HPV infection results in changes in host gene methylation which, in turn, are thought to contribute to the neoplastic progression of HPV-associated cancers. The objective of this study was to ...identify joint and disease-specific genome-wide methylation changes in anal and cervical cancer as well as changes in high-grade pre-neoplastic lesions. Formalin-fixed paraffin-embedded (FFPE) anal tissues (n = 143; 99% HPV+) and fresh frozen cervical tissues (n = 28; 100% HPV+) underwent microdissection, DNA extraction, HPV genotyping, bisulfite modification, DNA restoration (FFPE) and analysis by the Illumina HumanMethylation450 Array. Differentially methylated regions (DMR; t test q<0.01, 3 consecutive significant CpG probes and mean Δβ methylation value>0.3) were compared between normal and cancer specimens in partial least squares (PLS) models and then used to classify anal or cervical intraepithelial neoplasia-3 (AIN3/CIN3). In AC, an 84-gene PLS signature (355 significant probes) differentiated normal anal mucosa (NM; n = 9) from AC (n = 121) while a 36-gene PLS signature (173 significant probes) differentiated normal cervical epithelium (n = 10) from CC (n = 9). The CC progression signature was validated using three independent publicly available datasets (n = 424 cases). The AC and CC progression PLS signatures were interchangeable in segregating normal, AIN3/CIN3 and AC and CC and were found to include 17 common overlapping hypermethylated genes. Moreover, these signatures segregated AIN3/CIN3 lesions similarly into cancer-like and normal-like categories. Distinct methylation changes occur across the genome during the progression of AC and CC with overall similar profiles and add to the evidence suggesting that HPV-driven oncogenesis may result in similar non-random methylomic events. Our findings may lead to identification of potential epigenetic drivers of HPV-associated cancers and also, of potential markers to identify higher risk pre-cancerous lesions.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Models of bladder tumor progression have suggested that genetic alterations may determine both phenotype and clinical course.
We have applied expression microarray analysis to a divergent set of ...bladder tumors to further elucidate the course of disease
progression and to classify tumors into more homogeneous and clinically relevant subgroups. cDNA microarrays containing 10,368
human gene elements were used to characterize the global gene expression patterns in 80 bladder tumors, 9 bladder cancer cell
lines, and 3 normal bladder samples. Robust statistical approaches accounting for the multiple testing problem were used to
identify differentially expressed genes. Unsupervised hierarchical clustering successfully separated the samples into two
subgroups containing superficial (pT a and pT 1 ) versus muscle-invasive (pT 2 -pT 4 ) tumors. Supervised classification had a 90.5% success rate separating superficial from muscle-invasive tumors based on a
limited subset of genes. Tumors could also be classified into transitional versus squamous subtypes (89% success rate) and
good versus bad prognosis (78% success rate). The performance of our stage classifiers was confirmed in silico using data from an independent tumor set. Validation of differential expression was done using immunohistochemistry on tissue
microarrays for cathepsin E, cyclin A2, and parathyroid hormone–related protein. Genes driving the separation between tumor
subsets may prove to be important biomarkers for bladder cancer development and progression and eventually candidates for
therapeutic targeting.
Purpose We assessed whether an association exists between a change in prostate specific antigen and biopsy progression in men on active surveillance. Materials and Methods A cohort of patients ...undergoing active surveillance for prostate cancer was identified from the urological oncology database at our institution. Multivariate logistic regression was performed to determine whether prostate specific antigen velocity, defined as the change in ln(prostate specific antigen) per year, is associated with biopsy progression, defined as a Gleason upgrade or volume progression on repeat biopsy within 24 months of diagnosis. Results A total of 241 men with a mean ± SD age of 61 ± 7 years and mean prostate specific antigen 4.9 ± 2.2 ng/ml met study inclusion criteria. Median time to repeat biopsy was 10 months (IQR 6–13). Biopsy progression developed in 55 men (23%), including a Gleason score upgrade in 46 (19%), greater than 33% positive cores in 11 (5%) and greater than 50% maximum single core positive in 12 (5%). The median prostate specific antigen ratio per year was 1.0 (IQR 0.95–1.03), although 1 man had a ratio of greater than 1.26 (doubled over 3 years) and 7 had a ratio of less than 1/1.26 (halved over 3 years). On multivariate analysis prostate specific antigen doubling within 3 years was associated with a 1.4-fold increase in the odds of biopsy progression (OR 1.4, 95% CI 0.6–3.4, p = 0.46). Conclusions There is little change in prostate specific antigen during the first 24 months of surveillance in men with well staged, low risk prostate cancer. We believe that these findings highlight the importance of repeat biopsy during surveillance.
