Estimating the direct costs of oral cancer in Spain: a retrospective hospital data analysis.

BACKGROUND: Studies on the costs incurred from cancer in Spain are scarce and have focused on the most prevalent types such as colorectal, breast, and lung cancer. The aim of this study was to calculate the direct costs associated with the diagnostic, treatment and follow-up procedures for oral cancer in Spain. MATERIAL AND METHODS: Applying a bottom-up approach, we retrospectively analyzed the medical records of a cohort of 200 patients with oral cancer (C00-C10), diagnosed and treated in Spain between 2015 and 2017. For each patient, we collected their age, sex, degree of medical impairment (American Society of Anesthesiologists [ASA] classification), tumor extent (TNM classification), relapses and survival during the first 2 years of follow-up. The final calculation of the costs is expressed in absolute values in euros as the percentage of the gross domestic product per capita and in international dollars (I$). RESULTS: The total cost per patient rose to €16,620 (IQR, €13,726; I$11,634), and the total direct cost at the national level was €136,084,560 (I$95,259,192). The mean cost for oral cancer represented 65.1% of the gross domestic product per capita. The costs for the diagnostic and therapeutic procedures were determined by the ASA grade, tumor size, lymph node infiltration and presence of metastases. CONCLUSIONS: The direct costs for oral cancer are considerable compared with other types of cancer. In terms of gross domestic product, the costs were similar to those of countries neighboring Spain, such as Italy and Greece. The main determinants of this economic burden were the patient's degree of medical impairment and tumor extent.

ABCG2/BCRP gene expression is related to epithelial-mesenchymal transition inducer genes in a papillary thyroid carcinoma cell line (TPC-1).

Tumor malignancy is associated with the epithelial-mesenchymal transition (EMT) process and resistance to chemotherapy. However, little is known about the relationship between the EMT and the multidrug-resistance gene in thyroid tumor progression. We investigated whether the expression of the ABCG2/BCRP gene is associated with ZEB1 and other EMT inducer genes involved in tumor dedifferentiation. We established a subpopulation of cells that express the ABCG2/BCRP gene derived from the thyroid papillary carcinoma cell line (TPC-1), the so-called TPC-1 MITO-resistant subline. The most relevant findings in these TPC-1 selected cells were a statistically significant upregulation of ZEB1 and TWIST1 (35- and 15-fold change respectively), no changes in the relative expression of vimentin and SNAIL1, and no expression of E-cadherin. The TPC-1 MITO-resistant subline displayed a faster migration and greater invasive ability than parental cells in correlation with a significant upregulation of the survivin (BIRC5) gene (twofold change, P<0.05). The knockdown of ZEB1 promoted nuclear re-expression of E-cadherin, reduced expression of vimentin, N-cadherin, and BIRC5 genes, and reduced cell migration (P<0.05). Analysis of human thyroid carcinoma showed a slight overexpression of the ABCG2/BCRP at stages I and II (P<0.01), and a higher overexpression at stages III and IV (P<0.01). SNAIL1, TWIST1, and ZEB1 genes showed higher expression at stages III and IV than at stages I and II. E- and N-cadherin genes were upregulated at stages I and II of the disease (ninefold and tenfold change, respectively, P<0.01) but downregulated at stages III and IV (fourfold lower, P<0.01). These results could be a promising starting point for further study of the role of the ABCG2/BCRP gene in the progression of thyroid tumor.

Mutation at position -132 in the islet amyloid polypeptide ( IAPP) gene promoter enhances basal transcriptional activity through a new CRE-like binding site.

AIMS/HYPOTHESIS: Mutations in the islet amyloid polypeptide ( IAPP) gene may play a potential role in the abnormal regulation or expression of the peptide. The aim of this study was to determine the functional role of the -132 G/A mutation reported in the promoter region of the IAPP gene in a population of Spanish Type 2 diabetic patients. METHODS: We investigated the transcriptional activity using MIN6 cells and luciferase reporter plasmids in several culture conditions. Key regulatory elements of the IAPP promoter region were also analysed by electrophoretic mobility shift assays (EMSA). RESULTS: The mutant construct doubled IAPP transcriptional activity ( p<0.001). Both constructs showed severely reduced promoter activity (four-fold decrease) in the presence of verapamil and diazoxide. In contrast, IAPP promoter activity was doubled after incubation with forskolin or dexamethasone, regardless of the glucose concentrations in the culture media. EMSA revealed that the -132 G/A mutation increased the binding affinity through two DNA-protein complexes. In addition, a cAMP-responsive element binding protein (CREB) was identified by super-shift EMSA. CONCLUSIONS/INTERPRETATION: Our studies show that the wild-type and the mutant constructs are regulated in a similar pattern under all conditions, strongly indicating that the -132 G/A mutation increases basal but not inducible transcription. These results may be explained by new binding to the mutant region through CREB and other transcription factors not yet identified.

