Heterogeneity of cancer stem cell-related marker expression is associated with three-dimensional structures in malignant pleural effusion produced by lung adenocarcinoma.

INTRODUCTION: Cancer stem cells have been described in lung adenocarcinoma-associated malignant pleural effusion. They show clinically important features, including the ability to initiate new tumours and resistance to treatments. However, their correlation with the three-dimensional tumour structures in the effusion is not well understood. METHODS: Cell blocks produced from lung adenocarcinoma patients' pleural effusion were examined for cancer stem cell-related markers Nanog and CD133 using immunocytochemistry. The three-dimensional cancer cell structures and CD133 expression patterns were visualized with tissue-clearing technology. The expression patterns were correlated with tumour cell structures, genetic variants and clinical outcomes. RESULTS: Thirty-nine patients were analysed. Moderate-to-strong Nanog expression was detected in 27 cases (69%), while CD133 was expressed by more than 1% of cancer cells in 11 cases (28%). Nanog expression was more homogenous within individual specimens, while CD133 expression was detected in single tumour cells or cells within small clusters instead of larger structures in 8 of the 11 positive cases (73%). Although no statistically significant correlation between the markers and tumour genetic variants or patient survival was observed, we recorded seven cases with follow-up specimens after cancer treatment, and four (57%) showed a change in stem cell-related marker expression corresponding to treatment response. CONCLUSIONS: Lung adenocarcinoma cells in the pleural effusion show variable expression of cancer stem cell-related markers, some showing a correlation with the size of cell clusters. Their expression level is potentially correlated with cancer treatment effects.

Anti-Atopic Dermatitis Activity of Epi-Oxyzoanthamine Isolated from Zoanthid.

Atopic dermatitis (AD, eczema) is a condition that causes dry, itchy, and inflamed skin and occurs most frequently in children but also affects adults. However, common clinical treatments provide limited relief and have some side effects. Therefore, there is a need to develop new effective therapies to treat AD. Epi-oxyzoanthamine is a small molecule alkaloid isolated from Formosan zoanthid. Relevant studies have shown that zoanthamine alkaloids have many pharmacological and biological activities, including anti-lymphangiogenic functions. However, there are no studies on the use of epi-oxyzoanthamine on the skin. In this paper, epi-oxyzoanthamine has been shown to have potential in the treatment of atopic dermatitis. Through in vitro studies, it was found that epi-oxyzoanthamine inhibited the expression of cytokines in TNF-α/IFN-γ-stimulated human keratinocyte (HaCaT) cells, and it reduced the phosphorylation of MAPK and the NF-κB signaling pathway. Atopic dermatitis-like skin inflammation was induced in a mouse model using 2,4-dinitrochlorobenzene (DNCB) in vivo. The results showed that epi-oxyzoanthamine significantly decreased skin barrier damage, scratching responses, and epidermal hyperplasia induced by DNCB. It significantly reduced transepidermal water loss (TEWL), erythema, ear thickness, and spleen weight, while also increasing surface skin hydration. These results indicate that epi-oxyzoanthamine from zoanthid has good potential as an alternative medicine for treating atopic dermatitis or other skin-related inflammatory diseases.

Adult NTRK-rearranged spindle cell neoplasms of the viscera: with an emphasis on rare locations and heterologous elements.

