A novel entity of massive multifocal osteolyses in the elderly.

Osteolyses are common findings in elderly patients and most frequently represent malignant or locally aggressive bone tumors, infection, inflammatory and endocrine disorders, histiocytoses, and rare diseases such as Gorham-Stout syndrome. We here report on a novel entity of massive multifocal osteolyses in both shoulders, the right hip and left knee joint and the dens of an 83-year-old patient not relatable to any previously known etiopathology of bone disorders. The soft tissue mass is of myxoid stroma with an unspecific granulomatous inflammatory process, aggressively destroying extensive cortical and cancellous bone segments and encroaching on articulating bones in diarthrodial large joints. Radiological, nuclear medical, serological, histological, and immunohistochemical analyses were incapable of further classifying the disease pattern within the existing scheme of pathology. Quantitative polymerase chain reaction and next generation sequencing revealed that mutations are not suggestive of any known hereditary or acquired bone disease. Possible treatment options include radionuclide therapy for pain palliation and percutaneous radiation to arrest bone resorption while surgical treatment is inevitable for pathological fractures. This case study shall increase the awareness of the musculoskeletal community and motivate to collect further information on this rare but mutilating disorder.

Unusual Case of Splenic Metastasis in Adenosquamous Carcinoma of the Cervix Uteri: Diagnosis and Treatment Considerations.

BACKGROUND Due to several factors such as its specific cellular and biochemical microenvironment, the spleen is not a predestined organ of frequent metastatic colonization in the case of primary solid carcinoma. Hence, the mode of diagnosis and the preferred treatment of a lesion highly suspicious of splenic metastasis must be decided on a case-by-case basis, considering not only the biological tumor entity but also the stage of the primary disease. CASE REPORT In the present case, we demonstrate the clinical course of a 37-year-old female patient who initially presented to our clinic with irregular vaginal bleeding. A consecutive gynecological examination revealed a 3×3-cm large mass of the cervix uteri, and the subsequent histomorphological workup led to the diagnosis of an adenosquamous carcinoma of the cervix uteri. Therapeutically, the patient received multimodal treatment, namely radical hysterectomy with adjuvant radio-chemotherapy. After 1.5 years, the patient presented to our Emergency Department with intermittent left-sided abdominal pain. Subsequent abdominal imaging (computed tomography scan, magnetic resonance imaging, positron emission tomography) determined a metabolically active splenic lesion with a central necrosis - signs of malignancy in line with a splenic metastasis. Presentation and discussion of the case within our interdisciplinary tumor board led to the decision of splenectomy followed by chemotherapy, a procedure that could be considered as therapeutic treatment in such exceptional cases. CONCLUSIONS The collection and reporting of atypical clinical courses remains a key factor in precision medicine to enable the most evidence-based decision making in such cases.

THSD7A Positivity Predicts Poor Survival and Is Linked to High FAK Expression and FGFR1-Wildtype in Female Patients with Squamous Cell Carcinoma of the Lung.

Lung cancer is the leading cause of cancer-related deaths in the western world, with squamous cell carcinoma being one of the most common histological subtypes. Prognostic and predictive markers are still largely missing for squamous cell carcinoma of the lung (LSCC). Several studies indicate that THSD7A might at least play a role in the prognosis of different tumors. FAK seems to play an important role in lung cancer and is discussed as a potential therapeutic target. In addition, there is evidence that FAK-dependent signaling pathways might be affected by THSD7A. For that reason, we investigated the role of THSD7A as a potential tumor marker in LSCC and whether THSD7A expression has an impact on the expression level of FAK. A total of 101 LSCCs were analyzed by immunohistochemistry using tissue microarrays. THSD7A positivity was associated with poor overall survival in female patients and showed a relation to high FAK expression in this subgroup. To our knowledge, we are the first to report these correlations in lung cancer. The results might be proof of the assumed activation of FAK-dependent signaling pathways by THSD7A and that as a membrane-associated protein, THSD7A might serve as a putative therapeutic target in LSCC.

