Quality assessment of tissue samples stored in a specialized human lung biobank.

Human sample, from patients or healthy donors, are a valuable link between basic research and clinic. Especially in translational research, they play an essential role in understanding development and progression of diseases as well as in developing new diagnostic and therapeutic tools. Stored in biobanks, fast access to appropriate material becomes possible. However, biobanking in a clinical context faces several challenges. In practice, collecting samples during clinical routine does not allow to strictly adhere to protocols of sample collection in all aspects. This may influence sample quality to variable degrees. Time from sample draw to asservation is a variable factor, and influences of prolonged storage at ambient temperature of tissues are not well understood. We investigated whether delays between 5 minutes and 3 hours, and the use of RNAlater RNA-preserving reagent would lead to a relevant drop in sample quality, measured by quantitative mRNA expression analysis. Our findings suggest that even under ambient conditions, delays up to 3 hours do not have a major impact on sample quality as long as the tissue remains intact.

Comprehensive clinical profiling of the Gauting locoregional lung adenocarcinoma donors.

A comprehensive characterization of lung adenocarcinoma (LADC) clinical features is currently missing. We prospectively evaluated Caucasian patients with early-stage LADC. Patients with LADC diagnosed between 2011 and 2015 were prospectively assessed for lung resection with curative intent. Fifty clinical, pathologic, radiologic, and molecular variables were recorded. Patients were followed till death/study conclusion. The main findings were compared to a separate cohort from France. Of 1943 patients evaluated, 366 were enrolled (18.8%; 181 female; 75 never-smokers; 28% of registered Bavarian cases over the study period). Smoking and obstruction were significantly more prevalent in GLAD compared with adult Bavarians (P < 0.0001). Ever-smoker tumors were preferentially localized to the upper lobes. We observed 120 relapses and 74 deaths over 704 cumulative follow-up years. Median overall and disease-free survival were >7.5 and 3.6 years, respectively. Patients aged <45 or >65 years, resected >60 days postdiagnosis, with abnormal FVC/DLCO VA , N2/N3 stage, or solid histology had significantly decreased survival estimates. These were fit into a weighted locoregional LADC death risk score that outperformed pTNM7 in predicting survival in the GLAD and in our second cohort. We define the clinical gestalt of locoregional LADC and provide a new clinical tool to predict survival, findings that may aid future management and research design.

Comprehensive Biomarker Analyses in Patients with Advanced or Metastatic Non-Small Cell Lung Cancer Prospectively Treated with the Polo-Like Kinase 1 Inhibitor BI2536.

BACKGROUND: Polo like kinase 1 (PLK1) is frequently upregulated in tumors and is thus viewed as a promising therapeutic target in various cancers. Several PLK1 inhibitors have recently been developed and clinically tested in solid cancers, albeit with limited success. So far, no predictive biomarkers for PLK1 inhibitors have been established. To this end, we conducted a post-hoc biomarker analysis of tumor samples from non-small cell lung cancer (NSCLC) patients treated with the PLK1 inhibitor BI2536 in a phase II study. METHODS: We analyzed formalin-fixed paraffin-embedded surplus tumor tissue from 47 study patients using immunohistochemistry (IHC) and DNA sequencing of KRAS, EGFR, BRAF, and PIK3CA. RESULTS: KRAS-mutated patients showed numerically prolonged progression-free survival, but statistical significance was not established. Interestingly, when pathways rather than single genes were analyzed, a positive correlation between IHC staining of activated ERK (p-ERK) and mutated KRAS was detected, whereas KRAS mutation status was found to be negatively correlated with activated AKT (p-AKT). CONCLUSION: With this hypothesis-generating study in BI2531-treated patients, we could not establish a correlation between KRAS mutations and relevant clinical endpoints. Future clinical trials with concomitant systematic biosampling and comprehensive molecular analyses are required to identify biomarkers predictive for response to PLK1 inhibitors.

Metformin Triggers Autophagy to Attenuate Drug-Induced Apoptosis in NSCLC Cells, with Minor Effects on Tumors of Diabetic Patients.