NTRK-rearranged mesenchymal neoplasms mostly affect the soft tissues of pediatric patients. Given the responsiveness to selective NTRK inhibitors, it remains critical to identify those ultra-rare cases occurring in the viscera of adults. In five females and two males aged 18-53 years, we characterized visceral mesenchymal tumors harboring TPM3-NTRK1 [uterine cervix (N = 2), pleura, prostate], LMNA-NTRK1 (lung), SQSTM1-NTRK3 (heart), and NTRK3 rearrangement with unknown fusion partner (colon/mesocolon) with RNA sequencing, FISH, RT-PCR, and immunohistochemistry. The tumors exhibited spindled to ovoid/epithelioid or pleomorphic cells, often arranged in fascicles, and were low-to-intermediate-grade and high-grade in three and four cases, respectively. Keloid-like stromal collagen and perivascular hyalinization was noted in five. Adenosarcoma-like appearances were observed in two, manifesting frond-like protrusions in one cervical tumor and phyllodes-like architecture in the prostatic tumor. Abrupt high-grade transformation into pleomorphic liposarcoma was found in another cervical tumor, while the pleural tumor contained intermixed rhabdomyoblasts. Pan-TRK immunostaining was positive in all cases. All cases expressed CD34, while five were S100-positive. CDKN2A homozygous deletion with concomitant p16 loss occurred in 4/7. Whole-exome sequencing identified TP53 mutation (c.672+2T>C, involving a splice site, with concomitant protein loss) in a cervical sarcoma, limited to its heterologous liposarcomatous component. At least moderate pan-TRK immunoreactivity was present in varying proportions of potential pathologic mimics, with BCOR-positive sarcoma (56%, 5/9), undifferentiated uterine sarcoma (50%, 3/6), and spindle cell/sclerosing rhabdomyosarcoma (33%, 2/6) being among the most frequent. This underscored the unsatisfactory specificity of pan-TRK immunohistochemistry and warranted molecular confirmation in the diagnosis of adult NTRK-rearranged visceral mesenchymal neoplasms. The current report highlights the ever-expanding clinicopathologic and genetic spectrum of this entity by describing the unprecedented cardiac and pleural locations and heterologous differentiation, as well as the second NTRK-rearranged "prostatic stromal sarcoma," while substantiating CDKN2A deletion as a frequent occurrence.

Effect of Neferine on DNCB-Induced Atopic Dermatitis in HaCaT Cells and BALB/c Mice.

Atopic dermatitis (AD) is a chronic and persistent inflammatory skin disease characterized by eczematous lesions and itching, and it has become a serious health problem. However, the common clinical treatments provide limited relief and are accompanied by adverse effects. Therefore, there is a need to develop novel and effective therapies to treat AD. Neferine is a small molecule compound isolated from the green embryo of the mature seeds of lotus (Nelumbo nucifera). It has a bisbenzylisoquinoline alkaloid structure. Relevant studies have shown that neferine has many pharmacological and biological activities, including anti-inflammatory, anti-thrombotic, and anti-diabetic activities. However, there are very few studies on neferine in the skin, especially the related effects on inflammatory skin diseases. In this study, we proved that it has the potential to be used in the treatment of atopic dermatitis. Through in vitro studies, we found that neferine inhibited the expression of cytokines and chemokines in TNF-α/IFN-γ-stimulated human keratinocyte (HaCaT) cells, and it reduced the phosphorylation of MAPK and the NF-κB signaling pathway. Through in vivo experiments, we used 2,4-dinitrochlorobenzene (DNCB) to induce atopic dermatitis-like skin inflammation in a mouse model. Our results show that neferine significantly decreased the skin barrier damage, scratching responses, and epidermal hyperplasia induced by DNCB. It significantly decreased transepidermal water loss (TEWL), erythema, blood flow, and ear thickness and increased surface skin hydration. Moreover, it also inhibited the expression of cytokines and the activation of signaling pathways. These results indicate that neferine has good potential as an alternative medicine for the treatment of atopic dermatitis or other skin-related inflammatory diseases.

Anti-Allergic and Anti-Inflammatory Effects of Neferine on RBL-2H3 Cells.

Mast cells play a very important role in skin allergy and inflammation, including atopic dermatitis and psoriasis. In the past, it was found that neferine has anti-inflammatory and anti-aging effects on the skin, but its effect on mast cells has not yet been studied in detail. In this study, we used mast cells (RBL-2H3 cells) and mouse models to study the anti-allergic and inflammatory effects of neferine. First, we found that neferine inhibits the degranulation of mast cells and the expression of cytokines. In addition, we observed that when mast cells were stimulated by A23187/phorbol 12-myristate-13-acetate (PMA), the elevation of intracellular calcium was inhibited by neferine. The phosphorylation of the MAPK/NF-κB pathway is also reduced by pretreatment of neferine. The results of in vivo studies show that neferine can improve the appearance of dermatitis and mast cell infiltration caused by dinitrochlorobenzene (DNCB). Moreover, the expressions of barrier proteins in the skin are also restored. Finally, it was found that neferine can reduce the scratching behavior caused by compound 48/80. Taken together, our results indicate that neferine is a very good anti-allergic and anti-inflammatory natural product. Its effect on mast cells contributes to its pharmacological mechanism.

The expanding morphological and genetic spectrum of MYOD1-mutant spindle cell/sclerosing rhabdomyosarcomas: a clinicopathological and molecular comparison of mutated and non-mutated cases.

