Compositions and Methods for the Diagnosis and Therapy of BCL2-Associated Cancers
a technology of bcl2 and cancer, applied in the field of cancer diagnosis, can solve the problems of inability to demonstrate the consistent involvement of any of these genes in carcinogenesis, cancer is a significant source of mortality and morbidity, and chemotherapy agents employed are generally toxic to patients, so as to increase the sensitivity of cancer cells
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A 30 kb Deletion Region in Somatic Cell Hybrids of CLL Patients
[0158]The minimal region of loss at 13q14 in CLL patients has been unclear. Previously, various and relatively large (between 130 to 550 kb) regions deleted in 13q14 have been described in CLL (see FIG. 2B). LOH and Southern blot analyses were used to identify the centromeric boundary of homozygous loss at the Alu18 locus (FIG. 2D), which is located between D13S1150 and D13S272 less than 65 kb centromeric to exon 5 of the LEU2 gene. However, no small or overlapping homozygous deletions were found that allowed a better localization of the target tumor suppressor.
[0159]To better define the region of loss in CLL, somatic cell hybrids of mouse LM-TK− and CLL cells carrying 13q14 translocations and / or deletions were generated. PCR screening of resulting hybrid clones allowed the segregation of the two copies of chromosome 13 present in the tumors. In this manner, a 31.4 kb deletion was identified in one case, and the chromoso...
example 2
The miR15 and miR16 Genes are Localized in the Minimally Deleted Region of Chromosome 13 and are Highly Expressed in CD5+ Cells
[0161]Publicly available sequence information and databases were screened for new regulatory genes in the minimal region of loss at 13q14. A cluster of two recently cloned miRNA genes, miR15 and miR16, were located exactly in the deleted region (FIG. 2A). To evaluate the level of expression of miR15 and miR16 in normal tissues, Northern blot analysis of miR15 and miR16 RNA was performed on a panel of normal tissues, including CD5+ B cells isolated from tonsils of normal individuals (FIG. 3A). CD5+ B cells were used as controls, because B-CLL is characterized by a progressive accumulation of CD5+ B-lymphocytes. Ubiquitous expression of both miR15 and miR16 genes was found, with the highest level in normal CD5+ lymphocytes. In addition, miR16 was consistently expressed at higher levels than miR15 in normal tissues. These data indicated that the miR15 and miR16...
example 3
The miR15 and miR16 Genes are Frequently Deleted or Downregulated in CLL Samples with Deletions at 13q14
[0162]To investigate whether the miR15 and miR16 genes were involved in CLL pathogenesis, 60 CLL samples and 30 human cancer cell lines were analyzed for miR15 and miR16 expression by Northern blotting (FIG. 3B). 68% of CLL patients (41 / 60), as well as 5 out of 6 analyzed prostate cancer cell lines, showed a significant reduction in expression when compared with their normal tissue counterparts. These findings demonstrated that the miR15 and miR16 genes are down-regulated in the majority of B-CLL and prostate cancer cases tested.
[0163]In addition, 23 out of 60 CLL samples (38%) presented a clearly identifiable band of about 70 nt representing the miR15 precursor RNA. The 70 nt miR15 band was not found in any normal tissue analyzed except for bone marrow (FIG. 3A), which indicated that miR15 precursor RNA could be inefficiently processed in CLL.
[0164]To determine whether the observ...
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