Irinotecan and radiation in vitro and in vivo

Loss of chromosome sequences at 13q14 (Rbl) and 17p13 (p53) associated with squamous cell carcinoma of head and neck (SCCHN) was evaluated in 12 recurrent tumors and 51 primary tumors from 63 patients. The incidence of LOH at 17p13 was 19 of 50 (38%) tumors, and at 13q14 was 21 of 57 (37%). LOH affecting Rbl and/or p53 was observed in 30 of 63 (48%) SCCHN. Coincident LOH at Rbl and p53 was detected in 10 of 46 (22%) tumors. There were nine cases in which primary and metastatic tumors were obtained from the same patient. Of these, seven were informative and five of these (71%) manifested LOH at p53 in both primary and metastatic sites. Examination of Rbl in these same tumors showed LOH in six of the nine metastases, and of these six, only three revealed LOH in the primary tumor. LOH at p53 or Rbl alone showed no correlation with clinical outcome. However, tumors that manifested LOH at both loci were associated with poorer patient outcome and poorer histological differentiation.     

Loss of heterozygosity on chromosome 22q and 17p correlates with aggressiveness of meningiomas

According to reported cytogenetic studies, there is a significant association between chromosomal aberrations and aggressiveness in meningiomas. With the method of restriction fragment length polymorphism analysis (RFLP), we examined tumor specific LOH on chromosome 17p and 22q in 30 cases of intracranial meningiomas. There were eight cases of meningiomas with aggressive characteristics, such as invasive meningioma, malignant meningioma, hemangiopericytic meningioma, and multiple meningiomas with central neurofibromatosis. Twenty-five of 30 cases (83%) were constitutionally heterozygous for at least one of the chromosome 22q DNA markers and sixteen of 25 informative cases (64%) displayed loss of heterozygosity (LOH). All of the 8 informative cases (100%) of meningiomas with aggressive characteristics, showed LOH on chromosome 22q whereas non-aggressive cases revealed LOH in eight of 17 informative cases (47%). At the loci on chromosome 17p, only two cases of malignant meningionas showed LOH. Our results suggest that the inactivations of putative tumor suppressor genes on chromosome 22q and 17p may correlate with aggressiveness and malignant transformation of meningiomas.     

DNA microarray analysis identifies CKS2 and LEPR as potential markers of meningioma recurrence

Meningiomas are the most frequent intracranial tumors. Surgery can be curative, but recurrences are possible. We performed gene expression analyses and loss of heterozygosity (LOH) studies looking for new markers predicting the recurrence risk. We analyzed expression profiles of 23 meningiomas (10 grade I, 10 grade II, and 3 grade III) and validated the data using quantitative polymerase chain reaction (qPCR). We performed LOH analysis on 40 meningiomas, investigating chromosomal regions on 1p, 9p, 10q, 14q, and 22q. We found 233 and 268 probe sets to be significantly down- and upregulated, respectively, in grade II or III meningiomas. Genes downregulated in high-grade meningiomas were overrepresented on chromosomes 1, 6, 9, 10, and 14. Based on functional enrichment analysis, we selected LIM domain and actin binding 1 (LIMA1), tissue inhibitor of metalloproteinases 3 (TIMP3), cyclin-dependent kinases regulatory subunit 2 (CKS2), leptin receptor (LEPR), and baculoviral inhibitor of apoptosis repeat-containing 5 (BIRC5) for validation using qPCR and confirmed their differential expression in the two groups of tumors. We calculated ΔCt values of CKS2 and LEPR and found that their differential expression (C-L index) was significantly higher in grade I than in grade II or III meningiomas (p < .0001). Interestingly, the C-L index of nine grade I meningiomas from patients who relapsed in <5 years was significantly lower than in grade I meningiomas from patients who did not relapse. These findings indicate that the C-L index may be relevant to define the progression risk in meningioma patients, helping guide their clinical management. A prospective analysis on a larger number of cases is warranted.     

