WT1 expression as a potential biomarker of malignancy in canine breast tumor / La expresión de WT1 como un biomarcador potencial maligno en un tumor de mama canino

SUMMARY: Veterinary oncology is very important nowadays to get a better understanding of human carcinogenesis. Estrogen receptor, progesterone receptor and Human Epidermal Growth Factor receptor 2 are frequently evaluated by immunohistochemistry (HIC) in human breast tumor. WT1 is an oncogene, its overexpression has been detected in leukemia and diverse solid tumors like breast cancer, lung cancer and mesothelioma in humans. WT1 expression was evaluated in 15 canine breast tumors (CBT) diagnosed by histopathological analysis to find its relationship with neoplasia and malignancy. IHC and RT-PCR were performed in CBT tissues. Fisher´s test was used to analyze WT1 relationship with malignancy. Of the 15 tumors, 9 (60 %) were diagnosed as benign and 6 (40 %) were malignant. With IHC, WT1 expression was positive only in biopsies diagnosed as malignant. Expression of WT1 by RT-PCR was detected in 14 of the 15 tumors (93.33 %) as well as in control healthy mammary gland. Clinical significance: This study describes for the first time a close correlation between CBT and a positive result for WT1 expression with IHC; hence, it can be used as a biomarker for this neoplasia and as an indicator of malignancy. RT-PCR analysis also showed to be good option to detect WT1 expression. These results will be useful to further investigations to elucidate WT1-related signaling pathways in CBT. Also to know molecules that regulate the translation of this protein as a marker for tumor progression.

RESUMEN: La oncología veterinaria es muy importante hoy en día para comprender mejor la carcinogénesis humana. El receptor de estrógeno, el receptor de progesterona y el receptor 2 del factor de crecimiento epidérmico humano se evalúan con frecuencia mediante inmunohistoquímica (HIC) en tumores de mama humanos. WT1 es un oncogén, su sobreexpresión se ha detectado en leucemia y en diversos tumores sólidos como el cáncer de mama, cáncer de pulmón y mesotelioma en humanos. La expresión de WT1 se evaluó en 15 tumores de mama caninos (TCC) diagnosticados mediante análisis histopatológico para encontrar su relación con la neoplasia y la malignidad. IHC y RT-PCR se realizaron en tejidos CBT. La prueba de Fisher se utilizó para analizar la relación de WT1 con la malignidad. De los 15 tumores, 9 (60 %) fueron diagnosticados como benignos y 6 (40 %) fueron malignos. Con IHC, la expresión de WT1 fue positiva solo en biopsias diagnosticadas como malignas. La expresión de WT1 por RT-PCR se detectó en 14 de los 15 tumores (93,33 %), así como en el control de la glándula mamaria sana. Importancia clínica: este estudio describe por primera vez una estrecha correlación entre la TCC y un resultado positivo para la expresión de WT1 con IHC; por lo tanto, se puede utilizar como un biomarcador para esta neoplasia y como un indicador de malignidad. El análisis por RT-PCR también demostró ser una buena opción para detectar la expresión de WT1. Estos resultados serán útiles para futuras investigaciones para dilucidar las vías de señalización relacionadas con WT1 en la TCC. También para conocer moléculas que regulan la traducción de esta proteína como marcador de progresión tumoral.

Le néphroblastome au CHU de Brazzaville

Le néphroblastome est le deuxième cancer de l'enfant à Brazzaville. Nous avons mené cette étude afin d'en dégager les principales caractéristiques et les difficultés de leur prise en charge à Brazzaville au Congo.Il s'était agi d'une étude descriptive transversale sur une période de six ans (2008-2013). Nous avions ainsi colligé les patients atteints de néphroblastome dont nous avons décrits les aspects épidémiologiques, cliniques et thérapeutiques. Durant la période d'étude, 52 enfants, âgés de 0 à 14 ans avaient été colligés, dont 14 pour néphroblastome. Nous avions constaté une prédominance féminine et l'âge moyen au diagnostic était de 50,6 mois. La masse abdominale était la principale circonstance de découverte. Un cas de néphroblastome bilatéral avait été retrouvé. La survie globale à 6 mois, était de d'environ 65 %

AAcute WT1-positive promyelocytic leukemia with hypogranular variant morphology, bcr-3 isoform of PML-RARα and Flt3-ITD mutation: a rare case report / Leucemia promielocítica aguda WT1-positivo com morfologia variante hypogranular, isoforma bcr-3 da PML-RARα e mutação Flt3-ITD: relato de caso raro

