Xp22.33p22.13 Duplication in a Male Patient Carrying a Recombinant X Chromosome Derived from an Inherited Intrachromosomal Insertion.

Intrachromosomal insertions are complex structural rearrangements that are challenging to interpret using classical cytogenetic methods. We report a male patient carrying a recombinant X chromosome derived from a maternally inherited intrachromosomal insertion. The patient exhibited developmental delay, intellectual disability, behavioral disorder, and dysmorphic facial features. To accurately identify the rearrangements in the abnormal X chromosome, additional cytogenetic studies were conducted, including fluorescence in situ hybridization (FISH), multicolor-banding FISH, and array comparative genomic hybridization. The results showed a recombinant X chromosome, resulting in a 13.05 Mb interstitial duplication of segment Xp22.33-Xp22.13, which was inserted at cytoband Xq26.1. The duplicated region encompasses 99 genes, some of which are associated with the patient's clinical manifestations. We propose that the combined effects of the Xp-duplicated genes may contribute to the patient's phenotype.

Correction to: Use of multicolor fluorescence in situ hybridization to detect deletions in clinical tissue sections.

Figure 2 is incorrect in the original version of this article. The correct figure 2 is provided below.

Use of multicolor fluorescence in situ hybridization to detect deletions in clinical tissue sections.

A variety of laboratory methods are available for the detection of deletions of tumor suppressor genes and losses of their proteins. The clinical utility of fluorescence in situ hybridization (FISH) for the identification of deletions of tumor suppressor genes has previously been limited by difficulties in the interpretation of FISH signal patterns. The first deletion FISH assays using formalin-fixed paraffin-embedded tissue sections had to deal with a significant background level of signal losses affecting nuclei that are truncated by the cutting process of slide preparation. Recently, more efficient probe designs, incorporating probes adjacent to the tumor suppressor gene of interest, have increased the accuracy of FISH deletion assays so that true chromosomal deletions can be readily distinguished from the false signal losses caused by sectioning artifacts. This mini-review discusses the importance of recurrent tumor suppressor gene deletions in human cancer and reviews the common FISH methods being used to detect the genomic losses encountered in clinical specimens. The use of new probe designs to recognize truncation artifacts is illustrated with a four-color PTEN FISH set optimized for prostate cancer tissue sections. Data are presented to show that when section thickness is reduced, the frequency of signal truncation losses is increased. We also provide some general guidelines that will help pathologists and cytogeneticists run routine deletion FISH assays and recognize sectioning artifacts. Finally, we summarize how recently developed sequence-based approaches are being used to identify recurrent deletions using small DNA samples from tumors.

Assessing the cost of implementing the 2011 Society of Obstetricians and Gynecologists of Canada and Canadian College of Medical Genetics practice guidelines on the detection of fetal aneuploidies.

BACKGROUND: The Society of Obstetricians and Gynecologists of Canada and the Canadian College of Medical Genetics published guidelines, in 2011, recommending replacement of karyotype with quantitative fluorescent polymerase chain reaction when prenatal testing is performed because of an increased risk of a common aneuploidy. STUDY OBJECTIVE: This study's objective is to perform a cost analysis following the implementation of quantitative fluorescent polymerase chain reaction as a stand-alone test. RESULTS: A total of 658 samples were received between 1 April 2014 and 31 August 2015: 576 amniocentesis samples and 82 chorionic villi sampling. A chromosome abnormality was identified in 14% (93/658) of the prenatal samples tested. The implementation of the 2011 Society of Obstetricians and Gynecologists of Canada and the Canadian College of Medical Genetics guidelines in Edmonton and Northern Alberta resulted in a cost savings of $46 295.80. The replacement of karyotype with chromosomal microarray for some indications would be associated with additional costs. CONCLUSION: The implementation of new test methods may provide cost savings or added costs. Cost analysis is important to consider during the implementation of new guidelines or technologies. © 2017 John Wiley & Sons, Ltd.

Erratum to: Familial chromosomal translocation X; 22 associated with infertility and recurrent X mosaicism.

