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1.
Med Hypotheses ; 97: 22-25, 2016 Dec.
Article in English | MEDLINE | ID: mdl-27876123

ABSTRACT

Neuroblastoma (NB) is the most common extra cranial solid tumor of childhood and often lethal in childhood. Clinical and biologic characteristics that are independently prognostic of outcome in NB are currently used for risk stratification to optimally the therapy. It includes age at diagnosis, International Neuroblastoma Staging System tumor histopathology and MYCN amplification. However, even in patients with theoretically good prognosis, such as localized tumor and non-amplified MYCN, either disease progress or recurrence may occur. Potential genetic determinants of this unfavorable behavior are not yet fully clarified. The presence of elevated expression of AHCY, PKMYT1, and BLM has accompanied poor prognosis MYCN-amplified neuroblastoma patients. Considering the potential implication of these genes on the clinical management of NB, we hypothesize that the identification of genetic variations may have significant impact during development of the recurrent or progressive disease. Using targeted DNA sequencing, we analyzed the mutation profiles of the genes PKMYT1, AHCY, and BLM in tumor samples of five patients with MYCN amplified and 15 MYCN non-amplified NB. In our study, BLM germline variants were detected in two patients with MYCN-non-amplified neuroblastoma. Our data allow us to hypothesize that, regardless of MYCN status, these mutations partially abolish BLM protein activity by impairing its ATPase and helicase activities. BLM mutations are also clinically relevant because BLM plays an important role in DNA damage repair and the maintenance of genomic integrity. We also found a novel variant in our cohort, PKMYT1 mutation localized in the C-terminal domain with effect unknown on NB. We hypothesize that this variant may affect the catalytic activity of PKMYT1 in NB, specifically when CDK1 is complexed to cyclins. The prognostic value of this mutation must be further investigated. Another mutation identified was a nonsynonymous variant in AHCY. This variant may be related to the slow progression of the disease, even in more aggressive cases. It affects the maintenance of the catalytic capacity of AHCY, leading to the consequent functional effects observed in the NB patients studied. In conclusion, our hypothesis may provide that mutations in BLM, AHCY and PKMYT1 genes found in children with MYCN-amplified or MYCN-non amplified neuroblastomas, may be associated with the prognosis of the disease.


Subject(s)
Adenosylhomocysteinase/genetics , Brain Neoplasms/genetics , Germ-Line Mutation , Membrane Proteins/genetics , N-Myc Proto-Oncogene Protein/genetics , Neuroblastoma/genetics , Protein Serine-Threonine Kinases/genetics , Protein-Tyrosine Kinases/genetics , RecQ Helicases/genetics , Child , Cohort Studies , DNA Damage , DNA Repair , Disease Progression , Drug Resistance, Neoplasm , Gene Expression Regulation, Neoplastic , Genetic Variation , Genome, Human , Humans , Models, Theoretical , Neoplasm Recurrence, Local , Prognosis , Protein Domains , Risk Factors , Sequence Analysis, DNA
2.
Braz. j. med. biol. res ; 45(9): 851-855, Sept. 2012. ilus, tab
Article in English | LILACS | ID: lil-646332

ABSTRACT

The tumorigenesis of pituitary adenomas is poorly understood. Mutations of the PIK3CA proto-oncogene, which encodes the p110-α catalytic subunit of PI3K, have been reported in various types of human cancers regarding the role of the gene in cell proliferation and survival through activation of the PI3K/Akt signaling pathway. Only one Chinese study described somatic mutations and amplification of the PIK3CA gene in a large series of pituitary adenomas. The aim of the present study was to determine genetic alterations of PIK3CA in a second series that consisted of 33 pituitary adenomas of different subtypes diagnosed by immunohistochemistry: 6 adrenocorticotropic hormone-secreting microadenomas, 5 growth hormone-secreting macroadenomas, 7 prolactin-secreting macroadenomas, and 15 nonfunctioning macroadenomas. Direct sequencing of exons 9 and 20 assessed by qPCR was employed to investigate the presence of mutations and genomic amplification defined as a copy number ≥4. Previously identified PIK3CA mutations (exon 20) were detected in four cases (12.1%). Interestingly, the Chinese study reported mutations only in invasive tumors, while we found a PIK3CA mutation in one noninvasive corticotroph microadenoma. PIK3CA amplification was observed in 21.2% (7/33) of the cases. This study demonstrates the presence of somatic mutations and amplifications of the PIK3CA gene in a second series of pituitary adenomas, corroborating the previously described involvement of the PI3K/Akt signaling pathway in the tumorigenic process of this gland.