SCARA5 Is Overexpressed in Prostate Cancer and Linked to Poor Prognosis.

Prostate cancer is one of the most common malignancies worldwide, showing a wide range of clinical behaviors. Therefore, several treatment options arise out of the diagnosis "prostate cancer". For this reason, it is desirable to find novel prognostic and predictive markers. In former studies, we showed that THSD7A expression is associated with unfavorable prognostic parameters in prostate cancer and is linked to a high expression of focal adhesion kinase (FAK). Recently, scavenger receptor class A member 5 (SCARA5) was reported to be the downstream gene of THSD7A in esophageal squamous cell carcinoma. SCARA5 is believed to play an important role in the development and progression of several different tumor types. Most studies describe SCARA5 as a tumor suppressor. There is also evidence that SCARA 5 interacts with FAK. To examine the role of SCARA5 as a potential biomarker in prostate cancer, a total of 461 prostate cancers were analyzed via immunohistochemistry using tissue microarrays. Furthermore, we compared the expression level of SCARA5 with our previously collected data on THSD7A and FAK. High SCARA5 expression was associated with advanced tumor stage (p < 0.001), positive nodal status (p < 0.001) and high Gleason-score (p < 0.001). At least, strongly SCARA5-positive cancers were associated with THSD7A-positivity. There was no significant association between SCARA5 expression level and FAK expression level. To our knowledge, we are the first to investigate the role of SCARA5 in prostate cancer and we demonstrated that SCARA5 might be a potential biomarker in prostate cancer.

THSD7A Positivity Is Associated with High Expression of FAK in Prostate Cancer.

Prostate cancer is one of the most common malignancies, and there are a wide range of treatment options after diagnosis. Most prostate cancers behave in an indolent manner. However, a given sub-group has been shown to exhibit aggressive behavior; therefore, it is desirable to find novel prognostic and predictive (molecular) markers. THSD7A expression is significantly associated with unfavorable prognostic parameters in prostate cancer. FAK is overexpressed in several tumor types and is believed to play a role in tumor progression and metastasis. Furthermore, there is evidence that THSD7A might affect FAK-dependent signaling pathways. To examine whether THSD7A expression has an impact on the expression level of FAK in its unphosphorylated form, a total of 461 prostate cancers were analyzed by immunohistochemistry using tissue microarrays. THSD7A positivity and low FAK expression were associated with adverse pathological features. THSD7A positivity was significantly associated with high FAK expression. To our knowledge we are the first to show that THSD7A positivity is associated with high FAK expression in prostate cancer. This might be proof of the actual involvement of THSD7A in FAK-dependent signaling pathways. This is of special importance because THSD7A might also serve as a putative therapeutic target in cancer therapy.

Strongly Radioiodine-Positive Pancreatic Adenocarcinoma Mimicking Metastasis of Differentiated Thyroid Cancer.

We present an interesting image of a strikingly intense radioiodine accumulation of a histologically proven pancreatic adenocarcinoma mimicking metastasis of differentiated thyroid cancer in a 63-year-old woman with recurrence of papillary thyroid carcinoma undergoing radioiodine therapy. This interesting image should draw attention to considering pancreatic adenocarcinoma in radioiodine-positive pancreatic lesions.

A new tool for bile duct tissue sampling: ex vivo clinical evaluation of intraductal cryobiopsy for cholangioscopy.

