Memory-impairing effects of local anesthetics in an elevated plus-maze test in mice

Detailed deletion mapping of chromosome 6q has shown that the highest percentage of loss of heterozygosity (LOH) is located at 6q25-q27 and suggested that an ovarian cancer associated tumor suppressor gene may reside in this region. To further define the smallest region of common loss, we used 12 tandem repeat markers spanning a region no more than 18 cM, located between 6q25.1 and 6q26, to examine allelic loss in 54 fresh and paraffin embedded invasive ovarian epithelial tumor tissues. Loss of heterozygosity was observed more frequently at the loci defined by marker D6S473 (14 of 32 informative cases, 44%) and marker D6S448 (17 of 40 informative cases, 43%). Detailed mapping of chromosome 6q25-q26 in these tumor samples identified a 4 cM minimal region of LOH between markers D6S473 and D6S448 (6q25.1-q25.2). Loss of heterozygosity at D6S473 correlated significantly both with serous versus non-serous ovarian tumors (P=0.040) and with high grade versus low grade specimens (P=0.023). The results suggest that a 4 cM deletion unit located at 6q25.1-q25.2 may contain the putative tumor suppressor gene which may play a role in the development and progression of human invasive epithelial ovarian carcinomas (IEOC).     

Reduced exploratory activity of audiogenic seizures susceptible Wistar rats

To search for the existence of a tumour-suppressor gene (TSG) associated with oral squamous cell carcinoma (SCC), PCR analysis of microsatellite polymorphisms corresponding to 14 loci which map to chromosome 7q21.3-qter was performed to screen 35 patients with oral SCC for loss of heterozygosity (LOH). LOH was observed in at least one of the loci in 19 of 34 (55.9%) informative cases. Among the loci tested, frequent LOH was restricted at D7S522 on chromosome 7q31.1, which was measured within 1 cM. Furthermore, we detected microsatellite instability (MI) in 11 of 35 (31.4%) cases tested. Our observations indicate that alterations of chromosome 7q are associated with oral SCC tumorigenesis and that 7q31.1 might harbour at least one putative TSG.     

Allelotype of pediatric rhabdomyosarcoma

An allelotype covering all autosomes was constructed for the embryonal form of childhood rhabdomyosarcoma (ERMS) in order to identify regions encompassing tumorsuppressor genes (TSG) involved in ERMS. Thusfar most studies were focussed on chromosome 11p15.5, which frequently shows loss of heterozygozity (LOH) in embryonal tumors like RMS and Wilms' tumor (WT). In this study we show that, besides LOH of chromosome 11p15.5 (72%), LOH of chromosome 16q was present in 54% of the tumors analysed. Delineation of these two regions shows that the smallest region of overlap (SRO) for chromosome 11 was between D11S988 and D11S922. This region, estimated to be 7 cM and 3-5 Mb, is also the location of the putative Wilms' tumor WT2 TSG. It contains several genes including IGF2 and potential tumorsuppressor genes like H19 and p57kip2, which might contribute to the carcinogenesis of RMS. Analysis of chromosome 16q LOH defined the SRO between D16S752 and D16S413. LOH of chromosome 16 is also found in other tumors, including WT. Our data suggest that genes involved in the development of RMS and WT may not only be similar for chromosome 11 but also for chromosome 16.     

The relative bioavailability and in vivo-in vitro correlations for four marketed carbamazepine tablets

Neuroblastoma is a heterogeneous childhood tumour of the sympathetic nervous system, in which deletions of chromosomal region 1p and amplification of the MYCN oncogene correlate with aggressive tumour behaviour. However, the majority of neuroblastoma tumours show neither of these aberrations, indicating that other chromosomal regions may be involved in tumorigenesis. Here, we report findings of loss of heterozygosity (LOH) on chromosome 3. In our neuroblastoma material, nine of 59 (15.3%) tested tumours showed allelic loss of chromosome 3p markers. We found significant clinical and biological differences between tumours with the loss of one entire chromosome 3 vs tumours with partial loss in chromosome region 3p. All children with tumours with whole chromosome 3 loss are long-term survivors, whereas all children with tumours showing partial 3p LOH have died from tumour progression. A consensus region found to be deleted in all the tumours with 3p deletions was defined by markers D3S1286 and D3S1295, i.e. 3p25.3-p14.3, distal to the FHIT gene.     