Loss of 14q31-q32.2 in renal cell carcinoma is associated with high malignancy grade and poor survival

The present study was undertaken to further approach the importance of 14q deletions in renal cell carcinoma (RCC) development. The initial screening using 2 RFLP markers from distal 14q identified loss of heterozygosity (LOH) in 17 of 45 informative cases (38%). In addition, in 37 patients with primary RCCs, it was shown that cases with LOH at D14S1 had significantly shorter survival as compared to cases with-out LOH (p<0.005). Subsequently, 19 primary tumors and 6 metastases were genotyped for 20 polymorphic markers and the findings were evaluated in relation to the clinical characteristics of the primary tumor and the survival during follow-up. Overall LOH was identified in 11 of the primary tumors (58%) and 4 of the metastases (66%). In metastases as well as in primary tumors the highest frequency of LOH was detected with markers from the distal part of the chromosome i.e., 14q32. Five minimal regions of overlapping deletions were identified, three of which (II, IV and V) were defined from the primary RCCs. From centromere to telomere these include region I proximal of D14S259, region II between D14S255 and D14S588, region III in the D14S61-D14S617 interval, region IV between D14S617 and D14S260, and region V telomeric of D14S1007. For the primary tumors, losses in regions IV and V were each significantly associated with high tumor grade (i.e., grade 3; p<0.05). Furthermore, LOH within region IV was also associated with a significantly shorter survival (p=0.02). In conclusion, the high frequency of distal 14q LOH supports the relevance of this alteration for the development of RCC.     

Genetic pattern of prostate cancer progression

Genetic alterations in primary prostate cancer (CaP) have been extensively studied, yet little is known about the genetic mechanisms underlying progression of primary CaP to metastatic prostate cancer. As a result, it is not possible to distinguish clinically indolent localized disease from potentially life-threatening tumors with high metastatic potential. To address this question, we collected tissue from 34 autopsy-derived metastases, samples rarely analyzed in previous studies. These were compared to a separate set of 17 prostatectomy specimens containing 22 foci of CaP associated with 49 examples of high-grade prostatic intraepithelial neoplasia (PIN), a histological precursor of CaP. We compared the loss of heterozygosity (LOH) profiles of high-grade PIN, primary CaP and metastases by analyzing 33 microsatellite markers previously found to have high frequencies of LOH in primary CaP. These markers were on chromosomes 5q, 6q, 7q, 8p, 9p, 10q, 11p, 13q, 16q, 17, 18q and 21q. In addition, markers on chromosomes 4p, 11q, 14q and 20q with no reported LOH in primary CaP were analyzed to determine the frequency of background LOH. In PIN lesions, the rate of LOH was significant only at D5S806 (20%) and D16S422 (29%). In addition, different PIN lesions within the same prostate gland were genetically diverse, indicating divergent evolution of synchronous neoplastic precursor lesions. LOH frequency was progressively higher in primary CaP and metastatic lesions. In primary CaP, significant losses occurred at the 8p, 10q, 11p, 16q, 17p, 18q and 21q loci (range 17-43%). Distinct patterns of LOH frequencies were observed in primary CaP compared with metastases. Although some loci (D16S422, D17S960, D21S156) showed similar frequencies of LOH in primary CaP and metastatic CaP, most other loci showed up to 7-fold metastasis-related increases. The metastatic samples revealed previously unrecognized prostate cancer LOH at D5S806, D6S262, D9S157, D13S133 and D13S227. These significant stage-specific differences in LOH frequency specify genetic loci that may play key roles in CaP progression and could represent clinically useful biomarkers for CaP aggressiveness.     

Accumulation of genetic alterations in brain metastases of sporadic breast carcinomas is associated with reduced survival after metastasis.

Tumor progression is characterized by stepwise accumulation of genetic alterations. To identify alterations associated with breast cancer metastasis, an analysis of comparative loss of heterozygosity (LOH) was performed on 38 primary sporadic breast carcinomas and 16 distant metastases. Two loci at 5q21 and 18q21 were chosen because of their reported increased deletion frequency in metastatic tumors. LOH at 17q21, 13q12-13, 17p13.1 and 11q22-23 was analyzed to determine whether there is a specific involvement of these breast cancer-associated gene loci in the metastatic process. Our data show that distant metastases are characterized by markedly increased LOH frequency at all loci examined. In both gene locus groups, significantly more distant metastases are affected by combined LOH. Furthermore, a significantly reduced postmetastatic survival time has been observed in patients with brain metastases affected by synchronous allelic loss at the four breast cancer-associated gene loci. Our results suggest that cumulative LOH of breast cancer-related gene loci is associated with a more aggressive phenotype of metastatic breast tumors.     

