α-methylacyl-CoA racemase (AMACR) expression in chordomas differentiates them from chondrosarcomas.

AIMS: Chordomas and chondrosarcomas are malignant mesenchymal tumours with overlapping morphological and immunohistochemical (IHC) characteristics. Our aim was to evaluate the IHC expression of α-methylacyl-CoA racemase (AMACR/P504S), ß-catenin and E-cadherin in chordomas relative to chondrosarcomas and assess the utility of these markers for differential diagnosis. METHODS: Archival sections of 18 chordomas, 19 chondrosarcomas and 10 mature cartilage samples were immunostained and scored for AMACR, ß-catenin and E-cadherin and the relative differential capacity of each marker was calculated. In addition, AMACR mRNA level was assessed in 5 chordomas by RT-PCR and evaluated by comparative CT method. RESULTS: AMACR and ß-catenin stained 88.9% and 94.1% of the chordomas respectively, 21.1% and 10.5% of the chondrosarcomas correspondingly and none of the mature cartilage samples. E-cadherin stained positively 82.4% of the chordomas, 36.8% of the chondrosarcomas and 42.9% of the mature cartilage cases. Both AMACR and ß-catenin showed statistically significant difference between chordomas and chondrosarcomas (p < 0.001 for both), unlike E-cadherin. AMACR was detected at the mRNA level. CONCLUSIONS: AMACR is expressed in most of the chordomas but only in a minority of chondrosarcomas. AMACR may serve as IHC marker of chordoma with differentiating ability comparable to that of ß-catenin.

Connection between expression of inducible nitric oxide synthase (iNOS) in skull base chordoma and lower urinary tract symptoms.

OBJECTIVE: To provide first insights into the potential role of iNOS expressed by skull base chordoma, which causes brainstem compression in and around Barrington's nucleus, and its effect on the micturition center. METHODS: Urodynamic testing of 22 symptomatic patients was performed. All women and men with skull base chordoma treated in two hospitals in Germany between 1986 and 2007 were studied. Lower urinary tract symptoms (LUTS) were documented in patients with acute brainstem compression due to local chordoma growth positive for iNOS expression. Brain magnetic resonance (MRI) images of the lesions of the symptomatic patients were performed. RESULTS: Of 74 treated patients, 22 (7 women, 15 men) with a median age of 37 years were evaluated with voiding diaries and computer urodynamic investigation. Urodynamic testing of 22 symptomatic patients with positive iNOS expression of skull base chordoma revealed detrusor overactivity in 55 %, low-compliance bladder in 14 %, detrusor sphincter dyssynergia in 45 % and uninhibited sphincter relaxation in 27 %. There was a significant correlation between strong iNOS expression (score 3-6) in skull base chordoma and severe urinary symptoms (p = 0.003, Spearman ρ = 0.526). CONCLUSIONS: The expression of iNOS in skull base chordoma compressing the dorsolateral pons, in and around Barrington's nucleus, may influence the pontine micturition center (PMC) and be responsible for lower urinary tract symptoms. Nitric oxide may possibly act as a neurotransmitter. We assume that the high infiltration of chordoma with monocyte/macrophages enhances the release of nitric oxide, as monocyte/macrophages are the main source of iNOS.

Expression of aurora kinase A and B in chondrosarcoma and its relationship with the prognosis.

PURPOSE: To investigate the expression of aurora kinase A and B in patients with chondrosarcoma and consider it as a prognostic marker and molecular target of therapy. METHODS: To evaluate the relationship of the aurora kinase A and B and the clinical pathological parameters and prognosis of chondrosarcoma. 72 case chondrosarcoma and 42 case chondroma were performed immunohistochemistry on the tissue microarray paraffin sections. The survival time of patients was followed-up. RESULTS: The expression of aurora kinase A and B in chondrosarcoma was significantly higher than that in chondroma (p<0.01). There were differences about the expression of aurora kinase A and B in chondrosarcoma between the recurrence group and the non-recurrence group, metastatic group and non-metastatic group (p<0.05), but not age and gender (p>0.05). The expression of aurora kinase A and B were significantly lower in group low grade conventional chondrosarcoma than that in groups medium and high grade conventional chondrosarcoma (p<0.01). The expression of aurora kinase A and B in chondrosarcoma showed a positive correlation (p<0.01). According to the Kaplan Meier analysis and multivariate Cox regression analysis, the survival rate was significantly different between the patients with positive aurora kinase A and the patients with negative expression (p<0.05) and aurora kinase A expression was an independent risk marker of survival (HR=11.263, 95%CI: 2.317-54.748, P=0.003). CONCLUSION: Both the aurora kinase A and B might involve in the oncogenic, invasive and metastatic process of chondrosarcoma; however, the mechanism is still unclear. The aurora kinase A and B could be used as a new prognostic marker and molecular therapeutic target for chondrosarcoma. VIRTUAL SLIDE: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/9101494267377096.