[This corrects the article DOI: 10.1186/s13039-016-0249-5.].

Familial chromosomal translocation X; 22 associated with infertility and recurrent X mosaicism.

BACKGROUND: Individuals with apparently balanced translocations, often, show no clinical findings. However, in meiosis, translocations tend to cause errors on chromosome disjunction and the ones involving sex chromosomes have particular implications for the phenotype. Male carriers of balanced X-autosome translocations are almost invariably infertile due to interruption of the spermatogenesis, but the mechanism is not fully understood. CASE PRESENTATION: In this case report, we performed a combination of classical cytogenetics (G-banding), molecular cytogenetics (fluorescence in situ hybridization and X-chromosome inactivation study), and cytogenomics (microarray-based comparative genomic hybridization) techniques for characterization of an inherited (X;22) translocation in a family originally referred for infertility investigation. Both proband and his sister are infertile and present the maternally inherited translocation. Interestingly, the maternal grandmother was mosaic for X chromosome monosomy suggesting that the t(X;22) in the proband's mother arose by errors at oogenesis. The presence of the same mosaicism of the X chromosome in the proband's aunt is consistent with this consideration. Array- CGH analysis showed no constitutional pathogenic gains or losses in the translocation carriers. The X-chromosome inactivation studies revealed that the translocated X;22 was active in 99.3% of cells in the mother and in 88% of cells in the daughter. We suggest that incomplete skewing of X inactivation (>97 %) of the daughter could justify the infertility. This study is the first description of recurrent mosaicism of the X chromosome associated with a familial X-autosome translocation. CONCLUSIONS: The phenotype of infertility was probably caused by disruption of spermatogenesis due to gametogenesis specific errors resulted from meiotic pairing and segregation anomalies on the translocated chromosomes.

PTEN losses exhibit heterogeneity in multifocal prostatic adenocarcinoma and are associated with higher Gleason grade.

Prostatic adenocarcinoma is an epithelial malignancy characterized by marked histological heterogeneity. It most often has a multifocal distribution within the gland, and different Gleason grades may be present within different foci. Data from our group and others have shown that the genomic deletion of the phosphatase and tensin homolog deleted on chromosome 10 (PTEN) tumor suppressor gene and the disruption of the ETS gene family have a central role in prostate cancer and are likely to be associated with Gleason grade. In this study, prostate cancer samples were systematically analyzed to determine whether there was concordance between PTEN losses and TMPRSS2-ERG fusion rearrangements, within or between foci in multifocal disease, using well-annotated tissue microarrays (TMAs) consisting of 724 cores derived from 142 radical prostatectomy specimens. Three-color fluorescence in situ hybridization analysis of both the PTEN deletion and the TMPRSS2-ERG fusion was used to precisely map genetic heterogeneity, both within and between tumor foci represented on the TMA. PTEN deletion was observed in 56 of 134 (42%) patients (hemizygous=42 and homozygous=14). TMPRSS2-ERG fusion was observed in 63 of 139 (45%) patients. When analyzed by Gleason pattern for a given TMA core, PTEN deletions were significantly associated with Gleason grades 4 or 5 over grade 3 (P<0.001). Although TMPRSS2-ERG fusions showed a strong relationship with PTEN deletions (P=0.007), TMPRSS2-ERG fusions did not show correlation with Gleason grade. The pattern of genetic heterogeneity of PTEN deletion was more diverse than that observed for TMPRSS2-ERG fusions in multifocal disease. However, the marked interfocal discordance for both TMPRSS2-ERG fusions and PTEN deletions was consistent with the concept that multiple foci of prostate cancer arise independently within the same prostate, and that individual tumor foci can have distinct patterns of genetic rearrangements.

Loss of phosphatase and tensin homolog protein expression is an independent poor prognostic marker in lung adenocarcinoma.

