Iwao Nozawa

Identification and characterization of a 500-kb homozygously deleted region at 1p36.2-p36.3 in a neuroblastoma cell line

Loss of heterozygosity of the distal region of chromosome 1p where tumor suppressor gene(s) might harbor is frequently observed in many human cancers including neuroblastoma (NBL) with MYCN amplification and poor prognosis. We have identified for the first time a homozygously deleted region at the marker D1S244 within the smallest region of overlap at 1p36.2-p36.3 in two NBL cell lines, NB-1 and NB-C201 (MASS-NB-SCH1), although our genotyping has suggested the possibility that both lines are derived from the same origin. The 800-kb PAC contig covering the entire region of homozygous deletion was made and partially sequenced (about 60%). The estimated length of the deleted region was 500 kb. We have, thus far, identified six genes within the region which include three known genes (DFF45, PGD, and CORT) as well as three other genes which have been reported during processing our present project for the last 3½ years (HDNB1/UFD2, KIAA0591F/KIF1B-β, and PEX14). They include the genes related to apoptosis, glucose metabolism, ubiquitin-proteasome pathway, a neuronal microtubule-associated motor molecule and biogenesis of peroxisome. At least three genes (HDNB1/UFD2, KIAA0591F/KIF1B-β, and PEX14) were differentially expressed at high levels in favorable and at low levels in unfavorable subsets of primary neuroblastoma. Since the 1p distal region is reported to be imprinted, those differentially expressed genes could be the new members of the candidate NBL suppressor, although RT-PCR-SSCP analysis has demonstrated infrequent mutation of the genes so far identified. Full-sequencing and gene prediction for the region of homozygous deletion would elucidate more detailed structure of this region and might lead to discovery of additional candidate genes.

Hunting the subset-specific genes of neuroblastoma: Expression profiling and differential screening of the full-length-enriched oligo-capping cDNA libraries

BACKGROUND Neuroblastoma (NBL) has a distinct nature in different prognostic subgroups. PROCEDURE To understand the molecular mechanism of NBLs genesis and biology as well as that of the neural crest development, we constructed full-length-enriched cDNA libraries by an oligo-capping method from two different subsets of primary NBL, one with favorable biology and the other with MYCN amplification. RESULTS Sequencing analysis of these libraries revealed that the expression profile was markedly different between both subsets. To identify the genes differentially expressed between the subsets, semi-quantitative RT-PCR analyses are proceeding. CONCLUSION So far, 54 transcripts have been found to be expressed at high levels in favorable NBLs, and significantly at low levels in unfavorable NBLs.

Application of a thermo-responsive membrane to the transdermal delivery of non-steroidal anti-inflammatory drugs and antipyretic drugs

Abstract A thermo-responsive membrane was incorporated into a transdermal therapeutic system (TTS) which responds to thermal stimuli associated with skin temperature changes. The thermo-responsive membrane was prepared by pouring monooxyethylene trimethylolpropane tristearate (MTTS) between two porous polypropylene films. The TTS with this thermoresponsive membrane as a rate controlling membrane released non-steroidal anti-inflammatory drugs (indomethacin, ketoprofen) and antipyretic drugs (acetoaminophen, ethenzamide) in response to temperature changes between 32 and 38°C in the in vitro experiments. It was then shown that the temperature-activated TTS was operative in vivo in a rabbit model .

Identification of the homozygously deleted region at chromosome 1p36.2 in human neuroblastoma

BACKGROUND We have identified for the first time a homozygously deleted region within the smallest region of overlap at 1p36.2-3 in two neuroblastoma cell lines. PROCEDURE The 800-kb PAC contig covering the entire homozygously deleted region was made and sequenced. To date, approximately 70% of sequencing has been accomplished, and the estimated length of the deleted region was 500 kb. RESULTS Currently, we have found six genes within the region, which include three known genes as well as three other genes that have been reported during processing of our present project for the last 3(1/2) years. We report here the results of expression and mutation analyses of those genes. CONCLUSIONS Full sequencing for the region of homozygous deletion as well as further analyses of the genes mapped within the region may reveal whether or not there is a neuroblastoma suppressor gene as proposed by the Knudsons two-hit hypothesis.