Transcritos de fusión del gen PML/RARα en pacientes con leucemia promielocítica / Fusion transcripts of the PML/RARα gene in patients with promyelocytic leukemia

Introducción: La leucemia promielocítica se considera una enfermedad bien definida por sus peculiares características clínicas, morfológicas, citogenéticas y moleculares. El descubrimiento de los mecanismos oncogenéticos implicados en la génesis de la enfermedad hacen de esta variante de leucemia uno de los modelos más relevantes de investigación traslacional. Objetivo: Caracterizar los transcritos de fusión del gen PML/RARα en pacientes con leucemia promielocítica. Métodos: Se realizó una investigación observacional, ambispectiva, descriptiva, longitudinal, en pacientes con diagnóstico de leucemia promielocítica, en el Instituto de Hematología e Inmunología, entre enero de 2001 y diciembre de 2020. El universo estuvo constituido por 105 pacientes que cumplieron los criterios de inclusión y exclusión. Resultados: No existe relación entre los transcritos y la edad, sexo, color de piel y las características clínicas. La presencia del transcrito de fusión bcr3 se asoció a mayores cifras de hemoglobina y menor valor de plaquetas. La incidencia de recaída no se relacionó con los transcritos de fusión y no se comprobó que existiera influencia de éstos, sobre la supervivencia global en pacientes con leucemia promielocítica. Conclusiones: Las características de los transcritos de fusión del gen PML/RARα son similares a los reportes internacionales, sobre todo en poblaciones de origen latino(AU)

Introduction: Promyelocytic leukemia is considered a well-defined entity due to its peculiar clinical, morphological, cytogenetic and molecular characteristics. The discovery of the oncogenetic mechanisms involved in the genesis of the disease makes this variant of leukemia one of the most relevant models for translational research. Objective: To characterize the fusion transcripts of the PML/RARα gene in patients with promyelocytic leukemia. Methods: An observational, ambispective, descriptive, longitudinal investigation was carried out in patients diagnosed with promyelocytic leukemia at the Institute of Hematology and Immunology, between January 2001 and December 2020. The sample consisted of 105 patients who met the criteria for inclusion and exclusion. Results: There is no relationship between the transcripts and age, sex, skin color and clinical characteristics. The presence of the bcr3 fusion transcript was associated with higher hemoglobin levels and lower platelet counts. The incidence of relapse was not related to fusion transcripts and their influence on overall survival in patients with promyelocytic leukemia was not proven. Conclusions: The fusion transcripts´scharacteristicsof the PML/RARα gene are similar to international reports, especially from populations of Latin origin(AU)

Follow-up and outcome of the twelve-year experience in adult patients with acute promyelocytic leukemia

ABSTRACT Acute promyelocytic leukemia is a subtype of acute myeloid leukemia, characterized by the presence of neoplastic promyelocytes, due to the reciprocal balanced translocation between chromosomes 15 and 17. Currently, with the use of agents that act directly on this molecular change, such as all-trans retinoic acid and arsenic trioxide, APL has shifted from a highly mortal to a curable disease. However, some cases are still at high risk of death, especially early death, and acquiring a better understanding of the clinical and biological factors involving APL is needed to correctly identify and treat such cases. The early suspected diagnosis and prompt initiation of the target therapy are important for better response rates. The follow-up and outcomes, using real-life data from 44 consecutive APL patients, were studied between 2001 and 2013. The overall survival rate was 82.7% and early death was 16%. Almost all patient deaths were due to severe bleeding, which was confirmed by multivariate analysis, as the most important prognostic factor leading to death. A better understanding the pathogenesis of the hemorrhagic complications in APL is needed, as well as the risk factors associated with early death in APL patients, as this has become synonymous with overall mortality.

Detección del gen fusión PML-RARα en pacientes colombianos con leucemia mieloide aguda / Detection of the PML-RARα fusion gene in Colombian patients with acute myeloid leukemia

Resumen Introducción: la leucemia promielocítica aguda es un subtipo de leucemia mieloide aguda caracterizada por la presencia de una translocación entre los cromosomas 15 y 17 que provoca la formación de un gen fusión denominado PML/RARα. Determinar la presencia de este gen fusión es crítico para estos pacientes ya que su presencia hace el diagnóstico de la enfermedad, aún sin tener resultados de patología. Con esta investigación se busca ajustar e implementar una prueba altamente sensible y específica para la detección del reordenamiento PML/RARα. Métodos: a partir de sangre periférica se extrajo RNA de pacientes diagnosticados con leucemia mieloide aguda en dos instituciones de Antioquia (Colombia). Se realizó RT-PCR anidada para la detección de PML/RARα, ajustando un protocolo previamente publicado. Resultados: se ajustó y estandarizó un método para detectar mediante RT-PCR el gen fusión PML/RARα. Mediante esta técnica se logró identificar la traslocación en cuatro pacientes (22 %) de la cohorte estudiada. Conclusiones: los resultados están de acuerdo con estudios previos. La detección de esta y otras alteraciones citogenéticas mediante pruebas moleculares permitirá tener información valiosa a nivel de diagnóstico y pronóstico de los pacientes con leucemia mieloide aguda en Antioquia.