Subject(s)
Adolescent , Adult , Aged , Female , Humans , Male , Middle Aged , Young Adult , Adenoma/genetics , Gene Amplification/genetics , Mutation/genetics , /genetics , Pituitary Neoplasms/genetics , Immunohistochemistry , Signal Transduction
3.
Braz J Med Biol Res ; 45(9): 851-5, 2012 Sep.
Article in English | MEDLINE | ID: mdl-22782554

ABSTRACT

The tumorigenesis of pituitary adenomas is poorly understood. Mutations of the PIK3CA proto-oncogene, which encodes the p110-α catalytic subunit of PI3K, have been reported in various types of human cancers regarding the role of the gene in cell proliferation and survival through activation of the PI3K/Akt signaling pathway. Only one Chinese study described somatic mutations and amplification of the PIK3CA gene in a large series of pituitary adenomas. The aim of the present study was to determine genetic alterations of PIK3CA in a second series that consisted of 33 pituitary adenomas of different subtypes diagnosed by immunohistochemistry: 6 adrenocorticotropic hormone-secreting microadenomas, 5 growth hormone-secreting macroadenomas, 7 prolactin-secreting macroadenomas, and 15 nonfunctioning macroadenomas. Direct sequencing of exons 9 and 20 assessed by qPCR was employed to investigate the presence of mutations and genomic amplification defined as a copy number ≥4. Previously identified PIK3CA mutations (exon 20) were detected in four cases (12.1%). Interestingly, the Chinese study reported mutations only in invasive tumors, while we found a PIK3CA mutation in one noninvasive corticotroph microadenoma. PIK3CA amplification was observed in 21.2% (7/33) of the cases. This study demonstrates the presence of somatic mutations and amplifications of the PIK3CA gene in a second series of pituitary adenomas, corroborating the previously described involvement of the PI3K/Akt signaling pathway in the tumorigenic process of this gland.


Subject(s)
Adenoma/genetics , Gene Amplification/genetics , Mutation/genetics , Phosphatidylinositol 3-Kinases/genetics , Pituitary Neoplasms/genetics , Adolescent , Adult , Aged , Class I Phosphatidylinositol 3-Kinases , Female , Humans , Immunohistochemistry , Male , Middle Aged , Proto-Oncogene Mas , Signal Transduction , Young Adult
4.
J Endocrinol Invest ; 34(8): e214-8, 2011 Sep.
Article in English | MEDLINE | ID: mdl-21270509