AIMS: Spindle cell/sclerosing rhabdomyosarcomas (SC/SRMS) feature spindled and/or rounded rhabdomyosarcomatous cells within variably hyalinised stroma. Only 30-67% of SC/SRMSs harbour neomorphic MYOD1 p.L122R mutations, indicating heterogeneity in this RMS type. We compared MYOD1-mutant and non-mutant cases to characterise the histological and genetic spectrum of mutated SC/SRMS. METHODS AND RESULTS: Seventeen RMSs with spindled, sclerosing or hybrid histology were sequenced to identify MYOD1 and PIK3CA mutations and reappraised to assess histological features and myogenic immunophenotypes. Twelve SC/SRMSs harboured MYOD1 mutations, including homozygous p.L122R (n = 8), heterozygous p.L122R (n = 3) and heterozygous p.E118K (n = 1). MYOD1-mutant tumours affected nine females and three males aged 8-64 years (median = 22.5), had a median size of 4.2 cm (range = 2-22) and involved the head and neck (n = 7), extremities (n = 4) and mediastinum (n = 1). Fascicular/spindle histology was predominant in four cases, including one with heterologous lipoblasts in focally myxoid stroma. Four sclerosing cases mainly comprised rounded cells, including one with multinucleated tumour cells. Four cases were histologically hybrid. The only PIK3CA (p.H1047R) mutation was detected in a predominantly spindled MYOD1-p.L122R-mutated case, but not in its laser-microdissected lipoblast-containing area. All MYOD1-mutant cases exhibited diffuse MYOD1 expression but patchy myogenin reactivity. At final follow-up (median = 13.5 months), recurrences (n = 4), metastases (n = 2) or both (n = 1) occurred in seven MYOD1-mutant cases; one had died of disease. Five non-mutated cases were reclassified as spindle embryonal (n = 3), dense embryonal (n = 1) and unclassifiable (n = 1) RMSs. CONCLUSION: MYOD1-mutant RMSs are uncommonly mutated with PIK3CA and behave aggressively with an expanded morphological and genetic spectrum, including lipoblastic differentiation, multinucleated cells and the alternative p.E118K mutation.

Targeting tumor-infiltrating Ly6G+ myeloid cells improves sorafenib efficacy in mouse orthotopic hepatocellular carcinoma.

Sorafenib, a multikinase inhibitor with antiangiogenic activity, is an approved therapy for hepatocellular carcinoma (HCC). It is unclear whether the proinflammatory and immunosuppressive mechanisms may limit the therapeutic efficacy of sorafenib in HCC. We used a syngeneic mouse liver cancer cell line to establish orthotopic liver or subcutaneous tumors to study how proinflammatory and immunosuppressive mechanisms impact on the efficacy of sorafenib. We found sorafenib exhibited a potent therapeutic effect in subcutaneous tumors, but a less potent effect in orthotopic liver tumors. The protein levels of interleukin-6 (IL-6) and vascular endothelial growth factor A (VEGF-A) were persistently elevated in orthotopic liver tumors, but not in subcutaneous tumors, treated with sorafenib. Likewise, the tumor-infiltrating Ly6G+ myeloid-derived suppressor cells (MDSCs) and immune suppressors were increased in orthotopic liver tumors, not in subcutaneous tumors, treated with sorafenib. The tumor-infiltrating Ly6G+ MDSCs of sorafenib-treated orthotopic liver tumors significantly induced IL-10 and TGF-ß expressing CD4+ T cells, and downregulated the cytotoxic activity of CD8+ T cells. IL-6, but not VEGF-A, protected Ly6G+ MDSCs from sorafenib-induced cell death in vitro. The combination of anti-Ly6G antibody or anti-IL-6 antibody with sorafenib significantly reduced the cell proportion of Ly6G+ MDSCs in orthotopic liver tumors, enhanced the T cells proliferation and improved the therapeutic effect of sorafenib synergistically. Modulating tumor microenvironment through targeting tumor-infiltrating Ly6G+ MDSCs represents a potential strategy to improve the anti-HCC efficacy of sorafenib.

Giant cell tumor of soft tissue is genetically distinct from its bone counterpart.

