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1.  USP22 Regulates Oncogenic Signaling Pathways to Drive Lethal Cancer Progression 
Cancer research  2013;74(1):272-286.
Increasing evidence links deregulation of the USP22 deubitiquitylase to cancer development and progression in a select group of tumor types, but its specificity and underlying mechanisms of action are not well-defined. Here we show that USP22 is a critical promoter of lethal tumor phenotypes that acts by modulating nuclear receptor and oncogenic signaling. In multiple xenograft models of human cancer, modeling of tumor-associated USP22 deregulation demonstrated that USP22 controls androgen receptor (AR) accumulation and signaling, and that it enhances expression of critical target genes co-regulated by AR and MYC. USP22 not only reprogrammed AR function, but was sufficient to induce the transition to therapeutic resistance. Notably, in vivo depletion experiments revealed that USP22 is critical to maintain phenotypes associated with end-stage disease. This was a significant finding given clinical evidence that USP22 is highly deregulated in tumors which have achieved therapeutic resistance. Taken together, our findings define USP22 as a critical effector of tumor progression whcih drives lethal phenotypes, rationalizing this enzyme as an appealing therapeutic target to treat advanced disease.
PMCID: PMC3947329  PMID: 24197134
Androgen receptor; Castration-resistant prostate cancer; Oncogene; Deubiquitylase; prostate cancer
2.  Chk1 targeting reactivates PP2A tumor suppressor activity in cancer cells 
Cancer research  2013;73(22):10.1158/0008-5472.CAN-13-1002.
Checkpoint kinase Chk1 is constitutively active in many cancer cell types and new generation Chk1 inhibitors show marked antitumor activity as single agents. Here we present a hitherto unrecognized mechanism that contributes to the response of cancer cells to Chk1 targeted therapy. Inhibiting chronic Chk1 activity in cancer cells induced the tumor suppressor activity of protein phosphatase PP2A, which by dephosphorylating MYC serine 62, inhibited MYC activity and impaired cancer cell survival. Mechanistic investigations revealed that Chk1 inhibition activated PP2A by decreasing the transcription of CIP2A, a chief inhibitor of PP2A activity. Inhibition of cancer cell clonogenicity by Chk1 inhibition could be rescued in vitro either by exogenous expression of CIP2A or by blocking the CIP2A-regulated PP2A complex. Chk1-mediated CIP2A regulation was extended in tumor models dependent on either Chk1 or CIP2A. The clinical relevance of CIP2A as a Chk1 effector protein was validated in several human cancer types, including neuroblastoma where CIP2A was identified as a NMYC-independent prognostic factor. Since the Chk1-CIP2A-PP2A pathway is driven by DNA-PK activity, functioning regardless of p53 or ATM/ATR status, our results offer explanative power for understand how Chk1 inhibitors mediate single-agent anticancer efficacy. Further, they define CIP2A-PP2A status in cancer cells as a pharmacodynamic marker for their response to Chk1-targeted therapy.
PMCID: PMC3870284  PMID: 24072747
Claspin; PPP2R2A; Chk1 serine 345
4.  Mapping of the chromosomal amplification 1p21-22 in bladder cancer 
BMC Research Notes  2014;7(1):547.
The aim of the study was to characterize a recurrent amplification at chromosomal region 1p21-22 in bladder cancer.
ArrayCGH (aCGH) was performed to identify DNA copy number variations in 7 clinical samples and 6 bladder cancer cell lines. FISH was used to map the amplicon at 1p21-22 in the cell lines. Gene expression microarrays and qRT-PCR were used to study the expression of putative target genes in the region.
aCGH identified an amplification at 1p21-22 in 10/13 (77%) samples. The minimal region of the amplification was mapped to a region of about 1 Mb in size, containing a total of 11 known genes. The highest amplification was found in SCaBER squamous cell carcinoma cell line. Four genes, TMED5, DR1, RPL5 and EVI5, showed significant overexpression in the SCaBER cell line compared to all the other samples tested. Oncomine database analysis revealed upregulation of DR1 in superficial and infiltrating bladder cancer samples, compared to normal bladder.
In conclusions, we have identified and mapped chromosomal amplification at 1p21-22 in bladder cancer as well as studied the expression of the genes in the region. DR1 was found to be significantly overexpressed in the SCaBER, which is a model of squamous cell carcinoma. However, the overexpression was found also in a published clinical sample cohort of superficial and infiltrating bladder cancers. Further studies with more clinical material are needed to investigate the role of the amplification at 1p21-22.