Expression of matrix metalloproteinases 1, 2, 9 and their tissue inhibitor-1 in cartilage-forming osteal tumors.

The expression of MMP-1, -2, -9 and TIMP-1 was studied in 10 benign cartilage-forming osteal tumors (5 osteochondromas and 5 chondromas) and 39 chondrosarcomas (14 central, 4 periosteal, 7 dedifferentiated, and 14 secondary tumors). No expression of MMP and TIMP-1 was detected in benign cartilage-forming osteal tumors. In chondrosarcomas, the expression of MMP-1 was detected in 84.6%, of MMP-2 in 71.8, of MMP-9 in 97.4, and of TIMP-1 in 82.4% cases, the levels of expression of these markers varied from 10 to 60%. The expression of MMP-1 was not associated with patient gender, maximum size and degree of differentiation of the tumor, but was linked with age. The expression of MMP-1 was more often detected in central and dedifferentiated chondrosarcomas; the expression of MMP-1(+) was significantly associated with 3-year relapse-free and 5-year overall survival of the patients. The expression of MMP-1 in the tumor was associated with unfavorable course of the disease. The values of MMP-2 expression in chondrosarcomas did not reflect the main clinical morphological characteristics of the disease and its prognosis. The level of MMP-9 protein expression in chondrosarcomas ≥40% is prognostically unfavorable, while <40% is a favorable factor for 3-year relapse-free survival. The risk of disease relapse within 1 year after the beginning of therapy was maximum in T3 tumors with expression of MMP-9 protein ≥40%. No relationships between the parameters of TIMP-1 expression in chondrosarcomas and the main clinical morphological characteristics of the disease and its prognosis were detected.

Telomerase reverse transcriptase subunit expression is associated with chondrosarcoma malignancy.

Expression of the telomerase reverse transcriptase subunit telomerase reverse transcriptase gene is associated with most human malignancies. Because telomerase reverse transcriptase is rarely expressed in normal tissue, its presence in pathologic specimens is considered a marker of transformed cells. Moreover, high levels of expression have been correlated with poor prognosis in many cancers. Although telomerase activity has been found in chondrosarcomas, its prognostic significance in these malignant cartilage tumors is unknown. Malignancy in cartilage-derived tumors is assessed routinely by histomorphologic grading, but even well differentiated, low-grade lesions can metastasize. This unpredictable behavior greatly complicates the clinical treatment of cartilage tumors, making better prognostic indicators desirable. To address this issue we used immunohistochemistry to compare telomerase reverse transcriptase expression in a collection of 61 tumors consisting of malignant chondrosarcomas of varying grade and benign enchondromas. Associated case histories were reviewed to test the hypothesis that telomerase reverse transcriptase expression levels correlated with subsequent tumor recurrence. We found that the relative abundance of telomerase reverse transcriptase-expressing cells correlated significantly with grade and recurrence. These findings indicate that telomerase reverse transcriptase immunostaining may be a useful adjunct to the conventional three-level grading system.

Cyclooxygenase-2 expression in chondrosarcoma.

