Marker For Mesothelioma Research Articles

Long-term survival of gastric cancer patients with positive immunocytochemistry peritoneal washings.

134 Background: The prognosis in gastric cancer is highly dependent on tumor stage at presentation. Surgery still remains the main therapeutic option in gastric cancer patients. However, the efficacy of this treatment may be substantially limited by the risk of peritoneal dissemination. The introduction of intraperitoneal chemotherapy (HIPEC) may affect the long-term outcomes in this group of patients, but high morbidity associated with such treatment provides the rational to identify the correct population of patients for HIPEC. Methods: This is the long-term effect analysis of prospectively observed cohort of patients assessed with immunocytochemistry peritoneal lavage in the single reffereal center. Between January 2002 and November 2004, the total of 140 patients with histologically confirmed gastric cancer were enrolled to the study. Laparotomy and intraoperative peritoneal lavage for immunocytochemistry examination were performed prior to gastrectomy. The fluid recovered was centrifuged and the sediment was fixed in 10% buffered formalin for 24 h, embedded in paraffin and cut into 4 µm-thick sections. Microscopic slides were also stained against cytokeratin 19 (CK-19), cytokeratin AE1/AE3 (CK-AE1/AE3) and mesothelioma marker. All patients were followed up with endpoints of cancer recurrence and mortality. Results: Median overall survival (OS) in patients with immunocytochemical evidence of free cancer cells in peritoneal washes was significantly shorter than in those without (11 [range: 2-55] vs. 45 [range: 1-164] months). The two groups differed significantly in long-term survival (5-year OS rate: 0.0% [0/17] vs. 43.1% [53/123]; 10-year OS rate: 0.0% [0/17] vs. 29.3% [36/123]) (p < 0.001). Positive peritoneal washing immunocytochemistry was an independent poor outcome prognostic factor after correction for all major prognostic factors. Conclusions: Positive peritoneal washing immunocytochemistry correlated with clinical staging of gastric cancer and was associated with poor overall survival (OS). With all patients dying in 5 years, this subgroup of patients defines a clear population for early salvage HIPEC for prospective verification.

MUC4 immunohistochemistry is useful in distinguishing epithelioid mesothelioma from adenocarcinoma and squamous cell carcinoma of the lung

The differential diagnosis of epithelioid mesothelioma from lung adenocarcinoma and squamous cell carcinoma requires the positive and negative immunohistochemical markers of mesothelioma. The IMIG guideline has suggested the use of Calretinin, D2–40, WT1, and CK5/6 as mesothelial markers, TTF-1, Napsin-A, Claudin 4, CEA as lung adenocarcinoma markers p40, p63, CK5/6, MOC-31 as squamous cell markers. However, use of other immunohistochemical markers is still necessary. We evaluated 65 epithelioid mesotheliomas, 60 adenocarcinomas, and 57 squamous cell carcinomas of the lung for MUC4 expression by immunohistochemistry and compared with the previously known immunohistochemical markers. MUC4 expression was not found in any of 65 cases of epithelioid mesothelioma. In contrast, MUC4 expression was observed in 50/60(83.3%) cases of lung adenocarcinoma and 50/56(89.3%) cases of lung squamous cell carcinoma. The negative MUC4 expression showed 100% sensitivity, 86.2% specificity and accuracy rate of 91.2% to differentiate epithelioid mesothelioma from lung carcinoma. The sensitivity, specificity, and accuracy of MUC4 are comparable to that of previously known markers of lung adenocarcinoma and squamous cell carcinoma, namely CEA, Claudin 4 and better than that of MOC-31. In conclusion, MUC4 immunohistochemistry is useful for differentiation of epithelioid mesothelioma from lung carcinoma, either adenocarcinoma or squamous cell carcinoma.