Abstract Introduction: acute promyelocytic leukemia is a subtype of acute myeloid leukemia characterized by the presence of a translocation between chromosomes 15 and 17, which causes the formation of a fusion gene called PML/RARα. Determining the presence of this fusion gene is critical for these patients, since their presence makes the diagnosis of the disease, even with no pathology results This research seeks to adjust and implement a highly sensitive and specific test for the diagnosis of this cytogenetic abnormality. Methods: peripheral blood samples from patients diagnosed with acute myelocytic leukemia were collected in two institutions of Antioquia (Colombia), from which RNA was extracted and nested RT-PCR was performed, adjusting a previously published protocol. Results: we adjusted and standardized a method to detect the PML/RARα fusion gene by RT-PCR. Using this technique, translocation was identified in four patients (22%) of the studied cohort. Conclusions: our results agree with previous studies. The detection of this and other cytogenetic alterations by means of molecular tests will allow to have valuable information at the level of diagnosis and prognosis of patients with AML in Antioquia.

La biología molecular en la precisión diagnósticade las leucemias / Molecular biology in the diagnostic accuracy of leukemia

Los estudios de citogenética y biología molecular permiten correlacionar la presencia de determinadas anomalías cromosómicas y moleculares con tipos específicos de leucemias y linfomas. Este conocimiento ha hecho posible el perfeccionamiento progresivo del sistema de clasificación de las enfermedades oncohematológicas. Actualmente la presencia de ciertas anomalías citogenéticas o moleculares son suficientes para identificar algunas de estas entidades y en ocasiones, el diagnóstico cambia después de un análisis integrado de la citomorfología con la citogenética y la biología molecular. Este reporte pretende resaltar la importancia del estudio molecular cuando la citomorfología es compleja y propicia diagnósticos erróneos. Mediante la reacción en cadena de la polimerasa, previa reverso-transcripción del ARN aislado de sangre medular, se estudiaron cuatro biomarcadores: los genes de fusión AML1-ETO, BCR-ABL, CBFβ-MYH11 y PML-RARα. Fueron estudiados 14 pacientes con diagnósticos inicial citomorfológico de: leucemia promielocítica (n= 6), leucemia aguda de linaje indefinido (n= 3) y dudoso entre leucemia mieloide crónica en crisis blástica mieloide y leucemia mieloide aguda (LMA) (n= 5). Al culminar la caracterización molecular todos fueron diagnosticados como LMA. Los resultados ilustran la importancia del estudio molecular en la clasificación de las leucemias, lo cual redunda en que el paciente reciba el tratamiento más adecuado y alcance una mejor respuesta.

Cytogenetic and molecular studies have correlated the presence of certain chromosomal and molecular anomalies with leukemia and lymphomas specific types. These evidences have allowed the progressive improvement of the system of classification of the oncohematological entities. Currently, the presence of certain cytogenetic or molecular anomalies is sufficient to identify specific entities and, in some occasions, the diagnostic changes after an integral analysis of cytomorfologic, cytogenetic and molecular studies. T This report aims to highlight the importance of molecular study when cytomorphology is complex and leads to erroneous diagnoses. Through polymerase chain reaction, prior reverse transcription of the RNA isolated from medullary blood, four biomarkers were studied: the fusion genes AML1-ETO, BCR-ABL, CBFβ-MYH11 and PML-RARα. Fourteen patients with initial diagnosis of: promyelocytic leukemia (n= 6), acute leukemia without lineage definition (n= 3) and chronic myeloid leukemia in myeloid blastic crisis or acute myeloid leukemia (AML) (n= 5) were studied. At the end of the molecular characterization all were diagnosed as AML. These results enlightened the role of the molecular studies in the classification of leukemia, which permit the patient receives the more appropriate treatment and achieve a better response.