INTRODUCTION: Phosphatase and tensin homolog (PTEN) has been established as a tumor suppressor gene with an important role in regulating the phosphatidylinositol-3-kinase/AKT antiapoptotic and survival pathway. The prognostic role of PTEN in non-small-cell lung carcinoma has not been evaluated completely in the context of other molecular information. METHODS: Tissue microarrays containing 152 resected non-small-cell lung cancer specimens were used to investigate PTEN and p53 by immunohistochemistry and PTEN by fluorescence in situ hybridization. DNA was isolated and subjected to mutational profiling using the Sequenom Oncocarta v1.0 panel. Clinicopathological features were correlated with PTEN expression, gene copy number, and mutation status. RESULTS: PTEN staining was absent in 63 (41.4%) of the cases. Significantly more squamous cell carcinomas compared with adenocarcinomas demonstrated loss of (negative) PTEN staining (26 of 44 [59%] versus 32 of 94 [34%]; p = 0.009). PTEN gene copy deletion was present in only seven of 124 evaluable cases (5.6%); all deleted cases were immunohistochemistry negative. In univariate and multivariate (MV) analyses adjusted for sex, age, histology, and stage, loss of PTEN protein expression was associated with significantly shorter disease-free survival (MV hazard ratio: 1.78, 95% confidence interval: 1.01-3.14, p = 0.048), whereas no significant associations were seen with p53 or KRAS and epidermal growth factor receptor (EGFR) mutation status. Importantly, the prognostic value of absent PTEN staining was limited to adenocarcinomas, with MV disease-free survival hazard ratio of 2.68 (95% confidence interval: 1.35-5.32, p = 0.005), whereas no such association was seen in squamous cell carcinomas. CONCLUSION: Absence of PTEN protein expression is an independent prognostic marker in early-stage resected lung adenocarcinoma.

Multiple CDK/CYCLIND genes are amplified in medulloblastoma and supratentorial primitive neuroectodermal brain tumor.

Embryonal brain tumors, which include medulloblastoma and the more aggressive supratentorial primitive neuroectodermal tumor (sPNET), comprise one of the largest group of malignant pediatric brain tumors. We observed in high resolution array comparative genomic hybridization and polymerase chain reaction analyses that several different components of the CDK/CYCLIND/pRB regulatory complex, including the CDK4/6 and CCND1/2 loci, are targets of gene amplification in medulloblastoma and sPNET. CDK6 and CCND1 gene amplification were respectively most common and robust, and overall CDK/CYCLIND gene amplification was more commonly observed in sPNET (25%) than medulloblastoma (1-5%). CDK6 overexpression enhanced in vitro and in vivo oncogenicity and endogenous CDK6 or CCND1 knockdown decreased pRB phosphorylation and impaired cell cycle progression in both medulloblastoma and sPNET cell lines. Although animal models implicate the pRB tumor suppressor pathway in medulloblastoma and sPNET, mutations of RB1 or the related INK4 tumor suppressor loci are rare in primary human tumors. Our data suggest that CDK/CYCLIND gene amplification may represent important mechanisms for functional inactivation of pRB in medulloblastoma and sPNET.

Copy number alterations of c-MYC and PTEN are prognostic factors for relapse after prostate cancer radiotherapy.

Despite the use of PSA, Gleason score, and T-category as prognosticators in intermediate-risk prostate cancer, 20-40% of patients will fail local therapy. In order to optimize treatment approaches for intermediate-risk patients, additional genetic prognosticators are needed. Previous reports using array comparative genomic hybridization (aCGH) in radical prostatectomy cohorts suggested a combination of allelic loss of the PTEN gene on 10q and allelic gain of the c-MYC gene on 8q were associated with metastatic disease. We tested whether copy number alterations (CNAs) in PTEN (allelic loss) and c-MYC (allelic gain) were associated with biochemical relapse following modern-era, image-guided radiotherapy (mean dose 76.4 Gy). We used aCGH analyses validated by fluorescence in-situ hybridization (FISH) of DNA was derived from frozen, pre-treatment biopsies in 126 intermediate-risk prostate cancer patients. Patients whose tumors had CNAs in both PTEN and c-MYC had significantly increased genetic instability (percent genome alteration; PGA) compared to tumors with normal PTEN and c-MYC status (p < 0.0001). We demonstrate that c-MYC gain alone, or combined c-MYC gain and PTEN loss, were increasingly prognostic for relapse on multivariable analyses (hazard ratios (HR) of 2.58/p = 0.005 and 3.21/p = 0.0004; respectively). Triaging patients by the use of CNAs within pre-treatment biopsies may allow for better use of systemic therapies to target sub-clinical metastases or locally recurrent disease and improve clinical outcomes.