PMCID: PMC4143550  PMID: 25135188
Gene amplification; Bladder cancer; DR1; aCGH
5.  Loss of PTEN Is Associated with Aggressive Behavior in ERG-Positive Prostate Cancer 
The associations of ERG overexpression with clinical behavior and molecular pathways of prostate cancer are incompletely known. We assessed the association of ERG expression with AR, PTEN, SPINK1, Ki-67, and EZH2 expression levels, deletion, and mutations of chromosomal region 3p14 and TP53, and clinicopathologic variables.
The material consisted of 326 prostatectomies, 166 needle biopsies from men treated primarily with endocrine therapy, 177 transurethral resections of castration-resistant prostate cancers (CRPC), and 114 CRPC metastases obtained from 32 men. Immunohistochemistry, FISH, and sequencing was used for the measurements.
ERG expression was found in about 45% of all patient cohorts. In a multivariate analysis, ERG expression showed independent value of favorable prognosis (P = 0.019). ERG positivity was significantly associated with loss of PTEN expression in prostatectomy (P = 0.0348), and locally recurrent CRPCs (P = 0.0042). Loss of PTEN expression was associated (P = 0.0085) with shorter progression-free survival in ERG-positive, but not in negative cases. When metastases in each subject were compared, consistent ERG, PTEN, and AR expression as well as TP53 mutations were found in a majority of subjects.
A similar frequency of ERG positivity from early to late stage of the disease suggests lack of selection of ERG expression during disease progression. The prognostic significance of PTEN loss solely in ERG-positive cases indicates interaction of these pathways. The finding of consistent genetic alterations in different metastases suggests that the major genetic alterations take place in the primary tumor.
Interaction of PTEN and ERG pathways warrants further studies.
PMCID: PMC4086660  PMID: 24083995
6.  Bcl-2 associated athanogene 5 (Bag5) is overexpressed in prostate cancer and inhibits ER-stress induced apoptosis 
BMC Cancer  2013;13:96.
The Bag (Bcl-2 associated athanogene) family of proteins consists of 6 members sharing a common, single-copied Bag domain through which they interact with the molecular chaperone Hsp70. Bag5 represents an exception in the Bag family since it consists of 5 Bag domains covering the whole protein. Bag proteins like Bag1 and Bag3 have been implicated in tumor growth and survival but it is not known whether Bag5 also exhibits this function.
Bag5 mRNA and protein expression levels were investigated in prostate cancer patient samples using real-time PCR and immunoblot analyses. In addition immunohistological studies were carried out to determine the expression of Bag5 in tissue arrays. Analysis of Bag5 gene expression was carried out using one-way ANOVA and Bonferroni’s Multiple Comparison test. The mean values of the Bag5 stained cells in the tissue array was analyzed by Mann-Whitney test. Functional studies of the role of Bag5 in prostate cancer cell lines was performed using overexpression and RNA interference analyses.
Our results show that Bag5 is overexpressed in malignant prostate tissue compared to benign samples. In addition we could show that Bag5 levels are increased following endoplasmic reticulum (ER)-stress induction, and Bag5 relocates from the cytoplasm to the ER during this process. We also demonstrate that Bag5 interacts with the ER-resident chaperone GRP78/BiP and enhances its ATPase activity. Bag5 overexpression in 22Rv.1 prostate cancer cells inhibited ER-stress induced apoptosis in the unfolded protein response by suppressing PERK-eIF2-ATF4 activity while enhancing the IRE1-Xbp1 axis of this pathway. Cells expressing high levels of Bag5 showed reduced sensitivity to apoptosis induced by different agents while Bag5 downregulation resulted in increased stress-induced cell death.
We have therefore shown that Bag5 is overexpressed in prostate cancer and plays a role in ER-stress induced apoptosis. Furthermore we have identified GRP78/BiP as a novel interaction partner of Bag5.