Epidemiological and prospective studies indicate that comprehensive lifestyle changes may modify the progression of prostate cancer. However, the molecular mechanisms by which improvements in diet ...and lifestyle might affect the prostate microenvironment are poorly understood. We conducted a pilot study to examine changes in prostate gene expression in a unique population of men with low-risk prostate cancer who declined immediate surgery, hormonal therapy, or radiation and participated in an intensive nutrition and lifestyle intervention while undergoing careful surveillance for tumor progression. Consistent with previous studies, significant improvements in weight, abdominal obesity, blood pressure, and lipid profile were observed (all P < 0.05), and surveillance of low-risk patients was safe. Gene expression profiles were obtained from 30 participants, pairing RNA samples from control prostate needle biopsy taken before intervention to RNA from the same patient's 3-month postintervention biopsy. Quantitative real-time PCR was used to validate array observations for selected transcripts. Two-class paired analysis of global gene expression using significance analysis of microarrays detected 48 up-regulated and 453 down-regulated transcripts after the intervention. Pathway analysis identified significant modulation of biological processes that have critical roles in tumorigenesis, including protein metabolism and modification, intracellular protein traffic, and protein phosphorylation (all P < 0.05). Intensive nutrition and lifestyle changes may modulate gene expression in the prostate. Understanding the prostate molecular response to comprehensive lifestyle changes may strengthen efforts to develop effective prevention and treatment. Larger clinical trials are warranted to confirm the results of this pilot study.
Purpose: Bladder carcinogenesis is believed to follow alternative pathways of disease progression driven by an accumulation of genetic
alterations. The purpose of this study was to evaluate ...associations between measures of genomic instability and bladder cancer
clinical phenotype.
Experimental Design: Genome-wide copy number profiles were obtained for 98 bladder tumors of diverse stages (29 pT a , 14 pT 1 , 55 pT 2-4 ) and grades (21 low-grade and 8 high-grade superficial tumors) by array-based comparative genomic hybridization (CGH). Each
array contained 2,464 bacterial artificial chromosome and P1 clones, providing an average resolution of 1.5 Mb across the
genome. A total of 54 muscle-invasive cases had follow-up information available. Overall outcome analysis was done for patients
with muscle-invasive tumors having “good” (alive >2 years) versus “bad” (dead in <2 years) prognosis.
Results: Array CGH analysis showed significant increases in copy number alterations and genomic instability with increasing stage
and with outcome. The fraction of genome altered (FGA) was significantly different between tumors of different stages (pT a versus pT 1 , P = 0.0003; pT a versus pT 2-4 , P = 0.02; and pT 1 versus pT 2-4 , P = 0.03). Individual clones that differed significantly between different tumor stages were identified after adjustment for
multiple comparisons (false discovery rate < 0.05). For muscle-invasive tumors, the FGA was associated with patient outcome
(bad versus good prognosis patients, P = 0.002) and was identified as the only independent predictor of overall outcome based on a multivariate Cox proportional
hazards method. Unsupervised hierarchical clustering separated “good” and “bad” prognosis muscle-invasive tumors into clusters
that showed significant association with FGA and survival (Kaplan-Meier, P = 0.019). Supervised tumor classification (prediction analysis for microarrays) had a 71% classification success rate based
on 102 unique clones.