Giant cell tumors of bone are locally aggressive bone neoplasms with a predilection for young adults. Histologically, they are composed of histiocytoid to spindled mononuclear cells, admixed with numerous large osteoclastic giant cells. Giant cell tumors of soft tissue are rare tumors that bear striking histological resemblance to giant cell tumors of bone and might be regarded as a soft tissue analog thereof. Point mutations of the H3F3A gene (coding for a histone H3.3 protein) at the Gly34 codon, mostly G34W resulting from a GGG>TGG nucleotide change, have recently been identified in a vast majority of giant cell tumors of bone. To delineate the possible pathogenic linkage between both tumor types, we analyzed the H3F3A genotypes in a series of 15 giant cell tumors of soft tissue by Sanger sequencing and found no mutation in any case. We then sequenced cognate histone H3 genes with an identical nucleotide sequence ('GGG') at the codon Gly34, including the H3F3B, H3F3C, HIST2H3A, HIST2H3C, and HIST2H3D genes, and no somatic mutation was detected. These results reveal that giant cell tumors of soft tissue are probably genetically distinct from their bone counterparts and suggest that they might be pathogenically unrelated. Given the prominence of non-neoplastic cells in these tumors and the limitations of the current study, however, analyses using more sensitive techniques might be required to solve the issue.

Longitudinal Melanonychia: Differences in Etiology Are Associated with Patient Age at Diagnosis.

BACKGROUND: Longitudinal melanonychia (LM) may occur as a result of nail apparatus melanoma. Knowledge of etiology plays an important role in the management of LM. OBJECTIVES: The study is aimed to compare the diagnosis of LM in different age groups. METHODS: We collected 63 cases (45 adults and 18 children) with LM who underwent nail matrix biopsy or excision in a 21-year cohort and assessed their clinicopathological features. RESULTS: Melanomas in adults and children were 40% and none, while nevi accounted for 15.6% in adults and 94.4% in children. There was a statistically significant difference between the average age at diagnosis for melanoma (54.5 ± 13.3 years) and nevus (15.2 ± 18.5 years). Logistic regression related the occurrence of melanoma to older ages with a relative risk of 1.2 compared to nevus, but no cutoffs between age groups could be defined between LM-associated nevus and melanoma. CONCLUSION: The adult group has a significantly higher risk of melanoma, while children with LM show mostly nonmelanoma etiologies. Tissue proof is more warranted in adult cases, and it is needed in selected cases of children with LM.

Genomewide copy number analysis of Müllerian adenosarcoma identified chromosomal instability in the aggressive subgroup.

Müllerian adenosarcomas are malignant gynecologic neoplasms. Advanced staging and sarcomatous overgrowth predict poor prognosis. Because the genomic landscape remains poorly understood, we conducted this study to characterize the genomewide copy number variations in adenosarcomas. Sixteen tumors, including eight with and eight without sarcomatous overgrowth, were subjected to a molecular inversion probe array analysis. Copy number variations, particularly losses, were significantly higher in cases with sarcomatous overgrowth. Frequent gains of chromosomal 12q were noted, often involving cancer-associated genes CDK4 (six cases), MDM2, CPM, YEATS4, DDIT3, GLI1 (five each), HMGA2 and STAT6 (four), without association with sarcomatous overgrowth status. The most frequent losses involved chromosomes 13q (five cases), 9p, 16q and 17q (four cases each) and were almost limited to cases with sarcomatous overgrowth. MDM2 and CDK4 amplification, as well as losses of RB1 (observed in two cases) and CDKN2A/B (one case), was verified by FISH. By immunohistochemistry, all MDM2/CDK4-coamplified cases were confirmed to overexpress both encoded proteins, whereas all four cases with (plus an additional four without) gain of HMGA2 overexpressed the HMGA2 protein. Both cases with RB1 loss were negative for the immunostaining of the encoded protein. Chromothripsis-like copy number profiles involving chromosome 12 or 14 were observed in three fatal cases, all of which harbored sarcomatous overgrowth. With whole chromosome painting and deconvolution fluorescent microscopy, dividing tumor cells in all three cases were shown to have scattered extrachromosomal materials derived from chromosomes involved by chromothripsis, suggesting that this phenomenon may serve as visual evidence for chromothripsis in paraffin tissue. In conclusion, we identified frequent chromosome 12q amplifications, including loci containing potential pharmacological targets. Global chromosomal instability and chromothripsis were more frequent in cases with sarcomatous overgrowth. To our knowledge, this is the first time that evidence of chromothripsis has been demonstrated in paraffin-embedded clinical tissues and in adenosarcomas.