ABSTRACT CONTEXT: Acute promyelocytic leukemia (APL) accounts for 8% to 10% of cases of acute myeloid leukemia (AML). Remission in cases of high-risk APL is still difficult to achieve, and relapses occur readily. CASE REPORT: Here, we describe a case of APL with high white blood cell counts in blood tests and hypogranular variant morphology in bone marrow, together with fms-like tyrosine kinase-3 with internal tandem duplication mutations (FLT3-ITD), and bcr-3 isoform of PML-RARα. Most importantly, we detected high level of Wilms’ tumor gene (WT1) in marrow blasts, through the reverse transcription polymerase chain reaction (RT-PCR). To date, no clear conclusions about an association between WT1 expression levels and APL have been reached. This patient successively received a combined treatment regimen consisting of hydroxycarbamide, arsenic trioxide and idarubicin plus cytarabine, which ultimately enabled complete remission. Unfortunately, he subsequently died of sudden massive hemoptysis because of pulmonary infection. CONCLUSION: Based on our findings and a review of the literature, abnormal functioning of WT1 may be a high-risk factor in cases of APL. Further studies aimed towards evaluating the impact of WT1 expression on the prognosis for APL patients are of interest.

RESUMO CONTEXTO: Leucemia promielocítica aguda (LPA) compreende 8% a 10% dos casos de leucemia mieloide aguda (LMA). A remissão em casos de LPA de alto risco ainda é dificilmente conseguida, e recorrência é comum. RELATO DE CASO: Descrevemos aqui um caso de LPA com glóbulos brancos elevados no exame de sangue e a morfologia variante hipogranular na medula óssea, juntamente com fms-like tirosina-quinase-3 com mutações de duplicação em tandem interna (FLT3-ITD) e a isoforma bcr-3 de PML- RARα. Mais importante, detectamos alto nível de gene do tumor de Wilms (WT1) em blastos medulares por RT-PCR (reverse transcription polimerase chain reaction). Até agora, não há conclusões claras sobre a associação entre os níveis de expressão WT1 e APL. Este paciente recebeu sucessivamente regime de tratamento combinado, de hidroxicarbamida, trióxido de arsênico e idarrubicina e citarabina, alcançando finalmente a remissão completa. Infelizmente, em seguida, ele morreu de repente de hemoptise maciça devido a uma infecção pulmonar. CONCLUSÃO: Com base em nossos resultados e numa revisão da literatura, a função anormal de WT1 pode ser um fator de alto risco em casos de APL. Novos estudos, com o objetivo de avaliar o impacto da expressão de WT1 no prognóstico dos doentes com APL, são de interesse.

Expression of WT1 Gene in Bone Marrow of Patients with Acute Myeloid Leukemia and Its Influence on Prognosis / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the expression level of WT1 gene in bone marrow of patients with acute myeloid leukemia (AML) and its relationship with prognosis.</p><p><b>METHODS</b>The copy numbers of WT1 and internal reference gene in bone marrow samples from 75 newly diagnosed AML patients were detected by using real-time quantitative PCR. The gene WT1 expression level was determined by the ratio of the copy numbers of WT1 to reference gene. And the clinical characteristics, the complete remission (CR) rate after induction chemotherapy, 2-year overall survival (OS) rate and event-free survival (EFS) rate were calculated and analysed.</p><p><b>RESULTS</b>The expression level of WT1 did not significantly correlate with common clinical parameters such as age, sex, molecular abnormality, FAB classification and risk stratification. The CR rate in the high WT1 expression group before treatment was 65.4%, which was lower than that of 93.9% in the low expression group (χ2=8.25, P<0.01). The 2-year overall survival rate and event-free survival rate of the two groups were statistically significantly different (P<0.05), and the OS and EFS rates in high WT1 expression group were lower than those in low expression group. After the induction chamotheropy for about 1, 3 month and 6 months, the 2-year OS rate significantly increased in patients with decrease of WT1 gene expression level by one log or more (P<0.05).</p><p><b>CONCLUSION</b>The expression level of WT1 gene in bone marrow may be an effective marker to evaluate therapy efficacy and prognosis for AML patients (non APL).</p>

Recent advances in the research on mechanisms underlying podocyte-specific gene mutation-related steroid-resistant nephrotic syndrome / 中国当代儿科杂志

Steroid-resistant nephrotic syndrome poses a significant clinical challenge. Its pathogenesis has not been fully elucidated. In recent years, numerous studies have shown that podocyte-specific gene mutations may play important roles in the development of steroid-resistant nephrotic syndrome. Among the identified genes mutated in podocytes include NPHS2, NPHS1, WT1, TRPC6, MDR1, PLCE1, LMX1B, and LAMB2. This review aims to summarize the characteristics of these mutated genes in podocytes. The putative role for these podocyte-specific mutated genes in the pathogenesis, diagnosis, treatment and prognosis of steroid-resistant nephrotic syndrome is also discussed.