OBJECTIVE: In recent years it has become evident that tissue cyclooxygenase-2 (COX-2) may play a role in carcinogenesis and tumor malignancy. There is now a mounting body of information that strongly implies that COX-2 inhibitors may be of some value in the management of patients with carcinomas, and most recently several similar reports have appeared relating to sarcomas. METHODS: The authors studied 32 samples of cartilage tumors from our tumor tissue bank for the presence of COX-2 by a Western blot technique. There were 29 patients from whom the samples were obtained, including 8 with enchondromas and 21 with chondrosarcomas. RESULTS: Thirteen of the 24 chondrosarcoma samples and none of the 8 enchondromas were positive for COX-2. An attempt was made to correlate these results with clinical data including age, gender, staging according to the Musculoskeletal Tumor Society, anatomical site, ploidic pattern, presence of metastases and death rate but no statistically valid correlation could be found. CONCLUSION: It is evident that COX-2 may play some role in chondrosarcoma but not in the benign enchondroma and that further studies with COX-2 inhibitors are warranted.

Malignant transformation in human chondrosarcoma cells supported by telomerase activation and tumor suppressor inactivation.

Human chondrosarcomas do not respond to current chemotherapies or radiation therapy, and their size and histological appearance do not reliably predict the risk of local recurrence and metastases, making selection of surgical treatment difficult. Identifying mechanisms responsible for the proliferation and invasive behavior of these tumors would be of immense clinical value. We hypothesized that telomerase expression is one of these mechanisms. We detected telomerase expression in 7 of 16 chondrosarcomas, but cells cultured from telomerase-negative chondrosarcomas acquired strong telomerase activity and lost tumor suppressor activity after their establishment in culture. These changes were associated with accelerated indefinite cell proliferation, morphological transition, and increased invasive activity, indicating that telomerase activation and loss of cell cycle control leads to the emergence of aggressive cells from chondrosarcoma cell populations. These observations may lead to better understanding of the factors responsible for malignant transformation, local recurrence, and metastases of cartilage neoplasms.

Immunohistochemical analysis of matrix metalloproteinase-1, -3, and -13 in naturally occurring cartilaginous tumors of dogs.

OBJECTIVE: To determine immunoreactivity of matrix metalloproteinase (MMP)-1, -3, and -13 in cartilaginous tumors of dogs, correlate expression of MMP with histologic grade of tumors and clinical outcome of dogs, and compare MMP immunoreactivity between chondrosarcomas and chondromas. SAMPLE POPULATION: Formalin-fixed, paraffin-embedded tissues obtained from samples of naturally occurring chondrosarcomas (n = 31) and chondromas (8) of dogs that were submitted to our veterinary medical diagnostic laboratory. PROCEDURE: Histologic sections from each sample were stained with H&E and monoclonal antibody to MMP-1, -3, and -13 by use of an avidin-peroxidase immunohistochemical technique. For each section, histologic grade (I, II, or III) and immunohistochemical expression (0, 1, 2, or 3) were evaluated. Clinical outcome was obtained from medical records or interviews with referring veterinarians and scored as a good outcome, moderate outcome, or poor outcome. Correlations among variables and differences between chondrosarcomas and chondromas were analyzed. RESULTS: Samples from chondrosarcomas had significantly higher immunoreactivity of MMP-1 and -13, compared with immunoreactivity in samples from chondromas. In chondrosarcomas, a significant positive correlation (r, 0.386) was found between MMP-1 and -13 immunoreactivities, and a significant negative correlation (r, -0.390) was detected between MMP-3 and -13 immunoreactivities. CONCLUSIONS AND CLINICAL RELEVANCE: A significant increase in expression of collagenases (MMP-1 and -13) in chondrosarcomas, compared with expression in chondromas, suggests that collagenases may play an important role in tumor progression, and possibly metastasis, in chondrosarcomas of dogs.

Chondrosarcoma is not characterized by detectable telomerase activity.

Reactivation of telomerase, an enzyme which elongates human telomeres, is associated with cell immortilization. In approximately 90% of malignant tumours telomerase activity can be demonstrated, whereas in benign tumours it is mostly absent. Chondrosarcomas are relatively rare malignant cartilaginous neoplasms. A small number of chondrosarcomas located centrally in bone arise secondarily to an enchondroma, while the majority of chondrosarcomas developing from the surface arise within the cartilage cap of an osteochondroma. The histological distinction between a benign lesion and low-grade chondrosarcoma is generally considered difficult. To investigate whether the progression towards chondrosarcoma is characterized by reactivation of telomerase activity, this study determined telomerase activity in ten enchondromas, five osteochondromas, and 37 chondrosarcomas using the TRAP assay. In all tumour samples except one, telomerase activity was absent. By adding tumour lysates to the positive control, an increasing inhibition of telomerase activity was found with an increasing chondroid matrix, suggesting that it may contain inhibitory factors. Inhibition due to endogenous RNAse or Taq-polymerase inhibitors was excluded. The lack of detectable telomerase activity in the high-grade component of a dedifferentiated chondrosarcoma without matrix favours the possibility that telomerase is truly absent. Either its true absence or inhibitory effects disabling telomerase detection exclude the telomerase TRAP assay as a diagnostic tool in the differential diagnosis of benign and low-grade malignant cartilaginous tumours.