Genetic, clinical, and biochemical analysis of unrelated Spanish families with multiple endocrine neoplasia type I.

OBJECTIVE: (1) To study seven unrelated Spanish families with multiple endocrine neoplasia type I (MEN I), describing clinical features and investigating the presence of germline mutations in the MEN1 gene, and (2) to establish reference values for pancreatic polypeptide and gastrin after a standardized test meal in a healthy control group, analyzing the usefulness of this test for detecting neuroendocrine gastroenteropancreatic tumors in subjects with MEN I. METHODS: Two or three generations of 7 kindreds with MEN I, consisting of a total of 39 individual family members, were investigated. Three of the families were subjected only to genetic analysis, and the other four families were also assessed clinically. A group of 23 healthy control subjects were also studied. RESULTS: Mutations in the MEN1 gene were found in six of the seven families studied. Of the 4 families studied clinically, 12 family members were genetically affected. In these study subjects, hyperparathyroidism, adrenal adenomas, neuroendocrine gastroenteropancreatic tumors, and pituitary adenomas developed in 100%, 50%, 16%, and 12%, respectively. All demonstrated pancreatic tumors were associated with abnormal results after a test meal, but 75% of them also showed high basal hormonal measurements. CONCLUSION: Analysis of the MEN1 gene decreases the total number of subjects who need to undergo repeated clinical and biochemical studies, but genetic mutations are not detected in all families with MEN I. Hyperparathyroidism is the most common manifestation of the syndrome, but the presence of adrenal adenomas has probably been underestimated. Ingestion of a standardized test meal for stimulation of gastrin and pancreatic polypeptide could be a complementary procedure for diagnosing gastroenteropancreatic tumors in selected patients with MEN I in whom basal gastrin and pancreatic polypeptide levels are normal.

IDDM12 (CTLA4) on 2q33 and IDDM13 on 2q34 in genetic susceptibility to type 1 diabetes (insulin-dependent).

Type 1 diabetes (insulin-dependent) is a multifactorial disease with polygenic susceptibility. The major genetic component (IDDM1) resides within the HLA region, but several non-HLA loci have been implicated in the genetic susceptibility. In the present study, we have analysed two such loci, IDDM12 (CTLA4) on 2q33 and IDDM13 on 2q34, in Danish (n = 254) and Spanish (n = 39) type 1 diabetic multiplex families. No significant evidence of linkage of IDDM12 was observed in any of the two studied data sets. However, when the present data were combined with previously published data, they strengthened the evidence of linkage at this locus, p = 0.00002. For the IDDM13 region, we found some positive evidence of linkage of the D2S137-D2S164-D2S1471 markers (p-values 0.007, 0.02, and 0.007, respectively) using transmission disequilibrium testing (TDT) and the Tsp version of the TDT. Importantly, random transmission of all tested alleles was observed in unaffected offspring (p > 0.3). Stratification for HLA (high risk and non-high risk genotypes) in the Danish families did not reveal heterogeneity at IDDM12 or IDDM13. In conclusion, our data on an entirely new family data set did not support the existence of IDDM12 as a type 1 diabetes susceptibility locus in the Danish population. In addition, we found support for evidence of linkage and association of the IDDM13/D2S137-D2S1471 region (approximately 3.5 cM) to type 1 diabetes, however, further studies are needed to substantiate this observation.

Modulation of vasoconstrictor and dilator pancreatic metabolites in streptozotocine diabetic rats: effect of bradykinin blockage and NO inhibition.

This study was designed to investigate the effect of HOE 140 (a bradykinin beta2 receptor antagonist) and N(w)-nitro-L-arginine methyl-ester (L-NAME, a nitric oxide synthase inhibitor) on endothelial and beta-cell function in induced streptozotocine (Stz) diabetic rats. The decrease in the insulinogenic index after Stz effect (control 286.03+/-104.12 and Stz 18.22+/-10.77, P<0.001 vs. Control) was partially prevented by L-NAME (46.54+/-10.12, P<0.001) and HOE 140 (105.12+/-23.06, P<0.001). It was observed in diabetic rats: L-NAME increased the pancreatic endothelin-1 (ET-1) production and HOE 140 did not. L-NAME and HOE 140 decreased the nitric oxide (NO) synthesis, increased prostacyclin 1-2 (PGI2), and did not modify thromboxane A-2 (TxA2). These results indicate that L-NAME and HOE 140 had a protective effect on the development of diabetes in the rat. The protective effect of L-NAME and HOE 140 on the insulinogenic index could be related to ET-1, bradykinin, PGI2, and NO.

N-Ras induces alterations in Golgi complex architecture and in constitutive protein transport.

