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Find video protocols related to scientific articles indexed in Pubmed.
Higher metastatic efficiency of KRas G12V than KRas G13D in a colorectal cancer model.
FASEB J.
PUBLISHED: 11-01-2014
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Although all KRas (protein that in humans is encoded by the KRas gene) point mutants are considered to have a similar prognostic capacity, their transformation and tumorigenic capacities vary widely. We compared the metastatic efficiency of KRas G12V (Kirsten rat sarcoma viral oncogene homolog with valine mutation at codon 12) and KRas G13D (Kirsten rat sarcoma viral oncogene homolog with aspartic mutation at codon 13) oncogenes in an orthotopic colorectal cancer (CRC) model. Following subcutaneous preconditioning, recombinant clones of the SW48 CRC cell line [Kras wild-type (Kras WT)] expressing the KRas G12V or KRas G13D allele were microinjected in the mouse cecum. The percentage of animals developing lymph node metastasis was higher in KRas G12V than in KRas G13D mice. Microscopic, macroscopic, and visible lymphatic foci were 1.5- to 3.0-fold larger in KRas G12V than in KRas G13D mice (P < 0.05). In the lung, only microfoci were developed in both groups. KRas G12V primary tumors had lower apoptosis (7.0 ± 1.2 vs. 7.4 ± 1.0 per field, P = 0.02), higher tumor budding at the invasion front (1.2 ± 0.2 vs. 0.6 ± 0.1, P = 0.04), and a higher percentage of C-X-C chemokine receptor type 4 (CXCR4)-overexpressing intravasated tumor emboli (49.8 ± 9.4% vs. 12.8 ± 4.4%, P < 0.001) than KRas G13D tumors. KRas G12V primary tumors showed Akt activation, and ?5 integrin, vascular endothelial growth factor A (VEGFA), and Serpine-1 overexpression, whereas KRas G13D tumors showed integrin ?1 and angiopoietin 2 (Angpt2) overexpression. The increased cell survival, invasion, intravasation, and specific molecular regulation observed in KRas G12V tumors is consistent with the higher aggressiveness observed in patients with CRC expressing this oncogene.-Alamo, P., Gallardo, A., Di Nicolantonio, F., Pavón, M. A., Casanova, I., Trias, M., Mangues, M. A., Lopez-Pousa, A., Villaverde, A., Vázquez, E., Bardelli, A., Céspedes, M. V., Mangues, R. Higher metastatic efficiency of KRas G12V than KRas G13D in a colorectal cancer model.
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Circulating pEGFR is a Candidate Response Biomarker of Cetuximab Therapy in Colorectal Cancer.
Clin. Cancer Res.
PUBLISHED: 10-18-2014
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Purpose:The lack of secreted biomarkers measurable by non-invasive tests hampers the development of effective targeted therapies against cancer. Our hypothesis is that cetuximab (an anti-EGFR mAb) induces a specific secretome in colorectal cancer (CRC) cells that could be exploited for biomarker discovery. Experimental Design:Considering the strong correlation between mutated-KRAS and a lack of response to cetuximab therapy, we addressed whether performing secretome-based proteomics on isogenic CRC cells sharing the KRAS-mutations found on patients would yield candidate secreted biomarkers useful in the clinical setting. Since 2D-culture did not optimally model the sensitivity/resistance to cetuximab observed in CRC patients, we moved to 3D-spheroids, developing a methodology for both cell-based assays and quantitative proteomics. Results:A large comparative quantitative proteomic analysis of the 3D-secretomes of CRC isogenic cells treated with cetuximab uncovered an EGFR pathway-centric secretome found only when cells grow in 3D. The validation of the secretome findings in plasma of CRC patients, suggests that phosphorylated-EGFR (pEGFR) is a candidate secreted biomarker of response to cetuximab. Conclusions:We have proved that 3D-spheroids from CRC cells generate secretomes with a drug sensitivity profile that correlates well with CRC patients, illustrating molecular connections between intracellular and extracellular signaling. Furthermore, we show how the secretion of pEGFR is associated with the sensitivity of CRC cells to cetuximab and the response of CRC patients to the drug. Our work could allow the non-invasive monitoring of anti-EGFR treatment in CRC patients.
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Resistance to Anti-EGFR Therapy in Colorectal Cancer: From Heterogeneity to Convergent Evolution.
Cancer Discov
PUBLISHED: 10-07-2014
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The EGFR-targeted antibodies cetuximab and panitumumab are used to treat metastatic colorectal cancers. Mutations in KRAS, NRAS, and BRAF and amplification of ERBB2 and MET drive primary (de novo) resistance to anti-EGFR treatment. Recently, the emergence of alterations in the same genes was detected in patients who responded to EGFR blockade and then relapsed. These results illuminate a striking overlap between genes that, when mutated, drive primary and secondary resistance to anti-EGFR antibodies. Remarkably, although the mechanisms of resistance are genetically heterogeneous, they biochemically converge on key signaling pathways. This knowledge is being translated in the rational design of additional lines of therapy.
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Genotyping cell-free tumor DNA in the blood to detect residual disease and drug resistance.
Genome Biol.
PUBLISHED: 08-30-2014
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DNA fragments released from cancer cells into the blood can be used to generate molecular profiles of tumors. Non-invasive 'liquid biopsies' can be used to monitor minimal residual disease and detect the emergence of drug resistance.
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RAF suppression synergizes with MEK inhibition in KRAS mutant cancer cells.
Cell Rep
PUBLISHED: 06-12-2014
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KRAS is the most frequently mutated oncogene in human cancer, yet no therapies are available to treat KRAS mutant cancers. We used two independent reverse genetic approaches to identify components of the RAS-signaling pathways required for growth of KRAS mutant tumors. Small interfering RNA (siRNA) screening of 37 KRAS mutant colorectal cancer cell lines showed that RAF1 suppression was synthetic lethal with MEK inhibition. An unbiased kinome short hairpin RNA (shRNA)-based screen confirmed this synthetic lethal interaction in colorectal as well as in lung cancer cells bearing KRAS mutations. Compounds targeting RAF kinases can reverse resistance to the MEK inhibitor selumetinib. MEK inhibition induces RAS activation and BRAF-RAF1 dimerization and sustains MEK-ERK signaling, which is responsible for intrinsic resistance to selumetinib. Prolonged dual blockade of RAF and MEK leads to persistent ERK suppression and efficiently induces apoptosis. Our data underlie the relevance of developing combinatorial regimens of drugs targeting the RAF-MEK pathway in KRAS mutant tumors.
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TGF-? and amphiregulin paracrine network promotes resistance to EGFR blockade in colorectal cancer cells.
Clin. Cancer Res.
PUBLISHED: 06-12-2014
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Purpose: Targeted inhibition of EGFR with the monoclonal antibodies cetuximab or panitumumab is a valuable treatment for RAS wild type colorectal cancers. The efficacy of EGFR blockade is limited by the emergence of acquired resistance often attributed to secondary KRAS mutations. Remarkably, tumor biopsies from resistant patients show that only a fraction of the resilient cells carry KRAS mutations. We hypothesized that a paracrine crosstalk driven by the resistant subpopulation may provide in trans protection of surrounding sensitive cells. Experimental design: Conditioned medium assays and three dimensional co-cultures were used to assess paracrine networks between cetuximab sensitive and resistant cells. Production of EGFR ligands by cells sensitive to cetuximab and panitumumab was measured. The ability of recombinant EGFR ligands to protect sensitive cells from cetuximab was assessed. Biochemical activation of the EGFR signaling pathway was measured by western blotting. Results: CRC cells sensitive to EGFR blockade can successfully grow despite cetuximab treatment when in the company of their resistant derivatives. Media conditioned by resistant cells protect sensitive parental cells from cetuximab. EGFR blockade triggers increased secretion of TGF? and amphiregulin. Increased secretion of ligands by resistant cells can sustain EGFR/ERK signaling in sensitive cells. Conclusions: CRC cells that develop resistance to cetuximab and panitumumab secrete TGF-? and amphiregulin, which protect the surrounding cells from EGFR blockade. This paracrine protective mechanism might be therapeutically exploitable.
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Mutational profiling of kinases in glioblastoma.
BMC Cancer
PUBLISHED: 04-14-2014
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Glioblastoma is a highly malignant brain tumor for which no cure is available. To identify new therapeutic targets, we performed a mutation analysis of kinase genes in glioblastoma.
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Minimal residual disease in breast cancer: in blood veritas.
Clin. Cancer Res.
PUBLISHED: 03-21-2014
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A blood-based molecular test might direct recommendations for systemic therapies in patients with early-stage breast cancer undergoing surgery with curative intent. A new study suggests that droplet digital PCR (ddPCR) can be used to detect cancer-specific DNA alterations in plasma with sensitivity suitable for monitoring minimal residual disease.
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Blockade of EGFR and MEK intercepts heterogeneous mechanisms of acquired resistance to anti-EGFR therapies in colorectal cancer.