Background and study aims Indeterminate biliary strictures represent a major challenge in clinical diagnostics. Diagnostic yield of radiological, endoscopic imaging and histopathological diagnosis is insufficient. The cryobiopsy technique is a new method for tissue extraction already used in different clinical settings. The aim of this ex vivo clinical study was to investigate feasibility and tissue quality of cryobiopsy in the bile duct. Patients and methods We included 14 patients who underwent pancreaticoduodenectomy. Bile duct samples were taken with either a new prototype cryoprobe or one of two forceps types. Results were analyzed for general feasibility, specimen size, histological assessability as well as representativity of retrieved tissue. Results Feasibility of cholangioscopic forceps was poor compared to gastric biopsy forceps or cryobiopsy. Significantly larger tissue samples were obtained with cryobiopsy (5.6 ±â€Š4.5 mm 2 ) compared to gastric biopsy forceps (3.3 ±â€Š5.1 mm 2 , P  = 0.006). Furthermore, cryobiopsy was superior in histological assessment quality ( P  = 0.02) and concerning representativity ( P  = 0.03). Conclusions Cryobiopsy in the bile duct is feasible and the quality of the obtained tissue is high. Further investigation of bile duct cryobiopsy in vivo is warranted.

PSMA-Positive Follicular Thyroid Carcinoma Incidentally Detected by [68Ga]Ga-PSMA-11 PET/CT: Correlation with Immunohistology Confirms Neovascular PSMA-Expression.

We present an interesting image of an intense PSMA-positive follicular thyroid carcinoma incidentally detected by [68Ga]Ga-PSMA-11 PET/CT in a 76-year-old man with biochemical recurrence of prostate cancer. Immunohistochemical staining demonstrated PSMA expression in the endothelial cells of tumor tissue. This interesting image should remind colleagues to consider malignant thyroid neoplasia in PSMA-positive thyroid lesions.

Mixed Adenocarcinomatous and Neuroendocrine Tumor of the Urinary Bladder With Concomitant Carcinoma In Situ: A Case Report With a Comprehensive Immunohistochemical Analysis and Review of the Literature.

Mixed adenoneuroendocrine carcinomas are rare and usually occur in the gastrointestinal tract. Although there have been several investigations regarding their developmental mechanism, the molecular origin of these tumors remains unclear. In this article, we present an exceedingly rare case of a mixed tumor of the urinary bladder with an adenocarcinomatous and a neuroendocrine component and a concomitant urothelial carcinoma in situ (CIS). Due to this extraordinary combination of tumor components, our goal was to extensively examine the 3 tumor components with regard to a representable common origin. Therefore, a comprehensive immunohistochemical analysis and review of the literature was performed. Besides expected outcome, our examination also revealed surprising staining results. Urothelial CIS, like the adenocarcinomatous component, showed strong staining for CDX2. In addition, parts of the adenocarcinoma were positive for synaptophysin like the neuroendocrine tumor component. All 3 components showed a significant overexpression of p53 and a moderate to strong membranous and cytoplasmatic staining for ß-catenin. To our knowledge, we are the first to describe a case of a mixed tumor of the urinary bladder with an adenocarcinomatous and a neuroendocrine component and a concomitant CIS. The components share striking molecular features that argue for a common clonal origin and a development of the invasive tumor via the urothelial precursor lesion.

THSD7A expression in human cancer.

We recently described a case of a Thrombospondin Type-1 Domain containing 7A (THSD7A) associated membranous nephropathy in a female patient who was synchronously suffering from a THSD7A-positive malignancy. We here investigated the role of THSD7A as a new potential tumor antigen by evaluating over 20 000 tissue spots in more than 70 different tumor entities by immunohistochemistry using tissue microarrays. THSD7A expression was highly variable in different neoplasias with differing staining patterns. Both gain and loss of THSD7A expression compared to expression status in non-tumor tissue were linked to tumor-specific markers in the different tumor entities and were of prognostic value. The potential role of THSD7A in tumor development and therapy needs further investigation.

An Indirect Immunofluorescence Method Facilitates Detection of Thrombospondin Type 1 Domain-Containing 7A-Specific Antibodies in Membranous Nephropathy.

