Squamous cell papilloma of the hypopharynx with elongated villous projections.

Squamous cell papilloma with elongated villous projections may occur in the hypopharynx and present with symptoms observable on physical examination.

Parenchymatous glossitis: A case report.

We present a case of parenchymatous glossitis with unilateral severe inflammation of the hyoglossus muscle, resulting in laryngeal edema. The route of inflammation was unique. Contrast-enhanced CT was useful for diagnosis. Administration of sensitive antibiotics led to improvement. Creatine phosphokinase may be an indicator of diagnosis and treatment response.

Induction of lung lesions by bronchial administration using bronchoscope technique in mice.

This study aimed to establish an exposure method that can induce homogeneous lesions with minimal inter-individual variability. The distribution of lesions induced by bleomycin (BLM) administration was also analyzed. C57BL mice were intrabronchially administered 20 µL of BLM (3 mg/mL) using a bronchoscope in the left or right bronchus. The mice were sacrificed 14 days after administration, and their lungs were evaluated histopathologically. BLM-induced inflammatory lesions were widely observed in the lungs. In the left bronchus-treated group, lesions were uniformly observed throughout the lobe, and no individual differences were noted. Meanwhile, in the right bronchus-treated group, individual differences in the distribution of the pulmonary lesions were observed. The distribution of lesions differed among the four lobes of the right lung owing to their anatomical features. Administration into the left bronchus is recommended for highly homogeneous lung exposure and for establishing models that contribute to highly accurate toxicity and efficacy evaluations.

PRMT1 Sustains De Novo Fatty Acid Synthesis by Methylating PHGDH to Drive Chemoresistance in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) chemoresistance hampers the ability to effectively treat patients. Identification of mechanisms driving chemoresistance can lead to strategies to improve treatment. Here, we revealed that protein arginine methyltransferase-1 (PRMT1) simultaneously methylates D-3-phosphoglycerate dehydrogenase (PHGDH), a critical enzyme in serine synthesis, and the glycolytic enzymes PFKFB3 and PKM2 in TNBC cells. 13C metabolic flux analyses showed that PRMT1-dependent methylation of these three enzymes diverts glucose toward intermediates in the serine-synthesizing and serine/glycine cleavage pathways, thereby accelerating the production of methyl donors in TNBC cells. Mechanistically, PRMT1-dependent methylation of PHGDH at R54 or R20 activated its enzymatic activity by stabilizing 3-phosphoglycerate binding and suppressing polyubiquitination. PRMT1-mediated PHGDH methylation drove chemoresistance independently of glutathione synthesis. Rather, activation of the serine synthesis pathway supplied α-ketoglutarate and citrate to increase palmitate levels through activation of fatty acid synthase (FASN). Increased palmitate induced protein S-palmitoylation of PHGDH and FASN to further enhance fatty acid synthesis in a PRMT1-dependent manner. Loss of PRMT1 or pharmacologic inhibition of FASN or protein S-palmitoyltransferase reversed chemoresistance in TNBC. Furthermore, IHC coupled with imaging MS in clinical TNBC specimens substantiated that PRMT1-mediated methylation of PHGDH, PFKFB3, and PKM2 correlates with chemoresistance and that metabolites required for methylation and fatty acid synthesis are enriched in TNBC. Together, these results suggest that enhanced de novo fatty acid synthesis mediated by coordinated protein arginine methylation and protein S-palmitoylation is a therapeutic target for overcoming chemoresistance in TNBC. SIGNIFICANCE: PRMT1 promotes chemoresistance in TNBC by methylating metabolic enzymes PFKFB3, PKM2, and PHGDH to augment de novo fatty acid synthesis, indicating that targeting this axis is a potential treatment strategy.

SGX523 causes renal toxicity through aldehyde oxidase-mediated less-soluble metabolite formation in chimeric mice with humanized livers.