WT1 gene expression difference in leukemia and non-leukemia and its clinical significance / 中国实验血液学杂志

This study was aimed to investigate the expression level of Wilms' tumor 1( WT1) gene in hematologic neoplasm (leukemia, multiple myeloma and lymphoma) patients and its clinical significance. Real-time quantitative polymerase chain reaction (RQ-PCR) was used to detect the copy number of WT1 gene and reference gene (ALB) in bone marrow cells of 228 patients with hematologic neoplasm in our hospital. The gene expression level was determined by using the ratio of the copy number of WT1 gene and reference gene. The results showed that the WT1 expression level between male and female patients was not statistically significantly different (P > 0.05). All the patients were divided into 3 groups: the group aged under 19, the group aged between 19-50, and the group aged over 50; the WT1 expression level among the three groups were not statistically significantly different (P > 0.05) . The above-mentioned patients were redivided into the groups aged under 45 and over 45, the difference between them was not statistically significant (P > 0.05). The difference of WT1 expression level between newly diagnosed patients and treated patients with hematologic neoplasm was statistically significant (P < 0.01), but no statistically significant difference of WT1 expression was found (P > 0.05) at each stage within 3 years after treatment, however, among them the difference between newly diagnosed leukemia patients and treated leukemia patients was very statistically significant (P < 0.01), while the difference between newly diagnosed and treated non-leukemia patients was not statistically significant (P > 0.05). The expression difference of WT1 between leukemia and non-leukemia patients was very statistically significant (P < 0.01), the difference between the newly diagnosed leukemia and non-leukemia patients also was very statistically significant (P < 0.01). The difference of WT1 expression between treated leukemia and non-leukemia patients was not statistically significant (P > 0.05). It is concluded that the WT1 expression level in leukemia patients can be a reliable marker to evaluate the prognosis of newly diagnosed leukemia and the curative effect for minimal residual disease. No WT1 expression difference has been found before and after treatment among the patients with non-leukemia, such as multiple myeloma and lymphoma, therefore, which should be furtherly explored.

Promoter methylation of Wilms' tumor gene on the X- chromosome in gastric cancer / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the changes in methylation levels of the promoters of the tumor suppressor gene Wilms' tumor gene on the X-chromosome (WTX) and its possible role in gastric cancer.</p><p><b>METHODS</b>WTX promoter methylation levels were detected in 20 pairs of specimens of gastric cancer and matched normal tissues and in 3 gastric cancer cell lines (MGC803, SCG7901, and BGC823) using the Sequenom MassARRAY quantitative analysis system. The gastric cancer cell line BGC823 was treated with 5-aza-2'-deoxycytidine (5-aza-dC) for demethylation and the changes in the level of WTX promoter methylation were investigated.</p><p><b>RESULTS</b>WTX promoter methylation levels were very low and showed no significant differences among normal gastric tissues, gastric cancer tissues and the 3 gastric cancer cell lines. In BGC823 cells, treatment with 5-aza-dC did not obviously affect the promoter methylation levels of WTX.</p><p><b>CONCLUSION</b>High methylation levels of WTX promoters are rare in gastric cancer.</p>

Clinical and pathological features of Denys-Drash syndrome: report of 3 cases / 中华儿科杂志

<p><b>OBJECTIVE</b>To study the clinical and pathological features of Denys-Drash syndrome (DDS).</p><p><b>METHOD</b>Three DDS cases who were treated in our department from December 2009 to June 2011 were subjected to this study by reviewing of literature.</p><p><b>RESULT</b>Both case 1 and case 2 were female, with karyotype 46, XX. Case 3 was male with bilateral cryptorchidism. The ages of nephropathy onset of the three cases were 1 year and 9 months, 2 years and 8 moths, and 3 months respectively. Proteinuria in case 2 and case 3 were evidenced to be resistant to steroid. Case 1 was partially responsive to tacrolimus, plasma albumin and cholesterol were improved, although proteinuria was persistent after Tacrolimus was administered. Remission was achieved in case 2 after administration of cyclosporine A and later tacrolimus, and her renal function remains normal till present (4 years and 9 months). Residue renal histology revealed diffused mesangial sclerosis (DMS) in all three patients. All of the three patients had developed right unilateral Wilms tumor. A novel WT1 missense mutation exon 9 c.1213C > G was detected in case 1. WT1 exon 9 c.1168C > T nonsense mutation and exon 8 c.1130A > T missense mutation were detected in case 2 and case 3, respectively.</p><p><b>CONCLUSION</b>The clinical manifestation of nephropathy in DDS is variable. The majority present with early onset nephropathy and reach renal failure before the age of 4 years. But in a few patients, nephropathy can also be present much later and progress slowly. Proteinuria in DDS is resistant to steroid but is responsive to calcineurin inhibitors, including Cyclosporine A. The effectiveness of tacrolimus was also observed in this study. DDS is evidently caused by WT1 mutation. DMS is the characteristic renal pathological change in DDS.</p>