Allelic loss on chromosomes 1p32, 9p21, 13q14, 16q22, 17p,and 22q12 in meningiomas associated with meningioangiomatosis and pure meningioangiomatosis

Meningioangiomatosis (MA) is a rare lesion appearing sporadically or as a part of neurofibromatosis 2. The occurrences of meningiomas arising from MA (MA-M) have raised doubts about the traditional concept of a hamartomatous origin for MA. Cytogenetic or molecular studies on MA, with or without meningiomas, are limited because of the rarity of MA. The current study was to evaluate the loss of heterozygosity (LOH) in seven cases of MA-M and two cases of pure MA. LOH on six chromosomes (1p32, 9p21, 13q14, 16q22, 17p, and 22q12) were investigated using 13 sets of microsatellite markers, including D1S193, D1S463, D22S193, D22S929, D22S282, TP53, D17S796, D16S421, D16S512, D13S118, D13S153, D9S162, and D9S104. PCR was performed using each marker and polymorphic analysis was accomplished by silver staining. Immunohistochemical stain for Ki-67 was carried out and labeling index was measured by using a semiquantitative manual counting method. The meningioma portions of MA-Ms showed LOH for loci on chromosomes 22q12, 9p21, and 1p32 in 57.1% (4/7), 28.6% (2/7), and 28.6% (2/7) of cases, respectively. The MA portions of MA-M had a LOH for loci on 22q12 in 28.6% (2/7) of cases, whereas each pure MA harbored one LOH on either chromosome 22q12 or 9p21. The proliferation indices of MA-Ms were significantly higher in the meningioma than in the MA components. Our data suggest that both the meningioma and the MA undergo the same overlapping clonal process, with the MA-M while undergoing additional genetic alterations that confer a greater proliferative potential.     

Molecular Genetic Study of a Metastatic Oligodendroglioma

Extracranial spread of neuroectodermal tumors is an unusual event, most frequently expected from glioblastomas and medulloblastomas. Single cases of metastatic oligodendrogliomas have been described, but no genetic data are reported. Oligodendrogliomas are characterized by distinct genetic alterations, i.e. loss of heterozygosity (LOH) of 1p and 19q; therefore, molecular genetic analysis of metastatic cases is of considerable interest. It may be instrumental in defining the distant tumor as metastatic oligodendroglioma and give clues to the genetic events associated with the highly malignant transformation. We present the case of a patient with multiple bone metastases from a cerebral oligodendroglioma. Oligodendroglioma grade II was the diagnosis both at original and second operation, performed 7 and 1 years before the extracranial dissemination. The extraneural spread presented before the local intracranial recurrence. The patient received procarbazine, lomustine, vincristine chemotherapy and radiotherapy after the second surgery. The computed tomography-guided biopsy of the bone lesions revealed tumor cells positive for GFAP, S-100 and Leu-7 and negative for cytokeratin, LCA and EMA. The genetic analysis of DNA from the original tumor, the bone metastasis and the autoptic brain tumor showed LOH of 1p; heterozygous deletion of CDKN2A/p 16 was detected as additional alteration in the metastasis and in the intracranial tumor at autopsy. TP53, MDM2 and CDKN2A/p14ARF genes were unchanged. Repeated brain surgery and extended survival may have acted as promoter of extraneural dissemination. Loss of CDKN2A most probably played an important role in the malignant progression: its involvement in metastatic potential remains to be clarified. Our data confirm that malignant transformation of oliogodendrogliomas may be undetected by histology and underscore the importance of genetic analysis. Coincidentally with intensive anticancer therapy, chemotherapy included, employed in patients with oligodendroglioma and the ensuing long survival, the frequency of metastatic oliogodendrogliomas may increase.     