Aberrant glycosylation of proteins and lipids is a common feature of many tumor cell types, and is often accompanied by alterations in membrane traffic and an anomalous localization of Golgi-resident proteins and glycans. These observations suggest that the Golgi complex is a key organelle for at least some of the functional changes associated with malignant transformation. To gain insight into this possibility, we have analyzed changes in the structure and function of the Golgi complex induced by the conditional expression of the transforming N-Ras(K61) mutant in the NRK cell line. A remarkable and specific effect associated with this N-Ras-induced transformation was a conspicuous rearrangement of the Golgi complex into a collapsed morphology. Ultrastructural and stereological analyses demonstrated that the Golgi complex was extensively fragmented. The collapse of the Golgi complex was also accompanied by a disruption of the actin cytoskeleton. Functionally, N-Ras-transformed KT8 cells showed an increase in the constitutive protein transport from the trans-Golgi network to the cell surface, and did not induce the appearance of aberrant cell surface glycans. The Golgi complex collapse, the actin disassembly, and the increased constitutive secretion were all partially inhibited by the phospholipase A2 inhibitor 4-bromophenylacyl bromide. The results thus suggest the involvement of the actin cytoskeleton in the shape of the Golgi complex, and intracellular phospholipase A2 in its architecture and secretory function.

Molecular pathology of multiple endocrine neoplasia type I: two novel germline mutations and updated classification of mutations affecting MEN1 gene.

Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant disorder characterized by the combined development of tumors in several endocrine glands and other tissues. The MEN1 gene was recently identified and isolated by positional cloning. This gene was screened in two unrelated MEN1 Spanish kindreds (with four affected members and seven asymptomatic members) using single-strand conformation polymorphism, DNA sequencing, and restriction enzyme analysis. Two novel germline mutations were identified: a missense in exon 2 (H139R) and a splice-site in intron 9 (1461-2A>C). These findings allowed us to identify the MEN1 carriers among the seven asymptomatic members analyzed. An updated review of the mutations and polymorphisms found in the analysis of the MEN1 gene is provided. The report of all germline mutations causing MEN1 and easy access to this updated information are both of special diagnostic interest, because this greatly facilitates the task of attributing the disorder to a specific mutation found in a given MEN1 family. This is especially helpful in the critical differentiation of missense mutations from nonsynonymous polymorphisms that fit the pattern of segregation of the disease, but do not cause it.

A novel germline mutation in exon 5 of the multiple endocrine neoplasia type 1 gene.

The autosomal dominant multiple endocrine neoplasia type 1 (MEN1) syndrome is characterized by neoplasia of parathyroids, anterior pituitary, and gastrointestinal and pancreatic neuroendocrine tissues. Recently the gene responsible for the MEN1 syndrome has been identified on chromosome region 11q13. Most of the described mutations are nucleotide substitutions and small deletions affecting exons 2 and 3, causing protein truncation. Only one mutation in exon 5 has been found, and this corresponds to a MEN1 sporadic case. Small insertions are also rare. We studied a MENI family composed of five members, two of whom were clinically affected. We found a new germline 1 basepair insertional mutation affecting the exon 5 of the MEN1 gene in the two members affected in this MEN1 family.

Use of somatostatin analogue scintigraphy in the localization of recurrent medullary thyroid carcinoma.

Detection of recurrence of medullary thyroid carcinoma (MTC) remains a diagnostic problem. Increased serum tumour marker levels frequently indicate recurrence while conventional imaging techniques (CIT) are non-diagnostic. In this study, we performed indium-111 octreotide scintigraphy and CIT in a series of 20 patients with MTC presenting with elevated serum tumour markers after surgery. 111In-octreotide whole-body studies detected 15 pathological uptake foci in 11 of the 20 patients studied and CIT detected 17 lesions in 11 of the 20 patients. Ten patients underwent reoperation, five of them with positive 111In-octreotide scintigraphy and CIT and two with positive isotopic exploration and negative CIT. Surgical findings demonstrated that the results of isotopic study and CIT had been false-positive for MTC in one case (sarcoidosis). The six patients with true-positive 111In-octreotide studies had significantly higher basal calcitonin (CT) and carcinoembryonic antigen (CEA) levels than the patients with negative isotopic studies. The expression of somatostatin receptor (SSTR) subtypes by PC-PCR could be investigated in four cases with a positive isotopic study. Among the three cases with a true-positive study, SSTR2, the SSTR subtype that preferentially binds to the somatostatin analogue octreotide, was detected in two, SSTR5 was demonstrated in the three, and SSTR3 was detected in one. No subtype of SSTR was detected in the case with a final diagnosis of sarcoidosis. We conclude that 111In-octreotide has limited sensitivity in detecting recurrence in patients with MTC, although its sensitivity may improve with high serum CT levels. This radionuclide imaging technique should be employed when conventional imaging techniques are negative or inconclusive or when the presence of somatostatin receptors may provide the basis for treatment with somatostatin analogues.