Low-grade serous carcinoma (mullerian/ovarian type) of the paratestis presenting as diffuse metastatic disease of unknown origin: Case report of an uncommon tumor with an unusual presentation

Low-grade (papillary) serous carcinoma of ovarian type is rare in males and histologically identical to low-grade serous carcinomas in female patients. We present a case of paratesticular low-grade serous carcinoma in a 42year old male, with the highly unusual initial presentation as diffuse metastatic disease in the abdomen. Imaging revealed a cystic lesion of the right testis/hemiscrotum. Biopsy of the abdominal metastasis and subsequent right orchiectomy specimen showed invasive papillary serous carcinoma, low grade, with prominent psammomatous calcifications. Immunohistochemically, both tumors were positive for PAX8, WT-1, CA-125 and cytokeratin CK7, and negative for mesothelioma markers (calretinin, D2-40) and CD10. The main differential diagnoses for this tumor include mesothelioma of tunica vaginalis, adenocarcinoma of rete testis/epididymis and metastases. Although rare as an initial presentation, ovarian type serous carcinoma of the paratestis should be included in the differential diagnosis in males with abdominal metastases that show papillary architecture, psamommatous calcifications and PAX8 immunoreactivity.

Loss of BRCA associated protein 1 expression in malignant mesothelioma and its diagnostic application

Objective: To investigate the expression of BRCA-associated protein 1 (BAP1) in malignant mesothelioma, non-small cell lung cancer and carcinosarcoma, and its application in the differential diagnosis. Methods: Twenty-two cases of malignant mesothelioma including 17 epithelioid type, 2 sarcomatoid type and 3 biphasic type were collected.As the study control, 80 non-small cell lung cancers infringement pleural membrane(including 40 lung adenocarcinomas and 40 lung squamous cell carcinomas) and 15 carcinosarcomas were included. BAP1 expression was detected using immunohistochemical method. A differential diagnosis antibody panel, including calretinin, WT1, CK5/6, D2-40, CAM5.2, CEA, TTF1, Napsin A, p63 and p40 was tested in all cases. Results: All 80 cases of non-small cell lung cancer and 15 cases of carcinosarcoma were BAP1 positive. In contrast, 64% (14/22) of malignant mesotheliomas lost BAP1 expression (P<0.01). Addition of BAP1 to the mesothelioma marker panel, the diagnostic accuracy of malignant mesothelioma was enhanced to 93%. Focal expression of BAP1 in tumors suggested multiclonal evolution of mesothelioma. Conclusions: Loss of BAP1 expression helps to confirm the diagnosis of malignant mesothelioma whereas all non-small cell lung cancer expresses BAP1. It is therefore recommended that BAP1 can be used in conjunction with other immunohistochemical markers to improve the diagnostic accuracy of malignant mesothelioma.

Assessment of potential predictors of calretinin and mesothelin to improve the diagnostic performance to detect malignant mesothelioma: results from a population-based cohort study

ObjectivesMesothelin and calretinin are blood-based markers for malignant mesothelioma. The objective of this study was to analyse the markers in plasma samples from cancer-free men and to identify factors influencing...

An Unusual Presentation of Testicular Metastasis in a Patient With Esophageal Adenocarcinoma

Esophageal adenocarcinoma is the eighth most common cancer and sixth leading cause of cancer mortality worldwide. Incidence of Esophageal adenocarcinoma (EAC) is dramatically increasing over the past few decades and is now the most common form of esophageal cancer in United States. 54 year old male with no significant medical history presented to primary care clinic with complaints of poor appetite, early satiety, heartburn and 40lb weight loss. He denied any Alcohol and Smoking use. Physical examination and basic labs were unremarkable. Upper GI series showed irregular marginated mass with narrowing and ulceration of distal esophagus. Follow up EGD and Endoscopic ultrasound showed mass extending from 36 cm to 42 cm with portion of tumor invading pericardium. TNM staging was determined as T4aN0Mx. Biopsies obtained from esophageal mass was consistent with invasive poorly differentiated adenocarcinoma. Decision was made to start neoadjuvant chemotherapy with carboplatin and paclitaxel followed by esophagectomy. Interval follow up PET scan after chemotherapy showed improvement in diffuse circumferential mural thickening of esophagus showing favorable response to therapy. Later, he presented to oncology clinic as he started to experience testicular pain and was noted to have right testicular mass and also left sided skull mass. CT head showed 2.9 x3.2 cm mass in the left parietal skull area and Right scrotal ultrasound showed 1.5x1.6x1.8 cm mass in right testicle. The patient underwent craniotomy resection followed by right radical orchiectomy. Biopsies of skull mass and right testicle showed metastatic poorly differentiated esophageal adenocarcinoma positive for CK7, CAM 5.2, MOC 31 and EMA. All markers for mesothelioma of tunica vaginalis, germ cell tumor of testes, adenocarcinoma of rete testis, prostate adenocarcinoma were negative and they were excluded. Pathologist coreviewed the esophageal mass biopsy along with skull mass and testicular mass and it was found that they had many features in common confirming our diagnosis of Testicular metastasis. Testicular cancer most commonly metastasizes to liver, lungs, bone, and adrenal glands via hematogenous route. Only one case of testicular metastasis from esophageal cancer has been reported so far. The exact pathogenesis of metastatic spread to testes remains unknown. High degree of suspicion is indicated when the patient presents with testicular mass in the setting of esophageal cancer.