Concordant p53 and DCC alterations and allelic losses on chromosomes 13q and 14q associated with liver metastases of colorectal carcinoma

To identify genetic alterations associated with acquisition of metastatic ability in colorectal carcinoma, 31 liver metastases and 40 primary tumors of colorectal carcinoma from 55 patients were analyzed for loss of chromosomal heterozygosity using 46 polymorphic DNA markers covering 15 chromosomes. Loss of heterozygosity (LOH) and/or rearrangement at the TP53 and DCC loci were detected in all liver metastases (10 of 10 at TP53 and 19 of 19 at DCC), and were observed in 59% (10 of 17) at TP53 and 75% (18 of 24) at DCC respectively in the primary tumors. Furthermore, the incidence of LOH on chromosomes 13q and 14q was higher than that on other chromosomes in liver metastasis, and it was higher in liver metastases than in primary tumors (20/30 vs. 18/39, p = 0.072 on chromosome 13q and 21/31 vs. 16/40, p = 0.018 on chromosome 14q). In 4 cases, LOH or rearrangement at loci on chromosomes I3q. 14q and 18q not detected in primary tumors was observed in liver metastases from the same patients. These results suggest that concordant p53 and DCC alterations and inactivation of several other tumor-suppressor genes, especially those on chromosomes 13q and 14q, play important roles in the acquisition of metastatic potential of colorectal carcinoma.     

Chromosomal alterations in oligodendroglial tumours over multiple surgeries: is tumour progression associated with change in 1p/19q status?

BACKGROUND: Oligodendroglial neoplasms have morphologic and genotypic heterogeneity. Loss of heterozygosity (LOH) of 1p and/or 19q is associated with increased treatment responsiveness and overall survival. However, the pathogenesis of treatment-resistance is unknown. We sought to determine if tumour progression is due to a proliferating sub-population of tumour cells with intact 1p, or if recurrent tumours retain 1p/19q LOH. METHODS: 24 patients with oligodendroglial neoplasms, possessing biopsy samples taken at diagnosis and at progression, were identified. 53 tumour specimens were available for LOH analysis of 1p and 19q, using PCR amplification of multiple microsatellite markers. 40 were also tested for 9p and 10q. RESULTS: At diagnosis, the median age was 34 (24-66) years, 14 were male. 19 tumours were WHO Grade II, and 5 were high grade. The most common genomic status was 19q LOH (70%). 13 (54%) tumours were 1p LOH at diagnosis: of these, 12 were 19q LOH, and 1 was 19q uninformative. All 12 patients with 1p/19q LOH primary tumours had persistent co-deletion at progression. 9 (38%) tumours were 1p intact at diagnosis, and 8 remained 1p intact in the progressed tumours. There was little heterogeneity of 9p and 10q between tumours at diagnosis and progression. CONCLUSION: 100% of oligodendroglial tumours with 1p/19q LOH, demonstrated persistent 1p/19q LOH in the progressed tumour. Therefore, progression of these tumours is not due to a proliferating sub-population of treatment-resistant, 1p intact tumour cells. We propose that additional mutations contribute to this aggressive phenotype, however, 9p LOH or 10q LOH are unlikely to be involved.     

Loss of heterozygosity for chromosomes 1 or 14 defines subsets of advanced neuroblastomas.

Neuroblastomas have been characterized genetically by N-myc amplification and by deletions or loss of heterozygosity (LOH) for the short arm of chromosome 1. However, recent studies have suggested deletion or allelic loss involving at least three other chromosome arms, 11q, 14q, and 17p. Therefore, we undertook an analysis of allelic loss for these respective chromosomal arms to determine the frequency and pattern of LOH as well as the correlation of these findings with other biological and clinical variables. A group of 24 pairs of normal and tumor DNAs was chosen that were representative of patients of different ages and stages. A substantial frequency of LOH (greater than or equal to 20%) was found only for 1p and 14q, whereas LOH for the other chromosome arms occurred in less than or equal to 5% of cases. On the basis of these results, we extended the analysis to a total of 59 neuroblastomas, and we found 1p LOH in 15 of the 59 cases (25%) and 14q LOH in 10 of 43 informative cases (23%). N-myc amplification was found in 15 of the 59 cases (25%). This analysis confirmed that 1p LOH and 14q LOH occurred almost exclusively in patients with advanced stages of disease. Furthermore, LOH for 1p and 14q usually occurred independent of each other, and 1p LOH frequently was associated with N-myc amplification, whereas 14q LOH was not. Thus, our results demonstrate that neuroblastomas are complex genetically and that there are at least two distinct loci for putative suppressor genes that are deleted independently in this tumor, both of which are associated with advanced stages of disease.     