Somatostatin and somatostatin receptor subtype gene expression in medullary thyroid carcinoma.

The possible existence of an autocrine/paracrine role for SRIF in normal and neoplastic thyroid parafollicular C cells has supported the use of SRIF analogues in the treatment of patients with medullary thyroid carcinoma (MTC). In this study, we have investigated the expression of SRIF by immunohistochemistry and RT-PCR, and the expression of SRIF receptor (SSTR) subtypes by RT-PCR, in a series of 14 MTCs. SRIF messenger RNA was detected in all cases, although immunoreactive cells were only identified in 8. SSTR messenger RNA was present in 12 out of the 14 tumors. Expression of more than 1 SSTR subtype was detected in 10 tumors. SSTR2, the subtype that preferentially binds to the SRIF analogue octreotide, was the subtype most frequently detected, whereas SSTR4 was not detected in any case. These results confirm the frequent expression of both SRIF and its receptors in MTC. The presence of different combinations of SSTR subtypes in a given patient may explain the variable clinical response to SRIF analogues and may promote the search for more selective drugs with different affinities to the various receptor subtypes.

Circannual somatostatin gene and somatostatin receptor gene expression in the early post-natal rat pineal gland.

The origin of somatostatin in the pineal gland is controversial as a double origin - neural in pinealopetal nerve fibers and parenchymal corresponding to locally synthesized peptide - is suggested. We have investigated the ontogeny and possible circadian and circannual variations of somatostatin and somatostatin receptor gene expression in the rat pineal gland using RT-PCR. Somatostatin gene expression was observed in the pineal of animals up to 15 days of postnatal life in autumn and winter, but not in prepubertal or adult rats; no transcript was detected during spring and summer at any age and a circadian rhythm was only detected in 15-day-old rats. In situ hybridization confirmed these results. The transcript of the somatostatin receptor gene (SSTR2) was detected at all ages studied with no seasonal variations in its expression, indicating separate regulatory mechanisms. In conclusion, somatostatin and somatostatin receptor mRNAs are expressed in the neonatal rat pineal with circadian and seasonal variations.

In vivo and in vitro flow cytometry comparative analysis of somatostatin-positive cells in the pineal gland of the neonatal rat.

Flow cytometry was used for comparative in vivo and in vitro analysis of cell populations staining positively for somatostatin. Experiments were carried out with pineals obtained from neonatal, 8- and 15-day-old rats. Pineal cells were obtained by dispersion with collagenase and then processed in a flow cytometer or maintained in culture for 1 or 2 weeks. Identification of somatostatin-immunopositive cell populations was performed using a polyclonal somatostatin antibody and confirmed by indirect immunostaining of cytospun smears with the avidin-biotin-peroxidase method. In vivo, the percentage of somatostatin-positive cells was 60.6 +/- 4% in neonatal pineals and declined to 22.2 +/- 11% in 15-day-old animals (p < 0.04). The density of peptide immunostaining decreased in 8-day-old animals but recovered to the neonate levels in 15 day-old animals; homogeneity in the immunopositive population increased with age. Maintenance in culture for 1 week resulted in an increase in positive somatostatin staining in animals of 8 and 15 days with no changes in neonates; however, after 2 weeks of culture, the percent of immunopositive cells decreased from 53.3 +/- 6 to 12.2 +/- 4% in the older animals and remained unchanged in neonates. We conclude that somatostatin is found in pinealocytes and shows a declining pattern during the perinatal period; this probably implies that the peptide plays a paracrine role important for cell differentiation in these young animals, since maximal cellularity and a high mitotic index occur within the first 3 days of life, and pineal cell differentiation is completed before the end of the third week of extrauterine life.

Expression of the ret proto-oncogene in phaeochromocytoma. An in situ hybridization and northern blot study.

The expression and distribution of ret proto-oncogene mRNA were investigated in five phaeochromocytomas of both familial and sporadic types by in situ hybridization (ISH) using digoxigenin-labelled cRNA probes and Northern blot (NB) analysis with random priming labelled cDNA probes. The probes corresponded to the tyrosine kinase domain of the gene. An excellent correlation was found between the ISH and NB results. By both techniques, the expression of the ret proto-oncogene was detected in three of the five cases, two of four familial tumours, and the only sporadic tumour that was studied. The results confirm that ret is frequently expressed in phaeochromocytomas and suggest that it might be an important event in their development and/or progression.

Expression of somatostatin in rat pineal cells in culture.

The expression of somatostatin mRNA was investigated in rat pineal cells after 1 week in culture, using reverse transcription of mRNA into cDNA and the polymerase chain reaction. The positive expression in cultured pineal cells demonstrates the capacity of this gland to synthesize somatostatin in denervated cells. Thus, apart from the neural origin of pineal somatostatin, which has been described in detail in the bovine species, a parenchymal source is demonstrated.