ABSTRACT

BACKGROUND: Pituitary tumors account for approximately 10-15% of intracranial neoplasms. AIM: Using the cDNA microarray method, we have previously compared expression under two distinct conditions: a pool of 4 clinically non-functioning pituitary adenomas (NFPA) and a spinal cord metastasis of a non-functioning pituitary carcinoma, in order to gain biological insights into genomic changes of pituitary neoplasias. In the present study, we further investigated the mRNA expression of 3 selected genes previously described as being involved in other neoplasias based on a series of 60 pituitary adenomas: CRABP1 (cellular retinoic acid binding protein 1), GRP (gastrin-releasing peptide), and RERG (Ras-related, estrogen- regulated, growth inhibitor). MATERIAL AND METHODS: The expression of CRABP1, GRP, and RERG was determined by quantitative RT-PCR. RESULTS: A significantly higher content of CRABP1 mRNA was observed in NFPA compared to functioning adenomas, and PRL-secreting adenomas showed a lower expression of this gene compared to normal pituitary. A lower expression of GRP mRNA was detected in NFPA compared to normal pituitary and also to functioning adenomas. RERG mRNA was overexpressed in NFPA in comparison to functioning adenomas and to normal pituitary. Among the functioning adenomas, only the ACTH-secreting adenomas presented a higher expression of RERG mRNA compared to normal pituitary. CONCLUSIONS: The findings of differential expression of CRABP1 in prolactinomas and of RERG in NFPA compared to normal pituitary suggests that retinoic acid and estrogen receptor, respectively, could be involved in the tumorigenesis of these adenomas subtypes. Additional studies are required to further confirm this hypothesis.


Subject(s)
ACTH-Secreting Pituitary Adenoma/metabolism , Adenoma/metabolism , GTP Phosphohydrolases , Gastrin-Releasing Peptide , Pituitary Neoplasms/metabolism , RNA, Messenger/metabolism , Receptors, Retinoic Acid , ACTH-Secreting Pituitary Adenoma/genetics , Adenoma/genetics , Adolescent , Adult , Aged , GTP Phosphohydrolases/genetics , GTP Phosphohydrolases/metabolism , Gastrin-Releasing Peptide/genetics , Gastrin-Releasing Peptide/metabolism , Gene Expression , Humans , Male , Middle Aged , Oligonucleotide Array Sequence Analysis , Pituitary Gland/metabolism , Pituitary Neoplasms/genetics , Prolactin/metabolism , RNA, Messenger/genetics , Receptors, Retinoic Acid/genetics , Receptors, Retinoic Acid/metabolism , Young Adult
5.
Braz J Med Biol Res ; 42(9): 791-5, 2009 Sep.
Article in English | MEDLINE | ID: mdl-19738985

ABSTRACT

Amplification of the MYCN gene in neuroblastomas is a potent biological marker of highly aggressive tumors, which are invariably fatal unless sound clinical management is applied. To determine the usefulness of semi-quantitative differential PCR (SQ-PCR) for accurate quantification of MYCN gene copy number, we evaluated the analytical performance of this method by comparing the results obtained with it for 101 tumor samples of neuroblastoma to that obtained by absolute and relative real-time PCR. Similar results were obtained for 100 (99%) samples, no significant difference was detected between the median log10 MYCN copy number (1.53 by SQ-PCR versus 1.55 by absolute real-time PCR), and the results of the two assays correlated closely (r = 0.8, Pearson correlation; P < 0.001). In the comparison of SQ-PCR and relative real-time PCR, SQ-PCR versus relative real-time PCR concordant results were found in 100 (99%) samples, no significant difference was found in median log10 MYCN copy number (1.53 by SQ-PCR versus 1.27 by relative real-time PCR), and the results of the two assays correlated closely (r = 0.8, Pearson correlation; P < 0.001). These findings indicate that the performance of SQ-PCR was comparable to that of real-time PCR for the amplification and quantification of MYCN copy number. Thus, SQ-PCR can be reliably used as an alternative assay in laboratories without facilities for real-time PCR.


Subject(s)
Neuroblastoma/genetics , Nuclear Proteins/genetics , Oncogene Proteins/genetics , Polymerase Chain Reaction/methods , Female , Gene Amplification , Humans , Male , N-Myc Proto-Oncogene Protein
6.
Braz. j. med. biol. res ; 42(9): 791-795, Sept. 2009. ilus, graf
Article in English | LILACS | ID: lil-524321

