Alterations of RASSF1A in premalignant cervical lesions: clinical and prognostic significance.

This study aimed to understand the importance of RASSF1A and CACNA2D2, located in chromosomal 3p21.31 region, in the development of uterine cervical carcinoma (CACX). To this end, firstly the expression (RNA) profiles of RASSF1A and CACNA2D2 were screened in primary cervical carcinoma (CACX) samples which indicated highly reduced expression for both genes. Thereafter alterations (deletion/methylation) of these genes were analyzed in 23 cervical intraepithelial neoplasia (CIN) and 110 CACX samples. In CIN, deletion was observed only for RASSF1A (26%), whereas methylation was in the following order: RASSF1A (35%) > CACNA2D2 (9%). However, in CACX their deletion frequencies were the same (50%) and methylation frequencies were comparable RASSF1A (33%), CACNA2D2 (27%). The reduced expression and molecular alterations of these genes were concordant. Overall alterations of RASSF1A showed association with CIN lesions and CACNA2D2 with disease progression from CIN → stage I/II. Interestingly, alterations of these genes showed significant association in CACX suggesting possible functional synergism during tumor progression. Alterations of RASSF1A and CACNA2D2 predicted poor prognosis for the patients. Moreover, RASSF1A alterations along with multiparity (≥5 yr) and early sexual debut (<19 yr) were determinants of worse prognosis. Our data suggests the association of RASSF1A and CACNA2D2 in cervical carcinogenesis and its importance in early diagnosis and prognosis of the tumor.

RBSP3 is frequently altered in premalignant cervical lesions: clinical and prognostic significance.

To understand the importance of frequent deletion of 3p22.3 in cervical carcinogenesis, alterations (deletion/methylation/expression) of the candidate genes STAC, MLH1, ITGA9, and RBSP3, located in the region, were analyzed in 24 cervical intraepithelial neoplasia (CIN) and 137 uterine cervical carcinoma (CACX) samples. In CIN, RBSP3 deletion (48%) and methylation (26%) were high compared with the other genes (4-9%). In CACX, alterations of these genes were as follows: deletion: STAC (54%) > MLH1 (46%) > RBSP3 (45%) > ITGA9 (41%), methylation: RBSP3 (25%) > ITGA9 (24%) > STAC (19%) > MLH1 (13%). Overall, alterations of RBSP3 showed association with CIN, whereas for STAC and MLH1, this frequency increased significantly from CIN --> Stage I/II and for ITGA9 from CIN --> Stage I/II and also from Stage I/II --> Stage III/IV. Quantitative mRNA expression analysis showed differential reduced expression of these genes in CACX concordant to their molecular alterations. The more active RBSP3B splice variant was underexpressed in CACX. RB1 was infrequently deleted in CACX. Concordance was seen between (i) inactivation of RBSP3 and intense p-RB1 nuclear immunostaining and (ii) low/absence of MLH1 expression and its molecular alterations in CACX. In normal cervical epithelium, p-RB1 immunostaining was low in differentiated cells, whereas MLH1 staining was seen in both nucleus and cytoplasm irrespective of differentiation stage. Alterations of the genes were significantly associated with poor prognosis. High parity (>or=5)/early sexual debut (

Morpho-Physiological Traits and Functional Markers Based Molecular Dissection of Heat-Tolerance in Urdbean.

Urdbean (Vigna mungo L. Hepper) is one of the important pulse crops. Its cultivation is not so popular during summer seasons because this crop is unable to withstand excessive heat stress beside lack of humidity in the atmosphere. Therefore, a panel of 97 urdbean diverse genotypes was assessed for yield under stress and non-stress conditions with an aim to identify heat tolerant genotypes. This study identified 8 highly heat tolerant and 35 highly heat sensitive genotypes based on heat susceptibility index. Further, physiological and biochemical traits-based characterization of a group of six highly heat sensitive and seven highly heat tolerant urdbean genotypes showed genotypic variability for leaf nitrogen balance index (NBI), chlorophyll (SPAD), epidermal flavnols, and anthocyanin contents under 42/25°C max/min temperature. Our results showed higher membrane stability index among heat tolerant genotypes compared to sensitive genotypes. Significant differences among genotypes for ETR at different levels of PAR irradiances and PAR × genotypes interactions indicated high photosynthetic ability of a few genotypes under heat stress. Further, the most highly sensitive genotype PKGU-1 showed a decrease in different fluorescence parameters indicating distortion of PS II. Consequently, reduction in the quantum yield of PS II was observed in a sensitive one as compared to a tolerant genotype. Fluorescence kinetics showed the delayed and fast quenching of Fm in highly heat sensitive (PKGU 1) and tolerant (UPU 85-86) genotypes, respectively. Moreover, tolerant genotype (UPU 85-86) had high antioxidant activities explaining their role for scavenging superoxide radicals (ROS) protecting delicate membranes from oxidative damage. Molecular characterization further pinpointed genetic differences between heat tolerant (UPU 85-86) and heat sensitive genotypes (PKGU 1). These findings will contribute to the breeding toward the development of heat tolerant cultivars in urdbean.