Deletion mapping of chromosomes 14q and 1p in human neuroblastoma.

It has been suggested that loss of heterozygosity (LOH) on the short arm of chromosome 1 is a critical event for the development of neuroblastoma, and we have previously shown frequent LOH on chromosome 14 in neuroblastoma. To pursue these observations, especially to define further the regions which are commonly deleted in the tumor, we examined for allelic losses in 27 cases of neuroblastomas by using a number of polymorphic DNA markers for chromosomes 14q and 1p. LOH was observed in 10 out of the 25 informative cases (40%) on chromosome 14q and in eight out of the 21 informative cases (38%) on 1p. The commonly deleted regions were distal to the D14S13 locus (14q32-qter) on chromosome 14 and distal to the D1S112 locus (1p36.1-pter) on chromosome 1. These results strongly suggest that tumor-suppressor genes important in the pathogenesis of human neuroblastoma are located on the distal part of both chromosomes 14q and 1p.     

Chromosomal alterations during lymphatic and liver metastasis formation of colorectal cancer

Comparative genomic hybridization (CGH) was used to screen colorectal carcinomas for chromosomal aberrations that are associated with metastatic phenotype. In total, 63 tumor specimens from 40 patients were investigated, comprising 30 primary tumors, 22 systemic metastases (12 liver, 6 brain, and 4 abdominal wall metastases) and 11 lymph node tumors. Using statistical analysis and histograms to evaluate the chromosomal imbalances, overrepresentations were detected most frequently at 20q11.2-20q13.2, 7q11.1-7q12, 13q11.2-13q14, 16p12, 19p13, 9q34, and 19q13.1-19q13.2. Deletions were prominent at 18q12-18q23, 4q27-4q28, 4p14, 5q21, 1p21-1p22, 21q21, 6q16-6q21, 3p12, 8p22-8p23, 9p21, 11q22, and 14q13-14q21. Hematogenous metastases showed more alterations than lymph node tumors, particularly more deletions at 1p, 3, 4, 5q, 10q, 14, and 21q21 and gains at 1q, 7p, 12qter, 13, 16, and 22q. Comparing liver metastases with their corresponding primary tumors, particularly deletions at 2q, 5q, 8p, 9p, 10q, and 21q21 and gains at 1q, 11, 12qter, 17q12-q21, 19, and 22q were more often observed. The analysis suggested that the different pathways of tumor dissemination are reflected by a nonrandom accumulation of chromosomal alterations with specific changes being responsible for the different characteristics of the metastatic phenotype.     

Prognostic impact of multiple allelic losses on metastatic recurrence in hepatocellular carcinoma after curative resection

Loss of heterozygosity (LOH) on chromosomes 13q, 16q and 17p has been associated with the progression of hepatocellular carcinoma (HCC). To investigate the prognostic impact of such LOH, we examined the metastasis-free survival of curatively resected HCC cases, in whom these LOHs were analyzed. Among the 49 HCC patients examined, the frequency of LOHs was 28% on 13q, 33% on 16q and 40% on 17p. The patients were followed up for metastatic recurrence after surgery and for analysis of the relationship between chromosomal changes and patients' metastasis-free survival. Univariate survival analysis showed the presence of LOH on 16q, 17p and the number of chromosomes with LOH were significantly and negatively associated with metastasis-free survival, indicating that patients with LOH on multiple chromosomes had a poorer prognosis after surgery than those with LOH on a single chromosome or no LOH. Multivariate Cox survival analysis identified the presence of LOH on 16q and the number of chromosomes with LOH as the most significant independent negatively predictive factors for metastasis-free survival. These findings indicate that accumulation of chromosomal changes is associated with metastatic behavior, and that LOH on 16q was the most useful prognostic indicator for metastasis after curative resection of HCC.     