SGX523 is a c-Met tyrosine kinase inhibitor that failed in clinical trials because of renal toxicity caused by crystal deposits in renal tubules. SGX523 is metabolized by aldehyde oxidase (AOX) in a species-dependent manner to the considerably less soluble 2-quinolinone-SGX523, which is likely involved in the clinically observed obstructive nephropathy. This study investigated the metabolism and renal toxicity of SGX523 in chimeric mice with humanized livers (humanized-liver mice). The 2-quinolinone-SGX523 formation activity was higher in humanized-liver mouse and human hepatocytes than in mouse hepatocytes. Additionally, this activity in the liver cytosolic fraction from humanized-liver mice was inhibited by the AOX inhibitors raloxifene and hydralazine. After oral SGX523 administration, higher maximum concentrations, larger areas under the plasma concentration versus time curves, and higher urinary concentrations of 2-quinolinone-SGX523 were observed in humanized-liver mice than in non-humanized mice. Serum creatinine and blood urea nitrogen levels were elevated in humanized-liver mice following repeated oral SGX523 administration. The accumulation of amorphous material in the tubules and infiltration of inflammatory cells around tubules were observed in the kidneys of humanized-liver mice after repeated oral SGX523 administration. These findings demonstrate that humanized-liver mice are useful for understanding the metabolism and toxicity of SGX523.

Development of rat duodenal monolayer model with effective barrier function from rat organoids for ADME assay.

The in-depth analysis of the ADME profiles of drug candidates using in vitro models is essential for drug development since a drug's exposure in humans depends on its ADME properties. In contrast to efforts in developing human in vitro absorption models, only a limited number of studies have explored models using rats, the most frequently used species in in vivo DMPK studies. In this study, we developed a monolayer model with an effective barrier function for ADME assays using rat duodenal organoids as a cell source. At first, we developed rat duodenal organoids according to a previous report, but they were not able to generate a confluent monolayer. Therefore, we modified organoid culture protocols and developed cyst-enriched organoids; these strongly promoted the formation of a confluent monolayer. Furthermore, adding valproic acid to the culture accelerated the differentiation of the monolayer, which possessed an effective barrier function and apicobasal cell polarity. Drug transporter P-gp function as well as CYP3A activity and nuclear receptor function were confirmed in the model. We expect our novel monolayer model to be a useful tool for elucidating drug absorption processes in detail, enabling the development of highly absorbable drugs.

Survival Impact of Lingual Lymph Node Metastases in Tongue Cancer: An Analysis Regarding Anatomical Subsites.

BACKGROUND/AIM: Lingual lymph node (LLN) metastasis from tongue cancer occurs at four subsites. However, subsite-related prognosis is unknown. This study aimed to analyze the association between LLN metastases and disease-specific survival (DSS) with respect to these four anatomic subsites. PATIENTS AND METHODS: Patients with tongue cancer treated between January 2010 and April 2018 at our institute were reviewed. The four subgroups of LLNs were median, anterior lateral, posterior lateral, and parahyoid. DSS was evaluated. RESULTS: LLN metastases occurred in 16 of the 128 cases; six and 10 cases were identified during initial and salvage therapy, respectively. Zero, four, three, and nine cases were median, anterior lateral, posterior lateral, and parahyoid LLN metastases, respectively. The 5-year DSS of patients with LLN metastasis was significantly poor on univariate analysis; parahyoid LLN metastasis showed the worst prognosis. Multivariate analysis indicated that only advanced nodal stage and lymphovascular invasion were significant survival factors. CONCLUSION: Parahyoid LLNs may require the most caution in tongue cancer. The significance of LLN metastases alone for survival was not confirmed on multivariate analysis.

An integrative epigenomic approach identifies ELF3 as an oncogenic regulator in ASCL1-positive neuroendocrine carcinoma.

Neuroendocrine carcinoma (NEC) is a highly aggressive subtype of the neuroendocrine tumor with an extremely poor prognosis. We have previously conducted a comprehensive genomic analysis of over 100 cases of NEC of the gastrointestinal system (GIS-NEC) and unraveled its unique and organ-specific genomic drivers. However, the epigenomic features of GIS-NEC remain unexplored. In this study, we have described the epigenomic landscape of GIS-NEC and small cell lung carcinoma (SCLC) by integrating motif enrichment analysis from the assay of transposase-accessible chromatin sequencing (ATAC-seq) and enhancer profiling from a novel cleavage under targets and tagmentation (CUT&Tag) assay for H3K27ac and identified ELF3 as one of the super-enhancer-related transcriptional factors in NEC. By combining CUT&Tag and knockdown RNA sequencing for ELF3, we uncovered the transcriptional network regulated by ELF3 and defined its distinctive gene signature, including AURKA, CDC25B, CLDN4, ITGB6, and YWAHB. Furthermore, a loss-of-function assay revealed that ELF3 depletion led to poor cell viability. Finally, using gene expression of clinical samples, we successfully divided GIS-NEC patients into two subgroups according to the ELF3 signature and demonstrated that tumor-promoting pathways were activated in the ELF3 signature-high group. Our findings highlight the transcriptional regulation of ELF3 as an oncogenic transcription factor and its tumor-promoting properties in NEC.