Correlation between RAR-βgene methylation and p53 gene mutation in bronchoalveolar lavage fluid in non-small-cell lung cancer / 重庆医学

Objective To investigate the clinical significance and correlation between RAR-βgene methylation and p53 gene mutation in bronchoalveolar lavage fluid(BALF)in non-small-cell lung cancer.Methods BALF samples from 85 lung cancer pa-tients(lung cancer group)and 70 cases(benign lung diseases group)with benign lung diseases were collected.RAR-βgene methyla-tion in BALF samples was detected by methylation-specific PCR (MSP),and p53 gene mutation was detected by PCR and DNA se-quencing method.Results The rate of RAR-βmethylation and p53 mutation in BALF in lung cancer were 49.4% and 36.5%,re-spectively.Both were higher than in benign lung diseases group(P <0.01).RAR-βmethylation rate(32.5%)of patients with TNM stages(Ⅰ+Ⅱ)(32.5%)was higher than the p53 mutation rate(12.5%)over the same stages (P <0.05).RAR-βmethylation rate and p53 mutation rate of patients with stages(Ⅲ+Ⅳ)were higher than those with stages(Ⅰ+Ⅱ)(P <0.01).p53 mutation rate in lung cancer patients with RAR-βmethylation was higher than those with unmethylated(P <0.01).RAR-βmethylation rate of lung cancer patients with p53 mutation was higher than those without p53 mutation(P <0.01).Conclusion Detection of RAR-βmethyl-ation and p53 mutation in BALF contribute to the diagnosis of lung cancer.

Leukemia Cutis with PML-RAR-a Translocation.

Leukemia cutis is the infiltration by neoplastic leukocytes or their precursors into the epidermis, dermis or the subcutis resulting in clinically identifiable cutaneous lesions. The diagnosis is of great value in cases where leukemia cutis aids in the diagnosis of leukemia. This has a poor prognosis and strongly correlates with additional signs of extramedullary involvement. A 27-year-old male patient, known case of AML-M3, presented with cutaneous nodules on both lower limb, which on biopsy proved to be leukemic deposits. Immunohistochemistry confirmed the diagnosis and the patient was discharged with advice to continue chemotherapy.

Correlation between cervical lesion development and histone acetylation modification that regulates RAR-β2 expression / 中国肿瘤临床

Objective:To investigate the relationship between cervical lesion development and histone acetylation that regulates RAR-β2 expression. Methods:Immunohistochemistry and Western blot analysis were performed to detect AcH3, RAR-β2, and involu-crin expression in normal cervical tissues as well as in tissues with cervical intraepithelial neoplasia (CIN)Ⅱ-Ⅲand squamous cell cer-vical carcinoma. The relationship among histone acetylation level, RAR-β2 expression, and cervical lesion severity were analyzed. Re-sults:AcH3, RAR-β2, and involucrin expression were reduced or absent with the progression of cervical lesions;significant differences were noted between the groups (P<0.05). Histone acetylation level and RAR-β2 expression were positively correlated (r=0.797, P<0.05). AcH3 and RAR-β2 expression, which were both associated with the cervical lesions, were negatively correlated [r=-0.547(AcH3), r=-0.585(RAR-β2), P<0.05]. Conclusion:Histone acetylation modification is associated with the regulation of RAR-β2 expression. This pro-cess is also likely to participate in the carcinogenesis of cervical carcinoma.

Deficiencia de retinoides y consumo de estrógenos como cofactores de riesgo de cáncer cervical / Retinoid deficiency and estrogens as risk cofactors in cervical cancer

La infección persistente por el virus del papiloma humano de alto riesgo (HR-HPV) está relacionada con la aparición de cáncer cervical (CC), una de las principales causas de mortalidad por cáncer en todo el mundo. La infección se produce en la zona de transformación, la región más sensible del cérvix a estrógenos y retinoides. El CC afecta a un bajo porcentaje de mujeres infectadas por HR-HPV y tarda en desarrollarse hasta décadas después de la infección, lo que sugiere que el HR-HPV es necesario pero no suficiente para causar CC. Otros factores son necesarios para la progresión desde la infección por HR-HPV hasta el cáncer, como por ejemplo: uso de anticonceptivos orales por largos períodos, fumar, partos múltiples, falta de micronutrientes, particularmente una dieta baja en retinoides, los cuales alteran la diferenciación epitelial, el crecimiento celular y la apoptosis de las células malignas. La detección precoz del HR-HPV y el manejo de lesiones precancerosas, aunado a un conocimiento detallado de factores de riesgo adicionales, puede ser una estrategia para prevenir esta enfermedad. La presente revisión se enfoca en explicar el efecto de los estrógenos, la deficiencia de retinoides y el HR-HPV en la aparición del CC. Dichos cofactores pueden actuar en conjunto para inducir transformación neoplásica en el epitelio escamoso del cérvix, promoviendo un segundo evento genético o epigenético que lleve a la aparición del CC.