ABSTRACT

Amplification of the MYCN gene in neuroblastomas is a potent biological marker of highly aggressive tumors, which are invariably fatal unless sound clinical management is applied. To determine the usefulness of semi-quantitative differential PCR (SQ-PCR) for accurate quantification of MYCN gene copy number, we evaluated the analytical performance of this method by comparing the results obtained with it for 101 tumor samples of neuroblastoma to that obtained by absolute and relative real-time PCR. Similar results were obtained for 100 (99 percent) samples, no significant difference was detected between the median log10 MYCN copy number (1.53 by SQ-PCR versus 1.55 by absolute real-time PCR), and the results of the two assays correlated closely (r = 0.8, Pearson correlation; P < 0.001). In the comparison of SQ-PCR and relative real-time PCR, SQ-PCR versus relative real-time PCR concordant results were found in 100 (99 percent) samples, no significant difference was found in median log10 MYCN copy number (1.53 by SQ-PCR versus 1.27 by relative real-time PCR), and the results of the two assays correlated closely (r = 0.8, Pearson correlation; P < 0.001). These findings indicate that the performance of SQ-PCR was comparable to that of real-time PCR for the amplification and quantification of MYCN copy number. Thus, SQ-PCR can be reliably used as an alternative assay in laboratories without facilities for real-time PCR.


Subject(s)
Female , Humans , Male , Neuroblastoma/genetics , Nuclear Proteins/genetics , Oncogene Proteins/genetics , Polymerase Chain Reaction/methods , Gene Amplification
7.
J Neuroendocrinol ; 20(9): 1052-7, 2008 Sep.
Article in English | MEDLINE | ID: mdl-18624930

ABSTRACT

The neurotensin (NT) produced in the hypothalamus and in pituitary gonadotrophs and thyrotrophs participates in neuroendocrine regulation. Recently, the involvement of this peptide in normal and neoplastic cell proliferation has been postulated. In the present study, we evaluated the expression of NT and its receptors (NTR1, 2 and 3) in a series of 50 pituitary adenomas [11 growth hormone (GH)-, eight prolactin (PRL)-, four adrenocorticotrophic hormone (ACTH)- and 27 nonfunctioning adenomas]. NT mRNA expression was significantly higher in functioning compared to nonfunctioning adenomas and with normal pituitary. Nonfunctioning pituitary adenomas showed lower expression of NT mRNA than normal pituitary. In the immunohistochemical study of functioning adenomas, NT was colocalised with GH, PRL and ACTH secreting cells. In nonfunctioning adenomas, the NT immunoreactivity intensity was variable among the samples. NTR3 mRNA expression was observed in all examined samples and was higher in the adenomas, both functioning and nonfunctioning, compared to normal pituitary. By contrast, NTR1 and NTR2 mRNA were not detected in either pituitary adenomas or normal tissue. The higher expression of NTR3, as well as the expression of NT by tumoural corticotrophs, lactotrophs and somatotrophs, which are cells types that do not express this peptide in the normal pituitary, suggests that NT autocrine and/or paracrine stimulation mediated by NTR3 may be a mechanism associated with the tumourigenesis of functioning adenomas.


Subject(s)
Adenoma/genetics , Neurotensin/genetics , Pituitary Neoplasms/genetics , Receptors, Neurotensin/genetics , Adenoma/metabolism , Adenoma/pathology , Adult , Aged , Autocrine Communication/genetics , Autocrine Communication/physiology , Female , Gene Expression Regulation, Neoplastic , Humans , Male , Middle Aged , Neurotensin/metabolism , Paracrine Communication/genetics , Paracrine Communication/physiology , Pituitary Neoplasms/metabolism , Pituitary Neoplasms/pathology , RNA, Messenger/metabolism , Receptors, Neurotensin/metabolism , Tumor Cells, Cultured , Young Adult
8.
J Endocrinol Invest ; 29(9): 826-30, 2006 Oct.
Article in English | MEDLINE | ID: mdl-17114915