MicroRNAs secreted by human preimplantation embryos and IVF outcome.

OBJECTIVE: To generate an effective embryo prediction model and identify a non-invasive evaluation method by analyzing microRNAs (miRNAs) in embryo culture medium. DESIGN: Analysis of microRNA profiles from spent culture medium of blastocysts with good morphology that did or did not result in pregnancy. SETTING: Clinical and experimental research. PATIENTS: Sixty patients who underwent thawed embryo transfer of blastocysts after intracytoplasmic sperm injection. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): The association of miRNA abundance levels secreted by blastocysts in culture medium and implantation success. RESULTS: Our RNA sequencing analysis found a total of 53 differentially expressed miRNAs in the culture media of pregnancy and non-pregnancy groups. Twenty-one miRNAs were analyzed for their potential to predict implantation success. Eight miRNAs (hsa-miR-191-5p, hsa-miR-320a, hsa-miR-92a-3p, hsa-miR-509-3p, hsa-miR-378a-3p, hsa-miR-28-3p, hsa-miR-512-5p, and hsa-miR-181a-5p) were further extracted from the results of a logistic regression analysis of qPCR Ct values. A prediction model for high-quality blastocysts was generated using the eight miRNAs, with an average accuracy of 0.82 by 5-fold cross validation. CONCLUSION: We isolated blastocyst miRNAs that may predict implantation success and created a model to predict viable embryos. Increasing the number of investigated cases and further studying the effect of each miRNA on embryonic development is needed to refine the miRNA-based predictive model.

Interleukin 6/gp130 axis promotes neural invasion in pancreatic cancer.

BACKGROUND: Nerve invasion (N-inv) is an important prognostic factor in pancreatic ductal adenocarcinoma (PDAC). Elucidation of circulating N-inv stimulators could provide deeper insights and novel perspectives for PDAC therapy. The interleukin (IL)-6/gp130 axis was evaluated in this study as a candidate N-inv stimulator. METHODS: A human pancreatic cancer (PC) cell, Capan-1, was confirmed to have the stimulant activity of IL-6/gp130 axis through the evaluation of mRNA, cell surface protein and intracellular protein levels and chemotaxis and wound healing assay. The upregulation of IL-6/gp130 axis was evaluated using tumor-derived IL-6 level and intratumoral pSTAT3 expression in N-inv of murine sciatic nerves by intraneural injection of Capan-1 cell (N-inv model) and using resected pancreatic cancer tissue and clinical data from 46 PDAC patients. RESULTS: mRNA and protein expressions of IL-6 and IL-6 receptor were found in whole cell lysate and condition medium from PC cell. Cell surface protein expression of gp130 were clearly detected on PC cell. IL-6 promoted migration and chemotaxis of PC cell. Serum IL-6 and tumoral IL-6 mRNA levels in N-inv model mice were significantly higher than those in subcutaneous tumor mice (p = 0.004 and p = 0.002, respectively). Silencing of IL-6 and gp130 on PC cell and administration of an anti-IL-6 receptor antibody, tocilizumab, suppressed N-inv, compared to each control (p = 0.070, p = 0.118 and p = 0.122, respectively). In PDAC patients, the high-N-inv group showed poor prognosis (p =0.059) and elevated serum levels of IL-6 and C-reactive protein, synthesis of which is promoted by IL-6, compared to those in the low-N-inv group (p = 0.006 and p = 0.075, respectively). Tumoral gp130 expression at N-inv was higher than that in the primary pancreatic tumor (p = 0.026). CONCLUSION: Biological activity of IL-6/gp130 axis promoted N-inv in murine model and was upregulated in PDAC patients with severe N-inv. This study is the first evidence that the IL-6/gp130 axis offers a potential therapeutic target in PDAC with N-inv.

rasH2 mouse: reproducibility and stability of carcinogenicity due to a standardized production and monitoring system.

