Circulating tumor cells (CTC) released into blood from primary cancers and metastases reflect the current status of tumor genotypes, which are prone to changes. Here, we conducted the first ...comprehensive genomic profiling of CTCs using array-comparative genomic hybridization (CGH) and next-generation sequencing. We used the U.S. Food and Drug Administration-cleared CellSearch system, which detected CTCs in 21 of 37 patients (range, 1-202/7.5 mL sample) with stage IV colorectal carcinoma. In total, we were able to isolate 37 intact CTCs from six patients and identified in those multiple colorectal cancer-associated copy number changes, many of which were also present in the respective primary tumor. We then used massive parallel sequencing of a panel of 68 colorectal cancer-associated genes to compare the mutation spectrum in the primary tumors, metastases, and the corresponding CTCs from two of these patients. Mutations in known driver genes e.g., adenomatous polyposis coli (APC), KRAS, or PIK3CA found in the primary tumor and metastasis were also detected in corresponding CTCs. However, we also observed mutations exclusively in CTCs. To address whether these mutations were derived from a small subclone in the primary tumor or represented new variants of metastatic cells, we conducted additional deep sequencing of the primary tumor and metastasis and applied a customized statistical algorithm for analysis. We found that most mutations initially found only in CTCs were also present at subclonal level in the primary tumors and metastases from the same patient. This study paves the way to use CTCs as a liquid biopsy in patients with cancer, providing more effective options to monitor tumor genomes that are prone to change during progression, treatment, and relapse.
Deregulation of transcription factors (TFs) is an important driver of tumorigenesis, but non-invasive assays for assessing transcription factor activity are lacking. Here we develop and validate a ...minimally invasive method for assessing TF activity based on cell-free DNA sequencing and nucleosome footprint analysis. We analyze whole genome sequencing data for >1,000 cell-free DNA samples from cancer patients and healthy controls using a bioinformatics pipeline developed by us that infers accessibility of TF binding sites from cell-free DNA fragmentation patterns. We observe patient-specific as well as tumor-specific patterns, including accurate prediction of tumor subtypes in prostate cancer, with important clinical implications for the management of patients. Furthermore, we show that cell-free DNA TF profiling is capable of detection of early-stage colorectal carcinomas. Our approach for mapping tumor-specific transcription factor binding in vivo based on blood samples makes a key part of the noncoding genome amenable to clinical analysis.
Genomic alterations in metastatic prostate cancer remain incompletely characterized. Here we analyse 493 prostate cancer cases from the TCGA database and perform whole-genome plasma sequencing on 95 ...plasma samples derived from 43 patients with metastatic prostate cancer. From these samples, we identify established driver aberrations in a cancer-related gene in nearly all cases (97.7%), including driver gene fusions (TMPRSS2:ERG), driver focal deletions (PTEN, RYBP and SHQ1) and driver amplifications (AR and MYC). In serial plasma analyses, we observe changes in focal amplifications in 40% of cases. The mean time interval between new amplifications was 26.4 weeks (range: 5-52 weeks), suggesting that they represent rapid adaptations to selection pressure. An increase in neuron-specific enolase is accompanied by clonal pattern changes in the tumour genome, most consistent with subclonal diversification of the tumour. Our findings suggest a high plasticity of prostate cancer genomes with newly occurring focal amplifications as a driving force in progression.
The importance of long non-coding RNAs (lncRNAs) in the pathogenesis of various malignancies has been uncovered over the last few years. Their dysregulation often contributes to or is a result of ...tumour progression. In prostate cancer, the most common malignancy in men, lncRNAs can promote castration resistance, cell proliferation, invasion, and metastatic spread. Expression patterns of lncRNAs often change during tumour progression; their expression levels may constantly rise (e.g., HOX transcript antisense RNA, HOTAIR), or steadily decrease (e.g., downregulated RNA in cancer, DRAIC). In prostate cancer, lncRNAs likewise have diagnostic (e.g., prostate cancer antigen 3, PCA3), prognostic (e.g., second chromosome locus associated with prostate-1, SChLAP1), and predictive (e.g., metastasis-associated lung adenocarcinoma transcript-1, MALAT-1) functions. Considering their dynamic role in prostate cancer, lncRNAs may also serve as therapeutic targets, helping to prevent development of castration resistance, maintain stable disease, and prohibit metastatic spread.
Mechanical interaction between cells - specifically distortion of tensional homeostasis-emerged as an important aspect of breast cancer genesis and progression. We investigated the biophysical ...characteristics of mechanosensitive ion channels (MSCs) in the malignant MCF-7 breast cancer cell line. MSCs turned out to be the most abundant ion channel species and could be activated by negative pressure at the outer side of the cell membrane in a saturable manner. Assessing single channel conductance (GΛ) for different monovalent cations revealed an increase in the succession: Li(+) < Na(+) < K(+) ≈Rb(+) ≈ Cs(+). Divalent cations permeated also with the order: Ca(2+) < Ba(2+). Comparison of biophysical properties enabled us to identify MSCs in MCF-7 as ion channels formed by the Piezo1 protein. Using patch clamp technique no functional MSCs were observed in the benign MCF-10A mammary epithelial cell line. Blocking of MSCs by GsMTx-4 resulted in decreased motility of MCF-7, but not of MCF-10A cells, underscoring a possible role of Piezo1 in invasion and metastatic propagation. The role of Piezo1 in biology and progression of breast cancer is further substantiated by markedly reduced overall survival in patients with increased Piezo1 mRNA levels in the primary tumor.