Definição do perfil de alterações das sequências dos genes APC, CTNNB1, WT1, ETX e PLCG2 em tumores de Wilms / Wilms tumor genomic re-sequencing for evaluating gene loci of WTX, WT1, CTNNB1, APC and PLCG2

O tumor de Wilms (TW) origina-se das células precursoras do rim embrionário, e já foram observados eventos comuns a ambos os processos, tumorigênese e nefrogênese. A identificação de alterações moleculares durante esses processos é crucial para entender os eventos desencadeadores do TW. O gene WT1 codifica para um fator de transcrição com expressão finamente coordenada durante o desenvolvimento do rim, e mutações nesse gene são relatadas em 10% dos TWs esporádicos. Adicionalmente, desregulação no nível de expressão ou mutações de outros genes como a CTNNB1 também sugerem uma conexão entre TW e nefrogênese. Também são encontradas mutações de WTX em alguns casos de TWs. Atualmente, acredita-se que mutações de WTX, WT1 e CTNNB1 juntas estão associadas a cerca de 30% dos TWs, sendo que mutações gênicas não foram associados para a maioria dos casos. Resultados de um projeto desenvolvido em nosso laboratório apontaram os genes APC e PLCG2 como candidatos a estarem alterados nesse tumor. Nesse estudo anterior, o APC foi observado com localização predominantemente nuclear nos TWs, semelhante aos estágios iniciais do desenvolvimento normal do rim, e diferente do rim normal, que apresenta localização citoplasmática. O PLCG2 foi associado pela primeira vez a uma doença, sendo observada a ausência de marcação proteica na maioria dos TW, semelhante aos estágios iniciais do desenvolvimento, e diferente do rim diferenciado, que apresenta alta expressão da proteína. Com o advento de técnicas modernas de sequenciamento, velozes do ponto de vista de geração de bases e relativamente baratas, tornou-se factível a avaliação de toda a região que abrange o gene a um custo e tempo relativamente baixos. Com isso torna-se viável aplicar o método de sequenciamento de alta performance para avaliação de diversos pacientes...

Imagenes pediatricas inusuales: tumor de Wilms gigante / Unusual pediatric imaging: giant Wilms tumor

Paciente femenino de 7 años de edad, sin antecedentes perinatales ni patológicos relevantes. Ingresó al Hospital del Niño por un cuadro clínicode 7 meses de evolución caracterizado por dolor abdominal crónico recurrente, distención abdominal, hiporexia, pérdida de peso, disnea progresiva, compromiso del estado general y en el último tiempo hematuria macroscópica.

Establishment of BGC-823/WTX-EGFP gastric cancer cell line stably expressing Wilms tumor gene on X chromosome / 南方医科大学学报

<p><b>OBJECTIVE</b>To establish the BGC-823/WTX-EGFP gastric cancer cell line with stable expression of Wilms tumor gene on the X chromosome (MTX) for functional analysis of WTX gene.</p><p><b>METHODS</b>The full-length WTX cDNA was amplified from human embryonic kidney 293FT cells and cloned into the pEGFP-N1 vector containing the reporter gene of green fluorescence protein. The recombinant pEGFP-WTX expression vector, after digestion by restriction enzyme to identify the size of target gene fragment, was transfected into 293FT cells and the expression of fluorescent reporter gene was observed under fluorescence microscope. pEGFP-WTX vector was transfected into human gastric cancer BGC-823 cell line to establish BGC-823/WTX-EGFP cell line stably expressing WTX. Quantitative RT-PCR and immunocytochemical staining were used to detect the expression of WTX in both BGC-823/WTX-EGFP and control BGC-823 cells.</p><p><b>RESULTS</b>The recombinant pEGFP-WTX plasmid was successfully constructed and verified by PCR and sequencing. The mRNA and protein expressions of MTX were significantly increased in BGC-823/WTX-EGFP cells as compared with those in the control cells.</p><p><b>CONCLUSION</b>The full-length WTX expression vector (pEGFP-WTX) and BGC-823/WTX-EGFP gastric cancer cell line have been successfully established to facilitate further functional study of WTX gene.</p>