A method of dynamic foot-pressure measurement for the evaluation of pediatric orthopaedic foot deformities

We analysed 42 differentiated thyroid tumors including 15 follicular adenomas (FA), 13 papillary thyroid cancers (PTC) and 14 follicular thyroid carcinomas (FTC) with 13 microsatellite markers specific for the long arm of human chromosome 7 within 7q31; this region is deleted frequently in several other tumor types. Overall, 20 of the 42 samples analysed (48%) displayed LOH with one or more of the markers tested. LOH was detected most frequently (78%) in FTC, the most malignant of the thyroid tumors. A smallest common deleted region (SCDR) was defined in this tumor type flanked by markers D7S480 and D7S490. This SCDR is distinct from D7S522, the most commonly deleted locus in many other tumors, which was deleted in only one FTC. D7S522 did show LOH in two of six informative PTCs. None of the PTC and only two of the FAs showed LOH in the FTC SCDR. Since FA is considered a premalignant stage of FTC, our results suggest that inactivation of a putative tumor suppressor at 7q31.2 may be acquired during adenoma to carcinoma progression. The absence of LOH at this locus amongst PTC suggests that inactivation of this tumor suppressor is specific for FTC. In conclusion, LOH at 7q31 is a frequent event in differentiated thyroid cancer, and we have defined a 2 cM SCDR specific for FTC.     

Definition and refinement of a region of loss of heterozygosity at 11q23.3-q24.3 in epithelial ovarian cancer associated with poor prognosis

Previous cytogenetic and loss of heterozygosity (LOH) data suggest that disruption of chromosome 11q23-qter occurs frequently in epithelial ovarian cancer and is associated with an adverse clinicopathological phenotype. Ten polymorphic microsatellite repeat loci were analyzed by PCR from the 11q22-q25 region between D11S35 and D11S968 in 40 ovarian tumors (including 31 epithelial ovarian cancers). Two distinct regions of loss were detected, suggesting possible sites for genes involved in epithelial ovarian neoplasia: a large centromeric region between D11S35 and D11S933 (11q22-q23.3) and a telomeric 8.5-Mb region lying between D11S934 and D11S1320 (11q23.3-24.3) not previously defined. LOH of the latter region but not the former one was significantly associated with poor survival, despite all tumors in this study having LOH somewhere on chromosome 11. This analysis provides a starting point for positional cloning.     

Detailed deletion mapping of the long arm of chromosome 6 in adult T-cell leukemia

Previously, we have found that the loss of heterozygosity (LOH) was frequently observed on chromosome 6q in acute/lymphoma-type adult T-cell leukemia (ATL), suggesting a putative tumor-suppressor gene for ATL may be present on chromosome 6q. To further define a region containing this gene, we performed fine-scale deletional mapping of chromosome 6q in 22 acute/lymphomatous ATL samples using 24 highly informative microsatellite markers. LOH was found in 9 samples (40. 9%) at 1 or more of the loci examined. Of the 9 samples, 8 shared the same smallest commonly deleted region flanked by D6S1652 and D6S1644 (6q15-21). The genetic distance between these two loci is approximately 4 cM. These results suggest that a putative tumor-suppressor gene on chromosome 6q15-21 probably plays a very important role in the evolution of acute/lymphomatous ATL. Our map provides key information toward cloning the gene.     

Loss of heterozygosity on the long arm of human chromosome 7 in sporadic renal cell carcinomas