The biologic plausibility of an association between type 2 diabetes mellitus (T2D) and lung cancer has received increasing attention, but the results of investigations remain largely inconclusive. In the present study we investigated the influence of the anti-diabetic drug metformin on the cytotoxic effects of EGFR targeted therapy and chemotherapy in 7 non-small cell lung cancer (NSCLC) cell lines and a cohort of lung cancer patients with/without T2D. In vitro cell viability assays indicated that metformin didn't potentiate the growth inhibitory effects of erlotinib at different doses in cell lines that are of distinct genetic background. EGFR downstream signaling evaluation further demonstrated that metformin, at its IC50 value, modified apoptosis caused in erlotinib or chemotherapeutic agent-treated cells via AKT activation and the inhibition of caspase 3 and PARP cleavages. These regulations were driven independently from EGFR, LKB1, KRAS, PTEN and p53 status. Metformin triggered autophagy (LC3B expression) was identified to interplay with apoptosis to attenuate the drug effect and postpone cancer cell death. In the retrospective study of 8 NSCLC patients, the administration of metformin did not induce statistically significant changes as assessed by immunohistochemical staining of pERK, pAKT and cleaved PARP. Consequently, the application of metformin for T2D NSCLC patients receiving chemo or EGFR targeted therapy should be considered with caution.

EWSR1-fusion-negative, SMARCB1-deficient primary pulmonary myxoid sarcoma.

Primary pulmonary myxoid sarcoma (PPMS) is a recently defined rare neoplasm with histological and molecular similarity to extraskeletal myxoid chondrosarcoma. To date, 20 cases have been reported. A 48-year-old man presented with a huge mass filling the right hemithorax and extending into the tracheobronchial system. Histological findings were consistent with PPMS. Immunohistochemistry was positive for vimentin, CD10, and EMA, but other lineage-specific markers were negative. SMARCB1 (INI1) expression was lost in the tumour cells. FISH analysis (EWSR1, FUS, NR4A3, and SMARCB1) revealed no abnormalities. This case suggests SMARCB1 loss as a possible alternative molecular event driving EWSR1-negative PPMS.

Genetic changes of non-small cell lung cancer under neoadjuvant therapy.

BACKGROUND: Large scale sequencing efforts defined common molecular alterations in non-small cell lung cancer (NSCLC) and revealed potentially druggable mutations. Yet, systematic data on the changes of the respective molecular profiles under standard therapy in NSCLC are limited. RESULTS: 14 out of 68 observed coding mutations (21%) and 6 out of 33 (18%) copy number variations (CNV) were lost or gained during therapy. Mutational and CNV changes clustered in 6/37 (16%) and 3/37 (8%) patients. Changes in clinically relevant mutations were rare but present in single cases for genes such as BRAF and PIK3CA. The type of radiochemotherapy but not the duration of therapy impacted on the frequency of mutational changes. METHODS: We established a lung cancer specific next-generation sequencing panel covering ~7500 hotspots of 41 genes frequently mutated in NSCLC and performed ultradeep multigene sequencing of 37 corresponding pre- and post-therapeutic formalin fixed paraffin-embedded specimens to discover mutational changes and copy number variations under neo-adjuvant radio- (RTX) and/or chemotherapy (CTX). CONCLUSION: We unraveled changes in common driver gene candidates in NSCLC under neo-adjuvant therapy. Our data shed first light on the genetic changes of NSCLC under conventional therapy and might be taken into account when the relevance of sequential biopsy approaches is discussed.

Vascular Leiomyoma of the Pulmonary Artery.

Leiomyoma of the pulmonary artery represents a curiosity in the literature. We describe a case of a 54-year-old female patient who presented with recurrent cough of a few weeks' duration. Computed tomography of the thorax located a smooth, limited tumor in the left thorax near the interlobar space. Thoracoscopic exploration showed a tumor mass, fused with the pulmonary artery. After anterolateral thoracotomy, a complete resection of the tumor was performed. The histopathologic examination showed the presence of a vascular leiomyoma of the tunica media of the pulmonary artery.

Carcinoid tumours of the lung and the 'PEPPS' approach: evaluation of preoperative bronchoscopic tumour debulking as preparation for subsequent parenchyma-sparing surgery.