Alteration of the expression ratio between WT1 gene and its isomers during all-trans retinoic acid-induced differentiation of HL-60 cells / 中华病理学杂志

<p><b>OBJECTIVE</b>To explore expression ratio alteration between WT1 gene and its isomers during differentiation of leukemia cell line HL-60 induced by all-trans retinoic acid (ATRA) and the relationship existed between them.</p><p><b>METHODS</b>The degree of cellular maturation was verified by NBT reduction test and immunophenotyping. Expression of unspliced WT1, WT1 (17AA+) and WT1 (KTS+) were determined by real-time fluorescence quantitative RT-PCR during the induced differentiation of HL-60 cells. The relative ratio of four splicing variants WT1 (+/+), WT1 (+/-), WT1 (-/+), WT1 (-/-) were analyzed.</p><p><b>RESULTS</b>During the induced differentiation of HL-60 cells, NBT reduction rate and CD11b positive rate increased significantly (P < 0.05 and P < 0.001, respectively). The expression of WT1 gene decreased from (4.17 +/- 2.21) x 10(-3) at 0 hour to (7.53 +/- 2.30) x 10(-4) at the 96th hour. The ratio of 17AA+decreased from 0.60 +/- 0.05 at 0 hour to 0.42 +/- 0.08 at the 96th hour. The ratio of KTS+ decreased from 0.53 +/- 0.08 at 0 hour to 0.41 +/- 0.04 at the 96th hour. The ratio of WT1 (+/+) decreased gradually from 0.32 +/- 0.06 at 0 hour to 0.17 +/- 0.03 at the 96th hour. Change of ratios of other two isomers not significant.</p><p><b>CONCLUSIONS</b>During the induced differentiation of HL-60 cells, WT1 gene expression decreases gradually. The phenotype of the majority of uninduced HL-60 cells is WT1 (+/+), in contrast to WT1 (-/-) phenotype after the induction of cell differentiation, indicating that WT1 gene may participate in the regulation of hematopoietic cell differentiation through modulation of the expression ratios of its four spliced variants.</p>

High frequency of loss of allelic integrity at Wilms' tumor suppressor gene-1 locus in advanced breast tumors associated with aggressiveness of the tumor.

Background: The product of Wilms' tumor suppressor gene (WT1), a nuclear transcription factor, regulates the expression of the insulin-like growth factor (IGF) and transforming growth factor (TGF) systems, both of which are implicated in breast tumorigenesis and are known to facilitate angiogenesis. In the present study, WT1 allelic integrity was examined by Loss of Heterozygosity (LOH) studies in infiltrating breast carcinoma (n=60), ductal carcinoma in situ (DCIS) (n=10) and benign breast disease (n=5) patients, to determine its possible association with tumor progression. Methods: LOH at the WT1 locus (11p13) as determined by PCR-RFLP for Hinf1 restriction site and was subsequently examined for its association with intratumoral expression of various growth factors i.e. TGF-β1, IGF-II, IGF-1R and angiogenesis (VEGF and Intratumoral micro-vessel density) in breast carcinoma. Results: Six of 22 (27.2%) genetically heterozygous of infiltrating breast carcinoma and 1 of 4 DCIS cases showed loss of one allele at WT1 locus. Histologically, the tumors with LOH at WT1 were Intraductal carcinoma (IDC) and were of grade II and III. There was no correlation in the appearance of LOH at WT1 locus with age, tumor stage, menopausal status, chemotherapy status and lymph node metastasis. The expression of factor IGF-II and its receptor, IGF-1R was significantly higher in carcinoma having LOH at WT1 locus. A positive correlation was observed between the TGF-β1, VEGF expression and IMD scores in infiltrating carcinoma. Conclusions: The current study indicates that the high frequency of loss of allelic integrity at Wilms' tumor suppressor gene-1 locus in high-graded breast tumors is associated with aggressiveness of the tumor.

Four new cases with WT1 gene mutations in Chinese patients with Wilms' tumor / 中华儿科杂志