Sci Transl Med
PUBLISHED: 02-21-2014
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Colorectal cancers (CRCs) that are sensitive to the anti-epidermal growth factor receptor (EGFR) antibodies cetuximab or panitumumab almost always develop resistance within several months of initiating therapy. We report the emergence of polyclonal KRAS, NRAS, and BRAF mutations in CRC cells with acquired resistance to EGFR blockade. Regardless of the genetic alterations, resistant cells consistently displayed mitogen-activated protein kinase kinase (MEK) and extracellular signal-regulated kinase (ERK) activation, which persisted after EGFR blockade. Inhibition of MEK1/2 alone failed to impair the growth of resistant cells in vitro and in vivo. An RNA interference screen demonstrated that suppression of EGFR, together with silencing of MEK1/2, was required to hamper the proliferation of resistant cells. Indeed, concomitant pharmacological blockade of MEK and EGFR induced prolonged ERK inhibition and severely impaired the growth of resistant tumor cells. Heterogeneous and concomitant mutations in KRAS and NRAS were also detected in plasma samples from patients who developed resistance to anti-EGFR antibodies. A mouse xenotransplant from a CRC patient who responded and subsequently relapsed upon EGFR therapy showed exquisite sensitivity to combinatorial treatment with MEK and EGFR inhibitors. Collectively, these results identify genetically distinct mechanisms that mediate secondary resistance to anti-EGFR therapies, all of which reactivate ERK signaling. These observations provide a rational strategy to overcome the multifaceted clonal heterogeneity that emerges when tumors are treated with targeted agents. We propose that MEK inhibitors, in combination with cetuximab or panitumumab, should be tested in CRC patients who become refractory to anti-EGFR therapies.
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Detection of circulating tumor DNA in early- and late-stage human malignancies.
Sci Transl Med
PUBLISHED: 02-21-2014
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The development of noninvasive methods to detect and monitor tumors continues to be a major challenge in oncology. We used digital polymerase chain reaction-based technologies to evaluate the ability of circulating tumor DNA (ctDNA) to detect tumors in 640 patients with various cancer types. We found that ctDNA was detectable in >75% of patients with advanced pancreatic, ovarian, colorectal, bladder, gastroesophageal, breast, melanoma, hepatocellular, and head and neck cancers, but in less than 50% of primary brain, renal, prostate, or thyroid cancers. In patients with localized tumors, ctDNA was detected in 73, 57, 48, and 50% of patients with colorectal cancer, gastroesophageal cancer, pancreatic cancer, and breast adenocarcinoma, respectively. ctDNA was often present in patients without detectable circulating tumor cells, suggesting that these two biomarkers are distinct entities. In a separate panel of 206 patients with metastatic colorectal cancers, we showed that the sensitivity of ctDNA for detection of clinically relevant KRAS gene mutations was 87.2% and its specificity was 99.2%. Finally, we assessed whether ctDNA could provide clues into the mechanisms underlying resistance to epidermal growth factor receptor blockade in 24 patients who objectively responded to therapy but subsequently relapsed. Twenty-three (96%) of these patients developed one or more mutations in genes involved in the mitogen-activated protein kinase pathway. Together, these data suggest that ctDNA is a broadly applicable, sensitive, and specific biomarker that can be used for a variety of clinical and research purposes in patients with multiple different types of cancer.
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Intrinsic resistance to MEK inhibition in KRAS mutant lung and colon cancer through transcriptional induction of ERBB3.
Cell Rep
PUBLISHED: 02-17-2014
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There are no effective therapies for the ~30% of human malignancies with mutant RAS oncogenes. Using a kinome-centered synthetic lethality screen, we find that suppression of the ERBB3 receptor tyrosine kinase sensitizes KRAS mutant lung and colon cancer cells to MEK inhibitors. We show that MEK inhibition results in MYC-dependent transcriptional upregulation of ERBB3, which is responsible for intrinsic drug resistance. Drugs targeting both EGFR and ERBB2, each capable of forming heterodimers with ERBB3, can reverse unresponsiveness to MEK inhibition by decreasing inhibitory phosphorylation of the proapoptotic proteins BAD and BIM. Moreover, ERBB3 protein level is a biomarker of response to combinatorial treatment. These data suggest a combination strategy for treating KRAS mutant colon and lung cancers and a way to identify the tumors that are most likely to benefit from such combinatorial treatment.
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Somatic alterations as the basis for resistance to targeted therapies.
J. Pathol.
PUBLISHED: 02-08-2014
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Recent advances in genetics and genomics have revealed new genes and pathways that are somatically altered in human malignancies. This wealth of knowledge has translated into molecularly defined targets for therapy over the past two decades, serving as key examples that translation of laboratory findings can have great impact on the ability to treat patients with cancer. However, given the genetic instability and heterogeneity that are characteristic of all human cancers, drug resistance to virtually all therapies has emerged, posing further and future challenges for clinical oncology. Here we review the history of targeted therapies, including examples of genetically defined cancer targets and their approved therapies. We also discuss resistance mechanisms that have been uncovered, with an emphasis on somatic genetic alterations that lead to these phenotypes.
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The combination of IDH1 mutations and MGMT methylation status predicts survival in glioblastoma better than either IDH1 or MGMT alone.
Neuro-oncology
PUBLISHED: 02-06-2014
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Genetic and epigenetic profiling of glioblastomas has provided a comprehensive list of altered cancer genes of which only O(6)-methylguanine-methyltransferase (MGMT) methylation is used thus far as a predictive marker in a clinical setting. We investigated the prognostic significance of genetic and epigenetic alterations in glioblastoma patients.
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Reversible and adaptive resistance to BRAF(V600E) inhibition in melanoma.
Nature
PUBLISHED: 01-31-2014
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Treatment of BRAF(V600E) mutant melanoma by small molecule drugs that target the BRAF or MEK kinases can be effective, but resistance develops invariably. In contrast, colon cancers that harbour the same BRAF(V600E) mutation are intrinsically resistant to BRAF inhibitors, due to feedback activation of the epidermal growth factor receptor (EGFR). Here we show that 6 out of 16 melanoma tumours analysed acquired EGFR expression after the development of resistance to BRAF or MEK inhibitors. Using a chromatin-regulator-focused short hairpin RNA (shRNA) library, we find that suppression of sex determining region Y-box 10 (SOX10) in melanoma causes activation of TGF-? signalling, thus leading to upregulation of EGFR and platelet-derived growth factor receptor-? (PDGFRB), which confer resistance to BRAF and MEK inhibitors. Expression of EGFR in melanoma or treatment with TGF-? results in a slow-growth phenotype with cells displaying hallmarks of oncogene-induced senescence. However, EGFR expression or exposure to TGF-? becomes beneficial for proliferation in the presence of BRAF or MEK inhibitors. In a heterogeneous population of melanoma cells having varying levels of SOX10 suppression, cells with low SOX10 and consequently high EGFR expression are rapidly enriched in the presence of drug, but this is reversed when the drug treatment is discontinued. We find evidence for SOX10 loss and/or activation of TGF-? signalling in 4 of the 6 EGFR-positive drug-resistant melanoma patient samples. Our findings provide a rationale for why some BRAF or MEK inhibitor-resistant melanoma patients may regain sensitivity to these drugs after a 'drug holiday' and identify patients with EGFR-positive melanoma as a group that may benefit from re-treatment after a drug holiday.
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Acquired resistance to EGFR-targeted therapies in colorectal cancer.
Mol Oncol
PUBLISHED: 01-22-2014
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Cetuximab and panitumumab are anti-epidermal growth factor receptor (anti-EGFR) monoclonal antibodies used as therapies for metastatic colorectal cancer patients. Intrinsic mechanisms of resistance, such as RAS mutations, can prevent patients from having a response with clinical benefit. The clinical efficacy of EGFR targeted antibodies is limited by the development of acquired (secondary) resistance, which typically occurs within 3-12 months from the start of therapy. Preclinical models and analyses of clinical samples have uncovered some of the alterations that confer a selective advantage to tumor cells when under the pressure of anti-EGFR therapy. Molecular profiling of clinical specimens confirmed that genetic alterations of genes in the EGFR-RAS-RAF-MEK signaling pathway and of receptor tyrosine kinases are mechanisms of acquired resistance to anti-EGFR antibodies. The escape from anti-EGFR blockade appears to converge on the (re)activation of MEK-ERK or AKT as revealed in preclinical studies. Circulating tumor DNA and patient derived xenografts have proven useful tools to monitor patients for resistance to anti-EGFR therapy and test combination therapies to overcome or reverse resistance.
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Liquid biopsies: genotyping circulating tumor DNA.
J. Clin. Oncol.
PUBLISHED: 01-21-2014
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Genotyping tumor tissue in search of somatic genetic alterations for actionable information has become routine practice in clinical oncology. Although these sequence alterations are highly informative, sampling tumor tissue has significant inherent limitations; tumor tissue is a single snapshot in time, is subject to selection bias resulting from tumor heterogeneity, and can be difficult to obtain. Cell-free fragments of DNA are shed into the bloodstream by cells undergoing apoptosis or necrosis, and the load of circulating cell-free DNA (cfDNA) correlates with tumor staging and prognosis. Moreover, recent advances in the sensitivity and accuracy of DNA analysis have allowed for genotyping of cfDNA for somatic genomic alterations found in tumors. The ability to detect and quantify tumor mutations has proven effective in tracking tumor dynamics in real time as well as serving as a liquid biopsy that can be used for a variety of clinical and investigational applications not previously possible.
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Climbing RAS, the everest of oncogenes.
Cancer Discov
PUBLISHED: 01-10-2014
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Mutations that activate the small GTP-binding protein KRAS are the most common oncogenic event in human tumors. Thirty years after its discovery, mutant KRAS has yet to be therapeutically conquered.
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BRAF V600E Is a Determinant of Sensitivity to Proteasome Inhibitors.
Mol. Cancer Ther.