Identification of a novel FN1-FGFR1 genetic fusion as a frequent event in phosphaturic mesenchymal tumour.

Phosphaturic mesenchymal tumours (PMTs) are uncommon soft tissue and bone tumours that typically cause hypophosphataemia and tumour-induced osteomalacia (TIO) through secretion of phosphatonins including fibroblast growth factor 23 (FGF23). PMT has recently been accepted by the World Health Organization as a formal tumour entity. The genetic basis and oncogenic pathways underlying its tumourigenesis remain obscure. In this study, we identified a novel FN1-FGFR1 fusion gene in three out of four PMTs by next-generation RNA sequencing. The fusion transcripts and proteins were subsequently confirmed with RT-PCR and western blotting. Fluorescence in situ hybridization analysis showed six cases with FN1-FGFR1 fusion out of an additional 11 PMTs. Overall, nine out of 15 PMTs (60%) harboured this fusion. The FN1 gene possibly provides its constitutively active promoter and the encoded protein's oligomerization domains to overexpress and facilitate the activation of the FGFR1 kinase domain. Interestingly, unlike the prototypical leukaemia-inducing FGFR1 fusion genes, which are ligand-independent, the FN1-FGFR1 chimeric protein was predicted to preserve its ligand-binding domains, suggesting an advantage of the presence of its ligands (such as FGF23 secreted at high levels by the tumour) in the activation of the chimeric receptor tyrosine kinase, thus effecting an autocrine or a paracrine mechanism of tumourigenesis.

TERT promoter mutation is uncommon in acral lentiginous melanoma.

BACKGROUND: Melanoma is a heterogeneous group of diseases with distinct sets of genetic changes. Recurrent and mutually exclusive C>T or CC>TT transition mutations were identified in the promoter region of the reverse transcriptase catalytic subunit of the telomerase gene (TERT) in melanoma recently, and it was suggested that they enhanced the expression of TERT gene and played important roles in the melanoma pathogenesis. These mono or di-nucleotide transitions were ultraviolet (UV)-signature mutations. METHODS: In this study, polymerase chain reaction and direct sequencing of TERT promoter using formalin-fixed and paraffin-embedded tissue was performed to investigate whether these UV-signature mutations were also present in acral lentiginous melanoma. RESULTS: TERT promoter mutation was identified in only 2 of the 32 cases (6%) of acral lentiginous melanomas while it was identified in 3 of the 9 cases (33%) of non-acral cutaneous melanomas. The difference was statistically significant (p = 0.028). CONCLUSIONS: The rarity of TERT promoter mutation in the acral lentiginous melanoma was consistent with the supposed role of UV light in the melanoma pathogenesis and also corroborated the view that acral lentiginous melanomas have a different pathogenesis with the melanomas from sun-exposed sites.

Dedifferentiated liposarcoma with homologous lipoblastic differentiation: expanding the spectrum to include low-grade tumours.

AIMS: Dedifferentiated liposarcoma (DDLPS) is traditionally defined as a non-lipogenic high-grade sarcoma arising from a well-differentiated liposarcoma that confers metastatic potential. Recently, DDLPSs with lipoblastic differentiation, i.e. morphologically lipogenic DDLPSs, were reported. Because of the lipoblastic differentiation, these tumours caused confusion, and were reported under different names. However, cytogenetic and molecular studies have revealed their DDLPS nature. So far, the cases reported have been high-grade pleomorphic liposarcoma-like tumours. In this study we have collected another series that contains low-grade tumours, and expand the histological spectrum. METHODS AND RESULTS: Eighteen cases of DDLPS with lipoblastic differentiation from various anatomical locations were analysed by routine histology, immunohistochemistry, and MDM2 fluorescence in-situ hybridization. Two main histological patterns were seen: one featured a spindle cell sarcoma containing lipoblasts with variable nuclear pleomorphism, and the other a pleomorphic liposarcoma-like tumour including the epithelioid variant. Two cases showed low nuclear grade and lipogenic activity in the metastatic foci. CDK4, MDM2 and p16(INK) (4a) overexpression was seen in all except one case. MDM2 amplification was found in all 16 cases tested. CONCLUSIONS: We have expanded the spectrum of this variant of DDLPS to include low-grade tumours, in which a careful search for increased mitotic activity is essential. Like conventional DDLPS, these tumours are capable of metastasis.