BACKGROUND: Preoperative bronchoscopic tumour ablation has been suggested as a beneficial treatment for bronchopulmonary carcinoid tumours, although data regarding its effects and long-term outcome are lacking. METHODS: In our case-matched cohort study with 208 patients with bronchopulmonary carcinoid tumours we investigated the role of preoperative bronchoscopic interventions before subsequent surgery and analysed the safety of this Procedure of Endobronchial Preparation for Parenchyma-sparing Surgery (PEPPS) based on metastasis and recurrence rates as well as survival data from 1991 to 2010. The subsequent surgery was classified into parenchyma-sparing procedures and classical lobectomies, bilobectomies and pneumonectomies. Data were obtained from the tumour registry and medical reports. Outcomes were the frequency of parenchyma-sparing surgery after bronchoscopic treatment as well as rates of metastasis, recurrence and survival. RESULTS: 132 of 208 carcinoids were located centrally. Among them, 77 patients could be recanalised preoperatively. After bronchoscopic preparation, the rate of subsequent parenchyma-sparing surgery methods was higher (p=0.021). The effect was measured by the number of segments removed. The 10-year survival rate was 89% (typical carcinoids) and 68% (atypical carcinoids), respectively. After applying PEPPS, long-term survival was slightly higher (p=0.23). Metastasis and recurrence rates showed no relevant differences between the bronchoscopically treated or non-treated groups, or between the two types of surgery classes or between the PEPPS and non-PEPPS groups. CONCLUSIONS: After preoperative bronchoscopic treatment, parenchyma-sparing surgery techniques can be applied more frequently. Furthermore, we detected no negative effects after PEPPS based on metastasis, recurrence and survival rates.

Preparation of cell blocks for lung cancer diagnosis and prediction: protocol and experience of a high-volume center.

Minimally invasive diagnostic techniques are increasingly being used to obtain specimens for pathological diagnosis and prediction. Referring to lung cancer, both endobronchial and endoesophageal ultrasound are used worldwide as diagnostic routine methods. Consequently, an increasing number of pathological samples are cytological and fewer are histological. On the other hand, the requirements for specific and sensitive tumor subtyping complemented by predictive analyses are steadily increasing and are an essential basis for evidence-based treatment decisions. In this article we focus on the cell block method as a helpful tool for diagnostic and predictive analyses in lung cancer and point out its advantages and disadvantages in comparison to conventional cytological and biopsy specimens. Furthermore, we retrospectively analyze the diagnostic results of the cell block method in a high-volume center over 5 years. The main advantages of cell blocks are the availability of established and validated protocols, archiving and the opportunity to have serial sections from the same specimens to provide or repeat molecular analyses. Actually, in case of tumor progression, even additional biomarkers can be tested using the original cell block when re-biopsies are not feasible. The cell block method should be considered as a reliable, complimentary approach to conventional cytological or biopsy procedures, which is helpful to fulfill the increasing requirements of high-quality diagnostics and prediction.

Prevalence of Achromobacter xylosoxidans in pulmonary mucosa-associated lymphoid tissue lymphoma in different regions of Europe.

Extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) comprises 7-8% of B-cell lymphomas and commonly originates from a background of long-standing chronic inflammation. An association with distinct bacteria species has been confirmed for several anatomical sites of MALT lymphoma. For pulmonary MALT lymphoma, however, a clear link with an infectious agent or autoimmune disorder has not yet been reported. Using a 16S rRNA gene-based approach, we have recently identified Achromobacter (Alcaligenes) xylosoxidans in eight of nine cases of pulmonary MALT lymphoma. A. xylosoxidans is a gram-negative betaproteobacterium with low virulence, but high resistance to antibiotic treatment. To further examine a potential association with A. xylosoxidans, 124 cases of pulmonary MALT lymphoma and 82 control tissues from six European countries were analysed using a specific nested PCR. Although prevalence rates for A. xylosoxidans varied significantly from country to country, they were consistently higher for MALT lymphoma as compared to controls. Overall, 57/124 (46%) pulmonary MALT lymphomas and 15/82 (18%) control tissues were positive for A. xylosoxidans (P = 0·004). Whether the significant association of A. xylosoxidans with pulmonary MALT lymphoma demonstrated in our study points to a potential causal role in the pathogenesis of this lymphoma will require further studies.

Training increases concordance in classifying pulmonary adenocarcinomas according to the novel IASLC/ATS/ERS classification.