Amplification of CyclinL1 in uterine cervical carcinoma has prognostic implications.

The chromosomal 3q25.31 region was consistently amplified in primary cancer of cervix (CACX). CyclinL1 is a candidate gene of this region and already have been implicated as an oncogene in head and neck cancers. In this study, we aimed to investigate the involvement of CyclinL1 in cervical carcinogenesis and for this purpose its copy number variation (CNV) was studied in 23 cervical intraepithelial neoplasia (CIN) and 110 CACX samples. In CIN lesions CyclinL1 was not amplified; however, the amplification frequency was 16% (9/56) in stage I/II tumors which remained comparable during subsequent stages of tumorigenesis. This implied association of CyclinL1 amplification with development of early invasiveness. Quantitation of mRNA expression revealed 2.6 ± 1.53-fold overexpression of this gene in primary CACX. The amplification/copy number gain of CyclinL1 and its mRNA profile were concordant, in tumors. Immunohistochemical (IHC) analysis in primary CACX, cell lines: SiHa and HeLa revealed intense nuclear expression of cyclinL1, which was further confirmed by Western blot in the cell lines. However 47% (7/15) CACX samples expressed high/intermediate level of cyclin L1. Kaplan-Meier survival analysis indicated CyclinL1 amplification as a determinant of poor patient outcome. Tumor radio-resistance developed as a consequence of CyclinL1 amplification. Cox multivariate analysis revealed that multiparous (≥5) CACX patients with amplified CyclinL1 locus along with advanced tumor stage (III/IV) had worst prognosis. Our data suggest importance of CyclinL1 in cervical carcinogenesis with its associated pathways viz: pre-mRNA splicing, cell-cycle regulation (G0/G1 and G2/M) being potential targets of therapeutic interventions in CACX.

A Pooled Analysis to Compare the Clinical Characteristics of Human Papillomavirus-positive and -Negative Cervical Precancers.

Given that high-risk human papillomavirus (HPV) is the necessary cause of virtually all cervical cancer, the clinical meaning of HPV-negative cervical precancer is unknown. We, therefore, conducted a literature search in Ovid MEDLINE, PubMed Central, and Google Scholar to identify English-language studies in which (i) HPV-negative and -positive, histologically confirmed cervical intraepithelial neoplasia grade 2 or more severe diagnoses (CIN2+) were detected and (ii) summarized statistics or deidentified individual data were available to summarize proportions of biomarkers indicating risk of cancer. Nineteen studies including 3,089 (91.0%) HPV-positive and 307 (9.0%) HPV-negative CIN2+ were analyzed. HPV-positive CIN2+ (vs. HPV-negative CIN2+) was more likely to test positive for biomarkers linked to cancer risk: a study diagnosis of CIN3+ (vs. CIN2; 18 studies; 0.56 vs. 0.24; P < 0.001) preceding high-grade squamous intraepithelial lesion cytology (15 studies; 0.54 vs. 0.10; P < 0.001); and high-grade colposcopic impression (13 studies; 0.30 vs. 0.18; P = 0.03). HPV-negative CIN2+ was more likely to test positive for low-risk HPV genotypes than HPV-positive CIN2+ (P < 0.001). HPV-negative CIN2+ appears to have lower cancer risk than HPV-positive CIN2+. Clinical studies of human high-risk HPV testing for screening to prevent cervical cancer may refer samples of HPV test-negative women for disease ascertainment to correct verification bias in the estimates of clinical performance. However, verification bias adjustment of the clinical performance of HPV testing may overcorrect/underestimate its clinical performance to detect truly precancerous abnormalities.

Identification of Changes in the Human Papilloma Virus 16 (HPV16) Genome During Early Dissemination of Cervical Cancer Cells May Complement Histological Diagnosis of Lymph Node Metastasis.

