PKD1 is the most common site for mutations in human autosomal dominant polycystic kidney disease (ADPKD). ADPKD is characterized by progressive replacement of kidney tissue by epithelial cysts and eventual renal failure. Hepatic and pancreatic cysts are also common. The PKD1 protein, polycystin, is a cell-surface protein of unknown function that is widely expressed in epithelia and in vascular smooth muscle and myocardium. None of the genetic forms of murine polycystic disease map to the murine Pkd1 locus. We introduced into mice by homologous recombination a Pkd1 truncation mutation, Pkd1-, that mimics a mutation found in ADPKD. Pkd1- heterozygotes have no discernible phenotype, whereas homozygotes die during the perinatal period with massively enlarged cystic kidneys, pancreatic ductal cysts and pulmonary hypoplasia. Renal cyst formation begins at embryonic day 15.5 (E15.5) in proximal tubules and progresses rapidly to replace the entire renal parenchyma. The timing of cyst formation indicates that full-length polycystin is required for normal morphogenesis during elongation and maturation of tubular structures in the kidney and pancreas.

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Acquired imatinib resistance in advanced Philadelphia-positive acute lymphoblastic leukemia (Ph ؉ ALL) has been associated with mutations in the kinase domain (KD) of BCR-ABL. We examined the prevalence of KD mutations in newly diagnosed and imatinib-naive Ph ؉ ALL patients and assessed their clinical relevance in the setting of uniform frontline therapy with imatinib in combination with chemotherapy. Patients enrolled in the German Multicenter Study Group for Adult Acute Lymphoblastic Leukemia (GMALL) trial ADE10 for newly diagnosed elderly Ph ؉ ALL were retrospectively examined for the presence of BCR-ABL KD mutations by denaturing highperformance liquid chromatography (D-HPLC), cDNA sequencing, and allelespecific polymerase chain reaction (PCR). A KD mutation was detected in a minor subpopulation of leukemic cells in 40% of newly diagnosed and imatinib-naive patients. At relapse, the dominant cell clone harbored an identical mutation in 90% of cases, the overall prevalence of mutations at relapse was 80%. P-loop mutations predominated and were not associ- IntroductionIncorporation of the ABL kinase inhibitor imatinib into frontline treatment of Philadelphia-positive acute lymphoblastic leukemia (Ph ϩ ALL) has significantly improved the antileukemic efficacy of induction therapy. Several cooperative ALL study groups have demonstrated complete remission rates consistently above 90%, irrespective of whether imatinib is used alone or combined with multiagent chemotherapy. [1][2][3][4][5][6][7][8][9] These results are superior to previously reported complete remission (CR) rates of 65% to 90% in younger patients [10][11][12][13] and 40% to 60% in Ph ϩ ALL patients older than 60 to 65 years of age. [14][15][16][17] Although accumulating evidence suggests that imatinib-containing therapeutic regimens may also improve long-term outcome in these patients, 3,[6][7][8]14 relapse remains a predominant cause of treatment failure. 3,[7][8][9] Numerous point mutations in the kinase domain (KD) of BCR-ABL that impair imatinib binding to varying degrees have been identified as a major mechanism of acquired resistance in patients with chronic myeloid leukemia (CML). [18][19][20][21][22][23][24][25] Data on BCR-ABL mutations in patients with Ph ϩ ALL or lymphoid blast crisis of CML are more limited. Two studies of patients with advanced Ph ϩ lymphoid leukemias identified 5 different KD mutations in 14 of the 17 evaluated patients with acquired resistance to imatinib. 26,27 Preponderance of the E255K/V P-loop mutation, which occurred in 6 of 9 patients (67%) following their treatment with imatinib was suggested by one of these reports 26 but not by the other. 27 However, all point mutations arose at positions within the KD that are known to be important for drug binding and to confer significant resistance to imatinib in vitro. [18][19][20] This demonstrated that different mutations within the BCR-ABL KD can be responsible for refractoriness of Ph ϩ lymphoid leukemias to imatinib, and also suggested that KD mutations may be a f...

The murine female reproductive tract differentiates during postnatal development. This process of cytodifferentiation and morphogenesis is dependent upon specific mesenchymal-epithelial interactions as well as circulating steroid hormones (Cunha, G.R., 1976. Int. Rev. Cytol. 47, 137-194; Pavlova, A. et al., 1994. Development 120, 335-346). Members of the Wnt family of signaling molecules have been recently identified in this system (Pavlova, A. et al., 1994. Development 120, 335-346; Bui, T.D. et al., 1997. Br. J. Cancer 75, 1131-1136; Miller, C., Sassoon, D.A., 1998. Development, in press). We describe the expression patterns of Wnt genes in the developing and adult female reproductive tract. Additionally, we note that changes in the levels of expression occur during the estrous cycle. Wnt gene expression patterns are regulated by the presence of epithelium in tissue graft experiments, suggesting that Wnt genes may indeed play roles in the mesenchymal-epithelial interactions critical for female reproductive tract development and function.