PMCID: PMC3598994  PMID: 23448667
Unfolded protein response; Cell stress; Endoplasmic reticulum; Apoptosis; Molecular chaperones; Refolding
7.  The tumorigenic FGFR3-TACC3 gene fusion escapes miR-99a regulation in glioblastoma  
Fusion genes are chromosomal aberrations that are found in many cancers and can be used as prognostic markers and drug targets in clinical practice. Fusions can lead to production of oncogenic fusion proteins or to enhanced expression of oncogenes. Several recent studies have reported that some fusion genes can escape microRNA regulation via 3′–untranslated region (3′-UTR) deletion. We performed whole transcriptome sequencing to identify fusion genes in glioma and discovered FGFR3-TACC3 fusions in 4 of 48 glioblastoma samples from patients both of mixed European and of Asian descent, but not in any of 43 low-grade glioma samples tested. The fusion, caused by tandem duplication on 4p16.3, led to the loss of the 3′-UTR of FGFR3, blocking gene regulation of miR-99a and enhancing expression of the fusion gene. The fusion gene was mutually exclusive with EGFR, PDGFR, or MET amplification. Using cultured glioblastoma cells and a mouse xenograft model, we found that fusion protein expression promoted cell proliferation and tumor progression, while WT FGFR3 protein was not tumorigenic, even under forced overexpression. These results demonstrated that the FGFR3-TACC3 gene fusion is expressed in human cancer and generates an oncogenic protein that promotes tumorigenesis in glioblastoma.
PMCID: PMC3561838  PMID: 23298836
8.  Convergence of oncogenic and hormone receptor pathways promotes metastatic phenotypes 
Cyclin D1b is a splice variant of the cell cycle regulator cyclin D1 and is known to harbor divergent and highly oncogenic functions in human cancer. While cyclin D1b is induced during disease progression in many cancer types, the mechanisms underlying cyclin D1b function remain poorly understood. Herein, cell and human tumor xenograft models of prostate cancer were utilized to resolve the downstream pathways that are required for the protumorigenic functions of cyclin D1b. Specifically, cyclin D1b was found to modulate the expression of a large transcriptional network that cooperates with androgen receptor (AR) signaling to enhance tumor cell growth and invasive potential. Notably, cyclin D1b promoted AR-dependent activation of genes associated with metastatic phenotypes. Further exploration determined that transcriptional induction of SNAI2 (Slug) was essential for cyclin D1b–mediated proliferative and invasive properties, implicating Slug as a critical driver of disease progression. Importantly, cyclin D1b expression highly correlated with that of Slug in clinical samples of advanced disease. In vivo analyses provided strong evidence that Slug enhances both tumor growth and metastatic phenotypes. Collectively, these findings reveal the underpinning mechanisms behind the protumorigenic functions of cyclin D1b and demonstrate that the convergence of the cyclin D1b/AR and Slug pathways results in the activation of processes critical for the promotion of lethal tumor phenotypes.
PMCID: PMC3533295  PMID: 23257359
9.  Contribution of ARLTS1 Cys148Arg (T442C) Variant with Prostate Cancer Risk and ARLTS1 Function in Prostate Cancer Cells 
PLoS ONE  2011;6(10):e26595.
ARLTS1 is a recently characterized tumor suppressor gene at 13q14.3, a region frequently deleted in both sporadic and hereditary prostate cancer (PCa). ARLTS1 variants, especially Cys148Arg (T442C), increase susceptibility to different cancers, including PCa. In this study the role of Cys148Arg substitution was investigated as a risk factor for PCa using both genetic and functional analysis. Cys148Arg genotypes and expression of the ARLTS1 were explored in a large set of familial and unselected PCa cases, clinical tumor samples, xenografts, prostate cancer cell lines and benign prostatic hyperplasia (BPH) samples. The frequency of the variant genotype CC was significantly higher in familial (OR = 1.67, 95% CI = 1.08–2.56, P = 0.019) and unselected patients (OR = 1.52, 95% CI = 1.18–1.97, P = 0.001) and the overall risk was increased (OR = 1.54, 95% CI = 1.20–1.98, P = 0.0007). Additional analysis with clinicopathological data revealed an association with an aggressive disease (OR = 1.28, 95% CI = 1.05-∞, P = 0.02). The CC genotype of the Cys148Arg variant was also contributing to the lowered ARLTS1 expression status in lymphoblastoid cells from familial patients. In addition significantly lowered ARLTS1 expression was observed in clinical tumor samples compared to BPH samples (P = 0.01). The ARLTS1 co-expression signature based on previously published microarray data was generated from 1587 cancer samples confirming the low expression of ARLTS1 in PCa and showed that ARLTS1 expression was strongly associated with immune processes. This study provides strong confirmation of the important role of ARLTS1 Cys148Arg variant as a contributor in PCa predisposition and a potential marker for aggressive disease outcome.
PMCID: PMC3197657  PMID: 22028916
10.  Overall and worst gleason scores are equally good predictors of prostate cancer progression 
BMC Urology  2011;11:21.
