Lidia Averboukh

Identification of rCop-1, a New Member of the CCN Protein Family, as a Negative Regulator for Cell Transformation

ABSTRACT By using a model system for cell transformation mediated by the cooperation of the activated H-ras oncogene and the inactivated p53 tumor suppressor gene, rCop-1 was identified by mRNA differential display as a gene whose expression became lost after cell transformation. Homology analysis indicates that rCop-1 belongs to an emerging cysteine-rich growth regulator family called CCN, which includes connective-tissue growth factor, CYR61, CEF10 (v-src inducible), and the product of thenov proto-oncogene. Unlike the other members of the CCN gene family, rCop-1 is not an immediate-early gene, it lacks the conserved C-terminal domain which was shown to confer both growth-stimulating and heparin-binding activities, and its expression is lost in cells transformed by a variety of mechanisms. Ectopic expression of rCop-1 by retroviral gene transfers led to cell death in a transformation-specific manner. These results suggest that rCop-1 represents a new class of CCN family proteins that have functions opposing those of the previously identified members.

Regulation of S100P expression by androgen

The growth and function the normal prostate is dependent on the presence of androgen. As prostate tumors progress there is a loss of androgen‐dependent cell growth. The identification of the genes that are regulated by androgens may be of pathological and clinical significance.

Classification of breast cancer cells on the basis of a functional assay for estrogen receptor

BackgroundThe receptor (ER) for estrogen (E2) is routinely assayed as a marker to determine the feasibility of anti-hormone therapy against breast cancer because ER-positive (ER+) tumors are much more likely to respond to anti-hormone therapy than are ER-negative (ER−). However 40% of ER+ breast cancer patients do not respond to anti-hormone therapy. We suggest that this unpredictability of therapeutic responses lies in the current ER assays, which measure only an initial component of the E2-responsive pathway, and that the difference depends upon altered downstream processes. We propose a functional criterion that subclassifies breast cancers on the basis of specific binding of ER to its cognate DNA sequence, the estrogen response element (ERE).Materials and MethodsER was identified in breast cancer cell lines by immunofluorescence assay, Western blot analysis, identification of ER-specific mRNA, and by interaction of the ER-ERE complex with three different ER-specific antibodies. ER-ERE complex formation was measured by electrophoretic mobility shift assay (EMSA). Transactivation of the E2-responsive gene was studied by transfection of cells with fusion gene construct with the promoter-containing ERE sequence and assay of reporter gene activity in the cell extracts.ResultsThe growth of ER+ T47D cells was sensitive to tamoxifen, ICI-182,780, and ethynyl estradiol (EE2), whereas another ER+ breast cancer cell line, 21PT, was resistant to these compounds. The estrogen receptor (ER) in the nuclear extracts of MCF-7 and T47D demonstrated hormone-dependent interaction with the response element (ERE) and also downstream transactivation of the E2-responsive PS2 promoter. But in the 21PT cell line that was designated as ER− on the basis of ligand-binding assay and was found to be ER+ by all the other ER assays, ER-ERE interaction and PS2 promoter transactivation were independent of hormone.ConclusionsOn the basis of the downstream functional assay of ER interaction with ERE, ER+ breast tumor cells can be subclassified into two categories. The first is E2-dependent (ERd+) and these cells should respond to anti-hormone therapy. The second type of ER interacts with ERE independent of E2 (ERi+) and constitutively transactivates responsive genes. It is predicted that the latter type of breast cancers will not respond to antihormone therapy.

Identification of novel diagnostic markers by differential display.

Accurate and early diagnosis of a disease state such as a viral infection, or in a more complicated situation cancer, means live saving because proper medical interventions can be applied in a timely manner before it is too late to treat the disease. Thus it is crucial that good diagnostic markers for any relevant diseases can be obtained. The markers can be in the form of DNA such as viral integration or chromosomal DNA aberrations including deletions, translocations, and point mutations. As a result, these genetic abnormalities, in turn, may lead to altered gene expressions, such as new genes being turned on. Therefore, the markers can also be in the form of mRNAs or their protein products. DNA-based diagnosis is now done mostly with amplification technology breakthroughs such as polymerase chain reaction (PCR). However, protein based diagnosis such as a blood antibody test for a disease specific antigen (e.g., HIV and HPV virus infections; prostate-specific antigen for prostate cancer) are more accurate, convenient, and noninvasive. Traditionally, a good diagnostic marker for an infectious disease can be obtained by identifying the etiological agent such as a venues or a bacterium, which may not be always easy. But for noninfectious diseases such as cancer, a good marker may be even harder to come by because the alteration is more subtle and difficult to detect.