Heterogeneity of allelic deletions within melanoma metastases

During the initiation and progression of malignant melanoma, a series of different genetic events accumulate on several different chromosomes. The biological heterogeneity of tumour cells presents a major problem, preventing effective treatment of melanoma. To examine the degree of genetic heterogeneity, we searched for allelic losses (loss of heterozygosity; LOH) on chromosomes 9p, 9q, 1p and 17p, examining different areas within human melanoma metastases. All of the examined metastases were informative within at least one dissected area for at least one marker. Out of 29 areas in 11 melanoma metastases, 58% showed LOH with at least one marker. On chromosome 9p21-22, eight out of 26 informative loci (31%) showed LOH at D9S171 (three not informative), two out of 18 (11%) at IFNA (11 not informative) and seven out of 24 (29%) at D9S169 (five not informative). LOH on chromosome 9q22.3 was examined by the microsatellite marker D9S12; three out of 24 areas (12.5%) showed LOH, and five were not informative. Deletions on chromosome 1p were assessed using D1S450. Four out of 25 (16%) showed LOH; four were not informative. Deletions on chromosome 17p13 were examined with TP53; two out of 21 cases (9%) showed LOH, and eight were not informative. Our data demonstrate an impressive heterogeneity of allelic losses in the investigated chromosomal areas within the same metastatic lesion. This suggests that there is not one specific genetic alteration that accounts for melanoma progression to metastases. Rather there seem to be multiple genetic alterations accumulating even on the same chromosome, and progression from melanoma to metastases is paralleled by the accumulation of clones harbouring multiple genetic abnormalities.     

Genome-wide allelotype analysis of sporadic primary nasopharyngeal carcinoma from southern China

Nasopharyngeal carcinoma (NPC) is one of the most common malignant tumors in Southern China, especially in the Guangdong area. To demonstrate a comprehensive profile of loss of heterozygosity (LOH) in NPC, we applied a large panel of 382 microsatellite polymorphism markers covering all the 22 autosomes in 98 cases of sporadic primary NPC. Of the 335 informative markers, 83 loci showed high level of LOH (presence in equal to or more than 30% cases) and most of the high frequent loci were clustered to chromosome 1p36 and 1p34, 3p14-p21, 3p24-p26, 3q25-q26 and 3q27, 4q31 and 4q35, 5q15-21 and 5q32-q33, 8p22-p23, 9p21-p23 and 9q33-q34, 11p12-p14, 13q14-q13 and 13q31-q32, 14q13-q11, 14q24-q23 and 14q32. High frequency of LOH was found in chromosomes 3, 5, 9 and 11 (>/=50%), while medium frequency of LOH was found in chromosomes 1, 4, 6, 14, 17 and 19 (40-49%). Several new regions showing high frequency of LOH were found in chromosome 1p36, 3q25-q26, 3q27, 5q15-q21, 8p22-p23 and 11p12-14. The relationship between LOH and TNM stage of NPC was evaluated. Regions 6p23 (D6S289), 8p23.1 (D8S549) and 9q34.2 (D9S1826) showed higher frequency of LOH in later stages (III and IV) than in earlier stages (I and II) (P<0.05). Thus, our study provides a global view on allelic loss in the development of NPC and should shed light on the way for localization of putative tumor suppressor genes associated with the pathogenesis of NPC.     

Allelic losses associated with the metastatic potential of colorectal-carcinoma

The aim of this study was to define the association of allelic losses with the metastatic potential of colorectal carcinoma and to determine whether allelic losses can be genetic markers for the prognosis of patients with colorectal carcinoma. Eighty primary colorectal tumors and 31 liver metastases from 95 patients were examined for loss of heterozygosity (LOH) at the APC, p53, RB, DCC and chromosome 14q loci by using polymerase chain reaction-single strand conformation polymorphism analysis and restriction fragment length polymorphism analysis. The incidence of LOH at the DCC and RB loci and on chromosome 14q in liver metastases was significantly higher than that in primary tumors. DCC and RB alterations were detected more frequently in primary tumors with higher metastatic potential. Although no statistically significant association was found between these losses and survival or distant metastasis, patients with DCC losses showed poorer survival by multivariate analysis (p=0.056). Thus, inactivation of the DCC and RB genes and gene(s) on chromosome 14q seem to be critical genetic events for the acquisition of metastatic potential in colorectal carcinoma. However, further studies will be required to utilize these genetic alterations as valuable prognostic markers.     