GATA3 Immunohistochemistry for Distinguishing Sarcomatoid and Desmoplastic Mesothelioma From Sarcomatoid Carcinoma of the Lung.

The separation of sarcomatoid and desmoplastic malignant mesotheliomas from sarcomatoid carcinomas of the lung metastatic to the pleura may be difficult, since both types of tumor can be morphologically similar and are frequently positive only for pan-keratin. GATA binding protein 3 (GATA3) is most commonly used as an immunohistochemical marker of breast and urothelial carcinoma, but is also known to stain other types of tumors including some mesotheliomas. In this study we asked whether GATA3 stains could be used to distinguish sarcomatoid/desmoplastic malignant mesotheliomas (N=19) from sarcomatoid carcinomas of the lung (N=13). Tumor staining was scored for diffuseness and intensity, with a maximum possible score of 6. All 19 sarcomatoid/desmoplastic malignant mesotheliomas examined showed strong diffuse staining for GATA3 (no case scored <3, mean score±SD for all 19 cases 5.4±0.9), whereas only 2 of 13 sarcomatoid carcinomas of the lung stained positively for GATA3 and the staining was weak and patchy (score 2 for each case, mean±SD for all 13 cases 0.4±0.8). There was no correlation between the intensity and diffuseness of GATA-3 staining and staining for traditional mesothelioma markers. Overall, any positive staining for GATA3 was 100% sensitive and 85% specific for sarcomatoid/desmoplastic mesothelioma. We conclude that strong diffuse staining for GATA3 favors a diagnosis of sarcomatoid/desmoplastic malignant mesothelioma over metastatic sarcomatoid carcinoma of the lung; conversely, complete absence of GATA-3 staining is evidence against a diagnosis of sarcomatoid/desmoplastic malignant mesothelioma.

Secretary leukocyte peptide inhibitor (SLPI) in pleural fluid is useful as a diagnostic marker of malignant pleural mesothelioma.

e23098 Background: There is no established diagnostic marker for malignant pleural mesothelioma (MPM). In particular, the differentiation from benign asbestos pleural effusion (BAPE) is challenging. Secretary leukocyte peptide inhibitor (SLPI) is an enzyme encoded by SLPI gene. SLPI gene is reported to be overexpressed in MPM cells. Methods: SLPI in pleural fluid was determined using Quantikine ELISA Human SLPI kit (R&amp;D Systems). Results: Exploratory research revealed that SLPI value in pleural fluid of patients with MPM (n = 52) were significantly higher than those in lung cancer (LC) (n = 69) and BAPE (n = 50) ( P= 0.000). Prospective validation study included 12 pts of MPM, 24 pts of LC, 26 pts of BAPE. Median values of SLPI in MPM, LC, and BAPE were 159 .6 ng/ml, 90.5 ng/ml, and 43.2 ng/ml, respectively. SLPI value in patients with MPM were significantly higher than those in other groups ( P= 0.000). Receiver operating characteristics ROC) analysis was performed to examine the usefulness of differentiation of MPM and other diseases, and demonstrated that area under the curve (AUC) value was 0.758. With the cut of value of 88.6 ng/ml, the sensitivity was 83.3% and the specificity was 59.8%. Concerning the differentiation between MPM and BAPE, AUC value was 0.904 and with the cut of value of 81.8 ng/ml, the sensitivity was 83.3% and the specificity was 92.3%. Conclusions: Pleural fluid SLPI is a useful biomarker for the diagnosis of MPM, in particular, for the differentiation from BAPE.