PUBLISHED: 10-09-2013
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A critical step toward defining tailored therapy in patients with cancer is the identification of genetic interactions that may impair-or boost-the efficacy of selected therapeutic approaches. Cell models able to recapitulate combinations of genetic aberrations are important to find drug-genotype interactions poorly affected by the heterogeneous genetics of human tumors. In order to identify novel pharmacogenomic relationships, we employed an isogenic cell panel that reconstructs cancer genetic scenarios. We screened a library of 43 compounds in human hTERT-HME1 epithelial cells in which PTEN or RB1 were silenced in combination with the targeted knockin of cancer-associated mutations in EGFR, KRAS, BRAF, or PIK3CA oncogenes. Statistical analysis and clustering algorithms were applied to display similar drug response profiles and mutation-specific patterns of activity. From the screen, we discovered that proteasome inhibitors show selectivity toward BRAF V600E-mutant cells, irrespective of PTEN or RB1 expression. Preferential targeting of BRAF-mutant cells by proteasome inhibitors was corroborated in a second BRAF V600E isogenic model, as well as in a panel of colorectal cancer cell lines by the use of the proteasome inhibitor carfilzomib. Notably, carfilzomib also showed striking in vivo activity in a BRAF-mutant human colorectal cancer xenograft model. Vulnerability to proteasome inhibitors is dependent on persistent BRAF signaling, because BRAF V600E blockade by PLX4720 reversed sensitivity to carfilzomib in BRAF-mutant colorectal cancer cells. Our findings indicated that proteasome inhibition might represent a valuable targeting strategy in BRAF V600E-mutant colorectal tumors. Mol Cancer Ther; 12(12); 2950-61. ©2013 AACR.
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Monitoring tumor-derived cell-free DNA in patients with solid tumors: Clinical perspectives and research opportunities.
Cancer Treat. Rev.
PUBLISHED: 10-07-2013
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Circulating cell-free DNA represents a non-invasive biomarker, as it can be isolated from human plasma, serum and other body fluids. Circulating tumor DNA shed from primary and metastatic cancers may allow the non-invasive analysis of the evolution of tumor genomes during treatment and disease progression through liquid biopsies. The serial monitoring of tumor genotypes, which are instable and prone to changes under selection pressure, is becoming increasingly possible. The "liquid biopsy" provide novel biological insights into the process of metastasis and may elucidate signaling pathways involved in cell invasiveness and metastatic competence. This review will focus on the clinical utility of circulating cell free DNA in main solid tumors, including genetic and epigenetic alterations that can be detected.
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Targeted Knock-in of the Polymorphism rs61764370 Does Not Affect KRAS Expression but Reduces let-7 Levels.
Hum. Mutat.
PUBLISHED: 07-17-2013
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Understanding the role of single-nucleotide polymorphisms (SNPs) in the pathological process represents a unique experimental challenge especially when the variants occur outside of coding regions. The noncoding SNP rs61764370 located in the 3-untranslated region of Kirsten rat sarcoma viral oncogene homolog (KRAS) has been implicated as a risk factor for the development of cancer and the response to targeted therapies. This cancer-associated variant is thought to affect the binding of the microRNA let-7, which allegedly modulates KRAS expression. Using site-specific homologous recombination, we inserted the rs61764370:T>G KRAS gene variant in the colorectal cancer cell line SW48 (SW48 +SNP) and assessed the cellular and biochemical phenotype. We observed a significant increase in cellular proliferation, as well as a reduction in the levels of the microRNA let-7a, let-7b, and let-7c. Transcriptional and biochemical analysis showed no concomitant change in the KRAS protein expression or modulation of the downstream mitogen activated kinase or PI3K/AKT signaling. These results suggest that the cancer-associated rs61764370 variant exerts a biological effect not through transcriptional modulation of KRAS but rather by tuning the expression of the microRNA let-7.
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Liquid biopsy: monitoring cancer-genetics in the blood.
Nat Rev Clin Oncol
PUBLISHED: 07-09-2013
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Cancer is associated with mutated genes, and analysis of tumour-linked genetic alterations is increasingly used for diagnostic, prognostic and treatment purposes. The genetic profile of solid tumours is currently obtained from surgical or biopsy specimens; however, the latter procedure cannot always be performed routinely owing to its invasive nature. Information acquired from a single biopsy provides a spatially and temporally limited snap-shot of a tumour and might fail to reflect its heterogeneity. Tumour cells release circulating free DNA (cfDNA) into the blood, but the majority of circulating DNA is often not of cancerous origin, and detection of cancer-associated alleles in the blood has long been impossible to achieve. Technological advances have overcome these restrictions, making it possible to identify both genetic and epigenetic aberrations. A liquid biopsy, or blood sample, can provide the genetic landscape of all cancerous lesions (primary and metastases) as well as offering the opportunity to systematically track genomic evolution. This Review will explore how tumour-associated mutations detectable in the blood can be used in the clinic after diagnosis, including the assessment of prognosis, early detection of disease recurrence, and as surrogates for traditional biopsies with the purpose of predicting response to treatments and the development of acquired resistance.
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Amplification of the MET receptor drives resistance to anti-EGFR therapies in colorectal cancer.
Cancer Discov
PUBLISHED: 06-02-2013
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EGF receptor (EGFR)-targeted monoclonal antibodies are effective in a subset of metastatic colorectal cancers. Inevitably, all patients develop resistance, which occurs through emergence of KRAS mutations in approximately 50% of the cases. We show that amplification of the MET proto-oncogene is associated with acquired resistance in tumors that do not develop KRAS mutations during anti-EGFR therapy. Amplification of the MET locus was present in circulating tumor DNA before relapse was clinically evident. Functional studies show that MET activation confers resistance to anti-EGFR therapy both in vitro and in vivo. Notably, in patient-derived colorectal cancer xenografts, MET amplification correlated with resistance to EGFR blockade, which could be overcome by MET kinase inhibitors. These results highlight the role of MET in mediating primary and secondary resistance to anti-EGFR therapies in colorectal cancer and encourage the use of MET inhibitors in patients displaying resistance as a result of MET amplification.
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Mouse models of Kras-mutant colorectal cancer: valuable GEMMs for drug testing?
Clin. Cancer Res.
PUBLISHED: 04-23-2013
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The development of effective therapies for colorectal cancer depends on the ability of preclinical models to faithfully recapitulate the molecular and biologic behavior of human tumors. This study reports on the characterization of colorectal genetically engineered mouse models and their derivative cell lines carrying wild-type or oncogenic Kras with concomitant Apc and p53 loss.
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Tivantinib (ARQ197) displays cytotoxic activity that is independent of its ability to bind MET.
Clin. Cancer Res.
PUBLISHED: 03-26-2013
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MET, the high-affinity receptor for hepatocyte growth factor, is frequently deregulated in human cancer. Tivantinib (ARQ197; Arqule), a staurosporine derivative that binds to the dephosphorylated MET kinase in vitro, is being tested clinically as a highly selective MET inhibitor. However, the mechanism of action of tivantinib is still unclear.
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Oncogenes and angiogenesis: a way to personalize anti-angiogenic therapy?
Cell. Mol. Life Sci.
PUBLISHED: 03-25-2013
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The acquisition of oncogenic mutations and promotion of angiogenesis are key hallmarks of cancer. These features are often thought of as separate events in tumor progression and the two fields of research have frequently been considered as independent. However, as we highlight in this review, activated oncogenes and deregulated angiogenesis are tightly associated, as mutations in cancer cells can lead to perturbation of the pro- and anti-angiogenic balance thereby causing aberrant angiogenesis. We propose that normalization of the vascular network by targeting oncogenes in the tumor cells might lead to more efficient and sustained therapeutic effects compared to therapies targeting tumor vessels. We discuss how pharmacological inhibition of oncogenes in tumor cells restores a functional vasculature by bystander anti-angiogenic effect. As genetic alterations are tumor-specific, targeted therapy, which potentially blocks the angiogenic program activated by individual oncogenes may lead to personalized anti-angiogenic therapy.
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KRAS gene amplification in colorectal cancer and impact on response to EGFR-targeted therapy.
Int. J. Cancer
PUBLISHED: 01-21-2013
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KRAS mutations are the most common oncogenic event in colorectal cancer (CRC) progression and their occurrence is associated with lack of response to anti epidermal growth factor receptor (EGFR) targeted therapies. Using preclinical models and patients samples we recently reported that the emergence of KRAS mutations but also KRAS amplification is associated with acquired resistance to the EGFR inhibitors cetuximab or panitumumab. We reasoned that KRAS amplification may also be responsible for primary resistance to these agents. Furthermore, while the prevalence of KRAS mutations has been well established in CRC, little is known about the frequency of KRAS amplification in large CRC series. We performed a screening of 1,039 CRC samples to assess the prevalence of KRAS amplification in this tumor type and further evaluated the role of this genetic alteration on the sensitivity to anti EGFR therapies. We detected KRAS amplification in 7/1,039 (0.67%) and 1/102 evaluable CRC specimens and cell lines, respectively. KRAS amplification was mutually exclusive with KRAS mutations. Tumors or cell lines harboring this genetic lesion are not responsive to anti-EGFR inhibitors. Although KRAS amplification is an infrequent event in CRC, it might be responsible for precluding response to anti-EGFR treatment in a small proportion of patients.
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Mixed lineage kinase MLK4 is activated in colorectal cancers where it synergistically cooperates with activated RAS signaling in driving tumorigenesis.
Cancer Res.
PUBLISHED: 01-14-2013
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Colorectal cancers (CRC) are commonly classified into those with microsatellite instability and those that are microsatellite stable (MSS) but chromosomally unstable. The latter are characterized by poor prognosis and remain largely intractable at the metastatic stage. Comprehensive mutational analyses have revealed that the mixed lineage kinase 4 (MLK4) protein kinase is frequently mutated in MSS CRC with approximately 50% of the mutations occurring in KRAS- or BRAF-mutant tumors. This kinase has not been characterized previously and the relevance of MLK4 somatic mutations in oncogenesis has not been established. We report that MLK4-mutated alleles in CRC are constitutively active and increase the transformation and tumorigenic capacity of RAS-mutated cell lines. Gene expression silencing or targeted knockout of MLK4 impairs the oncogenic properties of KRAS- and BRAF-mutant cancer cells both in vitro and in xenograft models. In establishing the role of MLK4 in intracellular signaling, we show it directly phosphorylates MEK1 (MAP2K1) and that MEK/ERK (MAPK1) signaling is impaired in MLK4 knockout cells. These findings suggest that MLK4 inhibitors may be efficacious in KRAS- and BRAF-mutated CRCs and may provide a new opportunity for targeting such recalcitrant tumors.