Monoclonal antibodies targeting the Epidermal Growth Factor Receptor (EGFR), such as cetuximab and panitumumab, have evolved to important therapeutic options in metastatic colorectal cancer (CRC). ...However, almost all patients with clinical response to anti-EGFR therapies show disease progression within a few months and little is known about mechanism and timing of resistance evolution. Here we analyzed plasma DNA from ten patients treated with anti-EGFR therapy by whole genome sequencing (plasma-Seq) and ultra-sensitive deep sequencing of genes associated with resistance to anti-EGFR treatment such as KRAS, BRAF, PIK3CA, and EGFR. Surprisingly, we observed that the development of resistance to anti-EGFR therapies was associated with acquired gains of KRAS in four patients (40%), which occurred either as novel focal amplifications (n = 3) or as high level polysomy of 12p (n = 1). In addition, we observed focal amplifications of other genes recently shown to be involved in acquired resistance to anti-EGFR therapies, such as MET (n = 2) and ERBB2 (n = 1). Overrepresentation of the EGFR gene was associated with a good initial anti-EGFR efficacy. Overall, we identified predictive biomarkers associated with anti-EGFR efficacy in seven patients (70%), which correlated well with treatment response. In contrast, ultra-sensitive deep sequencing of KRAS, BRAF, PIK3CA, and EGFR did not reveal the occurrence of novel, acquired mutations. Thus, plasma-Seq enables the identification of novel mutant clones and may therefore facilitate early adjustments of therapies that may delay or prevent disease progression.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Patients with testicular germ cell tumors (TGCT) have an increased risk for venous thromboembolism (VTE). We identified risk factors for VTE in this patient cohort and developed a clinical risk ...model.
In this retrospective cohort study at the Medical University of Graz we included 657 consecutive TGCT patients across all clinical stages. A predictive model for VTE was developed and externally validated in 349 TGCT patients treated at the University Hospital Zurich.
Venous thromboembolic events occurred in 34 (5.2%) patients in the Graz cohort. In univariable competing risk analysis, higher clinical stage (cS) and a retroperitoneal lymphadenopathy (RPLN) were the strongest predictors of VTE (p<0.0001). As the presence of a RPLN with more than 5cm in greatest dimension without coexisting visceral metastases is classified as cS IIC, we constructed an empirical VTE risk model with the following four categories (12-month-cumulative incidence): cS IA-B 8/463 patients (1.7%), cS IS-IIB 5/86 patients (5.9%), cS IIC 3/21 patients (14.3%) and cS IIIA-C 15/70 patients (21.4%). This risk model was externally validated in the Zurich cohort (12-month-cumulative incidence): cS IA-B (0.5%), cS IS-IIB (6.0%), cS IIC (11.1%) and cS IIIA-C (19.1%). Our model had a significantly higher discriminatory performance than a previously published classifier (RPLN-VTE-risk-classifier) which is based on the size of RPLN alone (AUC-ROC: 0.75 vs. 0.63, p = 0.007).
According to our risk stratification, TGCT patients with cS IIC and cS III disease have a very high risk of VTE and may benefit from primary thromboprophylaxis for the duration of chemotherapy.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Summary Background Analysis of the Austrian Breast and Colorectal Cancer Study Group trial-12 (ABCSG-12) at 48 months' follow-up showed that addition of zoledronic acid to adjuvant endocrine therapy ...significantly improved disease-free survival. We have now assessed long-term clinical efficacy including disease-free survival and disease outcomes in patients receiving anastrozole or tamoxifen with or without zoledronic acid. Methods ABSCG-12 is a randomised, controlled, open-label, two-by-two factorial, multicentre trial in 1803 premenopausal women with endocrine-receptor-positive early-stage (stage I–II) breast cancer receiving goserelin (3·6 mg every 28 days), comparing the efficacy and safety of anastrozole (1 mg per day) or tamoxifen (20 mg per day) with or without zoledronic acid (4 mg every 6 months) for 3 years. Randomisation (1:1:1:1 ratio) was computerised and based on the Pocock and Simon minimisation method to balance the four treatment arms across eight prognostic variables (age, neoadjuvant chemotherapy, pathological tumour stage; lymph-node involvement, type of surgery or locoregional therapy, complete axillary dissection, intraoperative radiation therapy, and geographical region). Treatment allocation was not masked. The primary endpoint was disease-free survival (defined as disease recurrence or death) and analysis was by intention to treat. This trial is registered with ClinicalTrials.gov , number NCT00295646 ; follow-up is ongoing. Findings At a median follow-up of 62 months (range 0–114·4 months), more than 2 years after treatment completion, 186 disease-free survival events had been reported (53 events in 450 patients on tamoxifen alone, 57 in 453 patients on