The rasH2 mouse was developed as a model for carcinogenicity studies in regulatory science. Its phenotype is stable during high-volume production and over successive generations. To produce rasH2 mice, three strains of mice (C57BL/6J-TgrasH2, C57BL/6J, and BALB/cByJ) were maintained individually. Since the homozygous c-HRAS genotype is lethal, hemizygous transgenic mice were maintained by crossing with inbred C57BL/6J mice. After breeding, male B6-transgenic mice were mated with female BALB/cByJ mice to obtain transgenic mice. Pups that were rasH2-Tg (tg/wt) or rasH2-Wt (wt/wt) were confirmed by genotyping. Frozen embryos were preserved by the Central Institute for Experimental Animals (CIEA) and sent to two facilities, CLEA Japan and Taconic Biosciences, where the mice were produced. Production colonies are created in both facilities and supplied to customers worldwide. To prevent genetic drift, the colonies were renewed for up to 10 generations, and renewals were carried out four times every five years from 2005 to 2021. To ensure the uniformity and maintenance of the phenotype of rasH2 mice, the carcinogen susceptibilities were monitored in every renewal of colonies by CIEA based on a standard protocol of the short-term carcinogenicity study using the positive control compound N-methyl-N-nitrosourea (MNU). Furthermore, simple carcinogenicity monitoring targeting the forestomach, the organ most sensitive to MNU, was performed approximately once a year. Based on the optimally designed production and monitoring systems, the quality of rasH2 mice with reproducibility and stability of carcinogenicity is maintained and supplied globally.

Protruding structures with high expression of LGR5 are formed during regrowth phase after chemo-treatment in xenograft model of colorectal adenocarcinoma.

Recurrence after chemotherapy is one of the biggest obstacles in cancer therapy, along with metastasis. Although histopathological evaluation in preclinical models like the xenograft model help us to understand the pathophysiological process of tumor growth, there are not enough detailed histopathological analyses of such models. In this study, to learn how a tumor recovers the typical tumor structure after structural corruption during chemo-treatment, xenografted tumors originating from a patient-derived xenograft of colorectal cancer (CRC) were analyzed histopathologically over time. There were many Duct (Flattened) at Day 1 (one day after the final administration of Irinotecan), but the ratio of Duct (Columnar) and Cribriform-structures typically found in colorectal adenocarcinoma-increased over time. Finally, at Day 15 (15 days after the final administration of Irinotecan), tumor structure and size were once again the same as in the control group. LGR5, a known cancer stem cell (CSC) marker for CRC, was highly expressed on protruding structures observed from Duct (Flattened) during their transformation into Duct (Columnar) and Cribriform. In addition, these LGR5-expressing protruding structures were either Ki67 negative or positive. These results suggest that the formation of protruding structures on Duct (Flattened) is a pivotal first step in the regrowth of tumors.

Comprehensive Genomic Profiling of Neuroendocrine Carcinomas of the Gastrointestinal System.

The neuroendocrine carcinoma of the gastrointestinal system (GIS-NEC) is a rare but highly malignant neoplasm. We analyzed 115 cases using whole-genome/exome sequencing, transcriptome sequencing, DNA methylation assays, and/or ATAC-seq and found GIS-NECs to be genetically distinct from neuroendocrine tumors (GIS-NET) in the same location. Clear genomic differences were also evident between pancreatic NECs (Panc-NEC) and nonpancreatic GIS-NECs (Nonpanc-NEC). Panc-NECs could be classified into two subgroups (i.e., "ductal-type" and "acinar-type") based on genomic features. Alterations in TP53 and RB1 proved common in GIS-NECs, and most Nonpanc-NECs with intact RB1 demonstrated mutually exclusive amplification of CCNE1 or MYC. Alterations of the Notch gene family were characteristic of Nonpanc-NECs. Transcription factors for neuroendocrine differentiation, especially the SOX2 gene, appeared overexpressed in most GIS-NECs due to hypermethylation of the promoter region. This first comprehensive study of genomic alterations in GIS-NECs uncovered several key biological processes underlying genesis of this very lethal form of cancer. SIGNIFICANCE: GIS-NECs are genetically distinct from GIS-NETs. GIS-NECs arising in different organs show similar histopathologic features and share some genomic features, but considerable differences exist between Panc-NECs and Nonpanc-NECs. In addition, Panc-NECs could be classified into two subgroups (i.e., "ductal-type" and "acinar-type") based on genomic and epigenomic features. This article is highlighted in the In This Issue feature, p. 587.

Generation of a lung squamous cell carcinoma three-dimensional culture model with keratinizing structures.