The International Association for the Study of Lung Cancer (IASLC), the American Thoracic Society (ATS), and the European Respiratory Society (ERS) proposed a classification for lung adenocarcinomas (ADC) based on the predominant growth pattern. This classification has been shown to have prognostic and maybe even predictive impact. However, until now, the reproducibility of this classification has not been sufficiently demonstrated. Digital images of 40 selected ADC cases were shown twice to members of the Pulmonary Pathology Working Group of the German Society of Pathology. Each time a teledialogue-based survey on the classification was performed. Between the voting procedures, salient features of the novel classification were presented and discussed in detail by its members. The mean percentages of consensual votes per pattern ranged between 59.6 and 75 %, with lepidic and solid being the pattern with the most discordant and concordant votes, respectively. The other patterns ranged in between (papillary 65.8 %; acinar 67.8 %; micropapillary 74.2 %). The extent of disagreement decreased after the educational session. This decrease, however, was heterogeneous for the different patterns with acinar being the pattern with the strongest improvement. The overall number of abstentions decreased significantly after the educational session (p < 0.001) as well. The IASLC/ATS/ERS classification of lung ADC can be applied with reasonable consensus even for difficult cases in a nationwide context. The reproducibility evidently improves following educational sessions, even among experienced lung pathologists. Worldwide harmonization is clearly the next step on the way to a clinically meaningful, internationally accepted use of this novel prognostic and potentially predictive tool in lung pathology.

Bronchial stump coverage and postpneumonectomy bronchopleural fistula.

To prevent postpneumonectomy bronchopleural fistula, coverage of the bronchial stump is recommended, especially for patients treated with neoadjuvant and adjuvant chemotherapy or radiochemotherapy. We compared outcomes after proximal pericardial fat pad coverage and coverage with pleura and surrounding tissues, by retrospective analysis of the records of 243 patients. Postpneumonectomy bronchopleural fistula occurred in 7/143 (4.9%) patients who had pericardial fat pad coverage, and in 6/100 (6.0%) treated by pleural covering. Bronchopleural fistula occurred in 11 patients within 21 days, in one after 2 months, and one after 6 months. Univariate analysis of comorbidities and risk factors did not show any significant differences between the groups. Advanced T stage and carcinomatous lymphangiosis at the resection margin were associated with a higher risk of bronchopleural fistula development, independent of the technique. Reinforcement of the bronchial stump by proximal pericardial fat pad coverage appears to be safe and feasible. It is comparable to coverage with pleura and surrounding tissues.

Expression and mutational status of PDGFR in thymic tumours.

BACKGROUND: There is an ongoing search for new therapeutic targets in invasive non-resectable thymic tumours because of the low response rates in current chemotherapeutic treatment modalities. In this study, the possibility that platelet-derived growth factor receptor A (PDGFRA) and/ or PDGFRB may represent potential therapeutic targets in epithelial tumours of the thymus was investigated. PATIENTS AND METHODS: Tissue samples were obtained by thymectomy from 36 different patients with epithelial tumours of the thymus (26 thymomas types A, AB, B1-3 and 10 thymic carcinomas). Normal thymi from three young children were used as controls. The PDGFRA and PDGFRB protein expressions as well as the mutational statuses of exons 12, 14 and 18 of the PDGFRA gene were analyzed. RESULTS: All the subtypes of thymomas and the thymic carcinomas showed staining for PDGFRA, but no mutations in the known mutational hotspots were identified. Only about one third of the tumours stained for PDGFRB. PDGFRA and PDGFRB protein staining were slightly positively correlated. CONCLUSION: PDGFRA may represent a potential therapeutic target in thymic tumours.

Monitoring of glucocorticoid therapy by assessment of CD14(+)CD16(+) monocytes: a case report.

Bronchiolitis obliterans with organizing pneumonia (BOOP) is a disease affecting small airways and alveoli. It is characterized by interstitial inflammation rich in foamy macrophages and by fibroblastic connective tissue expanding into the airway and alveolar lumen. We report herein on a 54-year-old male BOOP patient who was treated with glucocorticoids (GCs) and who over a 5-year period had three relapses. At diagnosis the patient showed elevated CD14(+)CD16(+) monocyte numbers (85 cells/microl) and increased serum C-reactive protein (CRP) levels (29.4 mg/l). With GC therapy both parameters decreased within a few days. Diagnosis of relapse was preceded by a rise in CD14(+)CD16(+) monocyte numbers and in CRP levels which again responded to GC treatment. We conclude that determination of CD14(+)CD16(+) monocytes is a useful marker for monitoring of BOOP diagnosis and GC therapy.