anastrozole alone, 36 in 450 patients on tamoxifen plus zoledronic acid, and 40 in 450 patients on anastrozole plus zoledronic acid). Zoledronic acid reduced risk of disease-free survival events overall (HR 0·68, 95% CI 0·51–0·91; p=0·009), although the difference was not significant in the tamoxifen (HR 0·67, 95% CI 0·44–1·03; p=0·067) and anastrozole arms (HR 0·68, 95% CI 0·45–1·02; p=0·061) assessed separately. Zoledronic acid did not significantly affect risk of death (30 deaths with zoledronic acid vs 43 deaths without; HR 0·67, 95% CI 0·41–1·07; p=0·09). There was no difference in disease-free survival between patients on tamoxifen alone versus anastrozole alone (HR 1·08, 95% CI 0·81–1·44; p=0·591), but overall survival was worse with anastrozole than with tamoxifen (46 vs 27 deaths; HR 1·75, 95% CI 1·08–2·83; p=0·02). Treatments were generally well tolerated, with no reports of renal failure or osteonecrosis of the jaw. Bone pain was reported in 601 patients (33%; 349 patients on zoledronic acid vs 252 not on the drug), fatigue in 361 (20%; 192 vs 169), headache in 280 (16%; 147 vs 133), and arthralgia in 266 (15%; 145 vs 121). Interpretation Addition of zoledronic acid improved disease-free survival in the patients taking anastrozole or tamoxifen. There was no difference in disease-free survival between patients receiving anastrozole and tamoxifen overall, but those on anastrozole alone had inferior overall survival. These data show persistent benefits with zoledronic acid and support its addition to adjuvant endocrine therapy in premenopausal patients with early-stage breast cancer. Funding AstraZeneca; Novartis.
We hypothesize that measurement of gene expression related to estrogen receptor α (ER; gene name ESR1) within a breast cancer sample represents intrinsic tumoral sensitivity to adjuvant endocrine ...therapy.
A genomic index for sensitivity to endocrine therapy (SET) index was defined from genes coexpressed with ESR1 in 437 microarray profiles from newly diagnosed breast cancer, unrelated to treatment or outcome. The association of SET index and ESR1 levels with distant relapse risk was evaluated from microarrays of ER-positive breast cancer in two cohorts who received 5 years of tamoxifen alone as adjuvant endocrine therapy (n = 225 and 298, respectively), a cohort who received neoadjuvant chemotherapy followed by tamoxifen and/or aromatase inhibition (n = 122), and two cohorts who received no adjuvant systemic therapy (n = 208 and 133, respectively).
The SET index (165 genes) was significantly associated with distant relapse or death risk in both tamoxifen-treated cohorts (hazard ratio HR = 0.70, 95% CI, 0.56 to 0.88, P = .002; and HR = 0.76, 95% CI, 0.63 to 0.93, P = .007) and in the chemo-endocrine-treated cohort (HR = 0.19; 95% CI, 0.05 to 0.69, P = .011) independently from pathologic response to chemotherapy, but was not prognostic in two untreated cohorts. No distant relapse or death was observed after tamoxifen alone if node-negative and high SET or after chemo-endocrine therapy if intermediate or high SET.
The SET index of ER-related transcription predicted survival benefit from adjuvant endocrine therapy, not inherent prognosis. Prior chemotherapy seemed to enhance the efficacy of adjuvant endocrine therapy related to SET index.
Non-coding RNAs and especially microRNAs have been discovered to act as master regulators of cancer initiation and progression. The aim of our study was to discover and characterize the function of ...yet functionally uncharacterized microRNAs in human breast carcinogenesis.
In an unbiased approach, we utilized an established model system for breast cancer (BC) stem cell formation ("mammosphere assay") to identify whole miRNome alterations in breast carcinogenesis. Clinical samples of BC patients were used to evaluate the human relevance of the newly identified miRNA candidates. One promising candidate, miR-1287-5p, was further explored on its impact on several hallmarks of cancer. The molecular mode of action was characterized by whole transcriptome analysis, in silico prediction tools, miRNA-interaction assays, pheno-copy assays, and drug sensitivity assays.
Among several other microRNAs, miR-1287-5p was significantly downregulated in mammospheres and human BC tissue compared to normal breast tissue (p < 0.0001). Low expression levels were significantly associated with poor prognosis in BC patients. MiR-1287-5p significantly decreased cellular growth, cells in S phase of cell cycle, anchorage-independent growth, and tumor formation in vivo. In addition, we identified PIK3CB as a direct molecular interactor of miR-1287-5p and a novel prognostic factor in BC. Finally, PI3Kinase pathway chemical inhibitors combined with miR-1287-5p mimic increased the pharmacological growth inhibitory potential in triple negative BC cells.
Our data identified for the first time the involvement of miR-1287-5p in human BC and suggest a potential for therapeutic interventions in difficult to treat triple negative BC.
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