ersistent infection with high-risk human papillomaviruses (HR-HPVs) is involved in cervical cancer (CC),a major cause of cancer mortality worldwide. Infection occurs primarily at the transformation zone (TZ),the most estrogen- and retinoid-sensitive region of the cervix. Development of CC affects a small per-centage of HR-HPV-infected women and often takes decades after infection, suggesting that HR-HPVis a necessary but not sufficient cause of CC. Thus, other cofactors are necessary for progression fromcervical HR-HPV infection to cancer such as long-term use of hormonal contraceptives, multiparity, smo-king, as well as micronutrient depletion and in particular retinoid deficiency, which alters epithelial diffe-rentiation, cellular growth and apoptosis of malignant cells. Therefore, early detection of HR-HPV andmanagement of precancerous lesions together with a profound understanding of additional risk factorscould be a strategy to avoid this disease. In this review we focus on the synergic effect of estrogens,retinoid deficiency and HR-HPVs in the development of CC. These risk factors may act in concert to indu-ce neoplastic transformation in the squamous epithelium of the cervix, setting the stage for secondarygenetic or epigenetic events leading to cervical cancer.

Retinoides (RARβ y CRBP1) en carcinoma de pulmón a células no pequeñas / Retinoid expression (RARβ and CRBP1) in non-small-cell lung carcinoma

Aunque los pacientes con cáncer de pulmón a células no pequeñas en estadios tempranos (NSCLC) tienen buen pronóstico, el 20-30% recae, siendo relevante la identificación de biomarcadores pronósticos. Los retinoides regulan crecimiento y diferenciación, y pueden antagonizar la progresión tumoral. Su efecto depende del transporte citosólico mediado por moléculas como CRBP1, y de la unión a receptores específicos (RARβ). Alteraciones en esta vía se asociaron con cáncer. Nuestro objetivo fue estudiar la expresión, mediante inmunohistoquímica, de RARβ y CRBP1 en el tejido tumoral de 49 pacientes NSCLC Estadio I/II, obtenido durante la cirugía. La supervivencia se analizó mediante los test Log Rank y multivariado de Cox. El 44.9% de los tumores fueron positivos para RARβ con expresión a nivel citoplasmático, mientras que el 34.7% lo expresó a nivel nuclear. La tinción para CRBP1 se observó en el 61.2% de los tumores. No se encontró asociación entre la expresión de ambas moléculas y las características clinicopatológicas (sexo, tamaño tumoral, nódulos línfáticos comprometidos, histopatología y p53). Tampoco se encontró asociación con el hábito de fu-mar. La presencia de células tumorales en el lavado pleural se asoció significativamente con la expresión de CRBP1. Por otro lado, se demostró asociación entre la expresión elevada de RARβ citoplasmático y menor supervivencia global (LR 4.17, p=0.0412). El análisis multivariado no mostró asociación con otras variables de pronóstico en NSCLC. En conclusión, en este grupo de pacientes NSCLC Estadio I/II, RARβ pareciera predecir la supervivencia global en forma independiente.

Although early-stage non-small-cell lung carcinoma (NSCLC) patients have a relative by favorable prognosis, the risk of a bad outcome remains substantial. Identification of reliable prognostic markers for disease recurrence and death has meaningful clinical application. Retinoids are involved in cell growth and differentiation and may antagonize cancer progression. Their effects are mediated through nuclear receptors called Retinoic Acid Receptor (RAR) and regulated by molecules such as Cellular Retinol-Binding Protein 1 (CRBP1) that function in retinol storage. The aim of this work was to analyze by immunohistochemistry the expression patterns of RARβ and CRBP1, involved in retinoid-mediated signaling, in the tumoral tissue of a cohort of stage I/II NSCLC patients (n=49) who underwent a successful surgical resection. Prognostic evaluation was performed with the multivariate Cox proportional hazard model: 44.9% of tumors were positive for RARβ staining at cytoplasmic level, while 34.7% showed nuclear staining. CRBP1 staining was observed in 61.2% of the lung tumors. No relationship was found between the number of cells expressing the studied molecules and clinical pathological features, including sex, T and N (stage), histopathology and p53 expression. Univariate analysis showed a significant association between positive cytoplasmatic expression of RARβ with shorter overall survival (Log-rank test 4.17, p=0.0412). Multivariate studies indicated that RARβ expression was not influenced by other clinical pathological parameters. In conclusion, in this cohort of stage I and II NSCLC, only the expression of RARβ at cytoplasmatic level is a significant independent unfavorable prognostic factor.