Cellular localization of EMMPRIN predicts prognosis of patients with operable lung adenocarcinoma independent from MMP-2 and MMP-9.

Extracellular matrix metalloproteinase (MMP) inducer (EMMPRIN, CD147) is a multifunctional protein that has been implicated in cancer invasion and metastasis by the induction of MMPs. To address its role in primary tumors of human non-small-cell lung cancer we assessed whether EMMPRIN expression is associated with the expression of MMP-2 and MMP-9 and with patient survival. Primary tumors of 150 patients (65 adenocarcinomas, 58 squamous cell carcinomas, and 27 of other subtypes) with completely resected lung cancers were stained by immunohistochemistry. We assessed intensity, extent, and cellular localization of EMMPRIN staining and determined MMP-2 and MMP-9 expression. 145 tumors expressed EMMPRIN (strong expression in 61 tumors), which was predominantly localized at the tumor cell membranes in 102 (68%) patients. We could not determine any correlation between EMMPRIN expression and MMP-2 or MMP-9 expression. The prognostic relevance of EMMPRIN was evaluated by Kaplan-Meier and multivariate Cox regression analysis in patients with adenocarcinoma (n=57) and squamous cell carcinoma of the lung (n=56). The median follow-up period was 36.0 months (range 4-156 months). Staining scores for EMMPRIN and MMP-2 and MMP-9 derived from staining intensities and percentages of positive cells did not predict outcome of patients. In contrast, univariate survival analysis demonstrated that membranous localization of EMMPRIN was associated with shortened survival in patients with adenocarcinoma (P=0.03; log-rank test), but not in squamous cell carcinoma. For the former patients, membranous EMMPRIN expression was also an independent predictor of patient survival (P=0.04; Cox regression analysis). The findings point to a role of EMMPRIN for the progression of adenocarcinoma of the lung that is unrelated to its function as inducer of MMPs.

COX-2 upregulation in thymomas and thymic carcinomas.

The treatment of advanced stage thymomas and thymic carcinomas is a multimodal therapy. New therapeutic targets are currently under investigation, including the epidermal growth factor receptor (EGFR) as well as KIT. A number of studies have shown protumorigenic potential of Cyclooxygenase-2 (COX-2) in a variety of human malignancies, but so far it is unknown whether COX-2 is expressed in primary malignancies of the thymus. Using tissue microarrays, the expression of COX-2, microsomal-PGES-1 and -PGES-2 (mPGES-1 and mPGES-2), as well as EGFR was evaluated in different subtypes of thymoma and thymic carcinomas. COX-2 was expressed in all subtypes as determined by immunohistochemistry. Some cases of type B2 and thymic carcinomas had COX-2 staining levels classified as mild to moderate. However, when measuring the optical color intensity, no significant differences could be detected. Concerning the expression levels, a weak correlation between the expression of COX-2, mPGES-1 and mPGES-2 as well as EGFR was found. Furthermore, additional cases of thymomas and thymic carcinomas were analyzed by COX-2 Western immunoblot analysis and were compared to normal thymi. The analysis showed that thymomas and thymic carcinomas had a significantly stronger COX-2 expression than that of the normal thymi (p < 0.04). In summary, COX-2 is expressed in all subtypes of thymomas and thymic carcinomas and thus represents, in addition to EGFR and KIT, a potential therapeutic target. Further studies are needed in order to determine whether a combined therapy using COX-2 inhibitors in addition to the evolving anti-EGFR antibody therapy may be considered as a treatment option.

Expression of multiple epigenetically regulated cancer/germline genes in nonsmall cell lung cancer.

Cancer/germline (CG) antigens represent promising targets for widely applicable mono- and multiantigen cancer vaccines for nonsmall cell lung cancer (NSCLC). Since little is known about their composite expression in this tumor type, we analyzed 7 CG genes (MAGE-A3, NY-ESO-1, LAGE-1, BRDT, HOM-TES-85, TPX-1 and LDHC) in 102 human NSCLC specimens. About 81% of NSCLC express at least 1 and half of the specimen at least 2 CG genes. Activation of most of these genes occurs more frequently in squamous cell cancer than in adenocarcinomas. Even though we found all genes but one to be regulated by genomic methylation, not all of them are co-expressed. In particular, combining CG genes not localized on the X-chromosome may provide effective treatment for an extended number of patients.