ABSTRACT

INTRODUCTION: About a third of acromegalic patients is resistant to available SS analogs (SA), octreotide (OCT) and lanreotide (LAN). Such resistance is related to reduction of SS receptor (SSTR) density or to a different expression of SSTR subtypes. There are 5 known SSTR subtypes. SSTR2 and SSTR5 are usually expressed in GH-secreting pituitary tumors, and both SA bind preferentially to SSTR2 and, to a lesser extent, to SSTR5. We herein describe an acromegalic patient who presented impressive tumor shrinkage without hormonal normalization during primary therapy with SA. MATERIAL AND METHODS: This 23-yr-old male acromegalic patient was treated with slow-release LAN (LAN-SR), 30 mg every 10 days for six months, followed by OCT-LAR, 30 mg every 28 days for an additional six months with a 75% tumor volume reduction but without GH and IGF-I normalization. Subsequently, he underwent pituitary surgery and expression of SSTR in the removed tumor was performed by real time RT-PCR by the 2-deltaCt method, using GAPDH as internal control. All PCR products were confirmed by automated sequencing. RESULTS: SSTR expression revealed an unusual profile, with almost exclusively expression of SSTR3. CONCLUSIONS: These unusual clinical and receptor subtypes profile suggest an important role of SSTR3 on tumor shrinkage. The low affinity of LAN and OCT for this SSTR subtype could be compensated by its high expression in this GH-secreting pituitary macroadenoma.


Subject(s)
Acromegaly/drug therapy , Growth Hormone-Secreting Pituitary Adenoma/drug therapy , Human Growth Hormone/blood , Insulin-Like Growth Factor I/analysis , Octreotide/therapeutic use , Peptides, Cyclic/therapeutic use , Receptors, Somatostatin/metabolism , Somatostatin/analogs & derivatives , Acromegaly/blood , Acromegaly/diagnostic imaging , Acromegaly/etiology , Adult , Gene Expression , Growth Hormone-Secreting Pituitary Adenoma/blood , Growth Hormone-Secreting Pituitary Adenoma/complications , Growth Hormone-Secreting Pituitary Adenoma/diagnostic imaging , Humans , Male , Pituitary Gland/diagnostic imaging , Radiography , Remission Induction/methods , Somatostatin/therapeutic use
9.
Neuroendocrinology ; 82(3-4): 208-14, 2005.
Article in English | MEDLINE | ID: mdl-16601360

ABSTRACT

In order to search for candidate genes related to pituitary adenoma aggressiveness, the present investigation was intended to compare the mRNA expression profile from a pool of four nonfunctional pituitary adenomas (NFPA) with a spinal cord metastasis of a nonfunctional pituitary carcinoma (MNFPC). The metallothionein isoform 3 (MT3) gene was differentially expressed in nonfunctional adenomas in comparison to the metastasis of nonfunctional carcinoma. A microarray dataset comprising 19,881 probes was employed for comparing expression profiles of a spinal cord metastasis of a nonfunctional pituitary carcinoma with a pool of four nonfunctional pituitary adenomas. RT-qPCR confirmed the microarray findings and was used to investigate MT3 mRNA gene expression in tumor samples of a series of 52 different pituitary adenoma subtypes comprising 10 corticotropin (ACTH)-producing, 18 growth hormone (GH)-producing, 8 prolactin (PRL)-producing, and 16 nonfunctional adenomas. Microarray data analysis by GeneSifter program unveiled Gene Ontology terms related to zinc ion-binding activity closely related to MT3 function. MT3 mRNA expression was statistically significantly higher in ACTH-producing pituitary adenomas and in nonfunctional pituitary adenomas in comparison to the other pituitary adenoma subtypes. The more abundant expression of this gene in ACTH-producing pituitary adenomas suggests that MT3 could be related to distinct pituitary cell lineage regulating the activity of some transcription factor of importance in hormone production and/or secretion.