The Immunohistochemical Expression of Nuclear Retinoid Receptor and CREB in Gastric Adenocarcinoma

PURPOSE: Transcriptional factors of CREB (cAMP response element binding protein) are involved in regulating the gene expression in response to a variety of signaling pathways. The proteins produced by the CREB genes play key roles in many physiological processes, including memory and long-term potentiation. The retinoic acid receptor (RAR) axis mediates epithelial cell differentiation and proliferation in many tissues. This study examined the expressions of RAR and CREB and their relationship with the clinicopathologic factors and their significance. MATERIALS AND METHODS: The levels of the RAR and CREB expressions were measured in 150 gastric adenocarcinomas by performing immunohistochemical staining. RESULTS: 1. An RAR protein expression was found in 63.3% of the adenocarcinomas (95/150) and a CREB expression was found in 60.7% (91/150) of the adenocarcinomas. 2. An RAR protein expression was found in 72.2% (78/108) of the intestinal type adenocarcinomas and in 40.5% (17/42) of the diffuse type adenocarcinomas (P0.05). Based on the depth of invasion, a CREB expression was found in 50% (12/24) of the T1 adenocarcinomas, in 52.4% (11/21) of the T2 adenocarcinomas, in 64.6% (62/96) of the T3 adenocarcinomas, in 66.6% (6/9) of the T4 adenocarcinomas, in 71.1% (59/83) of the adenocarcinomas with lymph node metastasis and in 47.8% (32/67) of the adenocarcinomas without lymph node metastasis (P<0.01). 4. The RAR protein and CREB expressions coincided in 71.4% of the gastric adenocarcinomas, and a significant correlation between them was found (P<0.05). CONCLUSION: We found a significant relationship between the expression of RAR and CREB and the histology and lymph node metastasis of gastric cancer. Further studies are needed to confirm their biologic meaning in gastric carcinogenesis.

Do PML/RAR-alpha isoforms have clinical significance in patients with acute promyeloctyic leukemia? / 대한내과학회지

Acute promyelocytic leukemia (APL) is characterized by a specific t (15;17) translocation which produce a PML/RAR-alpha fusion messenger RNA and by effectiveness of all-trans retinoic acid (ATRA) differentiation therapy. Breakpoints within PML intron 3 (bcr 3) produce a short PML/RAR-alpha isoform (S-isoform), whereas breakpoints within PML intron 6 (bcr 1) result in a longer form (L-isoform). Additionally, breakpoints within PML exon 6 (bcr 2) make a variable length transcript (V-isoform) in a small number of patients. The influence of breakpoint site on patient outcome remains controversial. Previous reports showed that patients with S-isoform have an increased incidence of clinical relapse and shorter survival compared to those with L-isoform. Others reported no difference in DFS between these patients groups. In this issue, Lee et al. reported that there were 58 L-isoform (62.1%), 32 S-isoform (34.0%), 4 V-isoform (4.3%) and, no significant prognostic factor for EFS from induction therapy using anthracycline plus ATRA among 94 patients with APL. They concluded pretreatment clinical characteristics and treatment outcomes were not significantly different according to PML/RAR-alpha isoform types in this induction group. Recently, it was reported that FLT3/ITD mutation was frequently associated with S-isoform and with the M3v form of leukemia and CNS relapse in APL was mostly related to S-isoform. With previous studies including this article, outcomes of different types of PML/RAR-alpha isoforms are not conclusive. Future researches need to be focused not only on clinical outcomes of different types of PML/RAR-alpha isoforms, but also clinical relevance of PML/RARA-alpha mRNA isoforms with other prognostic factors and particular clinical characteristics.

Clinical significance of PML/RAR alpha isoforms in acute promyelocytic leukemia / 대한내과학회지

BACKGROUND/AIMS: There are three types of PML-RAR alpha mRNA fusion transcripts associated with acute promyelocytic leukemia (APL): the short (S)-form, the long (L)-form and the variable (V)-form. No study on the Korean population has addressed the clinical significance of the specific types of PML-RAR alpha mRNA fusion transcripts for APL patients who receive the combination therapy of all-trans-retinoic-acid and idarubicin (AIDA regimen). METHODS: We performed a retrospective analysis on 94 patients with APL to evaluate differences in the therapeutic outcomes, such as the response rate, an event-free survival (EFS), and overall survival (OS), after remission following the induction of chemotherapy. We also analyzed whether differences in the pretreatment clinical characteristics depend on the PML-RAR alpha isoform. RESULTS: The median age of the patients was 41 years (range 15-85). Among the 94 patients, there were 58 L-form cases (62.1%), 32 S-form cases (34.0%), and 4 V-form cases (4.3%). The CR rate following remission induction treatment was 84.9%. The CR rate was higher in patients with an initial WBC <10.0x109/L, as compared to patients with an initial WBC higher than 10.0X109/L (93.5% vs. 65.4%, p=0.001). The AIDA induction regimen was associated with a better EFS than non-AIDA induction regimens (81.9% vs. 49.6%, p=0.006). The induction group was also a significant prognostic factor for EFS in the multivariate analysis (p=0.020). There were no differences in OS and EFS in patients with either isoform L or isoform S in the AIDA induction group. CONCLUSIONS: This retrospective study demonstrated that pretreatment clinical characteristics and treatment outcomes were not significantly different among patients with varying PML-RAR alpha isoform types in the AIDA induction group.