Gleason scoring has experienced several modifications during the past decade. So far, only one study has compared the prognostic abilities of worst (WGS) and overall (OGS) modified Gleason scores after the ISUP 2005 conference. Prostatic needle biopsies are individually paraffin-embedded in 57% of European pathology laboratories, whereas the rest of laboratories embed multiple (2 - 6) biopsies per one paraffin-block. Differences in the processing method can have a far-reaching effect, because reporting of the Gleason score (GS) is different for individually embedded and pooled biopsies, and GS is one of the most important factors when selecting treatment for patients.
The study material consisted of needle biopsies from 236 prostate cancer patients that were endocrine-treated in 1999-2003. Biopsies from left side and right side were embedded separately. Haematoxylin-eosin-stained slides were scanned and analyzed on web-based virtual microscopy. Worst and overall Gleason scores were assessed according to the modified Gleason score schema after analyzing each biopsy separately. The compound Gleason scores (CGS) were obtained from the original pathology reports. Two different grade groupings were used: GS 6 or less vs. 7 vs. 8 or above; and GS 7(3 + 4) or less vs. 7(4 + 3) and 8 vs. 9-10. The prognostic ability of the three scoring methods to predict biochemical progression was compared with Kaplan-Meier survival analysis and univariate and multivariate Cox regression analyses.
The median follow-up time of the patients was 64.5 months (range 0-118). The modified GS criteria led to upgrading of the Gleason sums compared to the original CGS from the pathology reports 1999-2003 (mean 7.0 for CGS, 7.5 for OGS, 7.6 for WGS). In 43 cases WGS was > OGS. In a univariate analysis the relative risks were 2.1 (95%-confidence interval 1.8-2.4) for CGS, 2.5 (2.1-2.8) for OGS, and 2.6 (2.2-2.9) for WGS. In a multivariate analysis, OGS was the only independent prognostic factor.
All of the three Gleason scoring methods are strong predictors of biochemical recurrence. The use of modified Gleason scoring leads to upgrading of GS, but also improves the prognostic value of the scoring. No significant prognostic differences between OGS and WGS could be shown, which may relate to the apparent narrowing of the GS scale from 2-10 to 5-10 due to the recent modifications.
PMCID: PMC3193164  PMID: 21978318
11.  ETS1 Mediates MEK1/2-Dependent Overexpression of Cancerous Inhibitor of Protein Phosphatase 2A (CIP2A) in Human Cancer Cells 
PLoS ONE  2011;6(3):e17979.
EGFR-MEK-ERK signaling pathway has an established role in promoting malignant growth and disease progression in human cancers. Therefore identification of transcriptional targets mediating the oncogenic effects of the EGFR-MEK-ERK pathway would be highly relevant. Cancerous inhibitor of protein phosphatase 2A (CIP2A) is a recently characterized human oncoprotein. CIP2A promotes malignant cell growth and is over expressed at high frequency (40–80%) in most of the human cancer types. However, the mechanisms inducing its expression in cancer still remain largely unexplored. Here we present systematic analysis of contribution of potential gene regulatory mechanisms for high CIP2A expression in cancer. Our data shows that evolutionary conserved CpG islands at the proximal CIP2A promoter are not methylated both in normal and cancer cells. Furthermore, sequencing of the active CIP2A promoter region from altogether seven normal and malignant cell types did not reveal any sequence alterations that would increase CIP2A expression specifically in cancer cells. However, treatment of cancer cells with various signaling pathway inhibitors revealed that CIP2A mRNA expression was sensitive to inhibition of EGFR activity as well as inhibition or activation of MEK-ERK pathway. Moreover, MEK1/2-specific siRNAs decreased CIP2A protein expression. Series of CIP2A promoter-luciferase constructs were created to identify proximal −27 to −107 promoter region responsible for MEK-dependent stimulation of CIP2A expression. Additional mutagenesis and chromatin immunoprecipitation experiments revealed ETS1 as the transcription factor mediating stimulation of CIP2A expression through EGFR-MEK pathway. Thus, ETS1 is probably mediating high CIP2A expression in human cancers with increased EGFR-MEK1/2-ERK pathway activity. These results also suggest that in addition to its established role in invasion and angiogenesis, ETS1 may support malignant cellular growth via regulation of CIP2A expression and protein phosphatase 2A inhibition.
PMCID: PMC3062549  PMID: 21445343
12.  Allelic Variation at the 8q23.3 Colorectal Cancer Risk Locus Functions as a Cis-Acting Regulator of EIF3H 
PLoS Genetics  2010;6(9):e1001126.