Subject(s)
Adenoma/metabolism , Adrenocorticotropic Hormone/metabolism , Nerve Tissue Proteins/biosynthesis , Pituitary Neoplasms/metabolism , Adenoma/pathology , Gene Expression , Human Growth Hormone/metabolism , Humans , Metallothionein 3 , Nerve Tissue Proteins/genetics , Oligonucleotide Array Sequence Analysis , Pituitary Neoplasms/secondary , Prolactin/metabolism , Protein Isoforms/biosynthesis , RNA, Messenger/analysis , Reverse Transcriptase Polymerase Chain Reaction , Spinal Cord Neoplasms/secondary
10.
Braz J Med Biol Res ; 27(9): 2181-4, 1994 Sep.
Article in English | MEDLINE | ID: mdl-7787801

ABSTRACT

F9 mouse teratocarcinoma cells have a high capacity to adhere to laminin and we identified alpha 6/beta 1 integrin as the principal laminin-binding protein present in these cells. F9 cells differentiated into parietal endoderm when monolayer cultures were treated with retinoic acid and dibutyryl cyclic AMP. In this process a decreased adherence to laminin was observed due to a lower expression of alpha 6/beta 1 integrin on the cell surface.


Subject(s)
Down-Regulation , Integrins/physiology , Laminin/physiology , Receptors, Laminin/physiology , Tretinoin/pharmacology , Animals , Bucladesine/pharmacology , Cell Adhesion , Cell Differentiation/drug effects , Integrin alpha6beta1 , Integrins/metabolism , Laminin/metabolism , Mice , Protein Binding , Receptors, Laminin/metabolism , Tumor Cells, Cultured/drug effects
11.
Braz. j. med. biol. res ; 27(9): 2181-4, Sept. 1994. graf
Article in English | LILACS | ID: lil-144470

ABSTRACT

F9 mouse teratocarcinoma cells have a high capacity to adhere to laminin and we identified alpha6/beta1 integrin as the principal laminin-binding protein present in these cells. F9 cells differentiated into parietal endoderm when monolayer cultures were treated with retinoic acid and dibutyryl cyclic AMP. In this process a decreased adherence to laminin was observed due to a lower expression of alpha6/beta1 integrin on the cell surface


Subject(s)
Mice , Animals , Down-Regulation , Integrins/physiology , Laminin/physiology , Tretinoin/pharmacology , Cell Adhesion , Bucladesine/pharmacology , Cell Differentiation , Flow Cytometry , Integrins/metabolism , Laminin/metabolism , Protein Binding , Receptors, Laminin/metabolism , Receptors, Laminin/physiology , Tumor Cells, Cultured/drug effects
12.
Int J Clin Lab Res ; 24(3): 148-53, 1994.
Article in English | MEDLINE | ID: mdl-7529599

ABSTRACT

The integrin family of adhesion receptors is likely to be important for tumor cell invasion and dissemination. We have studied the effects of the differentiating agents retinoic acid on integrin expression by the human melanoma cell line MeWo. Our results show that this agent inhibits cellular proliferation, increases melanin content and induces morphological changes in MeWo cells. Functionally, these alterations are associated with an enhanced adhesion to matrix protein vitronectin and higher levels of expression of vitronectin receptor on the cell surface. This is accompanied by increased levels of alpha v integrin mRNA. Although the mechanism by which retinoic acid regulates the expression of vitronectin receptor in MeWo cells needs further examination, this system may represent a good model for understanding the role of this receptor in melanoma progression, as well the molecular basis for retinoic acid therapy in these tumors.


Subject(s)
Integrins/drug effects , Melanoma/metabolism , Receptors, Cytoadhesin/drug effects , Tretinoin/pharmacology , Blotting, Northern , Cell Adhesion/drug effects , Cell Division/drug effects , Humans , Integrins/biosynthesis , Melanins/metabolism , Melanoma/pathology , Receptors, Cytoadhesin/biosynthesis , Receptors, Vitronectin , Tumor Cells, Cultured
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