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Among the four isoforms of the calcitonin receptor (CTR) described in humans, two differ by the presence of h-CTR1 or absence of h-CTR2 of 16 amino acids in the first intracellular loop. Both receptors are biologically active. The TT cell line derived from a human medullary carcinoma of the thyroid is characterized by the secretion of large amounts of calcitonin. We have recently shown that this cell line expresses h-CTR2. In the present work we have studied the expression of CTR during TT cell proliferation and used dexamethasone to modify calcitonin expression in order to establish if an autocrine regulation involving calcitonin and its receptor was functional in the TT cells. The expression of this receptor and of calcitonin during TT cell proliferation was studied by reverse transcriptase-polymerase chain reaction (RT-PCR). Dexamethasone, a potent inhibitor of TT cell proliferation, levels (day 6 of culture) specifically increased receptor levels from day 8 onwards. CT peptide and CT mRNA levels decreased or were similar during experimental time. CTR regulation by glucocorticoids is suggested in TT cells. Autocrine regulation of CTR is also suggested by relation between CT mRNA levels and CTR mRNA.

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We recently reported the presence of a truncated form (h-CTR2) of the human calcitonin receptor (CTR) in TT cells, a cell line derived from medullary thyroid carcinoma (MTC). This form (h-CTR2), characterized by the absence of 16 amino acids in the first intracellular domain, was also detected in two cases of MTC. In the present study we determined the expression of CTR mRNA in a larger sample, representative of the different clinical forms of MTC, and in normal thyroid. h-CTR2 was expressed in all MTC specimens (both sporadic and familial) and in the normal thyroid samples. The expression of the receptor mRNA was higher in MTC compared with normal thyroid. Moreover, CT and CTR mRNA levels were modified significantly during proliferation. This result suggests that CT may be involved in proliferation of MTC via autocrine/paracrine regulation. Calcitonin secretion by MTC may play a role in the development and spread of these tumors.

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NAD+-dependent 15-hydroxyprostaglandin dehydrogenase (type-I 15-PGDH) inactivates prostaglandins. We recently reported an mRNA sequence coding for a predicted isomer (PGDH(rI)) of this enzyme. The TT cell line, derived from medullary thyroid carcinoma (MTC), expresses mRNAs for both isomers. We report here the expression by TT cells and MTC of a third 15-PGDH related mRNA (PGDH(rII)), 241 nt shorter than type-I 15-PGDH. RNase protection assays confirmed that TT cells expressed this mRNA (PGDH(rII)). Thus different splicing patterns could be involved in the post-transcriptional regulation of type-I 15-PGDH gene in MTC.

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Medullary thyroid carcinoma (MTC) is a rare endocrine tumour secreting the calcitonin (CT), its main tumoral marker, occuring as a sporadic or a familial disease. These diseases are associated with a 5-years prognostic from less than 50 to 100%, depending on the tumoral stage at the diagnosis time. The surgical management is demonstrated to be able to obtain a biological recovery in some patients. The other therapeutic means have no or poorly effects on the MTC evolution. Partial and transient responses are described for 15 to 20% of the patients with the use of multiple trials of chemotherapy. The external radiotherapy may have some little effects on local tumoral involvement without influence on the survival rates as compared to the surgery alone. The hormonal therapy with somatostatin analogues and the metabolic scintigraphies using MIBG, somatostatine analogues, radio-active iodine have no therapeutic effects demonstrated. The development of targeted therapy by the use of specific monoclonal antibodies, such as anti-CEA radiolabeled antibodies, is to be evaluated. The prognostic factors, as defined in large series of patients studied, does not necessary reflect the aggressiveness of the tumour. Thus, the therapeutic approach of MTC patients must take into consideration that the biologic activity of the disease does not implicate an aggressive disease. The evaluation of CT and CEA circulating levels remains of prognostic value and a useful tool for the practicians for the management of MTC patients.

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Calcitonin (CT), a hypocalcemic and hypophosphatemic hormone, is produced by the C-cells of the thyroid gland. It is the main tumoral marker of medullary thyroid carcinoma (MTC). Hypersecretion of CT is also associated with other types of tumors. Thus, heterogeneity of circulating CT can play an important role in the accurate determination of hormone levels in blood samples obtained from MTC patients. Further studies will be necessary to establish the predictive value of the several peptides coded by the calcitonin gene family. All of them specifically reflect the ways and the pattern of alternative splicing of the primary transcript of the Calc I gene. Such relations implicate further investigations concerning the relationship between calcitonin circulating levels, biosynthetic activity of C-cells and the expression of gene encoding for this hormone, in normal and neoplastic conditions.