Chromosomal imbalances in sporadic neuroendocrine tumours of the thymus.

Neuroendocrine (carcinoid) tumours of the thymus are rare neoplasms characterized by a highly malignant clinical behavior. Some of these tumors are associated with MEN1. In this study we evaluated 10 cases of sporadic thymic neuroendocrine tumours using immunohistochemistry and comparative genomic hybridization (CGH). All tumours showed a diffuse expression of neuron specific enolase (NSE) and synaptophysin. Chromosomal imbalances were detected in 8/10 cases, the most frequent gains were seen on chromosome Xp (3/10 cases), 7p, 7q, 11q, 12q, and 20q (2/10 each), losses were most frequently detected at 6q (5/10 each), 6p (3/10 each), 4q (3/10 each), 3p, 10q, 11q and 13 q (2/10 each). These CGH data show a degree of overlap with chromosomal imbalances commonly observed in advanced thymomas.

Oncogenetic tree models based on cytogenetic data: new insights into the development of epithelial tumors of the thymus.

Epithelial tumors of the thymus are rare neoplasms typically arising in the anterior mediastinum. There is an ongoing discussion whether thymomas of different histological subtypes form a biological continuum or represent distinct biological entities. To further investigate this question, we performed a statistical analysis of CGH data of 65 previously published cases. Losses of 3p, 6p, 6q, 13q, 16q, and 17p, as well as gains on 1q, were found in at least 10% of the cases. Comparing the data from B2, B3, and C thymomas, we noted an increasing complexity of karyotypes that may be well explained by a sequential order of these types. The frequencies of losses on 16q and 17p show a significant trend with respect to the sequence from B2 to B3 and C thymomas, indicating that these aberrations may be important events in the transition between these tumor types. To identify pathways of genetic development and progression of thymomas, we used oncogenetic tree models representing the dependencies between recurrent chromosomal aberrations. This analysis suggests that gains on the long arm of chromosome 1 occur early in tumor development and are correlated with losses on 6p and 6q. There is a weak correlation with losses on 16q and 17p, which appear to be late events. An independent pathway leads to losses on 3p and 13q, which are closely correlated. Our results indicate that the development of thymomas seems to be in some part a multistep mechanism. Oncogenetic tree models are a helpful means to determine developmental pathways of tumors arising from the same progenitor cell, as shown here for thymomas.

Atypical goblet cell hyperplasia in congenital cystic adenomatoid malformation as a possible preneoplasia for pulmonary adenocarcinoma in childhood: A genetic analysis.

Congenital cystic adenomatoid malformation (CCAM) of the lung is a congenital lesion that is sometimes complicated by bronchioloalveolar adenocarcinoma (BAC). In some cases foci of atypical goblet cell hyperplasia (AGCH) can be found within the cysts. It has been proposed that CCAM and AGCH predispose to the development of BAC. The present study used comparative genomic hybridization (CGH) to screen 22 cases of CCAM (epithelium, surrounding normal lung tissue, and both preneoplastic and neoplastic lesions) for chromosomal imbalances. Of these 22 cases, 10 were CCAM type 1, 10 were type 2, and 2 were type 3. Of the 10 cases of CCAM type 1, 2 were associated with AGCH, 1 was associated with atypical adenomatous hyperplasia (AAH) and associated tubular adenocarcinoma (AC), and 2 were associated with BAC (1 mucinous and 1 predominantly nonmucinous). The present study also involved immunohistochemistry for interleukin (IL)-13, IL-4 receptor-alpha (IL-4r alpha), cytokines involved in the differentiation of goblet cells, and mucin 2 protein (Muc2). Chromosomal aberrations were not detected in the epithelium or the surrounding normal lung tissue, whereas varying aberrations were found in the neoplastic lesions. The most frequent genomic imbalances observed in both AGCH and the carcinomas were gains in chromosomes 2 and 4. Interestingly, a predominance of gains was also reported in AC of nonsmokers. Chromosomal aberrations in AGCHs arising in CCAMs support their preneoplastic status. Nuclear expression of IL-13, IL-4r alpha, and Muc2 was detected in AGCH, whereas a cytoplasmic and nuclear reaction was seen in normal epithelium. This likely reflects an association with goblet cell differentiation, but it also drives proliferation in AGCH.