No abstract

Germ-line mutations in the adenomatous polyposis coli (APC) gene result in familial adenomatous polyposis coli (APC), an inherited syndrome that predisposes affected individuals to early onset of colorectal cancer. Somatic APC mutations also have been detected in sporadic colon tumors. We have used single strand conformational polymorphism (SSCP) analysis to scan a region of the APC gene that frequently is mutated in both APC and sporadic colorectal cancer. Four truncating mutations were found between codons 1060 and 1327 in 17 of 68 unrelated APC individuals. Fourteen of these persons carried either of two previously described five-nucleotide deletions which represent about 20% of APC mutations in these pedigrees. Patients with mutations in this region of exon 15 develop a classic APC colonic phenotype with multiple, diffuse adenomas developing by the second or third decade. However, the density of adenomas and the extracolonic disease manifestations associated with this syndrome are variable among individuals with identical APC mutations.

Pre-clinical data showed that priming CD34+ hematopoietic progenitor cells (HPC) with complement fragment 3a (C3a) improved homing and engraftment. Thus, we hypothesized that priming of UCB hematopoietic progenitors with C3a would facilitate homing and could potentially be used to address the need for improved engraftment after UCB transplantation. We primed one of two UCB for double UCB transplant after nonmyeloablative conditioning. This design provided adequate safety and the potential to observe skewed long-term chimerism in favor of the C3a primed unit as a surrogate measure of efficacy. C3a priming of one UCB unit did not result in infusional toxicity. Increased grades 1–3 hypertension to were the only infusional adverse events observed in 9 (30%) patients. We observed no activation of inflammatory or coagulation pathways downstream of C3a. As tested, C3a priming did not impair engraftment, but did not skew chimerism towards the treated unit. As compared to historical controls, mortality and survival were not adversely affected. Thus, before any additional clinical studies, C3a priming to promote engraftment will require further pre-clinical optimization.

Wilma tumor may be caused by loss of function of genes at different loci. A Wilms tumor suppressor gene, WTI, at chromosome 11 band p13, has recently been cloned and characterized. WTI has been implicated in the development of Wilms tumor by virtue of mutations in patients with genitourinary anomalies and susceptibility to Wilma tumor. (LOH) for markers at 11p13 (1-4). Molecular analysis of DNA mapping to a homozygously deleted region at 11p13 in two cases of sporadic Wilms tumor with apparent normal karyotype (5, 6) led to the identification and isolation of WTJ independently by three groups (6-8). The predicted WT1 protein contains four zinc fingers and a proline/glutaminerich region, suggesting a transcriptional regulatory function. The WT1 zinc finger region has been shown to bind the promoter consensus sequence of early growth response genes (9) and it seems likely that the WT1 protein has a regulatory function in the expression of genes in early embryonic differentiation.The role of WTI as a Wilms tumor susceptibility gene has been supported by reports demonstrating homozygous deletions of the region encompassing all or part of WTI (5, 6, 10-13), constitutional intragenic WTI deletions followed by 11p13 tumor , and by a case with early chromosomal reduplication followed by nondisjunction that preceded a somatic 25-bp WTI deletion spanning an exon-intron junction (21). Homozygous somatic point mutations in sporadic unilateral Wilms tumor have thus far not been reported. We report on two such cases as well as on one with a de novo germ-line point mutation as first event. In all three, loss ofthe wild-type allele is expected to result in inactivation of WTI. MATERIALS AND METHODSFifty Wilms tumors from as many patients were analyzed in this study. Five of these patients had bilateral disease, one hemihypertrophy, one aniridia, one the Prader-Willi syndrome, and one Bloom syndrome. None Isolation of constitutional and tumor DNA and determination of LOH by Southern blot hybridization of restriction enzyme digests of unamplified genomic DNA, as well as by direct visualization of genomic DNA restriction fragments amplified by the polymerase chain reaction (PCR), were performed as described (4).Single-Strand Conformational Polymorphism (SSCP) Analysis. Exonic sequences from genomic DNA were amplified by PCR and the PCR products were analyzed for SSCP (22) in order to detect possible deletions, insertions, or point mutations. In the analysis of WTI exon 7, the primers used were Luc7 (sense), 5'-ACCTACGTGAATGTTCACATGT-GCTTA-3', and LucAS7 (antisense), 5'-TCTTGAACCAT-GTTTGCCCAAGACTGGA-3'. For WTI exon 8, (A-2)8 and (S-2)8, and for WTI exon 9, (A-2)9 and (S-2)9, which have been described (16), were used. For WTI exon 10, Luc10 (sense), 5'-GTTGCAAGTGTCTCTGACTGGCAATTGT-3', and LucAS10 (antisense), 5'-TGAAAGCAGTTCACA-CACTGTGCTGCCT-3', were used. PCR was carried out in a GeneAmp PCR system 9600 (Perkin-Elmer) with a final volume of 100 ,u1, containing 250 ,M each dATP, dGTP, and dTTP; 20 AM dCTP; 1 mCi (37 MBq...

Studies were conducted to determine whether the progressive development of anemia associated with the antineoplastic drug cis-diamminedichloroplatinum (cDDP) was the consequence of decreased erythropoietin (Epo) production due to cDDP-induced nephrotoxicity or selective inhibition of erythroid progenitor cells. Five days after a single intraperitoneal injection of cDDP, hypoxia-induced Epo production was not decreased in mice and was increased significantly in rats in spite of severe multifocal tubular necrosis. In both species, colony-forming units-granulocyte macrophage (CFU-gm) and colony-forming units-erythroid (CFU-e) were reduced significantly, with a greater decrease in CFU-e. Studies of an anemic patient receiving cDDP also showed elevated Epo and decreased CFU-gm and CFU-e. In vitro exposure of mouse and human bone marrow to cDDP caused a dose-dependent inhibition of CFU-gm and CFU-e in both species, with human CFU-e showing greatest sensitivity. The results indicate that the primary hematologic toxicity of cDDP is directed at the hematopoietic stem cell compartment.