Interactions and relationships of PTEN, ERG, SPINK1 and AR in castration-resistant prostate cancer.

AIMS: Recently, ETS-related gene (ERG) gene rearrangements, phosphatase tensin homologue (PTEN) deletions and serine protease inhibitor Kazal type 1 (SPINK1) overexpression were investigated as potential markers for molecularly subtyping prostate cancer (PCA). However, their incidence and co-association in castration-resistant PCA (CRPC) has not been characterized fully. METHODS AND RESULTS: A cohort of 59 CRPC patients was investigated for ERG rearrangements, PTEN deletions and androgen receptor (AR) amplification by fluorescence in-situ hybridization. SPINK1 overexpression was assessed by immunohistochemistry. ERG rearrangements and PTEN deletions were detected in 22 of 53 (41.5%) and 35 of 55 (63.6%) of cases, with 15 of 22 (68.1%) of ERG rearrangements occurring through deletions. SPINK1 overexpression occurred in three of 51 (5.8%) of cases exclusively in non-ERG rearranged and AR amplification was detected in 12 of 49 (24.4%) of cases. Only PTEN deletions showed intrafocal heterogeneity occurring in nine of 35 (25.7%) of cases. PTEN deletions were significantly associated with each of ERG rearrangements occurring by deletions only (P = 0.001), AR amplification (P = 0.002) and SPINK1 overexpression (P = 0.002). None of the SPINK1 overexpressing tumours showed AR amplification (P = 0.005) and all occurred in PTEN deleted foci (P = 0.002). CONCLUSION: Te study supports the heterogeneous nature of CRPC and confirms a significant association between PTEN, ERG, AR and SPINK1. Characterizing combined markers will aid in defining PCA subgroups relevant to prognosis contributing to the design of improved therapeutic approaches for CRPC.

The 16p13.3 (PDPK1) Genomic Gain in Prostate Cancer: A Potential Role in Disease Progression.

BACKGROUND: Prostate cancer (PCa) is a leading cause of cancer death, and distinguishing aggressive from indolent tumors is a major challenge. Identification and characterization of genomic alterations associated with advanced disease can provide new markers of progression and better therapeutic approaches. METHODS: We performed fluorescence in situ hybridization to detect the copy number gain of chromosome 16p13.3 in 75 PCa samples including 10 lymph node (LN) metastases and their matched primary tumors, 9 samples of castration-resistant prostate cancer (CRPC), and 46 additional primary PCa specimens with clinicopathologic parameters. RESULTS: We detected the gain in 5 of 10 LN metastases and 3 of 5 matched primary tumors, 3 of 9 CRPC samples, and 9 of 46 (20%) primary tumors where the 16p13.3 alteration was associated with high Gleason score and elevated preoperative prostate-specific antigen levels. The level of 16p13.3 gain was higher in LN metastasis and CRPC specimens compared to primary PCa. Chromosome mapping revealed the gain spans PDPK1 encoding the 3-phosphoinositide-dependent protein kinase-1 (PDK1). Knockdown of PDK1 in three PCa cell lines reduced migration without affecting growth and re-expressing PDK1 rescued motility. CONCLUSION: Our findings support a prognostic value of the 16p13.3 gain and a role of PDK1 in PCa progression through migration.

PTEN genomic deletions that characterize aggressive prostate cancer originate close to segmental duplications.