Conclusions: Array-based CGH identified quantitative and qualitative differences in DNA copy number alterations at high resolution according
to tumor stage and grade. Fraction genome altered was associated with worse outcome in muscle-invasive tumors, independent
of other clinicopathologic parameters. Measures of genomic instability add independent power to outcome prediction of bladder
tumors.
Purpose: More than 1,300,000 prostate needle biopsies are done annually in the United States with up to 16% incidence of isolated
high-grade prostatic intraepithelial neoplasia (HGPIN). HGPIN has low ...predictive value for identifying prostate cancer on
subsequent needle biopsies in prostate-specific antigen–screened populations. In contemporary series, prostate cancer is detected
in ∼20% of repeat biopsies following a diagnosis of HGPIN. Further, discrete histologic subtypes of HGPIN with clinical implication
in management have not been characterized. The TMPRSS2-ERG gene fusion that has recently been described in prostate cancer has also been shown to occur in a subset of HGPIN. This may
have significant clinical implications given that TMPRSS2-ERG fusion prostate cancer is associated with a more aggressive clinical course.
Experimental Design: In this study, we assessed a series of HGPIN lesions and paired prostate cancer for the presence of TMPRSS2-ERG gene fusion.
Results: Fusion-positive HGPIN was observed in 16% of the 143 number of lesions, and in all instances, the matching cancer shared
the same fusion pattern. Sixty percent of TMPRSS2-ERG fusion prostate cancer had fusion-negative HGPIN.
Conclusions: Given the more aggressive nature of TMPRSS2-ERG prostate cancer, the findings of this study raise the possibility that gene fusion-positive HGPIN lesions are harbingers
of more aggressive disease. To date, pathologic, molecular, and clinical variables do not help stratify which men with HGPIN
are at increased risk for a cancer diagnosis. Our results suggest that the detection of isolated TMPRSS2-ERG fusion HGPIN would improve the positive predictive value of finding TMPRSS2-ERG fusion prostate cancer in subsequent biopsies.
Metagenomic next-generation sequencing (mNGS) of body fluids is an emerging approach to identify occult pathogens in undiagnosed patients. We hypothesized that metagenomic testing can be ...simultaneously used to detect malignant neoplasms in addition to infectious pathogens.
From two independent studies (n = 205), we used human data generated from a metagenomic sequencing pipeline to simultaneously screen for malignancies by copy number variation (CNV) detection. In the first case-control study, we analyzed body fluid samples (n = 124) from patients with a clinical diagnosis of either malignancy (positive cases, n = 65) or infection (negative controls, n = 59). In a second verification cohort, we analyzed a series of consecutive cases (n = 81) sent to cytology for malignancy workup that included malignant positives (n = 32), negatives (n = 18), or cases with an unclear gold standard (n = 31).
The overall CNV test sensitivity across all studies was 87% (55 of 63) in patients with malignancies confirmed by conventional cytology and/or flow cytometry testing and 68% (23 of 34) in patients who were ultimately diagnosed with cancer but negative by conventional testing. Specificity was 100% (95% CI 95-100%) with no false positives detected in 77 negative controls. In one example, a patient hospitalized with an unknown pulmonary illness had non-diagnostic lung biopsies, while CNVs implicating a malignancy were detectable from bronchoalveolar fluid.
Metagenomic sequencing of body fluids can be used to identify undetected malignant neoplasms through copy number variation detection. This study illustrates the potential clinical utility of a single metagenomic test to uncover the cause of undiagnosed acute illnesses due to cancer or infection using the same specimen.
The number of circulating tumor cells (CTCs) in metastatic prostate cancer patients provides prognostic and predictive information. However, it is the molecular characterization of CTCs that offers ...insight into the biology of these tumor cells in the context of personalized treatment.
