CGM2, a Member of the Carcinoembryonic Antigen Gene Family is Down- Regulated in Colorectal Carcinomas

We have determined the precise chromosomal location, the exon structure, and the expression pattern of CGM2, a member of the carcinoembryonic antigen (CEA) gene family. CGM2 cDNA was amplified by reverse transcription-polymerase chain reaction (RT/PCR) from the colon adenocarcinoma cell line, LS174T. A defective exon is missing from this cDNA clone, leading to a novel domain organization for the human CEA family with two immunoglobulin-like domains. The derived C-terminal domain predicts that the CGM2 protein is membrane-bound through a glycosyl phosphatidylinositol anchor. RT/PCR analyses identified CGM2 transcripts in mucinous ovarian and colonic adenocarcinomas as well as in adjacent colonic tissue, but not in other tumors including leukocytes from six chronic myeloid leukemia patients. Thus, unlike several other family members, CGM2 is not expressed in granulocytes but reveals a more CEA-like expression pattern. Northern blot analyses identified a 2.5-kilobase CGM2 mRNA that is strongly down-regulated in colonic adenocarcinomas compared with adjacent colonic mucosa, suggesting a possible tumor suppressor function. In addition, a 3.2- kilobase transcript was observed in a number of colon tumors that is not detectable in normal colonic tissue. This mRNA species could represent a tumor-specific CGM2 splice variant

Cloning of the Complete Gene for Carcinoembryonic Antigen

Carcinoembryonic antigen (CEA) is a widely used tumor marker, especially in the surveillance of colonic cancer patients. Although CEA is also present in some normal tissues, it is apparently expressed at higher levels in tumorous tissues than in corresponding normal tissues. As a first step toward analyzing the regulation of expression of CEA at the transcriptional level, we have isolated and characterized a cosmid clone (cosCEA1), which contains the entire coding region of the CEA gene. A close correlation exists between the exon and deduced immunoglobulin-like domain borders. We have determined a cluster of transcriptional starts for CEA and the closely related nonspecific cross-reacting antigen (NCA) gene and have sequenced their putative promoters. Regions of sequence homology are found as far as approximately 500 nucleotides upstream from the translational starts of these genes, but farther upstream they diverge completely. In both cases we were unable to find classic TATA or CAAT boxes at their expected positions. To characterize the CEA and NCA promoters, we carried out transient transfection assays with promoter-indicator gene constructs in the CEA-producing adenocarcinoma cell line SW403, as well as in nonproducing HeLa cells. A CEA gene promoter construct, containing approximately 400 nucleotides upstream from the translational start, showed nine times higher activity in the SW403 than in the HeLa cell line. This indicates that cis-acting sequences which convey cell type-specific expression of the CEA gene are contained within this region

Lack of functional and expression homology between human and mouse aldo-keto reductase 1C enzymes: implications for modelling human cancers

<p>Abstract</p> <p>Background</p> <p>Over recent years, enzymes of the aldo-keto reductase (AKR) 1C subfamily have been implicated in the progression of prostate, breast, endometrial and leukemic cancers. This is due to the ability of AKR1C enzymes to modify androgens, estrogens, progesterone and prostaglandins (PGs) in a tissue-specific manner, regulating the activity of nuclear receptors and other downstream effects. Evidence supporting a role for AKR1C enzymes in cancer derives mostly from studies with isolated primary cells from patients or immortalized cell lines. Mice are ideal organisms for <it>in vivo </it>studies, using knock-out or over-expression strains. However, the functional conservation of AKR1C enzymes between human and mice has yet to be described.</p> <p>Results</p> <p>In this study, we have characterized and compared the four human (AKR1C1,-1C2, -1C3 and -1C4) and the eight murine (AKR1C6, -1C12, -1C13, -1C14, -1C18, -1C19, -1C20 and -1C21) isoforms in their phylogeny, substrate preference and tissue distribution. We have found divergent evolution between human and murine AKR1C enzymes that was reflected by differing substrate preference. Murine enzymes did not perform the 11β-ketoreduction of prostaglandin (PG) D₂, an activity specific to human AKR1C3 and important in promoting leukemic cell survival. Instead, murine AKR1C6 was able to perform the 9-ketoreduction of PGE₂, an activity absent amongst human isoforms. Nevertheless, reduction of the key steroids androstenedione, 5α-dihydrotestosterone, progesterone and estrone was found in murine isoforms. However, unlike humans, no AKR1C isoforms were detected in murine prostate, testes, uterus and haemopoietic progenitors.</p> <p>Conclusions</p> <p>This study exposes significant lack of phylogenetic and functional homology between human and murine AKR1C enzymes. Therefore, we conclude that mice are not suitable to model the role of AKR1C in human cancers and leukemia.</p

Combined bezafibrate and medroxyprogesterone acetate: potential novel therapy for acute myeloid leukaemia