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We have recently identified in medullary thyroid carcinoma the existence of a second calcitonin messenger, generated by a splicing between the 3' coding region of exon 4 and exon 5 of Calc I gene. It differs from the first one in its 3' coding sequence and codes for a calcitonin precursor which generates the same N terminal peptide, calcitonin and a specific 21 amino acid carboxy terminal peptide differing from Katacalcin by its 8 last amino acids. We searched for the expression of this new messenger in normal human thyroid tissue by Northern and by polymerase chain reaction techniques. This second calcitonin messenger was expressed in 4/4 normal thyroids and 4/5 medullary thyroid carcinoma tissue samples. The expression of this second messenger is apparently a common occurrence in C cells whether normal or tumoral.

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OBJECTIVE: Treatment by octreotide has been suggested in medullary thyroid carcinoma patients with post-surgery metastases. The purpose of this study was to evaluate if the tumoral regression could be improved by a high dose and by prolonged octreotide treatment. DESIGN: Fourteen thyroidectomized patients were studied. All patients had persistently elevated plasma calcitonin levels with normal or elevated carcino-embryonic antigen levels. Five hundred micrograms/day of octreotide were administered by continuous subcutaneous infusion for 90 days. MEASUREMENTS: Plasma calcitonin and carcino-embryonic antigen levels were determined at days -30, -20, -2, -1, 0, +30, +60, +90, +120; morphological extension was evaluated every month. RESULTS: Continuous infusion of octreotide did not induce any significant decrease of calcitonin levels, or any morphological improvement, and had no major undesirable effect. However, in 4/14 patients calcitonin levels fell during treatment (-43, -50, -15, -20%), and in 9 patients calcitonin increased (+22 to +130%) after cessation of therapy. CONCLUSION: Biological or morphological parameters of medullary thyroid carcinoma are not significantly improved in a large series of patients treated by octreotide.

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Two genes code for calcitonin gene-related peptides (CGRPs). One expresses by tissue-specific alternate splicing calcitonin and CGRP I mRNAs, the other CGRP II mRNA. Calcitonin is the marker of sporadic or hereditary human medullary thyroid carcinoma (MTC). CGRP II expression is not well established in normal or tumoral thyroid. After amplification by polymerase chain reaction, CGRP I and II mRNAs were detected in six cases of MTC associated with other endocrine neoplasia (MEN IIa) and in two cases of isolated MTC. CGRP I was detected in all non-C cell tumoral thyroids (6 samples), CGRP II was barely detectable in three out of six cases. CGRP II could be a specific tumoral marker of MTC.

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The current work has been performed by the Cooperative French Group of Medullary Thyroid Carcinoma (GETC). A systematic evaluation of RIA somatostatin (SRIH) was performed in 34 medullary thyroid carcinomas (MTC) (25 inherited, seven sporadic). Plasma SRIH was measured by radioimmunoassay in parallel with calcitonin (CT) and carcinoembryonic antigen (CEA). Immunoassayable SRIH was tested in fresh tumoral tissue samples from the same 34 MTC and, for comparison, in 10 nontumoral thyroid extracts (less than 6 pmol/g wet). Although plasma SRIH was only slightly elevated in two of 20 cases, tumoral SRIH was elevated in 70.6% of our MTC (10 to 3973 pmol/g). The chromatography of two tumoral extracts showed that somatostatin 14 was the major molecular form. We found no correlation (P greater than 0.1) between tumoral SRIH and the following: (1) tumor size (r = 0.227); (2) epidemiologic form of MTC (r = 0.144); (3) plasma SRIH (r = 0.045), plasma CT (R = 0.095) or (4) plasma CEA (r = 0.032). Thus, in the authors' experience, SRIH appears as a major product of tumoral C-cell in human MTC, even when plasma SRIH is normal and SRIH immunohistochemical staining is scarce. Multiple hormonal production of these tumors may explain its presence but SRIH may act also as a regulator, since negative influence of SRIH on CT is demonstrated in normal as well in tumoral conditions.

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Medullary thyroid carcinoma is a rare and sometimes hereditary disease. The tumour can be diagnosed and followed up by measuring the amounts of calcitonin it secretes. The prognosis of this cancer largely depends on an early diagnosis and treatment. From the clinical and laboratory (calcitonin assays) data recorded in our department, we have endeavoured to determine the influence of a national multidisciplinary co-operative group (GETC: French medullary study group) on the diagnosis and prognosis of this malignancy. We are able to show that the number of medullary thyroid carcinomas detected (principally in their familial forms) has increased by 141 per cent after the GETC was created. Calcitonin levels at the time of diagnosis are significantly lower (P less than 0.05) in familial cases, which reflects an early detection. The same applies to post-operative calcitonin levels (P less than 0.005), so that in the long run a better prognosis can be expected. It seems therefore that together with a better knowledge of this cancer and its detection, the setting up of a national multidisciplinary co-operative group results in a better clinical and therapeutic approach of these patients, and particularly of the familial cases of medullary thyroid carcinoma.