Tumor-induced osteomalacia: clinical and basic studies.

A patient with classic clinical and biochemical features of tumor-induced osteomalacia (hypophosphatemia, phosphaturia, and undetectable serum concentrations of 1,25-dihydroxyvitamin D [1,25(OH)2D]) was studied before and after resection of a benign extraskeletal chondroma from the plantar surface of the foot. Presurgical laboratory evaluation was notable for normal serum concentrations of calcium, intact parathyroid hormone (PTH), parathyroid hormone-related protein (PTHrP), and osteocalcin, increased serum alkaline phosphate activity, and frankly elevated urinary cyclic adenosine monophosphate (cAMP) and pyridinium cross-link excretion. Quantitative histomorphometry showed severe osteomalacia and deep erosions of the cancellous surface by active osteoclasts. After resection, serum 1,25(OH)2D normalized within 24 h, while renal tubular phosphorus reabsorption and serum phosphorus did not normalized until days 2 and 3, respectively; serum Ca declined slightly, and serum intact PTH, osteocalcin, and urinary pyridinium cross-link excretion increased dramatically. Urinary cAMP excretion declined immediately after resection and then began to increase concomitant with the increase in serum intact PTH. A second bone biopsy taken 3 months after resection demonstrated complete resolution of the osteomalacia, increased mineral apposition rate (1.09 mu/day), resorption surface (9.2%), mineralizing surface (71%), and bone formation rate (0.83 mm3/mm2/day), and marked decrease in cancellous bone volume (13.1%) and trabecular connectivity compared with first biopsy. Tumor extracts did not affect phosphate transport in renal epithelial cell lines or 1 alpha-hydroxylase activity in a myelomonocytic cell line. The patient's course suggests that the normal 1,25(OH)2D and phosphorus metabolism is due to a tumor product that may be acting via stimulation of adenylate activity. Increased bone resorption prior to surgical resection suggests that the tumor may also produce an osteoclast activator. The rise in resorption surface and pyridinium cross-link excretion, increase in serum osteocalcin and bone mineralization, normalization of osteoid width, and fall in cancellous bone volume after resection are consistent with healing of osteomalacia by rapid remodeling.

Immunolocalization of matrix metalloproteinases and tissue inhibitors of metalloproteinases in human chondrosarcomas.

We have immunohistochemically examined the localization of matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) in human chondrosarcomas (CHS) (23 cases) and benign chondroid lesions (BCL) (16 cases of osteochondromas and 11 cases of enchondromas). In CHS, all the MMPs and TIMPs examined were positive. Among them, MMP-1 was immunolocalized in more than 90% of both CHS and BCL, but positive score of MMP-1 was significantly higher in CHS than that in BCL (p < 0.01). Compared with BCL, CHS expressed MMP-3 at a low level, and more often positive in MMP-9. It is possible that chondrosarcoma might have a tendency to lose the ability to secrete MMP-3, which is a metalloproteinase that can degrade cartilage proteoglycans and is related to normal cartilage turnover. MMP-2, TIMP-1 and TIMP-2 were immunolocalized in more than 70% of the cases of both BCL and CHS, but the positive scores of these were not statistically different between the two groups. Interestingly, in several cases of CHS, both MMP-1 and MMP-9 immunostains were observed preferentially within the cells at the marginal areas of cartilaginous lobules. These findings suggest that increased expression of MMP-1 and MMP-9 and decrease in MMP-3 expression are associated with the malignant phenotype of the cartilaginour tumors.