Deletion of PTEN at 10q23.3 occurs in ∼40% of human prostate cancers and is associated with aggressive metastatic potential, poor prognosis, and androgen-independence. This high frequency of recurrent PTEN deletions in prostate cancer suggests there may be unusual genomic features close to this locus that facilitate DNA alteration at 10q23.3. To explore possible mechanisms for deletions in the PTEN region, a meta-analysis of 311 published human genome array datasets was conducted and determined that the minimal prostate cancer-associated deletion at 10q23.3 corresponds to ∼2.06 MB region flanked by BMPR1A and FAS. On a separate cohort comprising an additional 330 tumors, four-color fluorescence in situ hybridization analysis using probes for BMPR1A, FAS, cen(10), and PTEN showed that 132 of 330 (40%) tumors had PTEN loss, 50 (15%) of which were homozygous losses (comprising in total 100 deletion events). Breakpoints between PTEN and BMPR1A or FAS were subsequently mapped in 100 homozygous and 82 hemizygous PTEN losses, revealing that 125/182 PTEN microdeletions occurred within the 940 kB interval between BMPR1A and PTEN. Furthermore, this breakpoint interval coincides with a repeat-rich region of 414 kB containing the SD17 and SD18 segmental duplications, which contain at least 13 homologous inverted repeat sequences. Together, these data suggest that a strong selective growth advantage for loss of PTEN and upregulation of PI3K/AKT, combined with the close proximity of PTEN to a large unstable segment of repeated DNA comprising SD17 and SD18, can lead to recurrent microdeletions of the PTEN gene in prostate cancer. © 2011 Wiley Periodicals, Inc.

Prostate cancer as a model system for genetic diversity in tumors.

This chapter will summarize novel understandings of the early molecular events in prostatic carcinogenesis that may underlie both the genetic and clinical heterogeneity. Areas covered include preneoplasia, stem cell concepts, telomere abnormalities, and the nature of tumor-stromal interactions. The oncogenomics of prostate cancer is reviewed with emphasis on androgen signaling, ETS gene family aberrations, and PTEN deletion. The notion that "field cancerization," coupled with genomic instability may explain both the occurrence of multifocal disease, and the recent observations of genetic diversity of ERG alteration in individual tumors are discussed. Collectively, genomic studies are rapidly moving human prostate cancer closer to the promise of personalized medicine, so that specific genetic profiles of individual tumors will determine the best therapeutic approaches.

Analysis of miRNA-gene expression-genomic profiles reveals complex mechanisms of microRNA deregulation in osteosarcoma.

Osteosarcoma is an aggressive sarcoma of the bone characterized by a high level of genetic instability and recurrent DNA deletions and amplifications. This study assesses whether deregulation of microRNA (miRNA) expression is a post-transcriptional mechanism leading to gene expression changes in osteosarcoma. miRNA expression profiling was performed for 723 human miRNAs in 7 osteosarcoma tumors, and 38 miRNAs differentially expressed ≥10-fold (28 under- and 10 overexpressed) were identified. In most cases, observed changes in miRNA expression were DNA copy number-correlated. However, various mechanisms of alteration, including positional and/or epigenetic modifications, may have contributed to the expression change of 23 closely linked miRNAs in cytoband 14q32. To develop a comprehensive molecular genetic map of osteosarcoma, the miRNA profiles were integrated with previously published array comparative genomic hybridization DNA imbalance and mRNA gene expression profiles from a set of partially overlapping osteosarcoma tumor samples. Many of the predicted gene targets of differentially expressed miRNA are involved in intracellular signaling pathways important in osteosarcoma, including Notch, RAS/p21, MAPK, Wnt, and the Jun/FOS pathways. By integrating data on copy number variation with mRNA and miRNA expression profiles, we identified osteosarcoma-associated gene expression changes that are DNA copy number-correlated, DNA copy number-independent, mRNA-driven, and/or modulated by miRNA expression. These data collectively suggest that miRNAs provide a novel post-transcriptional mechanism for fine-tuning the expression of specific genes and pathways relevant to osteosarcoma. Thus, the miRNA identified in this manner may provide a starting point for experimentally modulating therapeutically relevant pathways in this tumor.