Common genetic variation at human 8q23.3 is significantly associated with colorectal cancer (CRC) risk. To elucidate the basis of this association we compared the frequency of common variants at 8q23.3 in 1,964 CRC cases and 2,081 healthy controls. Reporter gene studies showed that the single nucleotide polymorphism rs16888589 acts as an allele-specific transcriptional repressor. Chromosome conformation capture (3C) analysis demonstrated that the genomic region harboring rs16888589 interacts with the promoter of gene for eukaryotic translation initiation factor 3, subunit H (EIF3H). We show that increased expression of EIF3H gene increases CRC growth and invasiveness thereby providing a biological mechanism for the 8q23.3 association. These data provide evidence for a functional basis for the non-coding risk variant rs16888589 at 8q23.3 and provides novel insight into the etiological basis of CRC.
Author Summary
Common inherited variation on human chromosome 8q23 influences the risk of developing colorectal cancer (CRC). To understand the basis of this association we have compared the frequency of common genetic variants at 8q23 in ∼2,000 CRC cases and ∼2,000 healthy controls. Functional analyses of variants strongly associated with CRC risk showed that the single nucleotide polymorphism rs16888589 underscores the 8q23.3 association. The region of the genome harboring rs16888589 increases the expression of the gene for eukaryotic translation initiation factor 3, subunit H. We show that increased expression of this gene increases CRC growth thereby providing a biological mechanism for the 8q23.3 association. This finding is of particular importance in elucidating the etiological basis of CRC.
PMCID: PMC2940760  PMID: 20862326
13.  Probabilistic analysis of gene expression measurements from heterogeneous tissues 
Bioinformatics  2010;26(20):2571-2577.
Motivation: Tissue heterogeneity, arising from multiple cell types, is a major confounding factor in experiments that focus on studying cell types, e.g. their expression profiles, in isolation. Although sample heterogeneity can be addressed by manual microdissection, prior to conducting experiments, computational treatment on heterogeneous measurements have become a reliable alternative to perform this microdissection in silico. Favoring computation over manual purification has its advantages, such as time consumption, measuring responses of multiple cell types simultaneously, keeping samples intact of external perturbations and unaltered yield of molecular content.
Results: We formalize a probabilistic model, DSection, and show with simulations as well as with real microarray data that DSection attains increased modeling accuracy in terms of (i) estimating cell-type proportions of heterogeneous tissue samples, (ii) estimating replication variance and (iii) identifying differential expression across cell types under various experimental conditions. As our reference we use the corresponding linear regression model, which mirrors the performance of the majority of current non-probabilistic modeling approaches.
Availability and Software: All codes are written in Matlab, and are freely available upon request as well as at the project web page∼erkkila2/. Furthermore, a web-application for DSection exists at
PMCID: PMC2951082  PMID: 20631160
15.  Generation of a Conditionally Replicating Adenovirus Based on Targeted Destruction of E1A mRNA by a Cell Type-Specific MicroRNA▿  
Journal of Virology  2008;82(22):11009-11015.
MicroRNAs have emerged as important players in tissue-specific mammalian gene regulation and have also been exploited in experimental targeting of gene expression. We have constructed a recombinant adenovirus that contains sequences complementary to the liver-specific microRNA 122 (miR122) in the 3′ untranslated region of the E1A gene. In Huh7 cells, which resemble normal hepatocytes in expressing high levels of miR122, this feature resulted in strongly reduced levels of E1A mRNA and protein. This property allowed us to generate a novel recombinant adenovirus that was severely attenuated in cells of hepatic origin but replicated normally in other cells. This strategy may be useful in circumventing liver toxicity associated with the systemic delivery of oncolytic adenoviruses. These data provide the first example of exploiting differential microRNA expression patterns to alter the natural tropism of a DNA virus. In addition, these results suggest that other microRNAs expressed in a tissue- or transformation-specific manner may also be used for the targeting of adenoviral replication and that the same principle may be applied to other viruses that have shown promise as oncolytic or gene delivery platforms.
PMCID: PMC2573287  PMID: 18799589
16.  Somatic mutation analysis of MYH11 in breast and prostate cancer 
BMC Cancer  2008;8:263.
MYH11 (also known as SMMHC) encodes the smooth-muscle myosin heavy chain, which has a key role in smooth muscle contraction. Inversion at the MYH11 locus is one of the most frequent chromosomal aberrations found in acute myeloid leukemia. We have previously shown that MYH11 mutations occur in human colorectal cancer, and may also be associated with Peutz-Jeghers syndrome. The mutations found in human intestinal neoplasia result in unregulated proteins with constitutive motor activity, similar to the mutant myh11 underlying the zebrafish meltdown phenotype characterized by disrupted intestinal architecture. Recently, MYH1 and MYH9 have been identified as candidate breast cancer genes in a systematic analysis of the breast cancer genome.