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Modeling tumor progression by the sequential introduction of genetic alterations into the genome of human normal cells.
Hum. Mutat.
PUBLISHED: 01-04-2013
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Cancer genomes display a complex blend of genetic lesions affecting oncogenes and tumor suppressor genes. Multiple modeling approaches indicate that 5-15 driver oncogenic events are required to achieve tumor progression in common epithelial cancers. In vitro, a lower number (2-3) of events is typically sufficient to achieve full transformation. We developed cellular models that closely resemble the occurrence of multiple genetic lesions to understand their role in tumor progression. Homologous recombination and transcriptional downregulation were used to recapitulate the co-occurrence of driver mutations targeting oncogenes and inactivation of tumor suppressor genes in human nontransformed epithelial cells. Knockdown of the tumor suppressor genes PTEN or RB1 was combined with mutagenic activation of individual oncogenes (EGFR, KRAS, BRAF, or PIK3CA), thus generating a combinatorial model. The simultaneous presence of oncogenic and tumor suppressive events resulted in distinct biochemical properties and anchorage-independent growth abilities. Notably, however, we found that even when up to four individual alterations were concomitantly present they were not sufficient to fully transform the target cells. Our results suggest that the close recapitulation of cancer lesions in not-transformed cells is essential to unveil their oncogenic potential and raise questions concerning the minimal requirements for neoplastic transformation of epithelial cells.
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Targeting oncogenic serine/threonine-protein kinase BRAF in cancer cells inhibits angiogenesis and abrogates hypoxia.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 12-27-2011
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Carcinomas are comprised of transformed epithelial cells that are supported in their growth by a dedicated neovasculature. How the genetic milieu of the epithelial compartment influences tumor angiogenesis is largely unexplored. Drugs targeted to mutant cancer genes may act not only on tumor cells but also, directly or indirectly, on the surrounding stroma. We investigated the role of the BRAF(V600E) oncogene in tumor/vessel crosstalk and analyzed the effect of the BRAF inhibitor PLX4720 on tumor angiogenesis. Knock-in of the BRAF(V600E) allele into the genome of human epithelial cells triggered their angiogenic response. In cancer cells harboring oncogenic BRAF, the inhibitor PLX4720 switches off the ERK pathway and inhibits the expression of proangiogenic molecules. In tumor xenografts harboring the BRAF(V600E), PLX4720 extensively modifies the vascular network causing abrogation of hypoxia. Overall, our results provide a functional link between oncogenic BRAF and angiogenesis. Furthermore, they indicate how the tumor vasculature can be "indirectly" besieged through targeting of a genetic lesion to which the cancer cells are addicted.
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Targeted therapies: how personal should we go?
Nat Rev Clin Oncol
PUBLISHED: 11-15-2011
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Despite the development of drugs inhibiting the oncogenic proteins that cancer cells are dependent on, attempts to match targeted therapies to the genetic makeup of individual tumors is proving more difficult than expected. Until now, the paradigm has been a binary correlation between a mutated cancer gene and response to a given therapy. However, recent evidence indicates that different genetic alterations, such as mutations in different codons of a cancer gene, might be related to distinct sensitivity to targeted therapies. An example is the divergent effect that individual EGFR, PIK3CA and KRAS mutations might have on response or resistance to tailored drugs. Furthermore, the idea that the presence of a specific mutation translates into sensitivity or resistance to a particular drug is likely too simplistic, since it does not capture the complexity of the signaling pathways in an individual cancer. Only the overall genetic milieu (alterations in upstream and/or parallel pathways) ultimately determines the response of individual tumors to therapy. We have critically analyzed data supporting the genetic, biological and biochemical differences of individual mutations within a single cancer gene. The role of cancer mutations as predictors of sensitivity and resistance to targeted therapies is discussed, together with the implications for the personalized treatment of cancer patients.
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KRAS mutations testing in colorectal carcinoma patients in Italy: from guidelines to external quality assessment.
PLoS ONE
PUBLISHED: 09-14-2011
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Monoclonal antibodies directed against the epidermal growth factor receptor (EGFR) have been approved for the treatment of patients with metastatic colorectal carcinoma (mCRC) that do not carry KRAS mutations. Therefore, KRAS testing has become mandatory to chose the most appropriate therapy for these patients.
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A molecularly annotated platform of patient-derived xenografts ("xenopatients") identifies HER2 as an effective therapeutic target in cetuximab-resistant colorectal cancer.
Cancer Discov
PUBLISHED: 09-02-2011
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Only a fraction of patients with metastatic colorectal cancer receive clinical benefit from therapy with anti-epidermal growth factor receptor (EGFR) antibodies, which calls for the identification of novel biomarkers for better personalized medicine. We produced large xenograft cohorts from 85 patient-derived, genetically characterized metastatic colorectal cancer samples ("xenopatients") to discover novel determinants of therapeutic response and new oncoprotein targets. Serially passaged tumors retained the morphologic and genomic features of their original counterparts. A validation trial confirmed the robustness of this approach: xenopatients responded to the anti-EGFR antibody cetuximab with rates and extents analogous to those observed in the clinic and could be prospectively stratified as responders or nonresponders on the basis of several predictive biomarkers. Genotype-response correlations indicated HER2 amplification specifically in a subset of cetuximab-resistant, KRAS/NRAS/BRAF/PIK3CA wild-type cases. Importantly, HER2 amplification was also enriched in clinically nonresponsive KRAS wild-type patients. A proof-of-concept, multiarm study in HER2-amplified xenopatients revealed that the combined inhibition of HER2 and EGFR induced overt, long-lasting tumor regression. Our results suggest promising therapeutic opportunities in cetuximab-resistant patients with metastatic colorectal cancer, whose medical treatment in the chemorefractory setting remains an unmet clinical need.
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Toll-like receptor 9 agonist IMO cooperates with cetuximab in K-ras mutant colorectal and pancreatic cancers.
Clin. Cancer Res.
PUBLISHED: 09-02-2011
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K-Ras somatic mutations are a strong predictive biomarker for resistance to epidermal growth factor receptor (EGFR) inhibitors in patients with colorectal and pancreatic cancer. We previously showed that the novel Toll-like receptor 9 (TLR9) agonist immunomodulatory oligonucleotide (IMO) has a strong in vivo activity in colorectal cancer models by interfering with EGFR-related signaling and synergizing with the anti-EGFR monoclonal antibody cetuximab.
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Increased detection sensitivity for KRAS mutations enhances the prediction of anti-EGFR monoclonal antibody resistance in metastatic colorectal cancer.
Clin. Cancer Res.
PUBLISHED: 06-01-2011
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KRAS mutations represent the main cause of resistance to anti-epidermal growth factor receptor (EGFR) monoclonal antibodies (MoAbs) in metastatic colorectal cancer (mCRC). We evaluated whether highly sensitive methods for KRAS investigation improve the accuracy of predictions of anti-EGFR MoAbs efficacy.
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Description of a novel Janus kinase 3 P132A mutation in acute megakaryoblastic leukemia and demonstration of previously reported Janus kinase 3 mutations in normal subjects.
Leuk. Lymphoma
PUBLISHED: 05-23-2011
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Gain-of-function (GOF) mutations of Janus kinase 2 (JAK2) are frequently seen in myeloproliferative disorders (MPDs). Meanwhile, JAK3 activating substitutions have been found in a few megakaryocytic cell lines and in primary myeloid leukemia (AMKL). Here, we sought to discover novel leukemogenetic mutations in de novo acute myeloid leukemia of non-Down syndrome (N-DS) by DNA sequencing. A total of 191 normal Caucasian individuals were studied to define single nucleotide polymorphisms (SNPs) within the JH2 and JH6 domains. Although known activating substitutions were observed in rare cases of acute myeloid leukemia (AML) (V722I [2/134] or P132T [1/119]), all samples were wild-type (WT) for the oncogenic A572V (119/119). Interestingly, a novel homozygous mutation (P132A) was discovered in a patient with acute megakaryoblastic leukemia and in vivo studies demonstrated that its ectopic expression was oncogenic in a mouse xenotransplant model. This study defines a novel JAK3 mutation among patients with N-DS AML and demonstrates that normal individuals can also display germline JAK3 substitutions, previously proven to have oncogenic properties, in vitro and in vivo. The discovery of these substitutions in normal donors encourages future studies to define new risk factors among patients with MPDs.
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Molecularly targeted therapies for colorectal cancer: Strategies for implementing translational research in clinical trials.
Curr. Opin. Mol. Ther.
PUBLISHED: 12-15-2010
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Few breakthroughs in preclinical research have translated into meaningful benefits, either in clinical terms or quality of life, for patients with advanced colorectal cancer, despite important preclinical discoveries regarding aberrant biological pathways associated with disease development and progression. The many reasons for the slow progress are diverse, ranging from the failure to codevelop biomarkers and targeted therapies, the regulatory burdens imposed on academic investigators, and the failure to collect serial tumor biopsies during clinical trials. This review discusses promising translational research that could help reduce the disparity between preclinical discovery and patient benefit, and advocate the concentration of efforts and resources on the most promising therapeutic targets in colorectal cancer, such as EGFR, VEGF and Fc? receptor.
