Icela Palma

Mitosis Is a Source of Potential Markers for Screening and Survival and Therapeutic Targets in Cervical Cancer

The effect of preventive human papillomavirus (HPV) vaccination on the reduction of the cervical cancer (CC) burden will not be known for 30 years. Therefore, it’s still necessary to improve the procedures for CC screening and treatment. The objective of this study was to identify and characterize cellular targets that could be considered potential markers for screening or therapeutic targets. A pyramidal strategy was used. Initially the expression of 8,638 genes was compared between 43 HPV16-positive CCs and 12 healthy cervical epitheliums using microarrays. A total of 997 genes were deregulated, and 21 genes that showed the greatest deregulation were validated using qRT-PCR. The 6 most upregulated genes (CCNB2, CDC20, PRC1, SYCP2, NUSAP1, CDKN3) belong to the mitosis pathway. They were further explored in 29 low-grade cervical intraepithelial neoplasias (CIN1) and 21 high-grade CIN (CIN2/3) to investigate whether they could differentiate CC and CIN2/3 (CIN2+) from CIN1 and controls. CCNB2, PRC1, and SYCP2 were mostly associated with CC and CDC20, NUSAP1, and CDKN3 were also associated with CIN2/3. The sensitivity and specificity of CDKN3 and NUSAP1 to detect CIN2+ was approximately 90%. The proteins encoded by all 6 genes were shown upregulated in CC by immunohistochemistry. The association of these markers with survival was investigated in 42 CC patients followed up for at least 42 months. Only CDKN3 was associated with poor survival and it was independent from clinical stage (HR = 5.9, 95%CI = 1.4–23.8, p = 0.01). CDKN3 and NUSAP1 may be potential targets for the development of screening methods. Nevertheless, further studies with larger samples are needed to define the optimal sensitivity and specificity. Inhibition of mitosis is a well-known strategy to combat cancers. Therefore, CDKN3 may be not only a screening and survival marker but a potential therapeutic target in CC. However, whether it’s indispensable for tumor growth remains to be demonstrated.

Participation of OCT3/4 and β-catenin during dysgenetic gonadal malignant transformation

Gonadoblastoma (GB) is an in situ tumor consisting of a heterogeneous population of mature and immature germ cells, other cells resembling immature Sertoli/granulosa cells, and Leydig/lutein-like cells, may also be present. GB almost exclusively affects a subset of patients with intersex disorders and in 30% of them overgrowth of the germinal component of the tumor is observed and the lesion is term dysgerminoma/seminoma. Several pathways have been proposed to explain the malignant process, and abnormal OCT3/4 expression is the most robust risk factor for malignant transformation. Some authors have suggested that OCT3/4 and beta-catenin might both be involved in the same oncogenic pathway, as both genes are master regulators of cell differentiation and, overexpression of either gene may result in cancer development. The mechanism by which beta-catenin participates in GB transformation is not completely clear and exploration of the E-cadherin pathway did not conclusively show that this pathway participated in the molecular pathogenesis of GB. Here we analyze seven patients with mixed gonadal dysgenesis and GB, in an effort to elucidate the participation of beta-catenin and E-cadherin, as well as OCT3/4, in the oncogenic pathways involved in the transformation of GB into seminoma/dysgerminoma. We conclude that the proliferation of immature germ cells in GB may be due to an interaction between OCT3/4 and accumulated beta-catenin in the nuclei of the immature germ cells.

Testosterone metabolites mediate its effects on myocardial damage induced by ischemia/reperfusion in male Wistar rats

The role of testosterone in cardiovascular (CV) homeostasis is in controversy, and the exact effects of testosterone on the cardiovascular system remain poorly understood. Testosterone is metabolized by aromatase into 17β-estradiol and by 5α-reductase into dihydrotestosterone (DHT). Thus, identification of these metabolites in the heart may help to explain the controversy regarding the cardiovascular effects of testosterone. We analyzed the expression patterns of these testosterone-metabolizing enzymes and assessed the effect of its enzymatic activity inhibition on ischemia (40 min)/reperfusion (4h, I/R) via the left anterior descendent coronary artery in intact and gonadectomized male rats. Myocardial damage was measured as percentage of infarcted area vs. area at risk. Aromatase and 5α-reductase protein expression was found in the left ventricle of intact and orchidectomized rats. Exogenous testosterone had no effect on I/R induced myocardial damage in intact male rats, meanwhile exogenous testosterone protects against I/R injury in orchidectomized rats. However, enzymatic inhibition of aromatase increased myocardial damage in the presence of testosterone, while enzymatic inhibition of 5α-reductase significantly decreased the level of myocardial damage. Our results also showed that sub-chronic inhibition of 5α-reductase resulted in myocardial protection in both groups. Furthermore, in orchidectomized and intact male rats IV treatment with DHT induces a significant increase in the myocardial damage induced by I/R. Thus, the effect of testosterone on cardiovascular pathophysiology could be related, at least in part to changes in the balance of testosterone 5α-reduction and aromatization.