Cytogenetic and molecular analysis of DNA sequences with highly polymorphic microsatellite markers have implicated allele loss in several chromosomal regions including 3p, 6p, 6q, 8p, 9p, 9q, 11p and 14q in the pathogenesis of sporadic renal cell carcinomas (RCCs). Deletions involving the long arm of chromosome 7 have not been described in RCCs although they have been seen in several other tumor types. However, there have been no detailed analysis of loss of heterozygosity (LOH) of 7q sequences in sporadic RCCs. We therefore studied LOH for DNA sequences on 7q with 10 highly polymorphic markers in 92 matched normal/tumor samples representing sporadic RCCs including papillary, nonpapillary, and oncocytomas in order to determine whether allelic loss could be detected in a tumor type with no visible 7q rearrangements at the cytogenetic level. We found chromosome 7q allele loss in 59 of 92 cases (64%) involving one, two, or more microsatellite markers. The most common allele loss included loci D7S522 (24%) and D7S649 (30%) at 7q31.1-31.2, a region that contains one of the common fragile sites, FRA7G. By comparative multiplex PCR analysis, we detected a homozygous deletion of one marker in the 7q 31.1-31.2 region in one tumor, RC21. These results support the idea that a tumor suppressor gene in 7q31 is involved in the pathogenesis of sporadic renal cell carcinomas.     

Analysis of PTEN and the 10q23 region in primary prostate carcinomas

Deletions involving chromosome 10q23 occur frequently in prostatic carcinomas. Recently, a novel tumour suppressor gene, PTEN, mapping to this interval, has been identified. Mutation or deletion of PTEN has been observed in a proportion of prostate cancer cell lines; however, primary prostate carcinomas have not been studied. We have investigated the involvement of PTEN in primary prostatic adenocarcinomas using a panel of 51 matched normal and prostate tumour DNAs. We first determined the proportion of tumours with allele loss at loci in 10q23 which span the region containing the PTEN gene. Our results show that LOH involving 10q23 is common in primary prostate carcinomas. Twenty-five of 51 (49%) tumours showed loss of heterozygosity (LOH) over the region spanning the PTEN locus. We next directly analysed the PTEN gene for mutations of the coding region using single strand conformation polymorphism (SSCP) and sequence analyses. Of those tumours with LOH, only a single tumour was found to carry a missense mutation in PTEN. No mutations in PTEN were identified in tumours without LOH. Our results suggest either that mutation of PTEN is a late event in prostate tumorigenesis, or that another tumour suppressor gene important in prostate cancer may lie close to PTEN in 10q23.     

Human bladder cancers and normal bladder mucosa present the same hot spot of heterozygous chromosome-9 deletion

Loss of heterozygosity (LOH) on chromosome 9 is the most frequent genetic alteration in bladder cancer identified to date, suggesting the presence of key gene(s) for this pathology. In this study, we examined 44 bladder tumors and 21 normal bladder samples for LOH on both arms of chromosome 9. Sixteen microsatellite markers, 12 on the short arm (encompassing 9p21-22) and 4 on the long arm (encompassing 9q33-34), were chosen for their highly frequent alterations in bladder cancer. LOH for at least one marker was identified in 42 tumor samples (95.5%), and 14 tumors (32%) displayed LOH for all informative tested markers. Detailed analysis showed that 2 markers on chromosome 9p (D9S157 and D9S156) had the highest frequencies of allelic loss (about 70%), independent of tumor grade and stage. The same study was performed on the 21 normal bladder mucosa samples: 50% of informative cases presented a single specific LOH at the D9S156 locus. Normal samples showing LOH at this locus were therefore screened with 3 novel microsatellite markers in the 810-kb region incorporating D9S156. Using this marker, we found no further heterozygous loss in this region. This result allows different interpretations of the D9S156 loss in normal bladder mucosa, and suggests that D9S156 may be more an indicator of bladder epithelium impairment than a tumor-initiation marker. Similarly, this unexpected result calls in question the interpretation of LOH studies.     

Carbohydrate-deficient glycoprotein syndrome type I: determination of the oligosaccharide structure of newly synthesized glycoproteins by analysis of calnexin binding

Familial predisposition to basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) of the skin are apparent in the autosomal dominant syndromes naevoid basal cell carcinoma syndrome (NBCCS) and multiple self-healing squamous epitheliomata (MSSE) respectively. The gene responsible for NBCCS has been proposed to be a tumour-suppressor gene and is mapped to the same 2 Mb interval on 9q22.3 as the MSSE gene ESS1. In an attempt to further map the NBCCS gene, we have examined loss of heterozygosity (LOH) in 16 sporadic BCCs and two familial BCCs using microsatellite markers located within the candidate gene region. The overall frequency of LOH observed was 67% in the BCCs and partial or interstitial deletions were found in eight tumours, with the highest LOH frequency at markers D9S280, D9S287 and D9S180. To determine if the same genomic region also shows frequent LOH in tumours with a squamous phenotype, we have examined 11 SCCs, four actinic keratoses and 13 cases of Bowen's disease for LOH at 9q22.3. An overall LOH frequency of 50% was observed at D9S180, and occurred in all types of squamous tumours. In contrast, a much lower LOH frequency of only 6% was found at the D9S287 locus. Our observation of different patterns of LOH at 9q22.3 in sporadic BCCs and SCCs implies that more than one tumour-suppressor gene might be located in this genomic region.     