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Targeting EGFR/HER2 pathways enhances the antiproliferative effect of gemcitabine in biliary tract and gallbladder carcinomas.
BMC Cancer
PUBLISHED: 11-18-2010
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Advanced biliary tract carcinomas (BTCs) have poor prognosis and limited therapeutic options. Therefore, it is crucial to combine standard therapies with molecular targeting. In this study EGFR, HER2, and their molecular transducers were analysed in terms of mutations, amplifications and over-expression in a BTC case series. Furthermore, we tested the efficacy of drugs targeting these molecules, as single agents or in combination with gemcitabine, the standard therapeutic agent against BTC.
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Association of KRAS p.G13D mutation with outcome in patients with chemotherapy-refractory metastatic colorectal cancer treated with cetuximab.
JAMA
PUBLISHED: 10-28-2010
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Patients with metastatic colorectal cancer who have KRAS codon 12- or KRAS codon 13-mutated tumors are presently excluded from treatment with the anti-epidermal growth factor receptor monoclonal antibody cetuximab.
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Broccoli, PTEN deletion and prostate cancer: where is the link?
Mol. Cancer
PUBLISHED: 08-18-2010
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The concept that vegetables and fruits are relevant sources of cancer-preventive substances is strongly supported by population studies. Among others, cruciferous vegetables like broccoli, cabbage, cauliflower and Brussels sprouts are thought to affect the development of various types of cancers and especially prostate tumors. Yet, the identification of the molecular mechanisms by which the active compounds contained in these vegetables mediate their anticancer activity has historically lagged behind. Accordingly, direct laboratory evidence of how individual nutrients affect cancer genes and the pathways they control remains the major obstacle to progress in this research field. Here we review a recent report investigating the interaction between sulforaphane, a dietary isothiocyanate derived from broccoli, and expression of the PTEN tumor suppressor gene in pre malignant prostate tissue.
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Effects of KRAS, BRAF, NRAS, and PIK3CA mutations on the efficacy of cetuximab plus chemotherapy in chemotherapy-refractory metastatic colorectal cancer: a retrospective consortium analysis.
Lancet Oncol.
PUBLISHED: 07-08-2010
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Following the discovery that mutant KRAS is associated with resistance to anti-epidermal growth factor receptor (EGFR) antibodies, the tumours of patients with metastatic colorectal cancer are now profiled for seven KRAS mutations before receiving cetuximab or panitumumab. However, most patients with KRAS wild-type tumours still do not respond. We studied the effect of other downstream mutations on the efficacy of cetuximab in, to our knowledge, the largest cohort to date of patients with chemotherapy-refractory metastatic colorectal cancer treated with cetuximab plus chemotherapy in the pre-KRAS selection era.
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Mutational profiling of kinases in human tumours of pancreatic origin identifies candidate cancer genes in ductal and ampulla of vater carcinomas.
PLoS ONE
PUBLISHED: 05-20-2010
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Protein kinases are key regulators of cellular processes (such as proliferation, apoptosis and invasion) that are often deregulated in human cancers. Accordingly, kinase genes have been the first to be systematically analyzed in human tumors leading to the discovery that many oncogenes correspond to mutated kinases. In most cases the genetic alterations translate in constitutively active kinase proteins, which are amenable of therapeutic targeting. Tumours of the pancreas are aggressive neoplasms for which no effective therapeutic strategy is currently available.
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Deregulation of the PI3K and KRAS signaling pathways in human cancer cells determines their response to everolimus.
J. Clin. Invest.
PUBLISHED: 05-19-2010
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Personalized cancer medicine is based on the concept that targeted therapies are effective on subsets of patients whose tumors carry specific molecular alterations. Several mammalian target of rapamycin (mTOR) inhibitors are in preclinical or clinical trials for cancers, but the molecular basis of sensitivity or resistance to these inhibitors among patients is largely unknown. Here we have identified oncogenic variants of phosphoinositide-3-kinase, catalytic, alpha polypeptide (PIK3CA) and KRAS as determinants of response to the mTOR inhibitor everolimus. Human cancer cells carrying alterations in the PI3K pathway were responsive to everolimus, both in vitro and in vivo, except when KRAS mutations occurred concomitantly or were exogenously introduced. In human cancer cells with mutations in both PIK3CA and KRAS, genetic ablation of mutant KRAS reinstated response to the drug. Consistent with these data, PIK3CA mutant cells, but not KRAS mutant cells, displayed everolimus-sensitive translation. Importantly, in a cohort of metastatic cancer patients, the presence of oncogenic KRAS mutations was associated with lack of benefit after everolimus therapy. Thus, our results demonstrate that alterations in the KRAS and PIK3CA genes may represent biomarkers to optimize treatment of patients with mTOR inhibitors.
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PIK3CA mutations associated with gene signature of low mTORC1 signaling and better outcomes in estrogen receptor-positive breast cancer.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 05-17-2010
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PIK3CA mutations are reported to be present in approximately 25% of breast cancer (BC), particularly the estrogen receptor-positive (ER+) and HER2-overexpressing (HER2+) subtypes, making them one of the most common genetic aberrations in BC. In experimental models, these mutations have been shown to activate AKT and induce oncogenic transformation, and hence these lesions have been hypothesized to render tumors highly sensitive to therapeutic PI3K/mTOR inhibition. By analyzing gene expression and protein data from nearly 1,800 human BCs, we report that a PIK3CA mutation-associated gene signature (PIK3CA-GS) derived from exon 20 (kinase domain) mutations was able to predict PIK3CA mutation status in two independent datasets, strongly suggesting a characteristic set of gene expression-induced changes. However, in ER+/HER2- BC despite pathway activation, PIK3CA mutations were associated with a phenotype of relatively low mTORC1 signaling and a good prognosis with tamoxifen monotherapy. The relationship between clinical outcome and the PIK3CA-GS was also assessed. Although the PIK3CA-GS was not associated with prognosis in ER- and HER2+ BC, it could identify better clinical outcomes in ER+/HER2- disease. In ER+ BC cell lines, PIK3CA mutations were also associated with sensitivity to tamoxifen. These findings could have important implications for the treatment of PIK3CA-mutant BCs and the development of PI3K/mTOR inhibitors.
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MET mutations in cancers of unknown primary origin (CUPs).
Hum. Mutat.
PUBLISHED: 05-04-2010
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Cancer of unknown primary origin (CUP) defines metastatic disease of unknown origin, accounting for 3-5% of all cancers. Growing evidence demonstrates that inappropriate execution of a genetic program named "invasive growth," driven by the MET oncogene, is implicated in the metastatic process. MET activation in cancers is mainly consequent to overexpression, whereas mutations are rarely found. We reasoned that the occurrence of MET somatic mutations might sustain premature occult dissemination of cancer cells, such as that observed in CUPs. We sequenced MET in genomic DNA obtained from 47 early metastatic cancers. By extensive immunohistochemical analysis a primary site was afterward postulated in 24 patients, whereas 23 cases remained of unknown primary (CUPs). MET somatic mutations were found in seven cases, all belonging to the CUP cohort. Mutational incidence (30%) was thus significantly higher than the expected one (4%), in the absence of high mutational background. Several nucleotide changes were novel and clustered either in the kinase domain or in the extracellular semaphorin domain. Mutated receptors were functional and sustained the transformed phenotype, suggesting that MET activating mutations are genetic markers associated with the CUP syndrome.
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International network of cancer genome projects.