Study PML/RAR alpha fusion gene on 21 patients diagnosed with acute promyelocytic leukemia (AML \u2013 M3)

Background: In Vietnam, there are a number of studies on the application of ATRA in treating acute promyelocytic leukemia (AML \u2013 M3) but they have still faced with certain difficulties. Objectives: (1). Study PML/RAR alpha fusion gene on the patients diagnosed with AML \u2013 M3. (2). Study the index of hematology of the PML/RAR alpha positive group. Subject and Method: 21 patients with acute promyelocytic leukemia (M3) were studied. All patients were examined with morphology, coagulation and cytogenetic tests and RNA were extracted from leukemic cells and PCR for PML/RAR alpha fusion transcript. Result and conclusion: PML/RAR alpha positive in 67% including 4 patients which were not discovered t(15; 17) by cytogenetic technique. Rates of three subtype (bcr1, bcr2 and bcr3) of PML/RAR alpha were 7 patients (50%), 3 patients (21,5%) and 4 patients (28,5%), respectively. WBC and bone marrow cells of PML/RAR alpha positive group were 5.08+/-3.87 and 155.82+/-106.21. D \u2013 Dimer level was 1954.89+/-1575.28; 93% of patients in the PML/RAR alpha positive group had DIC.

Study on the relationship between the RAR-β gene expressive defection and its methylation / 西安交通大学学报·英文版

Objective: To observe the expression of RAR-β gene in SiHa, HeLa, C33A and CasKi cell lines of cervical carcinoma and to investigate the role of methylated RAR-β in its expressive defection. Methods: Reverse transcription polymerase chain reaction (RT-PCR) was used to analyze the mRNA expression of RAR-β gene. Immunohistochemistry and Western Blot were used to analyze the protein expression of RAR-β gene in four cervical cancer cell lines as well as the influence of 5-Aza-cdR on gene expressive defection. Methylation specific PCR (MSP) was used to detect whether there was the methylation in RAR-β gene in four cell lines. The change of RAR-β gene methylation state was also observed by MSP. The cell proliferation rate influenced by the 5-Aza-cdR was observed by MTT assay. Results: The expression of RAR-β mRNA and protein in SiHa, HeLa and CasKi cell lines of cervical cancer was silent or decreased, whereas its expression was detected in C33A cell line. By using MSP method, it was found that there was RAR-β gene methylation in those three cell lines, whereas there was no RAR-β gene methylation in C33A cell line. After treated with the 5-Aza-cdR, methylated RAR-β gene was partly demethylated, and RAR-β mRNA and protein were re-expressed in the previous three cell lines in which RAR-β gene expression was silent or decreased. The 5-Aza-cdR treatment could supress cell proliferation as well. Conclusion: The RAR-β gene expressive defection plays an important role in the carcinogenesis of cervical cancer. The abnormal RAR-β gene methylation in the the promoter region has an important role in gene expressive defection. The cell proliferation can be supressed by demethylated treatment.

Study on the relationship between the RAR-β gene expressive defection and its methylation / 药物分析学报

Objective To observe the expression of RAR-β gene in SiHa, HeLa,C33A and CasKi cell lines of cervical carcinoma and to investigate the role of methylated RAR-β in its expressive defection. Methods Reverse transcription polymerase chain reaction (RT-PCR) was used to analyze the mRNA expression of RAR-β gene. Immunohistochemistry and Western Blot were used to analyze the protein expression of RAR-β gene in four cervical cancer cell lines as well as the influence of 5-Aza-cdR on gene expressive defection. Methylation specific PCR (MSP) was used to detect whether there was the methylation in RAR-β gene in four cell lines. The change of RAR-β gene methylation state was also observed by MSP. The cell proliferation rate influenced by the 5-Aza-cdR was observed by MTT assay. Results The expression of RAR-β mRNA and protein in SiHa, HeLa and CasKi cell lines of cervical cancer was silent or decreased, whereas its expression was detected in C33A cell line. By using MSP method, it was found that there was RAR-β gene methylation in those three cell lines, whereas there was no RAR-β gene methylation in C33A cell line. After treated with the 5-Aza-cdR, methylated RAR-β gene was partly demethylated, and RAR-β mRNA and protein were re-expressed in the previous three cell lines in which RAR-β gene expression was silent or decreased. The 5-Aza-cdR treatment could supress cell proliferation as well. Conclusion The RAR-β gene expressive defection plays an important role in the carcinogenesis of cervical cancer. The abnormal RAR-β gene methylation in the the promotor region has an important role in gene expressive defection. The cell proliferation can be supressed by demethylated treatment.