The aim of this study was to investigate the role of somatic MYH11 mutations in two common tumor types; breast and prostate cancers. A total of 155 breast cancer and 71 prostate cancer samples were analyzed for those regions in MYH11 (altogether 8 exons out of 42 coding exons) that harboured mutations in colorectal cancer in our previous study.
In breast cancer samples only germline alterations were observed. One prostate cancer sample harbored a frameshift mutation c.5798delC, which we have previously shown to result in a protein with unregulated motor activity.
Little evidence for a role of somatic MYH11 mutations in the formation of breast or prostate cancers was obtained in this study.
PMCID: PMC2562392  PMID: 18796164
17.  Androgen regulation of the androgen receptor coregulators 
BMC Cancer  2008;8:219.
The critical role of the androgen receptor (AR) in the development of prostate cancer is well recognized. The transcriptional activity of AR is partly regulated by coregulatory proteins. It has been suggested that these coregulators could also be important in the progression of prostate cancer. The aim of this study was to identify coregulators whose expression is regulated by either the androgens and/or by the expression level of AR.
We used empty vector and AR cDNA-transfected LNCaP cells (LNCaP-pcDNA3.1, and LNCaP-ARhi, respectively), and grew them for 4 and 24 hours in the presence of dihydrotestosterone (DHT) at various concentrations. The expression of 25 AR coregulators (SRC1, TIF2, PIAS1, PIASx, ARIP4, BRCA1, β-catenin, AIB3, AIB1, CBP, STAT1, NCoR1, AES, cyclin D1, p300, ARA24, LSD1, BAG1L, gelsolin, prohibitin, JMJD2C, JMJD1A, MAK, PAK6 and MAGE11) was then measured by using real-time quantitative RT-PCR (Q-RT-PCR).
Five of the coregulators (AIB1, CBP, MAK, BRCA1 and β-catenin) showed more than 2-fold induction and 5 others (cyclin D1, gelsolin, prohibitin, JMJD1A, and JMJD2C) less than 2-fold induction. Overexpression of AR did not affect the expression of the coregulators alone. However, overexpression of AR enhanced the DHT-stimulated expression of MAK, BRCA1, AIB1 and CBP and reduced the level of expression of β-catenin, cyclinD1 and gelsolin.
In conclusion, we identified 5 coactivators whose expression was induced by androgens suggesting that they could potentiate AR signaling. Overexpression of AR seems to sensitize cells for low levels of androgens.
PMCID: PMC2518564  PMID: 18673534
18.  Different gDNA Content in the Subpopulations of Prostate Cancer Extracellular Vesicles: Apoptotic Bodies, Microvesicles, and Exosomes 
The Prostate  2014;74(14):1379-1390.
Extracellular vesicles (EVs) are cell-derived membrane vesicles. EVs contain several RNAs such as mRNA, microRNAs, and ncRNAs, but less is known of their genomic DNA (gDNA) content. It is also unknown whether the DNA cargo is randomly sorted or if it is systematically packed into specific EV subpopulations. The aim of this study was to analyze whether different prostate cancer (PCa) cell-derived EV subpopulations (apoptotic bodies, microvesicles, and exosomes) carry different gDNA fragments.
EV subpopulations were isolated from three PCa cell lines (LNCaP, PC-3, and RC92a/hTERT) and the plasma of PCa patients and healthy donors, and characterized by transmission electron microscopy, nanoparticle tracking analysis and total protein content. gDNA fragments of different genes were detected by real time quantitative PCR and confirmed by DNA sequencing.
We report that the concentration of EVs was higher in the cancer patients than in the healthy controls. EV subpopulations differed from each other in terms of total protein and DNA content. Analysis of gDNA fragments of MLH1, PTEN, and TP53 genes from the PCa cell-derived EV subpopulations showed that different EVs carried different gDNA content, which could even harbor specific mutations. Altogether, these results suggest that both nucleic acids and proteins are selectively and cell-dependently packed into the EV subtypes.
EVs derived from PCa cell lines and human plasma samples contain double-stranded gDNA fragments which could be used to detect specific mutations, making EVs potential biomarkers for cancer diagnostics and prognostics.
PMCID: PMC4312964  PMID: 25111183

Results 1-18 (18)