Selective increase in expression of isoform PP1 gamma 1 of type-1 protein phosphatase in chondrosarcoma cells.

The expression of the two catalytic subunits of protein phosphatase (PP) type 1 PP1 gamma 1 and PP1 delta was examined in 4 cases of osteochondroma and 4 cases of enchondroma as a benign cartilaginous tumor, and 4 cases of chondrosarcoma as a malignant cartilaginous tumor using immunohistochemical analysis. The percentage of tumor cells stained positively with antiserum against PP1 catalytic subunit isoform PP1 gamma 1 were significantly higher in chondrosarcoma than in osteochondroma and enchondroma. Furthermore, chondrosarcoma showed markedly high S-phase fraction in the cell cycle of tumor cells, as compared to osteochondroma and enchondroma. These results suggest that PP1 gamma 1 is involved in the accelerated growth of malignant cells in chondrosarcoma.

Isozymes of glycogen phosphorylase in human cartilage and cartilage tumors.

Isozyme patterns of glycogen phosphorylase in normal fetal and adult human cartilage and benign and malignant cartilage tumors were investigated by means of polyacrylamide gel disc electrophoresis. In both normal cartilage and cartilage tumors four distinct bands were observable: these were of the liver, liver-like and fetal types and of a hybrid type between the muscle and fetal types judging by their mobilities in the small-pore gels containing 0.01% and 0.1% glycogen. The isozyme pattern of benign cartilage tumors was almost identical with that of normal cartilage. In normal fetal and adult cartilage and benign cartilage tumors, the liver and liver-like types were predominant, whereas in malignant cartilage tumors they were minor and the fetal type was predominant. These results indicate fetal deviation of phosphorylase isozymes in malignant transformation of cartilage cells.

Enzyme histochemical study on bone tumors.

A total of 19 cases with bone tumors, including six osteosarcomas. three giant cell tumors of bone, one malignant fibrous histiocytoma, four nonossifying fibromas, four chondromas and one chondrosarcoma, were examined as to enzyme histochemistry; the enzymes consisted of alkaline phosphatase (ALPase), acid phosphatase (ACPase), nonspecific esterase (NSE), adenosine triphosphatase (ATPase), 5'-nucleotidase (5'-Nucl) and beta-glucuronidase (beta-Gl). Osteosarcoma was strongly positive for ALPase followed by 5'-Nucl. Giant cell tumor, malignant fibrous histiocytoma and nonossifying fibroma showed enzyme histochemistry similar to each other: multinucleated giant cells and round cells in these tumors were strongly positive for ACPase, NSE, ATPase and 5'-Nucl simulating osteoclasts and histiocytes, whereas spindle cells were positive for ATPase and 5'-Nucl in their cytoplasm and weakly positive for ACPase. Chondroma and chondrosarcoma were focally positive for ACPase and NSE; the ACPase was sensitive to tartaric acid treatment. These observations showed that ALPase activity is very characteristic to osteosarcoma, and is useful for its diagnosis. From enzyme histochemistry, giant cell tumor, malignant fibrous histiocytoma and nonossifying fibroma can be regarded as a histiocyte-derived tumor of bone in contrast to osteosarcoma and cartilaginous tumors.

I-Naphthyl acetate esterase isoenzymes in synovial fluids and radiography of temporomandibular and knee joints.

Esterase isoenzymes of synovial fluids were presented in cases of pain-dysfunction syndrome, osteoarthrosis, osteochondroma, malignant fibrous histiocytoma of temporomandibular joint, and hemarthrosis, rheumatoid arthritis of the knee joint. Radiographic features of them were also comparatively presented in several cases. The electrophoretogram of I-Naphthyl acetate esterase of pain-dysfunction syndrome showed the esterase-I, while when inflammatory process developed at joints, electrophoretic pattern of synovial fluids revealed a rather similar feature of sera with variable stainabilities of esterase-I, and -III, irrespective of any inflammation. Osteochondroma showed two components of esterase-I and -III, on the other hand malignant fibrous histiocytoma presented esterase-I and -III in the early stage but a more complicated pattern in recurrence. Among these conditions, the most similarity between synovial fluid and serum was demonstrated in the case of hemarthrosis on the electrophoretogram of I-Naphthyl acetate esterases.