Deletion mapping of gliomas suggest the presence of two small regions for candidate tumor-suppressor genes in a 17-cM interval on chromosome 10q

The loss of genetic material on chromosome 10q is frequent in different tumors and particularly in malignant gliomas. We analyzed 90 of these tumors and found loss of heterozygosity (LOH) in >90% of the informative loci in glioblastoma multiforme (GBM). Initial studies restricted the common LOH region to 10q24-qter. Subsequently, the study of a pediatric GBM suggested D10S221 and D10S209, respectively, as centromeric and telomeric markers of a 4-cM LOH region. It is interesting to note that, in one subset of cells from this tumor, locus D10S209 seems involved in the allelic imbalance of a larger region, with D10S214 as telomeric marker. This 17-cM region contains the D10S587-D10S216 interval of common deletion recently defined on another set of gliomas.     

Effect of prednisolone on IgE-dependent biphasic cutaneous reaction in BALB/c mice

We have examined a series of 24 Merkel cell carcinoma (MCC) DNAs for loss of heterozygosity (LOH) at eight loci on chromosome 13. All patients were heterozygous for at least one locus. Overall, 18 of 24 (75%) patients showed LOH, among whom 10 patients demonstrated LOH at all informative loci. A single common region of loss was identified in all cases and included the marker D13S233 (13q14.3), which maps close to the retinoblastoma susceptibility gene RB1. The RB1 protein was not detected by Western blot analysis in any of nine MCC cell lines tested. These data indicate that 13q losses are the most common chromosomal losses observed to date in MCC and the likely target of these deletions is the RB1 locus.     

Increased expression of the NME1 gene is associated with metastasis in epithelial ovarian cancer

The genetic events involved in the development of metastases of epithelial ovarian cancer are largely unknown. One gene postulated to play a role in tumour metastasis suppression is NME1 (nm23-H1), and an inverse relationship between NME1 expression and metastatic potential has been observed for some solid tumours. In this study we have investigated the levels of mRNA expression of the 2 isoforms of the NME gene, NME1 and NME2. A maximum of 45 tumours samples from 33 patients were available for Northern blot analysis. We observed variable levels expression of NME1 and NME2 mRNA. The average level of NME1, but not NME2, mRNA expression was statistically higher in metastatic biopsies when compared with primary tumour biopsies. To examine the possible tumour suppressor gene role of NME1 in ovarian tumours, 76 patients were investigated by Southern blot analysis to determine the rate of allelic deletion. Allele loss at 5 other chromosome 17 loci (D17S5, TP53, NF1, D17S74, D17S4) was also evaluated for many of these 76 patients. Allele loss was observed in 22/30 (73%) informative patients at the NME1 locus. We also observed high rates of allele loss at the other loci evaluated. No correlations with clinical stage, histological subtype or patient survival were observed in either mRNA or DNA analyses. We have established that tumour progression in ovarian cancer is accompanied by over-expression of the NME1 gene; however, despite high rates of allele loss at the NME1 locus, the concept that NME1 may be a candidate tumour suppressor gene in ovarian cancer cannot be confirmed by this study.     