Relation between the expression of RAR?,RAR? and endometrial carcinoma / 重庆医科大学学报

Objective:To evaluate the role of the retinoic acid receptor ?(RAR?),retinoic acid receptor ?(RAR?) in the devel-opment and prognosis of the endometrial carcinoma by detecting the expression of these receptors. Methods:48 cases of paraffin-embedded tissues were selected, including 42 cases of adenocarcinoma and 6 cases of adeno-squamous carcinoma of endometrium. Immunohistochemistry was used to detect the level of RAR? and RAR?. The relation between the expression of these receptors and the clinical pathological parameters was evaluated. Results:The positive expression ratios of the RAR? and RAR? were 47.92%(23/48)and 25.00%(12/48),respectively. The level of RAR? was significantly higher than that of RAR?(P0.05). The level of the RAR? decreased with the increasing of the malignancy extent of the endome-trial adenocarcinoma(P0.05)respectively. Conclusion:There is close relation between the ex-pression of the RAR? and the development of the endometrial carcinoma. The defect of the RAR? and the high expression of the RAR? indicates a bad prognosis of the endometrial carcinoma. Detection of these two is benefited receptors together for us to estimate the prognosis of the carcinoma.

Immunohistochemical Expression of Nuclear Retinoid Receptor and CREB(cAMP Response Element Binding Protein) in Lung Cancers / 결핵및호흡기질환

BACKGROUND: Transcriptional factors of the CREB(cAMP Response Element Binding Protein) are involved in the regulation of gene expression in response to a variety of signaling pathways. Proteins produced by the CREB genes play key roles in many physiological processes, including memory and long term potentiation. The retinoic acid receptor (RAR) axis mediates epithelial cell differentiation and proliferation in many tissues including the lung. MATERIAL AND METHOD: The RAR and CREB expression levels were examined in 60 adenocarcinomas and 60 squamous cell carcinomas of the lung using immunohistochemical staining. RESULTS: 1)RAR protein expression was found in 58.3%(35/60) of adenocarcinomas and 36.7%(22/60) of squamous cell carcinomas(P0.05). 4)CREB expression was found in 61.7%(37/60) of adenocarcinomas and 40%(24/60) of squamous cell carcinomas(P0.05). 7)RAR and CREB expression was found in 68.5% of lung cancers, and there was a significant correlation between them(P<0.05). CONCLUSION: RAR and CREB expression can be used to indirectly determine the malignant potentiality of a cell.

Evaluation of kinetics of PML-RAR_? fusion gene in children with acute promyelocytic leukemia / 医学研究生学报

Objective: To investigate the clinical treatment results of combined Tretinoin-chemotherapy protocol and kinetics of PML-RAR ? fusion gene in childhood acute promyelocytic leukemia(APL). Methods: Ten children with APL were involved in this study. Induction therapy was Tretinoin alone(6 cases),Tretinoin plus chemotherapy(3 cases) and arsenic trixode(1 case). Postremission therapy consisted of three consolidation courses with DA,MA or HA and a monthly maintenance therapy over 4-5 years. Monitoring of minimal residual disease was performed regularly by RT-PCR assay for PML-RAR ? at differential clinical stages. Results: Clinical complete remission(CR) was obtained in 9 cases (90%).After a median follow-up of 42 months(14-156 months), the estimated 5-year event-free survival was (56? 16.5)%.Four cases relapsed at 14-42 months after achieving CR and 5 cases remained continuing CR. PML-RAR ? fusion gene was positive in all cases at CR and turned negative gradually during consolidation and maintenance treatment. The duration of conversion to RT-PCR negative status varied from 6 to 42 months.Four patients who were persistent positive(2 cases) or converted to positive(2 cases) for PML-RAR ? relapsed. Conclusion: Continuous negative RT-PCR results are associated with long-term disease-free survival and may be considered as potentially curative. RT-PCR assay for detection of PML-RAR ?should be performed regularly during post-remission period. The hematological relapse could potentially be averted through treatment modification according to molecular monitoring results of PML-RAR ?.