Loss of Cytochrome P450 17A1 Protein Expression in a 17α-Hydroxylase/17,20-Lyase-Deficient 46,XY Female Caused by Two Novel Mutations in the CYP17A1 Gene

Abstract17α-Hydroxylase deficiency (17OHD) is a rare form of congenital adrenal hyperplasia caused by mutations in the CYP17A1 gene. This condition shows considerable clinical and biochemical variation. Molecular characterization of novel mutations in the CYP17A1 gene and detailed study of their structural, enzymatic, and clinical consequences are required to fully understand enzyme behavior. Here, we present the first molecular characterization of two novel mutations in CYP17A1 in a 15-year-old female Mexican mestizo 46,XY female with primary amenorrhea and lack of pubertal development and severe hypertension that manifested only after surgery. A complete clinical and biochemical evaluation was compatible with 17OHD. Structural anomalies in the CYP17A1 gene were discovered by direct automated sequencing, which revealed a novel compound heterozygous K110X/R362H mutation that leads to a complete lack of enzyme activity. Immunohistochemical analyses performed to determine protein expression and localization showed that cytochrome P450 17A1 was completely absent in the patient’s testicular tissue. Studies of novel mutations, such as those described here, provide important information that allows us to better understand the effect of a given mutation on enzyme function and to observe the impact of the mutation on clinical phenotype.

Detection of Epstein-Barr Virus and Genotyping Based on EBNA2 Protein in Mexican Patients With Hodgkin Lymphoma: A Comparative Study in Children and Adults

BACKGROUND Epstein-Barr virus (EBV) is a member of the Herpesviridae family and is associated with Hodgkin lymphoma (HL). Isolates of EBV are classified according to sequence variation in the latency genes such as Epstein-Barr virus nuclear antigen (EBNA). EBNA2 contains the most divergent locus and is classified into type 1 and type 2 or EBNA2A and EBNA2B, respectively. We compared the frequency of EBV and the distribution of EBNA genotypes in Mexican children and adults with HL. PATIENTS AND METHODS Lymph node biopsy specimens from children and adults with HL were embedded in paraffin. EBV was identified by LMP1 amplification and Epstein-Barr-encoded RNA EBER by in situ hybridization (ISH) and genotyped as EBNA2A or EBNA2B using nested polymerase chain reaction (PCR) and specific primers for the detection of subtype. RESULTS Sixty-six samples were obtained from 3 hospitals-42 (63%) from children and 24 (37%) from adults with HL. Thirty-two of the 42 samples (76.1%) were positive for EBV in children and 16 of 24 (66.6%) samples were positive in adults (P = .41). In both children and adults, EBV was found more frequently in male patients. Thirty-four of 48 cases could be typed (70.8%). EBNA2A was found in 7/21 (33.3%) children and in 4/13 (30.8%) adults (P = 1.0), and EBNA2B was found in 10/21 (47.6%) children and in 9/13 (69.2%) adults (P = .22). A mix of subtypes was found in 4/21 (19%) children. CONCLUSION EBV was found frequently in both children and adults with HL. EBNA2B was the most frequent subtype, and a high frequency of mixed subtypes was found in children.

Utility of OCT3/4, TSPY and β-Catenin as Biological Markers for Gonadoblastoma Formation and Malignant Germ Cell Tumor Development in Dysgenetic Gonads

BACKGROUND: Gonadoblastoma (GB) is regarded as an in situ form of germ cell tumor in dysgenetic gonads, and 30% of patients with GB develop a dysgerminoma/seminoma tumor. OBJECTIVE: Determine whether OCT3/4 and β-catenin are expressed in dysgenetic gonads before GB development and whether TSPY participates in the OCT3/4-β-catenin pathways in the malignant invasive behavior. METHODS: dysgenetic gonads of Disorders of sex differentiation (DSD) patients with mixed gonadal dysgenesis were analyzed by immunohistochemistry and immunofluorescence for comparison with GB and dysgerminoma/seminoma. RESULTS: Our results suggest that the development of GB is secondary to the interaction of OCT3/4 and TSPY, that β-catenin does not participate in this process. CONCLUSIONS: The use of this biological markers detects the potential high risk gonads.

Williams' neural stem cells: new model for insight into microRNA dysregulation.