HEG1 is a novel mucin-like membrane protein that serves as a diagnostic and therapeutic target for malignant mesothelioma

The absence of highly specific markers for malignant mesothelioma (MM) has served an obstacle for its diagnosis and development of molecular-targeting therapy against MM. Here, we show that a novel mucin-like membrane protein, sialylated protein HEG homolog 1 (HEG1), is a highly specific marker for MM. A monoclonal antibody against sialylated HEG1, SKM9-2, can detect even sarcomatoid and desmoplastic MM. The specificity and sensitivity of SKM9-2 to MM reached 99% and 92%, respectively; this antibody did not react with normal tissues. This accurate discrimination by SKM9-2 was due to the recognition of a sialylated O-linked glycan with HEG1 peptide. We also found that gene silencing of HEG1 significantly suppressed the survival and proliferation of mesothelioma cells; this result suggests that HEG1 may be a worthwhile target for function-inhibition drugs. Taken together, our results indicate that sialylated HEG1 may be useful as a diagnostic and therapeutic target for MM.

Midkine is a potential novel marker for malignant mesothelioma with different prognostic and diagnostic values from mesothelin

BackgroundWe evaluated possible diagnostic and prognostic values of serum midkine in malignant pleural mesothelioma in comparison with those of serum mesothelin, a well-established diagnostic biomarker.MethodsSerum mesothelin and midkine levels were determined with an enzyme-linked immunosorbent assay. We examined specimens from 95 Turkish cases with malignant pleural mesothelioma, 56 metastatic cancers to pleura, 27 other types of benign pleural diseases and 20 benign asbestos pleurisy. The cut-off values were 1.5 nmol/L for mesothelin and 421 pg/mL for midkine.ResultsSensitivity and specificity of mesothelin were 51.6 and 71.4%, 51.6 and 85.2%, and 51.6 and 85% for differentiating mesothelioma from metastatic cancers to pleura, other benign pleural diseases and benign asbestos pleurisy, respectively. Sensitivity and specificity of midkine were 61.1 and 41.1%, 61.1 and 48.1%, and 61.1 and 75% to distinguish mesothelioma from metastatic cancers to pleura, other benign pleural diseases and benign asbestos pleurisy, respectively. Combination of both biomarkers did not improve the differential diagnostic efficacy. Mesothelin levels were elevated in the epitheloid type and in the advanced cases, but were not related to the prognosis. In contrast, elevated baseline levels of midkine were independently associated with a poor prognosis of mesothelioma patients after adjusting for the stage, the histological subtypes and treatment schedules (HR = 1.84; 95% CI: 1.09-3.09) (p = 0.022).ConclusionsSerum mesothelin showed moderate sensitivity and high specificity to differentiate malignant pleural mesothelioma from metastatic malignancy to pleura and from benign pleural diseases. In contrast, midkine was a useful marker for predicting prognosis of mesothelioma patients.

Identification of a new mesothelioma marker

Identification of a new mesothelioma marker

P3.03-047 Diagnostic Value of Secretary Leukocyte Peptide Inhibitor (SLPI) in Pleural Fluid in Malignant Pleural Mesothelioma: Topic: Mesothelioma Clinical

There is no established diagnostic marker for malignant pleural mesothelioma (MPM). The aim of this study was to evaluate the usefulness of secretary leukocyte peptide inhibitor (SLPI) in pleural fluid for the diagnosis of MPM. The study included 52 MPM patients, 69 patients of lung cancer with pleural effusion (LC), and 50 patients with benign asbestos pleural effusion (BAPE) that were included as a control group. Pleural fluid was collected from these patients and SLPI was determined using Quantikine ELISA Human SLPI (R＆D Systems). Pleural fluid hyaluronic acid (HA) and soluble mesothelin related peptide (SMRP) were also determined as comparison. Median values of pleural fluid SLPI in MPM, LC, and BAPE were 108.1 ng/ml, 87.4 ng/ml, and 48.6 ng/ml, respectively. SLPI value in patients with MPM were significantly higher than those in other groups (P=0.000). SLPI value was higher in epithelioid subtype of MPM than in other subtypes. Median values of pleural fluid HA in MPM, LC, and BAPE were 86950 ng/ml, 20700 ng/ml, and 31500 ng/ml, respectively. Median values of pleural fluid SMRP in MPM, LC, and BAPE were 15.38 ng/ml, 5.70 ng/ml, and 7.31 ng/ml, respectively. Pleural fluid HA and SMRP in MPM were also significantly higher than in other groups (P=0.000). Receiver operating characteristics analysis was performed to examine the usefulness of these 3 markers for the differentiation of MPM and BAPE, and demonstrated that area under the curve values were 0.823 for SLPI, 0.760 for HA, and 0.743 for SMRP. Pleural fluid SLPI is a useful biomarker for differential diagnosis of MPM.