Thrombospondin type 1 domain-containing 7A (THSD7A) is a target antigen identified in adult membranous nephropathy (MN) along with the major antigen phospholipase A2 receptor 1 (PLA2R1). The prevalence of THSD7A-Ab-positive patients is unknown, and it is unclear whether the clinical presentation differs between patients positive for PLA2R1-Ab or THSD7A-Ab. We screened serum samples of 1276 patients with MN from three different cohorts for the presence of THSD7A-Ab by Western blot analysis and a newly developed indirect immunofluorescence test (IFT). Compared with Western blot analysis, the IFT had a 92% sensitivity and a 100% specificity. The prevalence of THSD7A-associated MN in a prospective cohort of 345 patients with MN was 2.6%, and most were women. In this cohort, the percentage of patients with THSD7A-associated MN and malignant disease significantly exceeded that of patients with PLA2R1-associated MN and malignant disease. In all cohorts, we identified 40 patients with THSD7A-associated MN, eight of whom developed a malignancy within a median time of 3 months from diagnosis of MN. In one patient with THSD7A-associated MN and metastases of an endometrial carcinoma, immunohistochemistry showed THSD7A expression on the metastatic cells and within follicular dendritic cells of the metastasis-infiltrated lymph node. We conclude that the IFT allows sensitive and specific measurement of circulating THSD7A-Ab in patients with MN. Patients with THSD7A-associated MN differ in their clinical characteristics from patients with PLA2R1-associated MN, and more intensive screening for the presence of malignancies may be warranted in those with THSD7A-associated MN.

Determination of Tumor Heterogeneity in Colorectal Cancers Using Heterogeneity Tissue Microarrays.

Cancer is often heterogeneous both on a morphological and on a genetic level. Though resected tumors are often large, molecular tumor analysis is usually restricted to one tissue block. In this project we introduce a new tool for a high-throughput heterogeneity analysis of colorectal cancer. A heterogeneity tissue microarray (TMA) was manufactured from tissues of 340 patients with colorectal cancer. For this purpose 8 different tissue spots were taken from as many different cancer blocks per patient as possible (at least 4 different blocks). Additional tissue samples from 1 to 4 corresponding lymph node metastases were added from 134 patients. The system was then validated by analysing one parameter each known for minimal (p53) or substantial (HER2) heterogeneity in colorectal cancer. P53 alterations as detected by immunohistochemistry were seen in 174 (51.3 %) of 339 analyzable primary tumors of which 23 (13.2 % of positive cases) showed a heterogeneous distribution pattern. HER2 overexpression was seen in 18 (5.4 %) of 336 evaluable tumors. HER2 amplification occurred in 6 (33.3 %) of the 18 cases with HER2 overexpression. Genomic heterogeneity was more prevalent for HER2 alterations than for p53 alterations. For immunohistochemical expression analysis, 16 of 18 positive cases were heterogeneous (88.9 %) and for amplification 3 of 6 cases (50 %) were heterogeneous. Large section validation revealed, however a considerable fraction of heterogeneous cases were due to technical artifacts. In summary, our data suggest, that heterogeneity TMAs are a powerful tool to rapidly screen for molecular heterogeneity in colorectal cancer.

Heterogeneity of amplification of HER2, EGFR, CCND1 and MYC in gastric cancer.