Williams syndrome (WS) is a neurodevelopmental genetic disorder, due to a 7q11.23 hemizygous deletion. WS has a characteristic neurocognitive profile that includes intellectual disability (ID). Haploinsufficiency of some of the deleted genes is partially associated with the cognitive phenotype. The aim of this paper is to determine the differences in the microRNA (miRNA) expression in WS patients, using a neural cell model from the patients olfactory neuroepithelium (ONE), and to establish the relationship with those genes involved in neurodevelopment and neural function. To assess these goals, we made a comparative analysis of the miRNAs expression profile between WS patients and controls. Through an in silico analysis, we established potential pathways and targets associated with neural tissue. The expression profile shows 14 dysregulated miRNAs, including nervous system (NS)-rich miRNAs such as miR-125b, let-7c and miR-200. Most of these miRNAs have potential targets associated with NS functions while others have been reported to have specific neuronal functions. These data suggest that miRNAs widely contribute to the regulation of neurodevelopmental intrinsic processes, and that specific miRNAs could participate in WS neurobiology.

An overview of the infection of CMV, HSV 1/2 and EBV in Mexican patients with glioblastoma multiforme

Several risk factors are involved in glioblastoma, including cytomegalovirus (CMV). This research was carried out to determine the rate of CMV infection, as well as HSV 1/2 and EBV in brain tissue, in patients with glioblastomamultiforme (GBM). The tissues were tested using immunohistochemistry, PCR, in situ hybridization and real-time PCR. At least, one HHV was detected in 21/29 (72%) patients as follows: single infections with HSV-1/2 in 4/21 (19%), EBV in 6/21 (28.6%) and CMV in 1/21 (4.8%). Mixed viral infection, HSV-1/2 and EBV were detected in 4/21 patients (19%), CMV and EBV in 5/21 (23.8%), and HSV-1/2, EBV, and CMV in 1/21. The CMV viral load ranged from 3×102 to 4.33×105 genome/100ng of tissue. Genotype based on CMV gB was 3/7 where 2/3 was gB1 and 1/3 gB4. HSV, EBV and CMV were frequently found in brain tissues, more in mix in a population reported as highly seropositive.

Physical Activity Protects Men but Not Women for Sarcopenia Development

Objective: Sarcopenia is among the most deleterious effects of aging. The objective of this study was to analyze the relationship between performance tests and muscular volume over the life span of male and female participants. Method: A correlation study was conducted with healthy individuals (50 males and 47 females) between the ages of 20 and 94; the study group included active older people, sedentary younger people, and young athletes. Muscular volume was determined by tomography and muscular performance (4-meter speed tests [4 MSTs], chair test, and handgrip test), and a correlation analysis between the groups was performed. Results: Sex-related differences were observed between the variables; in males, muscle volume and functional parameters were closely related with age and physical activity, whereas in females, they were not related at all. Conclusion: Male and female muscle volume and performance demonstrate strong differences, which should be considered during clinical evaluations of sarcopenia.

Abstract A46: Expression of biomarkers in cervical intraepithelial neoplasia: Potential use screening.

Background: Persistent infection of high-risk Human papillomavirus (HPV) is recognized as a necessary cause of cervical cancer. Replication and viral integration into the cervix depend on the orderly expression of genes produced by viruses, by inducing the overexpression of multiple molecular proteins or biomarkers. Recently we have identified, 6 genes (CCNB2, CDC20, PRC1, SYCP2, NUSAP1 and CDKN3), which are proposed as universal molecular targets with a sensitivity and specificity close to100%, in cervical cancer. Objective: evaluate the expression of these genes in cervical intraepithelial neoplasia (CIN). Material and method: In this retrospective study we assessed the protein expression of 6 candidates tissue-based biomarkers (CCNB2, CDC20, PRC1, SYCP2, NUSAP1 and CDKN3) in a population based cohort of 200 cervical intraepithelial neoplasia that was collected over the period from 2008 to 2013. The tissue blocks were available and used for tissue microarray (TMA) construction, which included also samples of healthy epithelium. Quantitative RT-PCR was performed to validate expression of genes. Results: The expression of these genes was observed in cervical intraepithelial neoplasia with different pattern shown previously in tumors, but in progression with the advance of the lesion, also shown by IH, evaluated samples, finding a correlation in both methods. Conclusion: These results, may offer an additional strategy to current methods of the PAP, which is of greater use in our population. A test with greater sensitivity and specificity to identify lesions of high grade with more specificity can be a useful tool for cervical cancer screening. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A46. Citation Format: Ana Maria Espinosa Garcia, Ana Alfaro, Icela Palma, Marco Duran, Jaime Beumen. Expression of biomarkers in cervical intraepithelial neoplasia: Potential use screening. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A46.