We developed a novel approach to isolate CTCs away from hematopoietic cells with high purity, enabling genomic analysis of these cells. The isolation protocol involves immunomagnetic enrichment followed by fluorescence activated cell sorting (IE/FACS). To evaluate the feasibility of isolation of CTCs by IE/FACS and downstream genomic profiling, we conducted a pilot study in patients with metastatic castration resistant prostate cancer (CRPC). Twenty (20) sequential CRPC patients were assayed using CellSearch™. Twelve (12) patients positive for CTCs were subjected to immunomagnetic enrichment and fluorescence activated cell sorting (IE/FACS) to isolate CTCs. Genomic DNA of CTCs was subjected to whole genome amplification (WGA) followed by gene copy number analysis via array comparative genomic hybridization (aCGH).
CTCs from nine (9) patients successfully profiled were observed to have multiple copy number aberrations including those previously reported in primary prostate tumors such as gains in 8q and losses in 8p. High-level copy number gains at the androgen receptor (AR) locus were observed in 7 (78%) cases. Comparison of genomic profiles between CTCs and archival primary tumors from the same patients revealed common lineage. However, high-level copy number gains in the AR locus were observed in CTCs, but not in the matched archival primary tumors.
We developed a new approach to isolate prostate CTCs without significant leukocyte admixture, and to subject them to genome-wide copy number analysis. Our assay may be utilized to explore genomic events involved in cancer progression, e.g. development of castration resistance and to monitor therapeutic efficacy of targeted therapies in clinical trials in a relatively non-invasive manner.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Much work has been done to develop tumor-targeting antibodies by selecting a phage antibody library on cancer cell lines.
However, when tumor cells are removed from their natural environment, they ...may undergo genetic and epigenetic changes yielding
different surface antigens than those seen in actual cases of cancer. We developed a strategy that allows selection of phage
antibodies against tumor cells in situ on both fresh frozen and paraffin-embedded tissues using laser capture microdissection. By restricting antibody selection
to binders of internalizing epitopes, we generated a panel of phage antibodies that target clinically represented prostate
cancer antigens. We identified ALCAM/MEMD/CD166, a newly discovered prostate cancer marker, as the target for one of the selected
antibodies, demonstrating the effectiveness of our approach. We further conjugated two single chain Fv fragments to liposomes
and demonstrated that these nanotargeting devices were efficiently delivered to the interior of prostate cancer cells. The
ability to deliver payload intracellularly and to recognize tumor cells in situ makes these antibodies attractive candidates for the development of targeted cancer therapeutics.
The uncertainties inherent in clinical measures of prostate cancer (CaP) aggressiveness endorse the investigation of clinically validated tissue biomarkers. MUC1 expression has been previously ...reported to independently predict aggressive localized prostate cancer. We used a large cohort to validate whether MUC1 protein levels measured by immunohistochemistry (IHC) predict aggressive cancer, recurrence and survival outcomes after radical prostatectomy independent of clinical and pathological parameters.
MUC1 IHC was performed on a multi-institutional tissue microarray (TMA) resource including 1,326 men with a median follow-up of 5 years. Associations with clinical and pathological parameters were tested by the Chi-square test and the Wilcoxon rank sum test. Relationships with outcome were assessed with univariable and multivariable Cox proportional hazard models and the Log-rank test.
The presence of MUC1 expression was significantly associated with extracapsular extension and higher Gleason score, but not with seminal vesicle invasion, age, positive surgical margins or pre-operative serum PSA levels. In univariable analyses, positive MUC1 staining was significantly associated with a worse recurrence free survival (RFS) (HR: 1.24, CI 1.03-1.49, P = 0.02), although not with disease specific survival (DSS, P>0.5). On multivariable analyses, the presence of positive surgical margins, extracapsular extension, seminal vesicle invasion, as well as higher pre-operative PSA and increasing Gleason score were independently associated with RFS, while MUC1 expression was not. Positive MUC1 expression was not independently associated with disease specific survival (DSS), but was weakly associated with overall survival (OS).
In our large, rigorously designed validation cohort, MUC1 protein expression was associated with adverse pathological features, although it was not an independent predictor of outcome after radical prostatectomy.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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