Background: The majority of acute myeloid leukaemia (AML) patients are over sixty years of age. With current treatment regimens, survival rates amongst these, and also those younger patients who relapse, remain dismal and novel therapies are urgently required. In particular, therapies that have anti-leukaemic activity but that, unlike conventional chemotherapy, do not impair normal haemopoiesis. Principal Findings: Here we demonstrate the potent anti-leukaemic activity of the combination of the lipid-regulating drug bezafibrate (BEZ) and the sex hormone medroxyprogesterone acetate (MPA) against AML cell lines and primary AML cells. The combined activity of BEZ and MPA (B/M) converged upon the increased synthesis and reduced metabolism of prostaglandin D2 (PGD2) resulting in elevated levels of the downstream highly bioactive, anti-neoplastic prostaglandin 15-deoxy Δ12,14 PGJ2 (15d-PGJ2). BEZ increased PGD2 synthesis via the generation of reactive oxygen species (ROS) and activation of the lipid peroxidation pathway. MPA directed prostaglandin synthesis towards 15d-PGJ2 by inhibiting the PGD2 11β -ketoreductase activity of the aldo-keto reductase AKR1C3, which metabolises PGD2 to 9α11β-PGF2α. B/M treatment resulted in growth arrest, apoptosis and cell differentiation in both AML cell lines and primary AML cells and these actions were recapitulated by treatment with 15d-PGJ2. Importantly, the actions of B/M had little effect on the survival of normal adult myeloid progenitors. Significance: Collectively our data demonstrate that B/M treatment of AML cells elevated ROS and delivered the anti-neoplastic actions of 15d-PGJ2. These observations provide the mechanistic rationale for the redeployment of B/M in elderly and relapsed AML

From Vascular Smooth Muscle Cells to Folliculogenesis: What About Vasorin?

First described in 1988, vasorin (VASN) is a transmembrane glycoprotein expressed during early mouse development, and with a less extent, in various organs and tissues (e.g., kidney, aorta, and brain) postnatally. Vasn KO mice die after 3 weeks of life from unknown cause(s). No human disease has been associated with variants of this gene so far, but VASN seems to be a potential biomarker for nephropathies and tumorigenesis. Its interactions with the TGF-β and Notch1 pathways offer the most serious assumptions regarding VASN functions. In this review, we will describe current knowledge about this glycoprotein and discuss its implication in various organ pathophysiology

Low catalytic activity is insufficient to induce disease pathology in triosephosphate isomerase deficiency

Triosephosphate isomerase (TPI) deficiency is a fatal genetic disorder characterized by hemolytic anemia and neurological dysfunction. Although the enzyme defect in TPI was discovered in the 1960s, the exact etiology of the disease is still debated. Some aspects indicate the disease could be caused by insufficient enzyme activity, whereas other observations indicate it could be a protein misfolding disease with tissue-specific differences in TPI activity. We generated a mouse model in which exchange of a conserved catalytic amino acid residue (isoleucine to valine, Ile170Val) reduces TPI specific activity without affecting the stability of the protein dimer. TPIIle170Val/Ile170Val mice exhibit an approximately 85% reduction in TPI activity consistently across all examined tissues, which is a stronger average, but more consistent, activity decline than observed in patients or symptomatic mouse models that carry structural defect mutant alleles. While monitoring protein expression levels revealed no evidence for protein instability, metabolite quantification indicated that glycolysis is affected by the active site mutation. TPIIle170Val/Ile170Val mice develop normally and show none of the disease symptoms associated with TPI deficiency. Therefore, without the stability defect that affects TPI activity in a tissue-specific manner, a strong decline in TPI catalytic activity is not sufficient to explain the pathological onset of TPI deficiency

mfERG waveform characteristics in the RS1h mouse model featuring a 'negative' ERG

Several retinal disorders lead to a relatively greater attenuation of the b-wave compared to the a-wave of the electroretinogram (ERG), a constellation called `negative' ERG. To determine the waveform characteristics of multifocal ERGs (mfERGs) and their dependence on recording parameters in such a case, we studied the Rs1h−/Y mouse, the model for x-linked juvenile retinoschisis. mfERGs were recorded with a VERIS 4 system connected to a piggyback stimulator prototype that added the stimulus to the optical pathway of a HRA scanning-laser ophthalmoscope (SLO) by means of a wavelength-sensitive mirror. Real-time fundus visualization was achieved with the infrared laser of the SLO (835 nm). High-pass filter settings and the time interval used by the `artefact removal' feature were varied to study their influence on the waveform. The mfERG in the Rs1h−/Y mouse had a `negative' shape. However, the high-pass filter setting had to be lowered from the usual 10 Hz down to about 2 Hz in order to obtain that result, otherwise the negative shape was lost and mainly a positive peak remained. Similarly, a short time interval used by the `artefact removal' feature also removed the negative shape. The Rs1h−/Y mouse was found to be a valuable model of diseases with a `negative' waveform shape also in mfERG. Our results underline the importance of a lower high-pass filter cutoff frequency when recording mfERGs in such disorders. In addition, if the `artefact removal' feature is used, it should be verified that it doesn't distort the waveform shape