<p><b>OBJECTIVE</b>Wilms' tumor (WT) is the most common malignant renal tumor in childhood. The WT1 gene located at 11p13 was identified in 1990 as a tumor suppressor gene important in the development in WT. The WT1 gene consists of 10 exons, with exons 1 to 6 encoding an N-terminal proline- and glutamine-rich transactivational domain, and exons 7 to 10 encoding a C-terminal zinc-finger domain involved in DNA binding. In China we know little about the frequency and genotype of WT1 mutations in Chinese WT patients. This study aimed to determine the frequency and genotype of WT1 mutations in children with nonsyndromic WT in China.</p><p><b>METHODS</b>We collected peripheral blood of WT patients treated in Beijing Children's Hospital. Genomic DNA of 54 WT patients was isolated from blood samples. All coding WT1 exons and their flanking intronic sequences were amplified by PCR method. The amplified PCR products from all individuals were then subjected to automatic DNA sequencing.</p><p><b>RESULTS</b>Four different constitutional WT1 mutations were identified in four children. Three mutations are predicted to produce truncated protein. One mutation is missense. Of the four mutations, three had not been reported before. Patient 1 had a 1006 A > T transition in exon 7, which caused (336)Lys to become a stop codon (K336X). DNA sequence analyses in patient 2 indicated the point mutations in exon 9 which was a 1168 C > T substitution and caused (390)Arg to become a stop codon (R390X). It indicated a point mutations in exon 6 in patient 3 which was a 814 G > T substitution and resulted in (272)Glu to become a stop codon (E272X). In patient 4 there was a homozygous mutation in exon 10. The mutation was a 1228 A > G substitution and resulted in (410)Ser to become a Gly codon (S410G).</p><p><b>CONCLUSION</b>Constitutional WT1 mutations occur at a low frequency (7.4%) in Chinese patients with Wilms' Tumor. It is similar to the results of overseas study. Four WT1 gene mutations were confirmed, three were nonsense, one was missense.</p>

Human cytotoxic T lymphocyte responses specific to the DNA vaccine of Wilms' tumor gene product / 南方医科大学学报

<p><b>OBJECTIVE</b>To construct the eukaryotic expression vector harboring Wilms' tumor gene (WT1y) and assess the cytotoxic T lymphocyte (CTL) response specific to the DNA vaccine of WT1y product in Balb/c mice and in vitro cultured cells.</p><p><b>METHODS</b>Synthesized WT1y gene (321 bp including HLA-A*2402 anchoring residue) was cloned into the eukaryotic vector to construct the plasmid pCDNA3.1(+)/WT1y. WT1-specific antibody was detected in mouse serum by ELISA, and T lymphocyte proliferation was detected by flow cytometry. The CTL response was detected by co-culture of the murine spleen cells with the tumor cells.</p><p><b>RESULTS</b>The recombinant plasmid was confirmed using DNA sequencing. WT1-specific antibodies were detected in the serum of immunized mice, which showed significantly greater T lymphocyte proliferation than the control group (Plt;0.01). The CTLs resulted in specific lysis of WT1-expressing murine tumor cells.</p><p><b>CONCLUSION</b>Using HBVC as the vector for WT1y gene, wenave obtained the virus-like particles that induce high cellular immune response, which shed light on a new approach for treatment of Wilms' tumor.</p>

Clinical and genetic findings of five patients with WT1-related disorders / Achados clínicos e genéticos de cinco pacientes com anomalias relacionadas ao gene WT1

AIM: To present phenotypic variability of WT1-related disorders. METHODS: Description of clinical and genetic features of five 46,XY patients with WT1 anomalies. RESULTS: Patient 1: newborn with genital ambiguity; he developed Wilms tumor (WT) and chronic renal disease and died at the age of 10 months; the heterozygous 1186G>A mutation compatible with Denys-Drash syndrome was detected in this child. Patients 2 and 3: adolescents with chronic renal disease, primary amenorrhea and hypergonadotrophic hypogonadism; patient 2 had a gonadoblastoma. The heterozygous IVS9+4, C>T mutation, compatible with Frasier syndrome was detected. Patient 4: 9-year-old boy with aniridia, genital ambiguity, dysmorphisms and mental deficiency; a heterozygous 11p deletion, compatible with WAGR syndrome was detected. Patient 5: 2 months old, same diagnosis of patient 4; he developed WT at the age of 8 months. CONCLUSIONS: Constitutional abnormalities of WT1 cause gonadal and renal anomalies and predisposition to neoplasia and must be investigated in patients with ambiguous genitalia, chronic renal disease and(or) Wilms tumors; primary amenorrhea with chronic renal disease; and aniridia, genital ambiguity and dysmorphisms.

