Perry D. Nisen

Effects of the MYC oncogene antagonist, MAD, on proliferation, cell cycling and the malignant phenotype of human brain tumour cells

To investigate how overexpression of MAD, an antagonist of MYC oncogenes influences the malignant phenotype of human cancer cells, an adenovirus vector system was used to transfer the human MAD gene (AdMAD) into human astrocytoma cells. Decreased growth potential of AdMAD-infected cells was evidenced by a decrease in [3H]thymidine incorporation, an increase in cell doubling time and alteration of cell-cycle distribution. Diminished malignant potential of AdMAD-infected cells was manifested by their loss of anchorage-independent growth in soft agar and by their inability, in general, to induce tumorigenesis in a xenograft animal model. These studies indicate that adenovirus constructs encoding MAD dramatically inhibit the proliferation and tumorigenicity of human astrocytoma cells and support the use of MAD for gene therapy of human tumours.

Cytogenetics of a renal cell carcinoma in a 17-month-old child: Evidence for Xp11.2 as a recurring breakpoint☆

A renal cell carcinoma from a 17-month-old boy with a history of maternal hydrocarbon exposure was found to have a 46,Y,t(X;17)(p11.2;q25) karyotype. Although this translocation has not previously been reported, other translocations involving Xp11.2 have been described, suggesting that this may represent a non-random breakpoint involved in the pathogenesis of childhood renal cell carcinoma. Both chromosomes 3 in the tumor were normal by both karyotype and RFLP analysis.

Temperature-dependent modulation of lipopolysaccharide-induced interleukin-1 beta and tumor necrosis factor alpha expression in cultured human astroglial cells by dexamethasone and indomethacin.

In bacterial meningitis, LPS induces production in cerebrospinal fluid of the cytokines IL-1 beta and tumor necrosis factor alpha (TNF alpha), which are the principle mediators of meningeal inflammation. IL-1 beta and TNF alpha induce fever, and elevated temperature may affect cytokine expression. Dexamethasone treatment improves outcome in bacterial meningitis possibly by inhibiting IL-1 beta and TNF alpha. In this report, the effects of elevated temperature and dexamethasone on LPS-stimulated IL-1 beta and TNF alpha mRNA gene expression and protein synthesis were studied in human astrocytoma cell lines and primary cultures of human fetal astrocytes. Cells cultured at 40 degrees C exhibited smaller peaks of IL-1 beta and TNF alpha transcription and protein synthesis compared with cells cultured at 37 degrees C. The addition of dexamethasone before, during, or after exposure of the cells to LPS resulted in temperature-dependent inhibition of IL-1 beta transcription and protein synthesis. The most extensive inhibition occurred in pretreated cells cultured at 37 degrees C. Cotreatment with LPS and dexamethasone also inhibited TNF alpha mRNA transcription at both temperatures. The effects of another antiinflammatory agent, indomethacin, on LPS induction of IL-1 beta and TNF alpha mRNA were temperature and cell line dependent. These findings provide a possible explanation for the efficacy of dexamethasone treatment of bacterial meningitis and support the proposal that fever may be beneficial to the host in this disease.

Tumor suppression and inhibition of aneuploid cell accumulation in human brain tumor cells by ectopic overexpression of the cyclin-dependent kinase inhibitor p27KIP1.

To investigate how overexpression of p27KIP1, a downstream effector of TGF-beta and a universal cyclin-dependent kinase (CDK) inhibitor could influence the malignant phenotype of malignant human brain tumor cells, an adenovirus vector system was used to transfer the human p27KIP1 gene (Adp27KIP1) into the human astrocytoma cell line, U-373MG. Inhibition of CDK activity in Adp27KIP1-infected cells was indicated by inhibition of [3H]thymidine incorporation, an increase in cell doubling time and by cell cycle arrest in G1. Notably, ectopic overexpression of p27KIP1 was associated with a marked decrease in the accumulation of aneuploid cells. Diminished malignant potential of Adp27KIP1-infected cells was manifested by the loss of anchorage-independent growth in soft agar and by the inability to induce tumorgenesis in a xenograft model. These studies suggest that p27KIP1 is a tumor suppressor gene and supports the use of Adp27KIP1 for gene therapy of human brain tumors.