Molecular markers for malignant pleural mesothelioma in Mexico

Introduction: Malignant pleural mesothelioma (MPM) is a lethal cancer. There is an epidemic of MPM in Mexico with 500 cases per year. The diagnosis remains a challenge, so it is relevant to develop inexpensive and less invasive diagnostic tests. Methods: Case-control study from 3 different hospitals; MPM cases were confirmed by immunohistochemistry and histopathology. Asbestos exposure was assessed using a clinical and occupational questionnaire. Biomarkers levels were measured using ELISA. Based on ROC curves and the highest sensitivity, cut-off points were established for each biomarker. A logistic regression model was built to obtain odds ratios (OR) and 95% confidence intervals for each biomarker. Results: 101 MPM cases and 457 controls were included. Biomarkers mean levels were statistically different between cases and controls and between sexes. Area under the curve (AUC), sensitivity and cut-off points were determined: mesothelin 0.91, 90.9% and 1.59 nmol/l; calretinin 0.85, 94% and 0.69 nmol/l; and thrombomodulin 0.56, 55.8% and 0.68 nmol/l. In the logistic model for both sexes, based on cut-off points, the OR were: mesothelin &gt;0,74 nmol/l: OR=13.5 (5.7-33.5); calretinin &gt;0.23 ng/ml: OR=10 (4.6-21.7); thrombomodulin was discarded from the model. Models weighted by sex showed differences between men and women in calretinin levels, which showed a poor performance in women. Conclusions: Mesothelin and calretinin showed a good performance in men. Cut-off points support the diagnostic accuracy differentiated by sex in MPM cases. Cut-off points and biomarkers levels should be analyzed separately by sex.

Serum β-human chorionic gonadotropin as a tumor marker for malignant mesothelioma

Peritoneal mesotheliomas are aggressive tumors arising from the peritoneal mesothelium. Tumor markers are biochemical substances produced by tumor cells due to the effect of malignant process. They can be used for screening, diagnosis, and predicting the prognosis. β-human chorionic β-human chorionic gonadotrophin (β-hCG), is a marker of germ cell tumors and trophoblastic diseases. In some reports, a relationship between β-hCG and other malignant diseases has been described. Hereby, we present a case of malignant peritoneal mesothelioma associated with elevated serum β-hCG levels. A 49-year-old man was admitted to the hospital with ascites. Peritoneal mesothelioma was diagnosed with peritoneal biopsy. During the treatment period, serum β-hCG levels were elevated together with disease progression and tissue samples stained positive for β-hCG. Rising levels of serum β-hCG due to disease progression can predict the response to treatment. If these levels were elevated at the beginning of the treatment, it may be a prognostic marker for peritoneal mesothelioma. Thus, β-hCG may be an important marker for prognosis or prediction of treatment for peritoneal mesothelioma patients. Keywords: Mesothelioma, Human chorionic gonadotrophin, Tumor marker

Pathology reporting of malignant pleural mesothelioma first diagnosis: A population-based approach

Accurate pathologic diagnosis and reporting in malignant pleural mesothelioma are essential for clinical care, and cancer registration. Practical guidelines for pathologists are provided in publications and textbooks but it is unclear how these recommendations are applied in routine practice. We investigated the characteristics of pathology reports, and the extent to which they meet guideline standards. We reviewed 819 pathology reports relating to a first diagnosis of malignant pleural mesothelioma. Data sources were a regional section of the Italian network of the Mesothelioma Registry (2001–2014) and a pathology archive (1990–2000). We evaluated tumor characteristics, the diagnosis field including terminology and immunohistochemistry (IHC) workup, and report completeness (the proportion of items recorded). We investigated also two IHC panels identified by the most used markers in current practical guidelines, one best suited for epithelioid mesotheliomas (combinations of at least 2 positive and at least 2 negative mesothelioma markers) and the other best suited for sarcomatoid mesotheliomas (positive mesothelioma markers plus cytokeratins). Reports (753 histology, 66 cytology, IHC-confirmed 86%) were 74% complete and always narrative. Missing data were related to clinical history (76%), tumor laterality (61%), specimen size (38%), and histological subtype (23%). The proportion of cases with IHC was higher for epithelioid (90%) than sarcomatoid mesothelioma (87%). Compliance to IHC recommendations was higher for epithelioid (59%) than sarcomatoid mesothelioma (11%). The mean number of stains was significantly higher for sarcomatoid than epithelioid mesothelioma (p<0.000; Kruskal-Wallis test). Our findings show that although guidelines are designed to improve actual reporting practices, there is ample room for improvement in their application to standardize the diagnosis of mesothelioma. Synoptic pathology reporting needs to be implemented to better utilize pathology information.