OBJETIVO: Descrever a variabilidade fenotípica das anomalias relacionadas ao WT1. MÉTODOS: Descrição das características clínicas e genéticas de cinco pacientes 46,XY com anomalias no WT1. RESULTADOS: Paciente 1: Recém-nascido com ambigüidade genital desenvolveu tumor de Wilms (TW) e insuficiência renal crônica (IRC), com óbito aos 10 meses. Detectada a mutação 1186G>A em heterozigose, compatível com síndrome de Denys-Drash. Pacientes 2 e 3: Adolescentes com IRC, amenorréia primária e hipogonadismo hipergonadotrófico; a paciente 2 apresentava gonadoblastoma. Ambas apresentavam mutação IVS9+4, C>T em heterozigose, característica da síndrome de Frasier. Paciente 4: Idade 9 anos, aniridia, ambigüidade genital, dismorfismos e deficiência mental; deleção 11p, compatível com síndrome WAGR foi encontrada em heterozigose. Paciente 5: Dois meses, mesmo diagnóstico do paciente 4, desenvolveu TW aos 8 meses. CONCLUSÕES: Alterações constitucionais do WT1 determinam anomalias gonadais, renais e predisposição a neoplasias; devem ser pesquisadas em casos de ambigüidade genital associada a IRC e(ou) TW; de amenorréia primária com IRC; e aniridia, ambigüidade genital e dismorfismos.

Imunoexpressão de WT1, ß-cateinina, E-caderina e APC em tumor de Wilms esporádico / Expression of WT1, p53, Beta-catenina, E-cadherina and APC in sporadic Wilms tumor

O tumor de Wilms é uma neoplasia embrionária, constituída classicamente por três componentes histológicos: blastema, epitélio e mesênquima (estroma), os quais podem ser encontrados em diferentes proporções em cada tumor. ... Este estudo tem como principal objetivo determinar a freqüência da expressão das proteínas WT1, p53, Beta-catenina, E-caderina e APC nos diferentes componentes histológicos do tumor de Wilms, assim como correlacionar estes achados com o prognóstico. Para estudar a expressão dos marcadores, foi utilizado a imunoistoquímica, através da construção de Tissue Macroarray (TMA), o qual permite a análise de muitas amostras teciduais em uma única lâmina, otimizando custo e tempo. Nos nossos resultados, WT1 e p53 foram mais freqüentemente expressos no componente blastema (WT1: 97% e p53: 63% dos casos) e epitélio (WT1: 87% e p53: 58% dos casos), quando comparados com o estroma (WT1: 13% e p53: 22% dos casos) e com o tecido renal não tumoral (ausência de expressão em todos os casos estudados). ... A imunoexpressão nuclear ocorreu principalmente nos componentes blastematoso e estromal. Houve imunoexpressão de E-caderina na membrana em 47% dos casos, em citoplasma em 44% e em núcleo em 12%. O acúmulo em núcleo foi encontrado principalmente nos componentes blastema e mesênquima, 21 e 25% respectivamente. A imunoexpressão do APC ocorreu em núcleo em 95% dos tumores avaliados, em citoplasma em 9% e nenhum apresentou expressão em membrana. Nos componentes blastematoso e epitelial houve imunopositividade nuclear em 100% dos casos avaliados. No componente mesenquimal a positividade nuclear ocorreu em 89% dos casos. Os pacientes com estádios precoces tiveram mais freqüentemente positividade do WT1. Não houve associação dos outros fatores clínicos, epidemiológicos e evolução dos pacientes com expressão dos marcadores estudados.

Wilms tumor is a triphasic malignant neoplasm compromised of variable proportions of epithelial, blastemal and mesenchymal (stromal) elements. Different components have different clinical behavior. Patients with tumors predominant epithelial and/or stromal has more frequently low stage while predominant blastema has advanced disease. This study was undertaken to evaluate the expression of WT1, beta-catenins, E-cadherin and APC in Wilms tumor and correlate this expression with clinical characteristics, histologic cell type and prognosis. We studied the immunohistochemical expression of WT1, p53, beta-catenins, E-cadherin in different component of Wilms tumor using a tissue array. WT1 and p53 were more often expressed on blastema component (WT1: 97% and p53: 63% of cases) and on epithelial (WT1: 87% and p53: 58% of cases), when compare with the stromal component (WT1: 13% and p53: 22% of cases) and the normal renal tissue where it was not expressed. Immunopositivity of WT1 and p53 were significantly correlated. Immunopositivity of ß-catenina was seen in 85 % at the membrane (85%), 94% cytoplasm and 24% nuclear. Nuclear immunoexpression was detected specially on blastema and stromal component. Immunohistochemical expression of E-cadherina was detected in 47% at the membrane, 44% cytoplasm and 12% nuclear. Nuclear expression was seen more often at the blastema and stromal component (21%, 25% respectively). Immunopositivity of APC occurs at 95% on nucleus, cytoplasm (9%) and none at membrane. Among the blastema and the epithelial component 100% was positive on nucleus. On the stromal component 89% was positive at the nucleus. Immunoexpression of WT1 was correlated with local stage. The expression status of WT1, E-cadherin, ßcatenina, APC in this cohort was not of prognostic value.