PTEN deletion and heme oxygenase-1 overexpression cooperate in prostate cancer progression and are associated with adverse clinical outcome.

Overexpression of the pro-survival protein heme oxygenase-1 (HO-1) and loss of the pro-apoptotic tumour suppressor PTEN are common events in prostate cancer (PCA). We assessed the occurrence of both HO-1 expression and PTEN deletion in two cohorts of men with localized and castration-resistant prostate cancer (CRPC). The phenotypic cooperation of these markers was examined in preclinical and clinical models. Overall, there was a statistically significant difference in HO-1 epithelial expression between benign, high-grade prostatic intraepithelial neoplasia (HGPIN), localized PCA, and CRPC (p < 0.0001). The highest epithelial HO-1 expression was noted in CRPC (2.00 ± 0.89), followed by benign prostate tissue (1.49 ± 1.03) (p = 0.0003), localized PCA (1.20 ± 0.95), and HGPIN (1.07 ± 0.87) (p < 0.0001). However, the difference between HGPIN and PCA was not statistically significant (p = 0.21). PTEN deletions were observed in 35/55 (63.6%) versus 68/183 (37.1%) cases of CRPC and localized PCA, respectively. Although neither HO-1 overexpression nor PTEN deletions alone in localized PCA showed a statistically significant association with PSA relapse, the combined status of both markers correlated with disease progression (log-rank test, p = 0.01). In a preclinical model, inhibition of HO-1 by shRNA in PTEN-deficient PC3M cell line and their matched cells where PTEN is restored strongly reduced cell growth and invasion in vitro and inhibited tumour growth and lung metastasis formation in mice compared to cells where only HO-1 is inhibited or PTEN is restored. In summary, we provide clinical and experimental evidence for cooperation between epithelial HO-1 expression and PTEN deletions in relation to the PCA patient's outcome. These findings could potentially lead to the discovery of novel therapeutic modalities for advanced PCA.

PTEN genomic deletion is an early event associated with ERG gene rearrangements in prostate cancer.

OBJECTIVE: To investigate the interaction between, and significance of, ERG gene rearrangements and PTEN genomic deletions in relation to the development and progression of prostate cancer (PCA). PATIENTS AND METHODS: We interrogated an initial cohort of 220 men with localized PCA using fluorescence in situ hybridization for ERG rearrangements and PTEN genomic deletions. RESULTS: The incidences of ERG rearrangements and PTEN deletions in PCA were significantly higher than in high-grade prostatic intra-epithelial neoplasia (HGPIN) and benign prostate tissue (P < 0.001). ERG rearrangements and PTEN deletions were detected in 41.9 and 42.6% of patients' tumours, respectively. ERG rearrangements were never detected in benign prostate tissue, while PTEN aberrations were present at a basal level of 4.6%. PTEN hemizygous deletions showed higher frequency than homozygous deletions within each diagnostic category from benign prostate tissue to HGPIN and PCA (P ≤ 0.001). Furthermore, in 29 patients where all three tissues were available, PTEN genomic aberrations in PCA were significantly different from those in benign tissue (P = 0.005) and HGPIN (P = 0.02), reflecting the accumulation of genomic aberrations in the early stages of disease progression. Within this cohort, 71.4% of homozygous and 44.2% of hemizygous PTEN deletions occurred simultaneously with ERG rearrangements (P ≈ 0). Stratified according to Gleason score (GS), hemizygous PTEN deletions across various GS groups were observed at a higher frequency than homozygous deletions. However, PTEN homozygous deletions showed positive trends with higher GS, increasing in poorly differentiated PCA (GS 8-10) in comparison to moderately and well differentiated tumours (GS 6 and 7). CONCLUSION: We show significant association between ERG gene rearrangements and PTEN genomic aberrations in subset of PCA. Our analysis also provides further support for the observation that homozygous PTEN deletions can occur within the subset of HGPIN lesions, and shows accumulating genetic aberrations with disease progression, evidenced by higher detection in PCA than in HGPIN and more PTEN homozygous deletions in GS 8-10 than in 6-7.