, Thomas J Hudson, Warwick Anderson, Axel Artez, Anna D Barker, Cindy Bell, Rosa R Bernabé, M K Bhan, Fabien Calvo, Iiro Eerola, Daniela S Gerhard, Alan Guttmacher, Mark Guyer, Fiona M Hemsley, Jennifer L Jennings, David Kerr, Peter Klatt, Patrik Kolar, Jun Kusada, David P Lane, Frank Laplace, Lu Youyong, Gerd Nettekoven, Brad Ozenberger, Jane Peterson, T S Rao, Jacques Remacle, Alan J Schafer, Tatsuhiro Shibata, Michael R Stratton, Joseph G Vockley, Koichi Watanabe, Huanming Yang, Matthew M F Yuen, Bartha M Knoppers, Martin Bobrow, Anne Cambon-Thomsen, Lynn G Dressler, Stephanie O M Dyke, Yann Joly, Kazuto Kato, Karen L Kennedy, Pilar Nicolás, Michael J Parker, Emmanuelle Rial-Sebbag, Carlos M Romeo-Casabona, Kenna M Shaw, Susan Wallace, Georgia L Wiesner, Nikolajs Zeps, Peter Lichter, Andrew V Biankin, Christian Chabannon, Lynda Chin, Bruno Clément, Enrique De Alava, Françoise Degos, Martin L Ferguson, Peter Geary, D Neil Hayes, Amber L Johns, Arek Kasprzyk, Hidewaki Nakagawa, Robert Penny, Miguel A Piris, Rajiv Sarin, Aldo Scarpa, Marc van de Vijver, P Andrew Futreal, Hiroyuki Aburatani, Mònica Bayés, David D L Botwell, Peter J Campbell, Xavier Estivill, Sean M Grimmond, Ivo Gut, Martin Hirst, Carlos Lopez-Otin, Partha Majumder, Marco Marra, John D McPherson, Zemin Ning, Xose S Puente, Yijun Ruan, Hendrik G Stunnenberg, Harold Swerdlow, Victor E Velculescu, Richard K Wilson, Hong H Xue, Liu Yang, Paul T Spellman, Gary D Bader, Paul C Boutros, Paul Flicek, Gad Getz, Roderic Guigo, Guangwu Guo, David Haussler, Simon Heath, Tim J Hubbard, Tao Jiang, Steven M Jones, Qibin Li, Nuria López-Bigas, Ruibang Luo, Lakshmi Muthuswamy, B F Francis Ouellette, John V Pearson, Víctor Quesada, Benjamin J Raphael, Chris Sander, Terence P Speed, Lincoln D Stein, Joshua M Stuart, Jon W Teague, Yasushi Totoki, Tatsuhiko Tsunoda, Alfonso Valencia, David A Wheeler, Honglong Wu, Shancen Zhao, Guangyu Zhou, Mark Lathrop, Gilles Thomas, Teruhiko Yoshida, Myles Axton, Chris Gunter, Linda J Miller, Junjun Zhang, Syed A Haider, Jianxin Wang, Christina K Yung, Anthony Cros, Anthony Cross, Yong Liang, Saravanamuttu Gnaneshan, Jonathan Guberman, Jack Hsu, Don R C Chalmers, Karl W Hasel, Terry S H Kaan, William W Lowrance, Tohru Masui, Laura Lyman Rodriguez, Catherine Vergely, David D L Bowtell, Nicole Cloonan, Anna deFazio, James R Eshleman, Dariush Etemadmoghadam, Brooke B Gardiner, Brooke A Gardiner, James G Kench, Robert L Sutherland, Margaret A Tempero, Nicola J Waddell, Peter J Wilson, Steve Gallinger, Ming-Sound Tsao, Patricia A Shaw, Gloria M Petersen, Debabrata Mukhopadhyay, Ronald A DePinho, Sarah Thayer, Kamran Shazand, Timothy Beck, Michelle Sam, Lee Timms, Vanessa Ballin, Youyong Lu, Jiafu Ji, Xiuqing Zhang, Feng Chen, Xueda Hu, Qi Yang, Geng Tian, Lianhai Zhang, Xiaofang Xing, Xianghong Li, Zhenggang Zhu, Yingyan Yu, Jun Yu, Jörg Tost, Paul Brennan, Ivana Holcatova, David Zaridze, Alvis Brazma, Lars Egevard, Egor Prokhortchouk, Rosamonde Elizabeth Banks, Mathias Uhlén, Juris Viksna, Fredrik Ponten, Konstantin Skryabin, Ewan Birney, Ake Borg, Anne-Lise Børresen-Dale, Carlos Caldas, John A Foekens, Sancha Martin, Jorge S Reis-Filho, Andrea L Richardson, Christos Sotiriou, Giles Thoms, Laura van't Veer, Daniel Birnbaum, Hélène Blanché, Pascal Boucher, Sandrine Boyault, Jocelyne D Masson-Jacquemier, Iris Pauporté, Xavier Pivot, Anne Vincent-Salomon, Eric Tabone, Charles Theillet, Isabelle Treilleux, Paulette Bioulac-Sage, Thomas Decaens, Dominique Franco, Marta Gut, Didier Samuel, Jessica Zucman-Rossi, Roland Eils, Benedikt Brors, Jan O Korbel, Andrey Korshunov, Pablo Landgraf, Hans Lehrach, Stefan Pfister, Bernhard Radlwimmer, Guido Reifenberger, Michael D Taylor, Christof von Kalle, Partha P Majumder, Paolo Pederzoli, Rita A Lawlor, Massimo Delledonne, Alberto Bardelli, Thomas Gress, David Klimstra, Giuseppe Zamboni, Yusuke Nakamura, Satoru Miyano, Akihiro Fujimoto, Elias Campo, Silvia de Sanjosé, Emili Montserrat, Marcos Gonzalez-Díaz, Pedro Jares, Heinz Himmelbauer, Heinz Himmelbaue, Sílvia Beà, Samuel Aparicio, Douglas F Easton, Francis S Collins, Carolyn C Compton, Eric S Lander, Wylie Burke, Anthony R Green, Stanley R Hamilton, Olli P Kallioniemi, Timothy J Ley, Edison T Liu, Brandon J Wainwright.
Nature
PUBLISHED: 04-16-2010
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The International Cancer Genome Consortium (ICGC) was launched to coordinate large-scale cancer genome studies in tumours from 50 different cancer types and/or subtypes that are of clinical and societal importance across the globe. Systematic studies of more than 25,000 cancer genomes at the genomic, epigenomic and transcriptomic levels will reveal the repertoire of oncogenic mutations, uncover traces of the mutagenic influences, define clinically relevant subtypes for prognosis and therapeutic management, and enable the development of new cancer therapies.
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Integrated molecular dissection of the epidermal growth factor receptor (EGFR) [corrected] oncogenic pathway to predict response to EGFR-targeted monoclonal antibodies in metastatic colorectal cancer.
Target Oncol
PUBLISHED: 02-12-2010
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The introduction of KRAS testing as a diagnostic tool to select patients for epidermal growth factor receptor (EGFR)-targeted cetuximab- or panitumumab-based therapies for metastatic colorectal cancer is widely regarded as a key advance in the field of personalized cancer medicine. Oncologists are now facing emerging issues in the treatment of metastatic colorectal cancer, including: (i) the identification of additional genetic determinants of primary resistance to EGFR-targeted therapy for further improving selection of patients; (ii) the explanation of rare cases of patients carrying KRAS-mutated tumors who have been reported to respond to either cetuximab or panitumumab and (iii) the discovery of mechanisms of secondary resistance to anti-EGFR antibody therapies. Here we discuss the potential role of comprehensive dissection of the key oncogenic nodes in the EGFR signaling cascade to predict resistance and sensitivity to EGFR monoclonal antibodies in metastatic colorectal cancer. Current data suggest that, together with KRAS mutations, the evaluation of BRAF and PIK3CA/PTEN alterations could also be useful for selecting patients with reduced chance to benefit from EGFR-targeted therapy. Furthermore, measuring EGFR gene copy number also appears relevant to positively identify responders. Up until now, each of these markers has been mainly assessed as a single event, often in retrospective analyses and patients series. As these molecular alterations display overlapping patterns of occurrence, this adds considerable complexity to the drawing of an algorithm suitable for clinical decision-making. We suggest that in the near future comprehensive molecular analysis of the entire oncogenic pathway triggered by the EGFR should be performed, thus enhancing the prediction ability of individual markers.
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Tracking the genomic evolution of breast cancer metastasis.
Breast Cancer Res.
PUBLISHED: 02-02-2010
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Therapeutic choices for metastatic tumors are, in most cases, based upon the histological and molecular analysis of the corresponding primary tumor. Understanding whether and to what extent the genomic landscape of metastasis differs from the tumors from which they originated is critical yet largely unknown. A recent report tackled this key issue by comparing the genomic and transcriptional profile of a metastatic lobular breast tumor with that of the primary tumor surgically removed 9 years earlier. The extent of the differences suggests a high degree of mutational heterogeneity between primary and metastatic lesions and indicates that significant evolution occurs during breast cancer progression.
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Molecular mechanisms of resistance to cetuximab and panitumumab in colorectal cancer.
J. Clin. Oncol.
PUBLISHED: 01-25-2010
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Personalized cancer medicine based on the genetic milieu of individual colorectal tumors has long been postulated, but until recently this concept was not supported by clinical evidence. The advent of the epidermal growth factor receptor (EGFR) -targeted monoclonal antibodies cetuximab and panitumumab has paved the way to the individualized treatment of metastatic colorectal cancer (mCRC). Here we discuss the evidence that mCRCs respond differently to EGFR-targeted agents and that the tumor-specific response has a genetic basis. We outline how, from the initial observation that cetuximab or panitumumab as monotherapy is effective only in 10% to 20% of mCRCs, knowledge has being gained on the molecular mechanisms underlying primary resistance to these agents. The role of oncogenic activation of EGFR downstream effectors such as KRAS, BRAF, PIK3CA, and PTEN on response to therapy is discussed. We suggest that CRCs lacking oncogenic alterations in these four genes have the highest probability of response to anti-EGFR therapies and are defined as "quadruple negative." The rapid and effective translation of these findings into predictive biomarkers to couple EGFR-targeted antibodies to the patients who benefit from them is presented as a paradigm of modern clinical oncology. Finally, unresolved questions such as understanding the molecular basis of response as well the mechanisms of secondary resistance are presented as the future fundamental goals in this research field.
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The prognostic IDH1( R132 ) mutation is associated with reduced NADP+-dependent IDH activity in glioblastoma.
Acta Neuropathol.
PUBLISHED: 01-19-2010
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Somatic mutations in the isocitrate dehydrogenase 1 gene (IDH1) occur at high frequency in gliomas and seem to be a prognostic factor for survival in glioblastoma patients. In our set of 98 glioblastoma patients, IDH1 ( R132 ) mutations were associated with improved survival of 1 year on average, after correcting for age and other variables with Cox proportional hazards models. Patients with IDH1 mutations were on average 17 years younger than patients without mutation. Mutated IDH1 has a gain of function to produce 2-hydroxyglutarate by NADPH-dependent reduction of alpha-ketoglutarate, but it is unknown whether NADPH production in gliomas is affected by IDH1 mutations. We assessed the effect of IDH1 (R132 ) mutations on IDH-mediated NADPH production in glioblastomas in situ. Metabolic mapping and image analysis was applied to 51 glioblastoma samples of which 16 carried an IDH1 (R132 ) mutation. NADP+-dependent IDH activity was determined in comparison with activity of NAD+-dependent IDH and all other NADPH-producing dehydrogenases, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, malate dehydrogenase, and hexose-6-phosphate dehydrogenase. The occurrence of IDH1 mutations correlated with approx. twofold diminished NADP+-dependent IDH activity, whereas activity of NAD+-dependent IDH and the other NADP+-dependent dehydrogenases was not affected in situ in glioblastoma. The total NADPH production capacity in glioblastoma was provided for 65% by IDH activity and the occurrence of IDH1 (R132 ) mutation reduced this capacity by 38%. It is concluded that NADPH production is hampered in glioblastoma with IDH1 (R132 ) mutation. Moreover, mutated IDH1 consumes rather than produces NADPH, thus likely lowering NADPH levels even further. The low NADPH levels may sensitize glioblastoma to irradiation and chemotherapy, thus explaining the prolonged survival of patients with mutated glioblastoma.