BACKGROUND: Intra-tumor heterogeneity is a potential cause for failure of targeted therapy in gastric cancer, but the extent of heterogeneity of established (HER2) or potential (EGFR, CCND1) target genes and prognostic gene alterations (MYC) had not been systematically studied. METHODS: To study heterogeneity of these genes in a large patient cohort, a heterogeneity tissue microarray was constructed containing 0.6 mm tissue cores from 9 different areas of the primary gastric cancers of 113 patients and matched lymph node metastases from 61 of these patients. Dual color fluorescence in-situ hybridization was performed to assess amplification of HER2, EGFR, CCND1 and MYC using established thresholds (ratio ≥ 2.0). Her2 immunohistochemistry (IHC) was performed in addition. RESULTS: Amplification was found in 17.4% of 109 interpretable cases for HER2, 6.4% for EGFR, 17.4% for CCND1, and 24.8% for MYC. HER2 amplification was strongly linked to protein overexpression by IHC in a spot-by-spot analysis (p < 0.0001). Intra-tumor heterogeneity was found in the primary tumors of 9 of 19 (47.3%) cancers with HER2, 8 of 17 (47.0%) cancers with CCND1, 5 of 7 (71.4%) cancers with EGFR, and 23 of 27 (85.2%) cancers with MYC amplification. Amplification heterogeneity was particularly frequent in case of low-level amplification (<10 gene copies). While the amplification status was often different between metastases, unequivocal intra-tumor heterogeneity was not found in individual metastases. CONCLUSION: The data of our study demonstrate that heterogeneity is common for biomarkers in gastric cancer. Given that both TMA tissue cores and clinical tumor biopsies analyze only a small fraction of the tumor bulk, it can be concluded that such heterogeneity may potentially limit treatment decisions based on the analysis of a single clinical cancer biopsy.

MALDI imaging-based identification of prognostically relevant signals in bladder cancer using large-scale tissue microarrays.

OBJECTIVE: Although most patients with urinary bladder cancer present with noninvasive and low-malignant stages of the disease, about 20% eventually develop life-threatening metastatic tumors. This study was designed to evaluate the potential of matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) to identify molecular markers predicting the clinical course of bladder cancer. MATERIALS AND METHODS: We employed MALDI-MSI to a bladder cancer tissue microarray including paraffin-embedded tissue samples from 697 patients with clinical follow-up data to search for prognostically relevant associations. RESULTS: Analysis of our MALDI imaging data revealed 40 signals in the mass spectra (m/z signals) associated with epithelial structures. The presence of numerous m/z signals was statistically related to one or several phenotypical findings including tumor aggressiveness (stage, grade, or nodal status; 30 signals), solid (5 signals) or papillary (3 signals) growth patterns, and increased (6 signals) or decreased (12 signals) cell proliferation, as determined by Ki-67 immunohistochemistry. Two signals were linked with tumor recurrence in noninvasive (pTa category) tumors, of which one was also related to progression from pTa-category to pT1-category disease. The absence of one m/z signal was linked with decreased survival in the subset of 102 muscle-invasive cancers. CONCLUSION: Our data demonstrate the suitability of combining MSI and large-scale tissue microarrays to simultaneously identify and validate clinically useful molecular markers in urinary bladder cancer.

Heterogeneity and chronology of PTEN deletion and ERG fusion in prostate cancer.

TMPRSS2:ERG fusions, in combination with deletion of the phosphatase and tensin homolog (PTEN) tumor suppressor, have been suggested to cooperatively drive tumor progression in prostate cancer. We utilized a novel heterogeneity tissue microarray containing samples from 10 different tumor blocks of 189 prostatectomy specimens to study heterogeneity of genomic PTEN alterations in individual foci. PTEN alterations were found in 48/123 (39%) analyzable individual tumor foci, including 40 foci with deletions, 7 with deletion and rearrangement, and 1 focus with rearrangement only. PTEN was homogeneously aberrant in only 4 (8%) and heterogeneously in 44 (92%) of the foci. We found a specific sequence of molecular events from PTEN breakage followed by deletion of DNA sequences flanking the breakpoint, resulting in homozygous deletion. The observation that 16 of 19 foci with homogeneous ERG positivity had focal PTEN alterations but none of 10 foci with PTEN alterations had focal ERG positivity (P<0.0001) suggests that PTEN alterations typically develop subsequent to ERG fusions. We demonstrate a high level of intratumoral heterogeneity of PTEN alterations with deletions and rearrangements that challenges potential PTEN routine diagnosis testing in biopsies. The observation that PTEN alterations develop subsequent to ERG fusion strongly suggests that ERG expression may directly drive development of PTEN aberrations.