Detection of ERG gene rearrangements and PTEN deletions in unsuspected prostate cancer of the transition zone.

To assess the incidence of ERG rearrangements and PTEN deletions in a group of prognostically favorable PCA tumors of the transition zone (TZ). We interrogated 54 unsuspected PCA tumors using fluorescence in-situ hybridization for ERG rearrangements and PTEN deletions. ERG rearrangements were detected in 6/47 (12.7%) cases with 4/6 (66%) occurring by deletion. PTEN deletions were detected in 9/47 (19.1%) cases with only 3/47 (6.4%) having both PTEN losses and ERG rearrangements (p = 0.07). Only ERG rearrangements were associated with higher tumor volume > 5% (p = 0.04); PTEN deletions showed similar trends (p = 0.06). None of the markers showed association with Gleason score. In conclusion, although TZ and peripheral zone tumors share similar molecular etiologies, they exhibit differences in the rates of ERG and PTEN aberrations. The differences of ERG and /or PTEN genetic aberrations in TZ tumors may point toward a subset of tumors with adverse behavior. Additional studies implementing ERG and PTEN tissue-based FISH assays in tumors of the TZ are warranted to substantiate the potential clinical value of these biomarkers in the management of men with incidental PCA.

Cytogenetic molecular delineation of a terminal 18q deletion suggesting neo-telomere formation.

Deletion of the long arm of chromosome 18 is one of the most common segmental aneusomies compatible with life and usually involves a deletion of the terminal chromosomal region. However, the mechanisms implicated in the stabilization of terminal deletions are not well understood. In this study, we analyzed a girl with moderate mental retardation who had a cytogenetically visible terminal 18q deletion. In order to characterize the breakpoint in the terminal 18q region, we used fluorescence In situ hybridization (FISH) with bacterial artificial chromosomes (BACs) and pan-telomeric probes and also the array technique based on comparative genomic hybridization (array-CGH). FISH with pan-telomeric probes revealed no signal in the terminal region of the deleted chromosome, indicating the absence of normal telomere repeat (TTAGGG)n sequences in 18q. We suggest that neo-telomere formation by chromosome healing was involved in the repair and stabilization of this terminal deletion.

Development of metastatic and non-metastatic tumor lines from a patient's prostate cancer specimen-identification of a small subpopulation with metastatic potential in the primary tumor.

BACKGROUND: Metastasis is the major cause of prostate cancer deaths. Tumor heterogeneity in both primary and metastatic prostate cancers is a major hurdle in elucidating the mechanisms of metastasis in this disease. To circumvent this obstacle and improve our understanding of prostate cancer metastasis, we developed multiple tumor tissue lines from one patient's primary prostate cancer specimen to examine differences in metastatic ability. METHODS: Pieces of tissue from different foci of a patient's primary prostate tumor were grafted into subrenal capsules of NOD-SCID mice and transplantable tumor sublines were established by serial passage. The metastatic ability of each subline was tested via orthotopic grafting into mice. Chromosomal alterations exclusively presented in a metastatic subline were examined by SKY and investigated for their presence in parental tissues by fluorescence in situ hybridization. RESULTS: Three transplantable sublines were developed, resembling the primary tumor histologically, exhibiting poor differentiation, and different growth rates. Importantly, the LTL-220N and LTL-221N sublines were non-metastatic, whereas the LTL-220M subline was spontaneously metastatic in vivo. SKY analysis showed limited but unique chromosomal alterations in each subline. Some chromosomal alterations, exclusively present in the metastatic LTL-220M subline, were also observed in a small portion of the parental cancer tissues. CONCLUSION: The results indicate that in primary prostate tumors metastatic potential can be confined to a minority of cancer cells. Subrenal capsule xenograft methodology can be used to dissect heterogeneous cancer cells in a patient's primary tumor and sublines derived from such cells provide valuable tools for investigating mechanisms underlying prostate cancer metastasis.