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Screening for medullary thyroid cancer (MTC) in France is based on a protocol that has been widely distributed nationally. A network of coordinators utilizing a common questionnaire provides for an effective national screening program. Calcitonin stimulation procedures are systematically used for all first-degree relatives of MTC patients. Pathological studies utilize special immunopathologic techniques. Genealogic information is obtained on all index cases, and blood specimens are collected for establishing permanent cell lines. The data collected are used not only to establish the diagnosis of the hereditary or sporadic form of the disease but also to expand the screening as appropriate. This common protocol has benefited patients and their families by improving early detection of cases, increasing the number of families available for follow-up, and improving the prognosis of this cancer. Studies on these families have contributed significantly to the localization of the multiple endocrine neoplasia type 2 gene.

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Recent studies have suggested that somatostatin could reduce calcitonin plasma levels (CT) in normal subjects and in medullary thyroid carcinoma (MTC). The aim of this study was to examine the usefulness of the somatostatin analog, sandostatine (SMS 201.995) in MTC with elevated residual CT levels post-thyroidectomy with or without metastases. 18 patients (17-64 years, 12 men and 8 women) with CT greater than 850 pg/ml (N less than 150 pg/ml) and with metastases in 12 cases, were studied. MTC was sporadic in 11 cases, familial in 4 cases and of undefined form in 3. Initial posology was 300 micrograms/d of sandostatin (3 injections/day). It was then increased by 300 micrograms/d every 9 day till a maximum of 1500 micrograms/d. Treatment duration was 37 days in 11 cases and 60 days in 7 cases. Plasma CT and carcinoembryonic antigen levels (CEA) were measured before treatment and at the end of each dosage plateau. Morphologic evaluation of metastases was done at 0, 30, 60 days. 7/18 patients were reevaluated 2 to 8 months after with drawal of sandostatine. Treatment was well tolerated. Flushes improved in 4 out of 5 cases but diarrhea in only 2 out of 9 patients. Sandostatine was without any effect on plasma CEA. Heterogenous responses were observed for plasma CT levels (CT decreases greater than 20% in 8/18 patients when 900 to 1500 micrograms/day were administered). Patients were subdivised into 3 groups according to CEA levels and presence or absence of metastases. Group A (n = 9) had elevated CEA levels (greater than 10 mg/ml) and metastases.(ABSTRACT TRUNCATED AT 250 WORDS)

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MTC is characterized by multiple humoral and hormonal manifestations. Although calcitonin is the specific marker of the disease, somatostatin, the pro-opiomelanocortin derived peptides and bombesin--among hormones produced by the tumor--can represent an exacerbation of normal C cells potentialities through genome derepression induced by the cancer. In this paper, the functional polymorphism of princeps tumoral markers and the endocrinological aspects of this neoplasia are reviewed. Molecular biology has been instrumental in discovering new tumoral peptides ("ancestral" CT forms, cryptic peptide and CGRP) and methods of CT detection; therefore, the role of CT could be better evaluated. In addition to its calciotropic role, CT acts also as a neuromodulator on some hypophyseal hormones. Conversely, CT secretion is also regulated by amines and neuropeptides, providing the basis of potential hormonal treatment.

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DNA complementary to mRNA extracted from the thyroid glands of patients suffering from medullary carcinoma of the thyroid (MCT), a calcitonin-producing tumour, was inserted in the Pst site of pBR 322 by G-C tailing. The recombinant plasmids were used to transform Escherichia coli DP 50. Ampicillin-resistant clones were screened using a 32P-labelled cDNA to mRNA extracted from a case of MCT particularly rich in calcitonin (CT) mRNA. Positive clones were subsequently rescreened using a 32P poly(T) probe. Eighty clones were thus purified, and the inserts obtained by digestion with PstI were subjected to positive hybridization selection with subsequent translation in vitro. An insert stimulating synthesis of the protein and containing restriction sites compatible with the previously published complete sequence of calcitonin mRNA from rat was sequenced. This cDNA insert contained the entire coding region of 426 bp, 70 bp at the 5'-end, and 295 bp upstream from the poly(A) tail. The complete amino acid sequence of human preprocalcitonin could thus be deduced.

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