Neokyotorphin formation and quantitative evolution following human hemoglobin hydrolysis with cathepsin D

Gains of chromosome 7 and alterations of the 7q-arm have been frequently observed in multiple cancers using various cytogenetic and molecular genetic techniques. Using PCR analysis of microsatellite markers, we have previously reported that allelic imbalance of 7q31 is common in prostate cancer and is associated with higher tumor grade and advanced pathological stage. In an effort to better understand the chromosome 7 alterations in prostate cancer, we undertook a molecular cytogenetic study of 25 prostate specimens using fluorescence in situ hybridization (FISH) with DNA probes for the chromosome 7 centromere and for 5 loci mapped to 7q31 (D7S523, D7S486, D7S522, D7S480, and D7S490) and 1 locus at 7q11.23 (ELN). Six tumors had no apparent anomaly for any chromosome 7 probe. Nine tumors showed apparent simple gain of a whole chromosome 7, whereas one tumor had apparent simple loss of a whole chromosome 7. Four tumors had gain of the chromosome 7 centromere and additional overrepresentation of the 7q-arm. One tumor had overrepresentation of 7q31 without any apparent anomaly of the chromosome 7 centromere, and one tumor had apparent loss of the chromosome 7 centromere with no apparent anomaly of the 7q-arm. Three tumors had gain of the chromosome 7 centromere and loss of the 7q31 region. Gain of 7q31 was strongly correlated with tumor Gleason score. Multiplex PCR studies of these specimens supported these FISH observations. Mutation screening and DNA sequencing of the MET gene, which is mapped to 7q31, revealed only the presence of simple sequence polymorphisms but no apparent acquired disease-associated mutations. FISH analysis of metaphases from an aphidicolin-induced, chromosome 7 only, somatic cell hybrid demonstrated that the DNA probe for D7S522 spans the common fragile site FRA7G at 7q31. Our data indicate that the 7q-arm, particularly the 7q31 region, is genetically unstable in prostate cancer, and some of the gene dosage differences observed may be due to fragility at FRA7G.     

Frequent allelic losses on chromosome 13q in human male breast carcinomas

Loss of genetic material on chromosomes 13q and 17 has been suggested to be of importance in the initiation and progression of female breast cancer, but their involvement is less well illustrated in male breast carcinomas. The present study was designed to investigate the incidence of allelic loss and microsatellite instability for chromosomes 13q, 17p and 17q in 13 sporadic male breast carcinomas using matched normal-tumour DNA samples and seven polymorphic microsatellite markers. Genetic imbalance was found in one or more informative markers in 85% of the patients, with more frequent loss of heterozygosity and microsatellite instability at loci on chromosome 13q. Thus, a high incidence of allelic losses was observed at the retinoblastoma gene (4/6) and likewise at the D13S263 locus (7/12), which also exhibited the highest frequency of microsatellite instability. The intragenic microsatellite in intron 1 of the TP53 gene on chromosome 17p revealed loss of heterozygosity in 3 of 8 informative patients. The investigated proximal region of chromosome 13q is postulated to harbour several potential tumour suppressor genes associated with female breast cancer. The high incidence of allelic losses at the D13S263 microsatellite, located distal to both the BRCA2 and the Brush-1 loci but proximal to the retinoblastoma gene, possibly indicates the presence of an additional tumour suppressor gene which may be involved in male breast carcinomas. However, this hypothesis needs verification in an extended study of male breast carcinomas.     

Genetic imbalance on chromosome 17 in papillary serous carcinoma of the peritoneum

We extend the evaluation of allelic loss patterns on chromosome 17 to papillary serous carcinoma of the peritoneum (PSCP) which is histologically identical to papillary serous ovarian carcinoma (PSOC). DNA was obtained from 11 archival cases of PSCP, with 1-11 tumor sites per case. Using ten loci spanning chromosome 17, loss of heterozygosity (LOH) was identified in all 11 cases (100%). Furthermore, 75-100% of informative cases exhibited LOH at the loci p53, D17S1322 (intragenic to the tumor suppressor gene BRCA1), D17S1327 and MPO. PSCP cases exhibit a higher rate of LOH at most loci when compared with PSOC. Alternating allelic loss at different tumor sites was identified in three cases supporting a multifocal origin of PSCP. Microsatellite instability (MI) is an uncommon event which was identified in four cases. These data implicate chromosome 17 as a potential location of genetic events important in the pathogenesis of PSCP as well as ovarian cancer.     