Abstract 2976: Engineering next-generation anti-CD25 immunotoxins with improved cytotoxic activity and low immunogenicity

Abstract Objective: CD25 is expressed on many T cell malignancies, including adult T cell leukemia, and is a target for cancer therapy. Immunotoxins against CD25+ cells containing a portion of Pseudomonas exotoxin A (PE38) show extremely high cytotoxic activity. Our lab has previously created a de-immunized PE immunotoxin in which toxin domain II was removed. Removal of domain II confers two major advantages: i) the immunogenic sequences found in domain II are removed, generating a less immunogenic toxin; ii) the non-specific toxicity of domain II-truncated toxins is much lower, allowing much higher doses of immunotoxin to be given safely (7-10-fold higher safe dosage). However, removal of domain II, which does not affect or enhances the cytotoxic activity of immunotoxins targeting the B cell marker CD22 or the mesothelioma marker mesothelin, lowers the activity of anti-CD25 immunotoxins 30-fold. Because lowering toxin immunogenicity and animal toxicity are critical for the success of immunotoxins in a clinical setting, we have engineered domain II truncated immunotoxins with improved activity against CD25+ cells using a novel adaptation of an established screening system. Methods: Eighteen domain II truncation mutants were designed based upon the crystal structure of the toxin. To bypass the resource intensive protein refolding of standard immunotoxins, these domain II mutants were expressed as fusions with the ZZ protein, which is a small (13.6kD) derivative of protein A. Fusion to ZZ allowed easy production of these ZZ-domain II mutant fusion proteins in the E. coli periplasm and fast purification using IgG coated beads. The purified ZZ-domain II mutant toxins were then bound to anti-CD25 IgG by mixing (the ZZ-domain binds human Fc) and tested for cytotoxic activity on the CD25+ T-cell leukemia HUT-102 cell line. The most active domain II mutants were made into standard recombinant immunotoxins and retested. Results: Of the 18 tested ZZ-domain II mutants, 6 were completely inactive, 8 were active at near parental levels, and 4 were as active as the parental toxin. The 4 high-activity and 2 structurally-interesting, medium-activity mutants were then produced as standard recombinant immunotoxins and retested for cytotoxic activity. Two of the 4 high-activity mutants showed a 2-4-fold increase in activity compared to parental domain II truncated immunotoxin. While still being 8-fold less active than immunotoxins containing domain II (PE38), these high-activity mutant immunotoxins will most likely have lower immunogenicity than toxins with PE38 and much less non-specific toxicity, allowing higher doses and more treatment cycles to be given. Our engineered high-activity domain II mutant immunotoxins are improved candidates for treating CD25+ cancers and for eliminating regulatory immunosuppressive T cells. Citation Format: Gilad Kaplan, Fred Lee, Ira Pastan. Engineering next-generation anti-CD25 immunotoxins with improved cytotoxic activity and low immunogenicity. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2976.

Métastases révélatrices de mésothéliomes malins du péritoine : difficultés diagnostiques à propos de deux cas et conduite à tenir

Peritoneal malignant mesothelioma is a rare and extremely aggressive tumor that is sometimes difficult to diagnose. We report two cases of metastatic malignant peritoneal mesothelioma. In one case, malignant metastatic cells were identified in cervical lymph nodes while in the other case, the cells were found in the liver. In both cases, metastases were identified before discovering the primary tumor. This led to the misdiagnosis of carcinoma of unknown origin. Nevertheless, the histological and immuno-histochemical patterns were typical of malignant mesothelioma. Regarding metastasis of unknown origin, a differentiation of epithelioid peritoneal malignant mesothelioma and adenocarcinoma proved to be difficult. Therefore, we discuss the diagnostic usefulness of immuno-histochemical mesothelioma markers.