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The analysis of PIK3CA mutations in gastric carcinoma and metanalysis of literature suggest that exon-selectivity is a signature of cancer type.
J. Exp. Clin. Cancer Res.
PUBLISHED: 01-14-2010
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PIK3CA is one of the genes most frequently mutated in human cancers and it is a potential target for personalized therapy. The aim of this study was to assess the frequency and type of PIK3CA mutations in gastric carcinoma and compare them with their clinical pathological correlates.
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Biomarkers predicting clinical outcome of epidermal growth factor receptor-targeted therapy in metastatic colorectal cancer.
J. Natl. Cancer Inst.
PUBLISHED: 09-08-2009
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The monoclonal antibodies panitumumab and cetuximab that target the epidermal growth factor receptor (EGFR) have expanded the range of treatment options for metastatic colorectal cancer. Initial evaluation of these agents as monotherapy in patients with EGFR-expressing chemotherapy-refractory tumors yielded response rates of approximately 10%. The realization that detection of positive EGFR expression by immunostaining does not reliably predict clinical outcome of EGFR-targeted treatment has led to an intense search for alternative predictive biomarkers. Oncogenic activation of signaling pathways downstream of the EGFR, such as mutation of KRAS, BRAF, or PIK3CA oncogenes, or inactivation of the PTEN tumor suppressor gene is central to the progression of colorectal cancer. Tumor KRAS mutations, which may be present in 35%-45% of patients with colorectal cancer, have emerged as an important predictive marker of resistance to panitumumab or cetuximab treatment. In addition, among colorectal tumors carrying wild-type KRAS, mutation of BRAF or PIK3CA or loss of PTEN expression may be associated with resistance to EGFR-targeted monoclonal antibody treatment, although these additional biomarkers require further validation before incorporation into clinical practice. Additional knowledge of the molecular basis for sensitivity or resistance to EGFR-targeted monoclonal antibodies will allow the development of new treatment algorithms to identify patients who are most likely to respond to treatment and could also provide rationale for combining therapies to overcome primary resistance. The use of KRAS mutations as a selection biomarker for anti-EGFR monoclonal antibody (eg, panitumumab or cetuximab) treatment is the first major step toward individualized treatment for patients with metastatic colorectal cancer.
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Expression and functional regulation of myoglobin in epithelial cancers.
Am. J. Pathol.
PUBLISHED: 06-18-2009
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Myoglobin is a multifunctional heme protein that is thought to be expressed exclusively in myocytes. Its importance in both oxygen transport and free radical scavenging has been extensively characterized. We hypothesized that solid tumors could take advantage of proteins such as myoglobin to cope with hypoxic conditions and to control the metabolism of reactive oxygen and nitrogen species. We therefore sought to establish whether myoglobin might be expressed and functionally regulated in epithelial tumors that are known to face hypoxia and oxidative stress during disease progression. We analyzed the expression of myoglobin in human epithelial cancers at both transcriptional and protein levels; moreover, we investigated the expression levels of myoglobin in cancer cell lines subjected to different conditions, including hypoxia, oxidative stress, and mitogenic stimuli. We provide evidence that human epithelial tumors, including breast, lung, ovary, and colon carcinomas, express high levels of myoglobin from the earliest stages of disease development. In human cancer cells, myoglobin is induced by a variety of signals associated with tumor progression, including mitogenic stimuli, oxidative stress, and hypoxia. This study provides evidence that myoglobin, previously thought to be restricted to myocytes, is expressed at high levels by human carcinoma cells. We suggest that myoglobin expression is part of a cellular program aimed at coping with changed metabolic and environmental conditions associated with neoplastic growth.
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Selective cytotoxicity of a bicyclic Ras inhibitor in cancer cells expressing K-Ras(G13D).
Biochem. Biophys. Res. Commun.
PUBLISHED: 06-11-2009
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Mutation of RAS genes is a critical event in the pathogenesis of different human tumors and in some developmental disorders. Here we present an arabinose-derived bicyclic compound displaying selective cytotoxicity in human colorectal cancer cells expressing K-Ras(G13D), that shows high intrinsic nucleotide exchange rate. We characterize binding of bicyclic compounds by docking and NMR experiments and their inhibitory activity on GEF-mediated nucleotide exchange on wild-type and mutant Ras proteins. We demonstrate that the in vitro inhibition of Ras nucleotide exchange depends on the molar ratio between Ras and its GEF activator, suggesting that the observed in vivo selective effect may depend on biochemical parameters and actual intracellular concentration of the Ras protein and its regulators.
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Multi-determinants analysis of molecular alterations for predicting clinical benefit to EGFR-targeted monoclonal antibodies in colorectal cancer.
PLoS ONE
PUBLISHED: 06-05-2009
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KRAS mutations occur in 35-45% of metastatic colorectal cancers (mCRC) and preclude responsiveness to EGFR-targeted therapy with cetuximab or panitumumab. However, less than 20% patients displaying wild-type KRAS tumors achieve objective response. Alterations in other effectors downstream of the EGFR, such as BRAF, and deregulation of the PIK3CA/PTEN pathway have independently been found to give rise to resistance. We present a comprehensive analysis of KRAS, BRAF, PIK3CA mutations, and PTEN expression in mCRC patients treated with cetuximab or panitumumab, with the aim of clarifying the relative contribution of these molecular alterations to resistance.
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Molecular profiling of the "plexinome" in melanoma and pancreatic cancer.
Hum. Mutat.
PUBLISHED: 05-23-2009
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Plexins are transmembrane high-affinity receptors for semaphorins, regulating cell guidance, motility, and invasion. Functional evidences implicate semaphorin signals in cancer progression and metastasis. Yet, it is largely unknown whether plexin genes are genetically altered in human tumors. We performed a comprehensive gene copy analysis and mutational profiling of all nine members of the plexin gene family (plexinome), in melanomas and pancreatic ductal adenocarcinomas (PDACs), which are characterized by high metastatic potential and poor prognosis. Gene copy analysis detected amplification of PLXNA4 in melanomas, whereas copy number losses of multiple plexin genes were seen in PDACs. Somatic mutations were detected in PLXNA4, PLXNB3, and PLXNC1; providing the first evidence that these plexins are mutated in human cancer. Functional assays in cellular models revealed that some of these missense mutations result in loss of plexin function. For instance, c.1613G>A, p.R538H mutation in the extracellular domain of PLXNB3 prevented binding of the ligand Sema5A. Moreover, although PLXNA4 signaling can inhibit tumor cell migration, the mutated c.5206C>T, p.H1736Y allele had lost this activity. Our study is the first systematic analysis of the "plexinome" in human tumors, and indicates that multiple mutated plexins may be involved in cancer progression.
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Mutational profile of GNAQQ209 in human tumors.
PLoS ONE
PUBLISHED: 05-18-2009
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Frequent somatic mutations have recently been identified in the ras-like domain of the heterotrimeric G protein alpha-subunit (GNAQ) in blue naevi 83%, malignant blue naevi (50%) and ocular melanoma of the uvea (46%). The mutations exclusively affect codon 209 and result in GNAQ constitutive activation which, in turn, acts as a dominant oncogene.
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Mutational profiling of cancer candidate genes in glioblastoma, melanoma and pancreatic carcinoma reveals a snapshot of their genomic landscapes.
Hum. Mutat.
PUBLISHED: 04-11-2009
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A recent systematic analysis of 18.191 well annotated coding sequences (RefSeq) in breast and colorectal cancers has led to the identification of somatic mutations in 1.718 genes (Wood et al., 2007). Based on statistical parameters 280 of these have been denominated candidate cancer (CAN) genes. This analysis has provided an interesting snapshot of the landscape of tumor genomes by showing that they contain a few frequently mutated genes (denominated mountains). On the contrary, the large majority of CAN genes are altered at low frequency (designated hills). Whether hill type CAN genes are tumor specific is largely unknown. To address this question we evaluated the mutational profiles of 27 hill CAN genes in glioblastoma, melanoma and pancreatic carcinoma by sequencing the exons previously found mutated by Wood and colleagues. Only 4 of the breast/colorectal hill type CAN genes (SMAD4, MYO18B, NAV3 and MMP2) were also mutated in melanoma and pancreatic carcinoma, while none was altered in glioblastoma. These results suggest that hill type CAN genes are not frequently shared by different tumor types and that their mutation patterns are tissue specific. Tumor-specific genome wide mutational profiling will be required to identify hill type CAN genes that characterize the genomic landscapes of each cancer lineage.
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Absence of AKT1 mutations in glioblastoma.
PLoS ONE
PUBLISHED: 04-09-2009
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Oncogenic activation of the PI3K signalling pathway plays a pivotal role in the development of glioblastoma multiforme (GBM). A central node in PI3K downstream signalling is controlled by the serine-threonine kinase AKT1. A somatic mutation affecting residue E17 of the AKT1 gene has recently been identified in breast and colon cancer. The E17K change results in constitutive AKT1 activation, induces leukaemia in mice, and accordingly, may be therapeutically exploited to target the PI3K pathway. Assessing whether AKT1 is activated by somatic mutations in GBM is relevant to establish its role in this aggressive disease.