Infrequency of p53 gene mutations in ependymomas

SummaryEpendymomas, which comprise 5% of central nervous system tumors, have not been extensively characterized genetically. The p53 tumor suppressor gene is frequently mutated in human cancer, and is important in the pathogenesis of other central nervous system (CNS) tumors. Chromosomal DNA corresponding to the p53 tumor suppressor gene was amplified by the polymerase chain reaction (PCR) from 31 archival ependymoma specimens. DNA was screened for the presence of p53 mutations by single strand conformational polymorphism (SSCP) analysis; samples with altered mobility were further tested for the presence of mutation by direct DNA sequence analysis. Of the 31 ependymomas tested, one contained a detectable DNA sequence change in the p53 gene. Sequencing revealed a silent mutation in exon 6, at codon 213, which represents a known p53 sequence polymorphism. These findings suggest that in contrast to many other human cancers, p53 mutation is not important in the pathogenesis or progression of ependymomas.

Genetic alterations of chromosome band 9p21-22 in head and neck cancer are not restricted to p16INK4a

Although genetic alterations of chromosome band 9p21 – 22 occur frequently in head and neck squamous cell carcinoma (HNSCC) cell lines, alterations of the cyclin-dependent kinase inhibitor p16INK4a located in this region are less common in corresponding primary tumors. To further investigate genetic alterations at 9p21 – 22 and p16INK4a in primary HNSCC, a paired set of 21 tumors and blood specimens that were shown previously to exhibit allelic loss at 3p and elsewhere, were tested for LOH at 9p21 – 22 using eight different highly polymorphic marker. Sixteen of the samples (81%) exhibited LOH for at least one marker. Frequent LOH was found surrounding p16INK4a and at three additional non-contiguous regions of 9p21 – 22. No homozygous deletions were identified. SSCP screening and direct sequence analysis led to the identification of mutations the p16INK4a gene in two tumors. p16INK4a was not hypermethylated in any of the samples studied. Furthermore, there was no correlation between LOH at 9p21 – 22 and the RB1 tumor suppressor gene. These findings indicate that in the set of tumors that we tested, LOH at 9p21 – 22 is common in primary HNSCC but that genetic alterations of p16INK4a located in this region are unusual. Additional tumor suppressor genes at 9p21 – 22 may therefore be involved in the pathogenesis of this tumor.

p53 mutations, O6-alkylguanine DNA alkyltransferase activity, and sensitivity to procarbazine in human brain tumors

Background. In human brain tumors, sensitivity to procarbazine as measured by sensitivity in a xenograft tumor model correlated inversely with amounts of the DNA repair enzyme O6‐alkylguanine DNA alkyltransferase(AT).

Nonrandom X chromosome DNA methylation patterns in hemophiliac females.

Molecular X chromosome inactivation analysis was used to characterize three females (and their families) with severe hemophilia. First, the maternal and paternal X chromosomes were distinguished by restriction fragment length polymorphisms (RFLPs). Second, the patterns of methylation of X chromosome genes using methylation-sensitive restriction endonucleases were determined. Of the six X chromosome probes tested, only the phosphoglycerol-kinase (PGK) and hypoxanthine-phosphoribosyl-transferase (HPRT) clones were informative, indicating that other X chromosome probes are not useful for X inactivation analysis. After digestion with Hpa II or Hha I, the hybridization intensity of the RFLPs of all three mothers and an unaffected sister were diminished by 50%, consistent with random X chromosome inactivation. The methylation patterns of the X chromosomes of the affected females, however, were clearly nonrandom. Depending upon the probe and the patient, HPRT and PGK sequences were either completely methylated or unmethylated. These findings are extremely suggestive that nonrandom X chromosome inactivation (lyonization) is the basis for severe hemophilia in these females.

The molecular basis of severe hemophilia B in a girl

Severe phenotypic expression of recessive X-linked disorders in females is rare, although examples have been reported. Cases have been documented in females who were homozygous for a recessive alle...

Molecular characterization of congenital mesoblastic nephroma and its distinction from Wilms tumor

Background. Congenital mesoblastic nephroma (CMN) is a rare tumor of the neonatal kidney. It was once thought to be a variant of Wilms tumor that also arises from primitive renal cells.