Several xenobiotic (organic cation and anion) transporters have recently been identified, although their endogenous substrates, if such exist, remain unknown. When we initially identified NKT, also known as OAT1, the first member of the organic anion transporter (OAT) family (Lopez-Nieto CE, You G, Bush KT, Barros EJ, Beier DR, and Nigam SK. J Biol Chem 272: 6471-6478, 1997), we noted its expression in the embryonic kidney. We have now demonstrated its transporter function and more fully examined the spatiotemporal expression patterns of representative organic ion transporters, [NKT (OAT1), Roct, OCT1, and NLT, also known as OAT2] during murine development. In the kidney, NKT (OAT1), OCT1, and Roct transcripts appeared at midgestation, coinciding with proximal tubule differentiation, and gradually increased during nephron maturation. A similar pattern was observed for NLT (OAT2) in the liver and kidney, although, in the kidney, NLT (OAT2) transcription did not increase as dramatically. The roughly cotemporal expression of these related transporters in the developing proximal tubule may indicate common transcriptional regulation. Expression during embryogenesis in extrarenal sites could suggest a role in the formation and maintenance of nonrenal tissues. Importantly, all four genes were expressed in unexpected places during nonrenal organogenesis: Roct in the fetal liver (temporally coinciding with the onset of hematopoiesis) and neural tissue; NKT (OAT1) in the fetal brain; OCT1 in the ascending aorta and atrium; and NLT (OAT2) in the fetal lung, intestine, skin, and developing bone. Because these gene products mediate the transport of a broad range of metabolites and toxins, it seems likely that, apart from their known functions, these transporters play a role in transport of organic molecules, perhaps including those with morphogenetic activity. These genes could also play important developmental roles independent of transport function.

PKD1, the gene that is mutated in approximately 85% of autosomal dominant polycystic kidney disease (ADPKD) cases in humans, has recently been identified (Eur. PKD Consortium. Cell 77: 881-894, 1994; also, erratum in Cell 78: 1994). The longest open-reading frame of PKD1 encodes polycystin, a novel approximately 460-kDa protein that contains a series of NH2-terminal adhesive domains (J. Hughes, C. J. Ward, B. Peral, R. Aspinwall, K. Clark, J. San Millan, V. Gamble, and P. C. Harris. Nat. Genet. 10: 151-160, 1995; and Int. PKD Consortium. Cell 81: 289-298, 1995) and several putative transmembrane segments. To extend studies of polycystin to an experimentally accessible animal, we have isolated a cDNA clone encoding the 3' end of Pkd1, the mouse homologue of PKD1, and raised a specific antibody to recombinant murine polycystin. This antibody was used to determine the subcellular localization and tissue distribution of the protein by Western analysis and immunocytochemistry. In the mouse, polycystin is an approximately 400-kDa molecule that is predominantly found in membrane fractions of tissue and cell extracts. It is expressed in many tissues including kidney, liver, pancreas, heart, intestine, lung, and brain. Renal expression, which is confined to tubular epithelia, is highest in late fetal and early neonatal life and drops 20-fold by the third postnatal week, maintaining this level into adulthood. Thus the temporal profile of polycystin expression coincides with kidney tubule differentiation and maturation.

Deficiencies of natural anticoagulant proteins including antithrombin (AT), protein C (PC) and protein S (PS) are important causes of inherited thrombophilia. This study aimed to report on the practical experience gained in performing genetic analyses of a large cohort of patients with AT, PC and PS deficiencies and to relate this knowledge to clinical application. We genotyped a large cohort of 709 unrelated patients with AT (231), PC (234) and PS (244) deficiencies referred to us by physicians throughout Germany. Mutations were detected by direct sequencing and multiplex ligation-dependent probe amplification (MLPA). The highest mutation detection rate (MDR) was found for the SERPINC1 gene (83.5%), followed by the PROC (69%) and PROS1 (43%) genes. Even at AT activities close to the normal range (75%), the MDR was 70%. Contrastingly, for PC and PS deficiencies, the MDR dropped significantly and mildly lowered to subnormal values. At PS activities >55% for PS no mutations were detected. Mutation profiles of all three genes were similar with the highest prevalence for missense mutations (63-78%), followed by nonsense (7-11%), splice-site mutations (7-13%), small deletions (1-8%), small insertions/duplications (1-4%) and large deletions (3-6%). In conclusion, genetic testing is a useful diagnostic tool for diagnosing thrombophilia. Based on our data, genetic analysis for patients with AT deficiency is indicated for all subnormal activities. In contrast, genotyping is not advisable for PC activities >70% and for PS activities >55%.