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PIK3CA mutations in colorectal cancer are associated with clinical resistance to EGFR-targeted monoclonal antibodies.
Cancer Res.
PUBLISHED: 02-17-2009
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The monoclonal antibodies (moAb) panitumumab and cetuximab target the epidermal growth factor receptor (EGFR) and have proven valuable for the treatment of metastatic colorectal cancer (mCRC). EGFR-mediated signaling involves two main intracellular cascades: on one side KRAS activates BRAF, which in turn triggers the mitogen-activated protein kinases. On the other, membrane localization of the lipid kinase PIK3CA counteracts PTEN and promotes AKT1 phosphorylation, thereby activating a parallel intracellular axis. Constitutive activation of KRAS bypasses the corresponding signaling cascade and, accordingly, patients with mCRC bearing KRAS mutations are clinically resistant to therapy with panitumumab or cetuximab. We hypothesized that mutations activating PIK3CA could also preclude responsiveness to EGFR-targeted moAbs through a similar mechanism. Here, we present the mutational analysis of PIK3CA and KRAS and evaluation of the PTEN protein status in a cohort of 110 patients with mCRC treated with anti-EGFR moAbs. We observed 15 (13.6%) PIK3CA and 32 (29.0%) KRAS mutations. PIK3CA mutations were significantly associated with clinical resistance to panitumumab or cetuximab; none of the mutated patients achieved objective response (P = 0.038). When only KRAS wild-type tumors were analyzed, the statistical correlation was stronger (P = 0.016). Patients with PIK3CA mutations displayed a worse clinical outcome also in terms of progression-free survival (P = 0.035). Our data indicate that PIK3CA mutations can independently hamper the therapeutic response to panitumumab or cetuximab in mCRC. When the molecular status of the PIK3CA/PTEN and KRAS pathways are concomitantly ascertained, up to 70% of mCRC patients unlikely to respond to EGFR moAbs can be identified.
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IDH1 mutations at residue p.R132 (IDH1(R132)) occur frequently in high-grade gliomas but not in other solid tumors.
Hum. Mutat.
PUBLISHED: 01-02-2009
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Systematic sequence profiling of the Glioblastoma Multiforme (GBM) genome has recently led to the identification of somatic mutations in the isocitrate dehydrogenase 1 (IDH1) gene. Interestingly, only the evolutionarily conserved residue R132 located in the substrate binding site of IDH1 was found mutated in GBM. At present, the occurrence and the relevance of p.R132 (IDH1(R132)) variants in tumors other than GBMs is largely unknown. We searched for mutations at position R132 of the IDH1 gene in a panel of 672 tumor samples. These included high-grade glioma, gastrointestinal stromal tumors (GIST), melanoma, bladder, breast, colorectal, lung, ovarian, pancreas, prostate, and thyroid carcinoma specimens. In addition, we assessed a panel of 84 cell lines from different tumor lineages. Somatic mutations affecting the IDH1(R132) residue were detected in 20% (23 of 113) high-grade glioma samples. In addition to the previously reported p.R132H and p.R132S alleles, we identified three novel somatic mutations (p.R132C, p.R132G, and p.R132L) affecting residue IDH1(R132) in GBM. Strikingly, no IDH1 mutations were detected in the other tumor types. These data indicate that cancer mutations affecting IDH1(R132) are tissue-specific, and suggest that it plays a unique role in the development of high-grade gliomas.
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Emergence of KRAS mutations and acquired resistance to anti-EGFR therapy in colorectal cancer.
Nature
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A main limitation of therapies that selectively target kinase signalling pathways is the emergence of secondary drug resistance. Cetuximab, a monoclonal antibody that binds the extracellular domain of epidermal growth factor receptor (EGFR), is effective in a subset of KRAS wild-type metastatic colorectal cancers. After an initial response, secondary resistance invariably ensues, thereby limiting the clinical benefit of this drug. The molecular bases of secondary resistance to cetuximab in colorectal cancer are poorly understood. Here we show that molecular alterations (in most instances point mutations) of KRAS are causally associated with the onset of acquired resistance to anti-EGFR treatment in colorectal cancers. Expression of mutant KRAS under the control of its endogenous gene promoter was sufficient to confer cetuximab resistance, but resistant cells remained sensitive to combinatorial inhibition of EGFR and mitogen-activated protein-kinase kinase (MEK). Analysis of metastases from patients who developed resistance to cetuximab or panitumumab showed the emergence of KRAS amplification in one sample and acquisition of secondary KRAS mutations in 60% (6 out of 10) of the cases. KRAS mutant alleles were detectable in the blood of cetuximab-treated patients as early as 10 months before radiographic documentation of disease progression. In summary, the results identify KRAS mutations as frequent drivers of acquired resistance to cetuximab in colorectal cancers, indicate that the emergence of KRAS mutant clones can be detected non-invasively months before radiographic progression and suggest early initiation of a MEK inhibitor as a rational strategy for delaying or reversing drug resistance.
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Activation of ?-catenin by oncogenic PIK3CA and EGFR promotes resistance to glucose deprivation by inducing a strong antioxidant response.
PLoS ONE
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Glucose is an essential fuel for cell survival and its availability limits aberrant cellular proliferation. We have hypothesized that specific cancer mutations regulate metabolic response(s) to glucose deprivation (GD). By means of somatic knock-in cellular models, we have analyzed the response to glucose deprivation in cells carrying the frequent (delE746-A750)EGFR, (G13D)KRAS or (E545K)PIK3CA cancer alleles. We demonstrate that, in mammary epithelial cells, glucose has an essential antioxidant function and that these cells are very sensitive to GD. Conversely, isogenic cells carrying the (delE746-A750)EGFR or the (E545K)PIK3CA, but not the (G13D)KRAS allele, display high tolerance to GD by stimulating the expression of anti-oxidant genes (MnSOD and catalase). This adaptive transcriptional response is mediated by the activation of WNT/?-catenin and FOXO4 signalling. Our data highlights a new functional synergism between oncogenic EGFR and PIK3CA with WNT/?-catenin conferring high tolerance to oxidative stress generated by nutrient deprivation.
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Active PI3K pathway causes an invasive phenotype which can be reversed or promoted by blocking the pathway at divergent nodes.
PLoS ONE
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The PTEN/PI3K pathway is commonly mutated in cancer and therefore represents an attractive target for therapeutic intervention. To investigate the primary phenotypes mediated by increased pathway signaling in a clean, patient-relevant context, an activating PIK3CA mutation (H1047R) was knocked-in to an endogenous allele of the MCF10A non-tumorigenic human breast epithelial cell line. Introduction of an endogenously mutated PIK3CA allele resulted in a marked epithelial-mesenchymal transition (EMT) and invasive phenotype, compared to isogenic wild-type cells. The invasive phenotype was linked to enhanced PIP(3) production via a S6K-IRS positive feedback mechanism. Moreover, potent and selective inhibitors of PI3K were highly effective in reversing this phenotype, which is optimally revealed in 3-dimensional cell culture. In contrast, inhibition of Akt or mTOR exacerbated the invasive phenotype. Our results suggest that invasion is a core phenotype mediated by increased PTEN/PI3K pathway activity and that therapeutic agents targeting different nodes of the PI3K pathway may have dramatic differences in their ability to reverse or promote cancer metastasis.
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Inhibition of MEK and PI3K/mTOR suppresses tumor growth but does not cause tumor regression in patient-derived xenografts of RAS-mutant colorectal carcinomas.
Clin. Cancer Res.
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Gene mutations along the Ras pathway (KRAS, NRAS, BRAF, PIK3CA) occur in approximately 50% of colorectal cancers (CRC) and correlate with poor response to anti-EGF receptor (EGFR) therapies. We assessed the effects of mitogen-activated protein (MAP)/extracellular signal-regulated kinase (ERK) kinase (MEK) and phosphoinositide 3-kinase (PI3K)/mTOR inhibitors, which neutralize the major Ras effectors, in patient-derived xenografts from RAS/RAF/PIK3CA-mutant metastatic CRCs (mCRC).
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Unresponsiveness of colon cancer to BRAF(V600E) inhibition through feedback activation of EGFR.
Nature
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Inhibition of the BRAF(V600E) oncoprotein by the small-molecule drug PLX4032 (vemurafenib) is highly effective in the treatment of melanoma. However, colon cancer patients harbouring the same BRAF(V600E) oncogenic lesion have poor prognosis and show only a very limited response to this drug. To investigate the cause of the limited therapeutic effect of PLX4032 in BRAF(V600E) mutant colon tumours, here we performed an RNA-interference-based genetic screen in human cells to search for kinases whose knockdown synergizes with BRAF(V600E) inhibition. We report that blockade of the epidermal growth factor receptor (EGFR) shows strong synergy with BRAF(V600E) inhibition. We find in multiple BRAF(V600E) mutant colon cancers that inhibition of EGFR by the antibody drug cetuximab or the small-molecule drugs gefitinib or erlotinib is strongly synergistic with BRAF(V600E) inhibition, both in vitro and in vivo. Mechanistically, we find that BRAF(V600E) inhibition causes a rapid feedback activation of EGFR, which supports continued proliferation in the presence of BRAF(V600E) inhibition. Melanoma cells express low levels of EGFR and are therefore not subject to this feedback activation. Consistent with this, we find that ectopic expression of EGFR in melanoma cells is sufficient to cause resistance to PLX4032. Our data suggest that BRAF(V600E) mutant colon cancers (approximately 8-10% of all colon cancers), for which there are currently no targeted treatment options available, might benefit from combination therapy consisting of BRAF and EGFR inhibitors.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.