Clinical Significance of Quantitation of WT1 Gene Expression for Minimal Residual Disease Monitoring of Acute Myelogenous Leukemia / 대한진단검사의학회지

BACKGROUND: Following induction chemotherapy for AML, a sensitive determination of minimal residual disease (MRD) in patients achieving complete remission (CR) should enable the detection of early relapse. This study was designed to verify if quantitative assessment of the Wilms' tumor (WT1) gene by real time polymerase chain reaction (RQ-PCR) can be used as a marker for MRD detection during the monitoring of AML. METHODS: WT1 gene expression was quantified by RQ-PCR in 31 patients with AML at diagnosis (27 patients) and during follow-up (29 patients) relative to ABL control gene. In four patients, the WT1 gene expression was analyzed in comparison to a second PCR marker, PML-RARA fusion transcript. Prognostic significance of WT1 gene expression was analyzed at diagnosis and at the primary CR evaluation. Longitudinal WT1 gene analysis was performed in 17 AML patients. RESULTS: At diagnosis, WT1 gene expression exceeded the control level in all of the patients. Higher levels of WT1 gene expression were not associated with shorter event free survival or overall survival at diagnosis. Higher levels of WT1 gene expression were associated with shorter event free survival after induction chemotherapy. Relapse was observed in eight of 17 patients analysed longitudinally, and an increase of WT1 gene expression preceded morphologic relapse in four patients with the fusion transcript negative. Concomitant monitoring of PML-RARA fusion transcript reveals the lack of a significant correlation withWT1 gene expression. CONCLUSIONS: Quantitation of WT1 gene expression could be used for MRD monitoring of AML and for the early detection of relapse, especially in patients lacking specific molecular markers.

Mutations in SRY and WT1 genes required for gonadal development are not responsible for XY partial gonadal dysgenesis

The WT1 transcription factor regulates SRY expression during the initial steps of the sex determination process in humans, activating a gene cascade leading to testis differentiation. In addition to causing Wilms' tumor, mutations in WT1 are often responsible for urogenital defects in men, while SRY mutations are mainly related to 46,XY pure gonadal dysgenesis. In order to evaluate their role in abnormal testicular organogenesis, we screened for SRY and WT1 gene mutations in 10 children with XY partial gonadal dysgenesis, 2 of whom with a history of Wilms' tumor. The open reading frame and 360 bp of the 5' flanking sequence of the SRY gene, and the ten exons and intron boundaries of the WT1 gene were amplified by PCR of genomic DNA. Single-strand conformation polymorphism was initially used for WT1 mutation screening. Since shifts in fragment migration were only observed for intron/exon 4, the ten WT1 exons from all patients were sequenced manually. No mutations were detected in the SRY 5' untranslated region or within SRY open-reading frame sequences. WT1 sequencing revealed one missense mutation (D396N) in the ninth exon of a patient who also had Wilms' tumor. In addition, two silent point mutations were found in the first exon including one described here for the first time. Some non-coding sequence variations were detected, representing one new (IVS4+85A>G) and two already described (-7ATG T>G, IVS9-49 T>C) single nucleotide polymorphisms. Therefore, mutations in two major genes required for gonadal development, SRY and WT1, are not responsible for XY partial gonadal dysgenesis.

Expression of WT1 gene in patients with myelodysplastic syndrome and acute leukemia / 中国实验血液学杂志

To study the expression and significance of WT1 gene in patients with myelodysplastic syndrome (MDS) and acute leukemia (AL), RT-PCR was applied to monitor WT1 gene expression in 22 patients with MDS and in 69 patients with AL. The results showed that the positive rate of WT1 mRNA in MDS-RA and MDS-RAS was lower than that in MDS-RAEB and MDS-RAEB-t (10% versus 91.7%, P < 0.01). WT1 mRNA could be expressed in all subtype of AL, It was detected in 69% of newly diagnosed and relapsed patients, and in 12.5% patients CR. There was no difference at the relative expression level between newly diagnosed AL patients and relapsed patients, while the relative level of WT1 in MDS-RAEB and MDS-RAEB-t was lower than that in newly diagnosed AL. The CR rate in AML patients with positive expression was lower than that in patients with negative expression (41% versus 78%, P </= 0.05). AML patients with relative level of WT1 mRNA >/= 1 had lower CR rate (18%) than those with relative level < 1 (55%). It is concluded that the expression of WT1 gene in patients with MDS-RAEB and RAEB-t was higher than that in patients with RA and RAS. The detection of WT1 gene may be useful for assessing disease progress of patients with MDS. The expression of WT1 gene and its expression level have associated with the prognosis of newly diagnosed patients with AL, that WT1 gene may be an independent prognostic factor in AML.