Cancer of the uterine cervix (CACX) is one of the most common carcinoma affecting women worldwide. During treatment, histologically lymph node (LN) metastasis and presence of HPV DNA in blood plasma act as a major prognostic factor. Despite the lack of apparent LN involvement, some early-invasive CACX patients have shown recurrences and poor survival. This is suggestive of undetected early dissemination of cancer cells characterized by presence of HPV DNA in histologically non-metastatic LNs which finally progresses into histologically visible metastasis. This present study investigated the status and origin of HPV genome during early dissemination by molecular analysis in primary tumor (PT), histologically non-metastatic pelvic lymph nodes (LNs) and blood plasma (BP) of same patient. First, CACX patients showing signs of early dissemination was identified by detection of HPV in PT (n = 22) and their corresponding histologically non-metastatic pelvic LNs (n = 45) and BP (n = 18) followed by typing of HPV16/18. This was followed by comparative analysis of the physical, copy number and methylation (enhancer/early/late) status of HPV16 genome present in LNs and BP with that of PT. Our study revealed for the first time that the HPV16 genome were frequently present in the integrated form though the copy number was low in both non-metastatic LNs and BP. However, the methylation pattern of PT was discordant with that of corresponding LNs and BP in majority of the cases. Critical assessment of HPV16 profiles established that the presence of hrHPV may be due to the early dissemination of PT cells having significant pathological implications.

Inactivation of SLIT2-ROBO1/2 pathway in premalignant lesions of uterine cervix: clinical and prognostic significances.

The SLIT2-ROBO1/2 pathways control diverse biological processes, including growth regulation. To understand the role of SLIT2 and ROBO1/2 in cervical carcinogenesis, firstly their RNA expression profiles were screened in 21 primary uterine cervical carcinoma (CACX) samples and two CACX cell lines. Highly reduced expressions of these genes were evident. Concomitant alterations [deletion/methylation] of the genes were then analyzed in 23 cervical intraepithelial neoplasia (CIN) and 110 CACX samples. In CIN, SLIT2 was deleted in 22% samples compared to 9% for ROBO1 and none for ROBO2, whereas comparable methylation was observed for both SLIT2 (30%) and ROBO1 (22%) followed by ROBO2 (9%). In CACX, alteration of the genes were in the following order: Deletion:ROBO1 (48%) > SLIT2 (35%) > ROBO2 (33%), Methylation:SLIT2 (34%) > ROBO1 (29%) > ROBO2 (26%). Overall alterations of SLIT2 and/or ROBO1 (44%) and SLIT2 and/or ROBO2 (39%) were high in CIN followed by significant increase in stage I/II tumors, suggesting deregulation of these interactions in premalignant lesions and early invasive tumors. Immunohistochemical analysis of SLIT2 and ROBO1/2 in CACX also showed reduced expression concordant with molecular alterations. Alteration of all these genes predicted poor patient outcome. Multiparous (≥ 5) women with altered SLIT2 and ROBO1 along with advanced tumor stage (III/IV) and early sexual debut (<19 years) had worst prognosis. Our data suggests the importance of abrogation of SLIT2-ROBO1 and SLIT2-ROBO2 interactions in the initiation and progression of CACX and also for early diagnosis and prognosis of the disease.

Congenital spinal deformity: a comprehensive assessment at presentation.

STUDY DESIGN: A series of 126 consecutive patients with congenital spinal deformity is presented. OBJECTIVE: To assess the incidence of intraspinal anomaly and other organic defects associated with different types of spine deformity at presentation. SUMMARY OF BACKGROUND DATA: A high incidence of intraspinal abnormalities and other organ defects is reported in relation to congenital spine deformity. The prevalence of these problems with different types of deformities is to be determined. METHODS: All patients had MRI, echocardiography, renal ultrasound, and a thorough clinical assessment. RESULTS: Intraspinal abnormalities were found in 47 patients (37%). These abnormalities were significantly more common in patients with congenital kyphosis ( = 0.0048), and in those with scoliosis resulting from mixed and segmentation defects. Scoliosis patients with cervical and thoracic hemivertebrae had significantly more intraspinal abnormalities ( = 0.0253) than those with lumbar hemivertebrae. In 64 (55%) patients other organic defects were found. These defects were more common in patients with congenital scoliosis resulting from mixed defects ( = 0.002). Cardiac defects were detected in 26% and urogenital anomalies in 21% of the patients. CONCLUSIONS: Magnetic resonance imaging and echocardiography should be an essential part in the evaluation of patients with congenital spinal deformity, and special attention should be paid to patients with segmentation abnormalities, mixed defects, and kyphosis.