern. Immunohistochemistry revealed that the atypical lymphoid cells were positive for CD79a, CD20, and CD30. In addition, in situ hybridization for Epstein-Barr virus (EBV) revealed expression of EBV-encoded small RNAs. The patient was diagnosed with MTX-associated LPD (lymphomatoid granulomatosis), owing to her history of MTX treatment, the expression of the atypical lymphocytes for B-cell markers and EBV-encoded small RNA, and the angiocentric infiltrating pattern. The lesion reportedly disappeared after withdrawal of MTX.

A Biotin Tagging Immunoelectron Microscopy for Paraffin-embedded Sections.

We herein introduce a novel method of biotin tagging immunoelectron microscopy for formalin-fixed, paraffin-embedded sections. This method was developed to utilize the antigenicity of biotin on epoxy-embedded ultrathin sections that could readily be recovered by a previously established antigen retrieval method as most monoclonal antibodies failed to recognize their targets by immunoelectron microscopy following antigen retrieval. The biotin tagging method was composed of preembedding immunostaining, epoxy-embedding and sectioning, and postembedding immunostaining steps. The preembedding step utilized the streptavidin-biotin-peroxidase complex method for immunohistochemistry to tag every antigen with a biotin in 3-µm thick paraffin-embedded sections. Next, fixation and processing for transmission electron microscopy (TEM) were performed on sections on glass slides, and ultrathin sections were prepared in epoxy-embedded blocks. In the postembedding step, antigen retrieval was followed by serial incubations with an antibiotin monoclonal antibody and anti-mouse IgG-labeled gold particles. The results obtained using antibodies against a variety of intracellular targets were satisfactory; positive gold particles were observed corresponding to targeted intracellular structures. This study demonstrated that the biotin tagging method was a convenient approach for successful labeling of paraffin-embedded sections for TEM using monoclonal antibodies, although it has relatively poor subcellular labeling quality.

Difference in transducin-like enhancer of split 1 protein expression between basal cell adenomas and basal cell adenocarcinomas - an immunohistochemical study.

BACKGROUND: Basal cell adenoma (BCA) and basal cell adenocarcinoma (BCAC) are benign and malignant, basaloid salivary gland neoplasms, respectively. These tumors show a dual-cell proliferation of inner luminal/ductal cells and outer abluminal/myoepithelial or basal cells. The only difference between them is defined as a malignant morphology such as invasion. Recently, the nuclear expression of ß-catenin and a catenin beta-1 (CTNNB1) mutation were found in BCA. Transducin-like enhancer of split 1 (TLE1) belongs to the Groucho/TLE family, and it functions in the "off" state in the Wnt/ß-catenin signaling pathway. We hypothesized that if the dysregulation of the Wnt/ß-catenin signaling pathway could be attributed to the tumorigenesis of BCA/BCAC, there might be differences in TLE1 expression between BCA and BCAC. METHOD: The study included 35 BCA and 4 BCAC cases. We performed immunohistochemistry to detect TLE1 and ß-catenin and investigated the catenin beta-1 (CTNNB1) mutational profile among BCA and BCAC cases. RESULTS: In BCA, the expression of TLE1 was confined to luminal cells of glandular structures, in contrast to the expression of ß-catenin in abluminal cells. The BCA cases harbored CTNNB1 gene mutations (12/35). In BCAC, luminal cell staining of TLE1 was identical to BCA in non-invasive areas (4/4) but indistinct in invasive areas (3/4). The BCAC cases were ß-catenin positive for abluminal cells in both areas. The BCAC cases had CTNNB1 mutation (2/4) and the laser-captured microdissection allowed the separate collection of infiltrative and non-infiltrative areas to detect the same mutation. CONCLUSIONS: Immunohistochemical analysis for TLE1 can identify BCA and BCAC by luminal cell staining difference, especially indistinct luminal cell expression for TLE1 in invasive areas of BCAC. Moreover, TLE1 can be luminal/ductal cell markers.