A Case of Acute Promyelocytic Leukemia with Phenotypic Change at Relapse / 대한진단검사의학회지

Early diagnosis of acute promyelocytic leukemia (APL) depends primarily on morphological recognition before the presence of t(15;17) or PML-RAR gene rearrangement is confirmed. But the diagnosis is difficult to be made, if typical APL morphologic features are not found. Here, we describe a 32- year old man who had been diagnosed as APL. He relapsed with AML M1 like phenotype, lacking the typical features of APL. At relapse, t(15;17) and PML-RAR alpha gene rearrangement were detected. After 14 days of chemotherapy and all-trans retinoic acid, the phenotype changed from the AML M1 like features to the typical hypergranular APL. Awareness of atypical morphologic subtypes found in APL is important. And identification of t(15;17) or PML/RAR alpha rearrangement will be helpful in diagnosis of atypical APL.

Leucemia promielocitica aguda: utilidad de la citometría de flujo en el diagnóstico / Acute promyelocytic leukemia: usefulness of flow cytometry in diagnosis

La Leucemia Promielocítica Aguda (LPA) requiere una rápida identificación ya que deben tomarse decisiones terapéuticas tempranas. Habitualmente el estudio morfológico es suficiente, pero la existencia de falsos positivos y negativos hace necesario buscar métodos más objetivos y rápidos que la confirmación molecular de la t(15;17) (PMLRARα) patognomónica. Objetivo: Evaluar la utilidad del inmunofenotipo por Citometría de Flujo (CF) como método diagnóstico temprano a LPA ya que presenta un patrón fenotípico aberrante característico. Materiales y Métodos: Se evaluaron 85 pacientes con diagnóstico presuntivo de Leucemia Mieloide Aguda, de los cuales 10 presentaron perfil inmunofenotipico de LPA. Procedimientos diagnósticos en el aspirado medular: 1) observación microscópica e histoquímica para mieloperoxidasa; 2) análisis multiparamétrico con anticuerpos monoclonales por CF, que cubrió el espectro de los distintos linajes. Los datos se adquirieron y analizaron en un FACScalibur (Becton Dickinson). Los parámetros hemostáticos evaluados fueron: Tiempo de Protrombina y Tiempo de Tromboplastina Parcial Activado, Fibrinógeno, Dímero D y Productos de Degradación del Fibrinógeno. Resultados: Las características fenotípicas de probable LPA M3 t(15,17) la presentaron 10 de los pacientes. El perfil inmunofenotípico tuvo correlación con la morfología en 6 casos; en 2 pacientes, la morfología no fue suficiente para clasificarlos como portadores de LMA M2 o M3. Pacientes (n=2), que presentaban inicialmente morfología monocítica, el diagnóstico definitivo fue M3v hipogranular. Patrón inmunofenotípico: única población de blastos, autofluorescencia, CD33++ homogéneo, expresión heterogénea de CD13, ausencia de expresión de HLA DR, patrón CD15/ CD34 definido. Conclusiones: el inmunofenotipo aporta gran valor para un screening rápido de LPA con rearreglo PML/RARα.

Acute Promyelocytic Leukemia (APL) requires rapid identification since early therapeutic decisions should be made. In general, morphological screening is enough. However, the existence of false positive and false negative results requires a search for methods that are more objective and rapid than molecular confirmation of pathognomonic t(15;17) (PMLRARa+). The objective of this study was to assess the usefulness of immu- nophenotype by means of Flow Cytometry (FC) as an early diag- nosis method for APL since it presents a characteristically aberrant phenotypical pattern. The study consisted in the assessment of 85 patients presu- mably with a diagnosis for Acute Myeloid Leukemia (AML). Among these patients, 10 presented an APL immunophenotypical profi- le. The diagnostic procedures performed in the medullar aspi- ration were the following: 1) microscopic and histochemical observation for myeloperoxidase; 2) multi-parameter analysis with monoclonal antibodies by means of FC, that covered the spectrum for the different strains. Data were acquired and analyzed in a FACScalibur (Becton Dickinson). The hemostatic parameters assessed were the following: Prothrombine Time and Partially Activated Thromboplastine Time, Fibrinogen, D Dimer, and Fibrinogen Degradation Products. The following results were obtained: 10 of the patients showed phenotypical characteristics of probable APL M3 t(15,17). The immunophenotypical profile showed correlation with the morphology in 6 cases; in 2 patients, the morphology was not enough to classify them as AML M2 or M3 carriers. For those patients (n=2), presenting monocytic morphology at the on-set, the final diagnosis was hypogranular M3v. The immunophenotypical pattern found was only one blast population, auto-fluorescence, homogeneous CD33++, heterogeneous CD13 expression, lack of expression of HLA DR, defined CD15/CD34 pattern. In conclusion, the immunophenotype is very useful for rapid screening of APL with PML/RAR rearrangement.