Prognostic value of PD-L1 expression on immune cells or tumor cells for locally advanced esophageal squamous cell carcinoma in patients treated with neoadjuvant chemoradiotherapy.

PURPOSE: Programmed death-ligand 1 (PD-L1) expression may influence the prognosis of patients with localized esophageal cancer. The current study compared the prognostic value of PD-L1 expression between tumor cells and immune cells. METHODS: Archival esophageal tumor tissue samples were collected from patients who received paclitaxel and cisplatin-based neoadjuvant chemoradiotherapy (CRT) for locally advanced esophageal squamous cell carcinoma (ESCC) in three prospective phase II trials. PD-L1 expression on tumor and immune cells was examined immunohistochemically by using the SP142 antibody and scored by two independent pathologists. The association of PD-L1 expression with patient's outcomes was analyzed using a log-rank test and Cox regression multivariate analysis. RESULTS: A total of 100 patients were included. PD-L1 expression on tumor cells was positive (≥ 1%, TC-positive) in 55 patients; PD-L1 expression on immune cells was high (≥ 5%, IC-high) in 30 patients. TC-positive status was associated with poor overall survival (OS) (HR: 1.63, P = 0.035), whereas IC-high status was associated with improved OS (HR: 0.44, P = 0.0024). Multivariate analysis revealed that TC-positive, IC-high, and performance status were independent prognostic factors for progression-free survival and that IC-high and performance status were independent factors for OS. Furthermore, the combination of IC-high and TC-negative status was associated with the optimal OS, whereas that of TC-positive and IC-low status was associated with the worst OS. CONCLUSION: PD-L1 expression on tumor and immune cells may have different prognostic value for patients with locally advanced ESCC receiving neoadjuvant CRT. A combination of these two indexes may further improve the prognostic prediction.

Deep Convolutional Neural Networks Detect Tumor Genotype from Pathological Tissue Images in Gastrointestinal Stromal Tumors.

Gastrointestinal stromal tumors (GIST) are common mesenchymal tumors, and their effective treatment depends upon the mutational subtype of the KIT/PDGFRA genes. We established deep convolutional neural network (DCNN) models to rapidly predict drug-sensitive mutation subtypes from images of pathological tissue. A total of 5153 pathological images of 365 different GISTs from three different laboratories were collected and divided into training and validation sets. A transfer learning mechanism based on DCNN was used with four different network architectures, to identify cases with drug-sensitive mutations. The accuracy ranged from 87% to 75%. Cross-institutional inconsistency, however, was observed. Using gray-scale images resulted in a 7% drop in accuracy (accuracy 80%, sensitivity 87%, specificity 73%). Using images containing only nuclei (accuracy 81%, sensitivity 87%, specificity 73%) or cytoplasm (accuracy 79%, sensitivity 88%, specificity 67%) produced 6% and 8% drops in accuracy rate, respectively, suggesting buffering effects across subcellular components in DCNN interpretation. The proposed DCNN model successfully inferred cases with drug-sensitive mutations with high accuracy. The contribution of image color and subcellular components was also revealed. These results will help to generate a cheaper and quicker screening method for tumor gene testing.

Improving Algorithm for the Alignment of Consecutive, Whole-Slide, Immunohistochemical Section Images.

BACKGROUND: Accurate and precise alignment of histopathology tissue sections is a key step for the interpretation of the proteome topology and cell level three-dimensional (3D) reconstruction of diseased tissues. However, the realization of an automated and robust method for aligning nonglobally stained immunohistochemical (IHC) sections is still challenging. In this study, we aim to assess the feasibility of multidimensional graph-based image registration on aligning serial-section and whole-slide IHC section images. MATERIALS AND METHODS: An automated, patch graph-based registration method was established and applied to align serial, whole-slide IHC sections at ×10 magnification (average 32,947 × 27,054 pixels). The alignment began with the initial alignment of high-resolution reference and translated images (object segmentation and rigid registration) and nonlinear registration of low-resolution reference and translated images, followed by the multidimensional graph-based image registration of the segmented patches, and finally, the fusion of deformed patches for inspection. The performance of the proposed method was formulated and evaluated by the Hausdorff distance between continuous image slices. RESULTS: Sets of average 315 patches from five serial whole slide, IHC section images were tested using 21 different IHC antibodies across five different tissue types (skin, breast, stomach, prostate, and soft tissue). The proposed method was successfully automated to align most of the images. The average Hausdorff distance was 48.93 µm with a standard deviation of 14.94 µm, showing a significant improvement from the previously published patch-based nonlinear image registration method (average Hausdorff distance of 93.89 µm with 50.85 µm standard deviation). CONCLUSIONS: Our method was effective in aligning whole-slide tissue sections at the cell-level resolution. Further advancements in the screening of the proteome topology and 3D tissue reconstruction could be expected.