Tumor nests in lung squamous cell carcinoma (LUSC) have a hierarchical structure resembling squamous epithelium. The nests consist of basal-like cells on the periphery and layers of keratinocyte-like cells that differentiate towards the center of the nest, forming keratin pearls. Reproducing this spatial heterogeneity in in vitro models would be useful for understanding the biology of LUSC. Here, we established a three-dimensional (3D) culture model with a squamous epithelial structure using LUSC cell lines PLR327F-LD41 and MCC001F, established in-house. When PLR327F-LD41 cells were cultured in a mixture of Matrigel and collagen I, they generated 3D colonies (designated cancer organoids, or COs) with involucrin (IVL)-positive keratinizing cells in the center (IVLinner COs). COs with uniform size were generated by seeding PLR327F-LD41 cells in a form of small cell aggregates. Since Notch signaling induces the differentiation of squamous epithelium, we confirmed the effect of γ-secretase inhibitor in inhibiting Notch signaling in IVLinner COs. Surprisingly, γ-secretase inhibitor did not block induction of IVL-positive cells; however, cells residing between the CK5-positive basal-like layer and IVL-positive layer decreased significantly. Thus, our 3D culture model with uniform size and structure promises to be a useful tool for elucidating the biology of LUSC and for screening drug-candidates.

Metastasis to the lateral lingual lymph node located behind the submandibular gland: A case of squamous cell carcinoma of the tongue.

Inconsistency in the definition of LLNs may be a hurdle in ensuring the accuracy of the evidence. Refinements in the classification of LLNs, based on the fascial anatomy and lymphatic vessels, are warranted.

Continuous formation of small clusters with LGR5-positive cells contributes to tumor growth in a colorectal cancer xenograft model.

New cancer characteristics can be discovered by focusing on the process of tumor formation. Cancer stem cells (CSCs) are a key subpopulation, as they are theorized to be at the apex of the tumor hierarchy. We can better understand their function in the tumor hierarchy by using sectioned samples to observe the growth of tumors from their origins as CSCs. In this study, we evaluated the growth of moderate differentiated colorectal cancer from LGR5-positive cells, which is a CSC marker of colorectal cancer, using xenograft and three-dimensional culture models spatiotemporally. These cells express LGR5 at high levels and show CSC phenotypes. To detect them, we used a previously generated antibody that specifically targets LGR5, and were therefore able to observe LGR5-positive cells aggregating into small clusters (sCLs) over the course of tumor growth. Because these LGR5-expressing sCLs formed continuously during growth mainly in the invasive front, we concluded that the structure must contribute significantly to the expansion of CSCs and to tumor growth overall. We confirmed the formation of sCLs from gland structures using a three-dimensional culture model. In addition, sCLs exhibited upregulated genes related to stress response and partial/hybrid epithelial-mesenchymal transition (EMT), as well as genes reported to be prognosis factors. Finally, sCLs with high LGR5 expression were identified in clinical samples. Based on these results, we elucidate how sCLs are an important contributors to tumor growth and the expansion of CSCs.

Examination of clinical factors affecting intrauterine microbiota.

PURPOSE: Following reports of an increase in implantation and pregnancy continuation rates by a higher percentage of Lactobacillus in the intrauterine microbiota, it has received attention in infertility treatment. This study aimed to examine Japanese women for intrauterine microbiota. METHODS: The clinical background factors in women that influence the abundance of Lactobacillus in the bacterial microbiota were examined. We included 147 patients (31 and 116 in the follicular and luteal phase, respectively), from June 2018 to June 2020, who underwent their first intrauterine microbiota test and had not used antibiotics for at least 4 weeks before the test. In the luteal phase, we compared the background factors of women in cases with 90% or more and less than 90% of Lactobacillus. Differences in the intrauterine microbiota were examined during the follicular and luteal phases. RESULTS: The proportion of Lactobacillus tended to be low among women aged 36 years and older with a history of childbirth (P = 0.0631). Some bacteria were only detected during the follicular and luteal phases, and the bacterial microbiota may change during the menstrual cycle. CONCLUSION: Bacterial microbiota in the uterus may differ between the follicular and luteal phases. Furthermore, it was shown that the rate of Lactobacillus may be lower in women (older than 36 years) who had given birth, indicating that intrauterine microbiological testing may be considered for these women in clinical practice. LAY SUMMARY: Good implantation and pregnancy continuation rates have been reported when the proportion of the bacteria Lactobacillus is high in the uterus (intrauterine) bacterial population (microbiota). In this study, we assessed whether the clinical background of Japanese women (age, history of pregnancy and childbirth, and presence of gynecological or hormonal disorders) affect the proportion of intrauterine microbiota. Intrauterine samples were collected and sequenced to evaluate the intrauterine microbiota and the composition ratio of each bacterium. Comparing the percentage of Lactobacillus in the latter phase of the menstrual cycle with the clinical background, it was found that the percentage tended to be lower in women with a history of childbirth. We compared the intrauterine microbiota between the first phase and latter phase of the menstrual cycle and revealed that it may differ between the two phases. Advances in the development of criteria for assessing intrauterine microbiota are expected.