Molecular profiling reveals primary mesothelioma cell lines recapitulate human disease

Malignant mesothelioma (MM) is an aggressive, fatal tumor strongly associated with asbestos exposure. There is an urgent need to improve MM patient outcomes and this requires functionally validated pre-clinical models. Mesothelioma-derived cell lines provide an essential and relatively robust tool and remain among the most widely used systems for candidate drug evaluation. Although a number of cell lines are commercially available, a detailed comparison of these commercial lines with freshly derived primary tumor cells to validate their suitability as pre-clinical models is lacking. To address this, patient-derived primary mesothelioma cell lines were established and characterized using complementary multidisciplinary approaches and bioinformatic analysis. Clinical markers of mesothelioma, transcriptional and metabolic profiles, as well as the status of p53 and the tumor suppressor genes CDKN2A and NF2, were examined in primary cell lines and in two widely used commercial lines. Expression of MM-associated markers, as well as the status of CDKN2A, NF2, the ‘gatekeeper' in MM development, and their products demonstrated that primary cell lines are more representative of the tumor close to its native state and show a degree of molecular diversity, thus capturing the disease heterogeneity in a patient cohort. Molecular profiling revealed a significantly different transcriptome and marked metabolic shift towards a greater glycolytic phenotype in commercial compared with primary cell lines. Our results highlight that multiple, appropriately characterised, patient-derived tumor cell lines are required to enable concurrent evaluation of molecular profiles versus drug response. Furthermore, application of this approach to other difficult-to-treat tumors would generate improved cellular models for pre-clinical evaluation of novel targeted therapies.

Identification of cis- and trans-acting elements regulating calretinin expression in mesothelioma cells.

Calretinin (CALB2) is a diagnostic marker for epithelioid mesothelioma. It is also a prognostic marker since patients with tumors expressing high calretinin levels have better overall survival. Silencing of calretinin decreases viability of epithelioid mesothelioma cells. Our aim was to elucidate mechanisms regulating calretinin expression in mesothelioma. Analysis of calretinin transcript and protein suggested a control at the mRNA level. Treatment with 5-aza-2′-deoxycytidine and analysis of TCGA data indicated that promoter methylation is not likely to be involved. Therefore, we investigated CALB2 promoter by analyzing ~1kb of genomic sequence surrounding the transcription start site (TSS) + 1 using promoter reporter assay. Deletion analysis of CALB2 proximal promoter showed that sequence spanning the −161/+80bp region sustained transcriptional activity. Site-directed analysis identified important cis-regulatory elements within this −161/+80bp CALB2 promoter. EMSA and ChIP assays confirmed binding of NRF-1 and E2F2 to the CALB2 promoter and siRNA knockdown of NRF-1 led to decreased expression of calretinin. Cell synchronization experiment showed that calretinin expression was cell cycle regulated with a peak of expression at G1/S phase. This study provides the first insight in the regulation of CALB2 expression in mesothelioma cells.

Clinical utility of diagnostic markers for malignant pleural mesothelioma

Malignant mesothelioma has a very dismal prognosis with very few patients surviving one year after diagnosis. Early multimodal treatment, however, is expected to improve the outcome. Today, there is a strong need to have disease markers which could be used for screening, diagnosing, and/or monitoring tumour response to treatment. Old markers such as hyaluronic acid, various cytokeratin fragments (CYFRA 21.1, TPA) and other cancer antigens (CA 15.3, CA 125 or CA 19.9 or CEA) are not sensitive or specific enough and cannot be used in practice. More recently new molecules, such as soluble mesothelin and osteopontin, have been proposed for diagnostic purposes. Soluble mesothelin has a good specificity but has a sub-optimal sensitivity being negative in all sarcomatoid and in up to one half of epithelioid mesothelioma. On the contrary osteopontin has an inadequate specificity. Combining different markers together does not lead to an improvement in diagnostic accuracy. Neither marker can be used for screening purposes, the main limitation being the very low incidence of the disease in the at-risk, asbestos exposed population. Mesothelin is also a promising marker for monitoring response to treatment but published data is still insufficient to make recommendations. There is still a strong need for research is this area both in order to discover new markers as well as to correct the positioning of each existing molecule (alone or in combination) is the evaluation of the patients with a mesothelioma.