MALDI imaging on tissue microarrays identifies molecular features associated with renal cell cancer phenotype.

AIM: To identify molecular features associated with clinico-pathological parameters in renal cell cancer. MATERIALS AND METHODS: Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging was employed for a kidney cancer tissue microarray containing tissue samples from 789 patients for which clinical follow-up data were available. RESULTS: A comparison of mass spectrometric signals with clinico-pathological features revealed significant differences between papillary and clear cell renal cell cancer. Within the subgroup of clear cell RCC, statistical associations with tumor stage (seven signals, p<0.01 each), Fuhrman grade (seven signals, p<0.0001 each), and presence of lymph node metastases (10 signals, p<0.01 each) were found. In addition, the presence of one signal was significantly linked to shortened patient survival (p=0.0198). CONCLUSION: Our data pinpoint towards various molecules with potential relevance in renal cell cancer. They also demonstrate that the combination of the MALDI mass spectrometry imaging and large-scale tissue microarray platforms represents a powerful approach to identify clinically-relevant molecular cancer features.

Continuous tissue microarray based identification of cancers with homogeneous target expression for successful targeted therapy in clinical routine practice.

In cancer therapy, the number of drugs targeting cells with characteristic molecular aberrations is continuously rising. However, application of these new drugs still is limited to a few tumor entities. The aim of this study was to test the concept of routinely identifying all possible cancer patients who might eventually benefit from targeted therapy. Therefore, all malignant tumors routinely submitted to our Institute of Pathology over a period of 4 months were brought into a tissue microarray format. Using "in situ" methods, tumors were analyzed for HER2, EGFR, and KIT status as examples for potential therapeutic target genes. In positive cases, target heterogeneity was excluded by analyzing all available large sections. Outside of tumor entities for which targeted drugs are already approved, the study revealed six tumors with homogeneously distributed HER2 overexpression/amplification (bladder, esophageal and colorectal) and seven tumors with homogeneous EGFR amplification (vulvar, ovarian, breast, esophageal and laryngeal, and adenocarcinoma of unknown primary). A total of 151 tumors showed KIT overexpression but none of seven sequenced cases showed KIT mutations. We furthermore report on a 69-year-old patient with homogeneously HER2-amplified metastatic colorectal cancer who is successfully treated by trastuzumab monotherapy. This study demonstrates that tissue microarray based screening for therapeutic target genes in tumors outside established indications represents a feasible approach suitable for routine application. The successful treatment of one patient with homogeneously HER2 positive metastatic colorectal cancer argues for the clinical utility of this approach at least in carefully selected, homogeneous cancers.

MALDI imaging on large-scale tissue microarrays identifies molecular features associated with tumour phenotype in oesophageal cancer.

AIMS: Matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI) and tissue microarray (TMA) technologies were jointly utilized to search for molecular features associated with clinicopathological parameters in oesophageal cancer. METHODS AND RESULTS: Two TMAs from formalin-fixed tissue samples, including 300 adenocarcinomas and 177 squamous cell carcinomas with clinical follow-up data, were analysed. MALDI-MSI analysis revealed 72 distinct mass per charge (m/z) signals associated with tumour cells, 48 of which were found in squamous cell carcinomas only, and 12 of which were specific for adenocarcinomas. In adenocarcinomas, six signals were linked to early-stage (pT1-T2) tumours (two signals) and the presence (one signal) or absence (three signals) of lymph node metastasis. In squamous cell carcinomas, 24 signals were strongly linked to different phenotypic features, including tumour stage (four signals), histological grade (four signals), and lymph node metastasis (three signals). CONCLUSIONS: The high number of m/z signals that were found to be significantly linked to one or more phenotypic features of oesophageal cancer highlights the power of MALDI-MSI in the analysis of high-density TMAs. The data also emphasise substantial biological differences between adenocarcinomas and squamous cell carcinomas.