E74-Like Factor 3 Is a Key Regulator of Epithelial Integrity and Immune Response Genes in Biliary Tract Cancer.

The transcription factor E74-like factor 3 (ELF3) is inactivated in a range of cancers, including biliary tract cancer (BTC). Here, we investigated the tumor-suppressive role of ELF3 in bile duct cells by identifying several previously unknown direct target genes of ELF3 that appear to be implicated in biliary duct carcinogenesis. ELF3 directly repressed ZEB2, a key regulator of epithelial-mesenchymal transition, and upregulated the expression of CGN, an integral element in lumen formation. Loss of ELF3 led to decreased cell-cell junctions and enhanced cell motility. ALOX5 and CXCL16 were also identified as additional direct targets of ELF3; their corresponding proteins 5-lipoxygenase and CXCL16 play a role in the immune response. Conditioned medium from cells overexpressing ELF3 significantly enhanced the migration of natural killer cells and CD8+ T cells toward the conditioned medium. Gene expression profiling for BTC expressing high levels of ELF3 revealed significant enrichment of the ELF3-related genes. In a BTC xenograft model, activation of ELF3 increased expression of ELF3-related genes, enhanced the tubular structure of the tumors, and led to a loss of vimentin. Overall, our results indicate that ELF3 is a key regulator of both epithelial integrity and immune responses in BTC. SIGNIFICANCE: Thease finding shows that ELF3 regulates epithelial integrity and host immune responses and functions as a tumor suppressor in biliary tract cancer. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/2/489/F1.large.jpg.

Patient-derived xenograft (PDX) models: characteristics and points to consider for the process of establishment.

Tumor research has largely relied on xenograft models created by the engraftment of cultured cell lines derived from tumor tissues into immunodeficient mice for in vivo studies. Like in vitro models, such models retain the ability of tumor cells to continuously proliferate, so they have been used to predict the clinical relevance of studies on proliferating cells. However, these models are composed of a limited population of tumor cells, which include only those tumor cells that are able to adapt to culture conditions, and thus they do not reflect the diversity and heterogeneity of tumors. This, at least in part, explains the poor predictivity of non-clinical data in the research and development of molecularly targeted drugs. Recently, research focus has been directed towards patient-derived xenograft (PDX) models created by directly engrafting tumor tissues, which have not been cultured in vitro, into immunodeficient mice. PDX models reflect the diversity and heterogeneity of tumors, and the evidence they provide can be verified in the patient tissues from which they were derived originally. PDX models are anticipated to efficiently bridge non-clinical and clinical data in translational research. Based on the evidence obtained from our research experience, this review describes the characteristics of PDX models for acting as tumor models, and elucidates the points to consider when attempting to establish these models.

Tissue-specific effects of an anti-desmoglein-3 ADCC antibody due to expression of the target antigen and physiological characteristics of the mouse vagina.

Desmoglein-3 (DSG3) is a potential target of cytotoxic antibody therapy for squamous cell carcinomas but is also expressed in various normal squamous epithelia. We obtained information about DSG3 distribution in mouse tissues by immunohistochemistry and conducted an intravenous multiple-dose study in mouse to estimate the toxic potential of anti-DSG3 therapy. DSG3 was expressed in the squamous epithelium of several organs including the skin, esophagus, tongue, forestomach, eye, and vagina. It was expressed at all estrous cycles of the vagina with changes in distribution patterns along with the structural changes in each cycle, and expression was reduced in ovariectomized (OVX) mice. On the administration of the antibody, there was disarrangement of the vaginal mucosal epithelium with formation of miroabscess, increased granulocyte infiltration, and single cell necrosis. Despite similar expression levels of DSG3 in other tissues, histopathological changes were limited to the vagina. The severity of the changes was reduced by ovariectomy. From these findings, the lesions were thought to be related to the drastic change in the histological structure of the vaginal mucosa accompanying the estrous cycle. Thus, we have shown that the changing expression of target antigen distribution and its relationship with physiological changes in tissue structure are important features for estimating the toxic potential of cytotoxic antibody therapy.