Destabilization of chromosome 9 in transitional cell carcinoma of the urinary bladder.

The most frequent genetic alteration in transitional cell carcinoma of the urinary bladder (TCC) is loss of chromosome 9 which targets CDKN2A on 9p. The targets on 9q are not confirmed. Here, 81 advanced TCC specimens were investigated for loss of heterozygosity (LOH) and homozygous deletions (HD) on chromosome 9q using multiplex analysis of microsatellite markers. 41/81 tumours (51%) showed LOH on 9q, with LOH at all markers in 33 cases. Eight partial losses involved three regions in 9q12, 9q22.3, and 9q33- 9q34. No mutations were identified in the candidate tumour suppressor gene DBCCR1 in three tumours showing restricted LOH at 9q32-33. 22% of the specimens had HD at CDKN2A, but no HD was found on 9q. Two tumours had lost 9p only and five 9q only. 9q LOH was not related to tumour grade or stage and present or absent with equal frequency in recurrent TCC. LOH on 9q correlated with the extent of genome-wide hypomethylation (P < 0.0001) which extended into satellite sequences located in 9q12 juxtacentromeric heterochromatin. While the high frequency of chromosome 9q loss in TCC may reflect destabilization of the chromosome related to hypomethylation of repetitive DNA, the data are compatible with the existence of tumour suppressor genes on this chromosome arm.     

Deletion Mapping of Chromosome 1p and 22q in Pheochromocytoma

To identify the localization of tumor suppressor genes, 22 pheochromocytomas (9 hereditary and 13 sporadic) were examined for loss of heterozygosity (LOH) on the short arm of chromosome 1 and on the long arm of chromosome 22 by using 11 polymorphic DNA markers on each chromosome arm. LOH on 1p was observed in 12 of 22 informative cases (55%) and on 22q in 8 of 20 informative cases (40%). There was no significant difference in the frequency of LOH on 1p or 22q between hereditary and sporadic cases. We could localize the commonly deleted regions as distal to D1S73 and proximal to D1S63 on 1p and distal to D22S24 and proximal to D22S1 on 22q. In addition, the relationship between LOH on 1p and 22q was studied in 20 pheochromocytomas which were informative for probes on both chromosome arms. Of eight tumors that showed LOH on 22q, allelic loss on 1p was also detected in seven. Thus, LOH on 22q was correlated significantly with LOH on 1p ( P = 0.0249; Fisher's exact test). These results suggest that inactivation of multiple tumor suppressor genes may be required for development and progression of hereditary and non-hereditary pheochromocytoma.     

NF2 gene mutations and allelic status of 1p, 14q and 22q in sporadic meningiomas.

Formation of meningiomas and their progression to malignancy may be a multi-step process, implying accumulation of genetic mutations at specific loci. To determine the relationship between early NF2 gene inactivation and the molecular mechanisms that may contribute to meningioma tumor progression, we have performed deletion mapping analysis at chromosomes 1, 14 and 22 in a series of 81 sporadic meningiomas (54 grade I (typical), 25 grade II (atypical) and two grade III (anaplastic)), which were also studied for NF2 gene mutations. Single-strand conformational polymorphism analysis was used to identify 11 mutations in five of the eight exons of the NF2 gene studied. All 11 tumors displayed loss of heterozygosity (LOH) for chromosome 22 markers; this anomaly was also detected in 33 additional tumors. Twenty-nine and 23 cases were characterized by LOH at 1p and 14q, respectively, mostly corresponding to aggressive tumors that also generally displayed LOH 22. All three alterations were detected in association in seven grade II and two grade III meningiomas, corroborating the hypothesis that the formation of aggressive meningiomas follows a multi-step tumor progression model.