Frequent loss of heterozygosity on the long arm of chromosome 6: identification of two distinct regions of deletion in childhood acute lymphoblastic leukemia

Cytogenetic analysis of childhood acute lymphoblastic leukemia (ALL) identified nonrandom chromosomal abnormalities of the long arm of chromosome 6. Most of the alterations are deletions that are thought to be indicative of the presence of a tumor suppressor gene that is mutated on the remaining allele. These observations led us to consider whether 6q loss may contribute to the pathogenesis of childhood ALL. To define further a region containing this gene, we analyzed the loss of heterozygosity (LOH) of chromosome 6 in 113 primary ALL samples with matched normal DNA using 34 highly informative microsatellite markers. LOH was found in 17 (15%) samples at one or more of the loci, and partial or interstitial deletions of 6q were detected in 11 of these tumors. On the basis of these results, we performed a detailed deletional map and identified two distinct regions of deletion. The first region is flanked by D6S283 and D6S302 loci at 6q21-22. The second region is flanked by D6S275 and D6S283 loci at 6q21. Clinical analysis determined that LOH of 6q was demonstrated both in precursor-B cell ALLs (15 of 93; 16%) and in T cell ALLs (2 of 19; 11%). In addition, 19 patients have been studied at diagnosis and relapse; 18 showed the same 6q21-22 structural abnormality at relapse (normal, 16 patients; LOH, 2 patients) as their initial presentation, suggesting, albeit with a small patient sample size, that 6q21-22 deletions may be an initial event in leukemogenesis and may occur less frequently during the progression of childhood ALL. These data suggest the presence of putative tumor suppressor genes on chromosome arm 6q that are important in the development of both T and precursor-B childhood ALLs. Our map provides important information toward cloning putative ALL tumor suppressor genes.     

Loss of heterozygosity at 7q31 is a frequent and early event in prostate cancer

It is widely accepted that an accumulation of genetic alterations plays an important role in the genesis of human cancers, but little is known about prostate cancer in this respect. Recent studies have identified regions on chromosome arms 8p, 10q, 16q, and 18q that are frequently deleted in human prostate cancer. We have previously described a loss of heterozygosity (LOH) at the Met locus on chromosome band 7q31 in a study of 20 localized prostate tumors. To determine whether a region on the 7q arm is important in the initiation and/or progression of prostate cancer, prostate tissue from 13 patients with confined prostate tumors, 17 with local extracapsular extension, and 13 with metastatic forms were analyzed for LOH, using a DNA probe for RFLP (pMetH) and 8 CA microsatellite repeats (7 on 7q21-q33 and 1 on 7p). Twenty (47%) of the 43 cases studied showed LOH at one or more 7q loci. The most frequently deleted region was chromosome 7q31.1-7q31.2, whereas the centromeric locus on 7q21 was generally conserved. The percentage of LOH was normally distributed around the D7S480 locus. Moreover, the rate of LOH in the 7q31 region was lower in metastatic tumors than in localized tumors. These results strongly suggest the presence of a tumor suppressor gene on the chromosome band 7q31 with an important role in the early stages of prostate cancer.     

Allelic imbalance study of 16q in human primary breast carcinomas using microsatellite markers

The high incidence of allelic imbalance on the long arm of chromosome 16 in breast cancer suggests its involvement in the development and progression of the tumor. Several loss of heterozygosity (LOH) studies have led to the assignment of commonly deleted regions on 16q where tumor suppressor genes may be located. The most recurrent LOH regions have been 16q22.1 and 16q22.4-qter. The aim of this study was to gain further insight into the occurrence of one or multiple "smallest regions of overlap" on 16q in a new series of breast carcinomas. Hence, a detailed allelic imbalance map was constructed for 46 sporadic breast carcinomas, using 11 polymorphic microsatellite markers located on chromosome 16. Allelic imbalance of one or more markers on 16q was shown by 30 of the 46 tumors (65%). Among these 30 carcinomas, LOH on the long arm of chromosome 16 was detected at all informative loci in 19 (41%); 13 of them showed allelic imbalance on the long but not on the short arm, with the occurrence of variable "breakpoints" in the pericentromeric region. The partial allelic imbalance in 11 tumors involved either the 16q22.1-qter LOH region or interstitial LOH regions. A commonly deleted region was found between D16S421 and D16S289 on 16q22.1 in 29 of the 30 tumors. The present data argue in favor of an important involvement of a tumor suppressor gene mapping to 16q22.1 in the genesis or progression of breast cancer.