Loss of SFRP1 expression is a key progression event in gastrointestinal stromal tumor pathogenesis.

The mechanism of high-grade transformation in gastrointestinal stromal tumors (GISTs) remains to be clarified. We aim to discover the key progression events by studying biphasic GISTs. The study group included 101 GISTs. Nineteen of these had been screened from 263 GISTs to represent the early stage of GIST high-grade transformation, characterized by juxtaposed low-grade and high-grade regions in the same tumor (so-called biphasic GISTs). Mutational analyses, fluorescence in situ hybridization (FISH), NanoString analyses, telomere analysis, and gene expression profiling were carried out, followed by in silico analyses, cell line study, and immunohistochemical validation. Using gene expression analysis, downregulation of SFRP1 was revealed to be the main event in GIST high-grade transformation (p = 0.013), accompanied by upregulation of EZH2. In silico analyses revealed that downregulation of SFRP1 was a common feature in GIST progression across several different series. Immunohistochemically, the expression of SFRP1 was validated to be significantly lower in high-grade GISTs (WHO risk group 3a or higher) than in low-grade GISTs (p < 0.001), and attenuation/loss of SFRP1 was associated with GIST tumor progression (p < 0.001). By NanoString and FISH analyses, chromosomal 9/9p loss was the only recurrent large-scale chromosome aberration in biphasic GISTs, with a correlation with SFRP1 downregulation. Subclones containing chromosome 9/9p loss could be appreciated in the low-grade parts of biphasic GISTs. TP53 mutation, RB1 loss, KIT/PDGFRA mutation, and alternative lengthening of telomeres did not play a significant role in GIST high-grade transformation. In conclusion, high-grade transformation of GISTs features SFRP1 downregulation and chromosome 9/9p loss.

Trisomy 18 is a consistent cytogenetic feature in pilomatricoma.

Pilomatricoma, also known as 'calcifying epithelioma of Malherbe', is a common skin adnexal tumor that mimics hair growth. Its proliferating cells seem distinctly programmed to undergo terminal differentiation and death. We report the first cytogenetic investigations of pilomatricoma. Trisomy 18 was shown, in an index case, by G-banded karyotyping. This aberration was corroborated by interphase fluorescence in situ hybridization, using a chromosome 18 pericentromeric probe, in the basaloid epithelial component of 7 of 11 pilomatricomas, including the index case. Trisomy 18 was present in a small subset of cells, suggesting a role in pilomatricoma progression, rather than in tumor initiation. We conclude that trisomy 18 is a consistent feature in pilomatricoma, suggesting that genes carried on this chromosome, such as that for the antiapoptotic oncoprotein BCL2, may have a role in the growth and differentiation of this benign self-limited tumor.

The utility of discovered on gastrointestinal stromal tumor 1 (DOG1) antibody in surgical pathology-the GIST of it.

DOG1 (discovered on GIST 1), known also as TMEM16A and ANO1, has emerged in recent years as a promising biomarker for gastrointestinal stromal tumors (GIST). It was originally discovered through microarray expression profiling analysis as gene that is highly expressed in GIST, and subsequent immunohistochemical studies have shown its use in its diagnosis. The results from several series have shown a high overall sensitivity and specificity for DOG1 in the detection of GISTs and about 6% of GISTs overall exhibiting a DOG1+/KIT-immunoprofile. DOG1 antibodies are more sensitive than KIT antibodies in detecting tumors of gastric origin, tumors with epithelioid morphology, and tumors harboring PDGFRA mutation. Furthermore, DOG1 immunoreactivity is rarely observed in other mesenchymal and nonmesenchymal tumor types. These results support the use of DOG1 as a diagnostic biomarker for GIST. When used in combination with KIT, this panel of diagnostic biomarkers can help pathologists and clinicians to identify more patients who may benefit from targeted therapies.