Multiplex PCR/LDR for detection of K-ras mutations in primary colon tumors

Khanna, Marilyn; Park, Pat; Zirvi, Monib; Cao, Weiguo; Picon, Antonio I.; Day, John; Paty, Philip B.; Barany, Francis

doi:10.1038/sj.onc.1202291

Cited by 158 publications

(159 citation statements)

References 71 publications

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“…[23][24][25] As a control cell line, HT29 (91072201 ECACC, Porton Down, UK) cell with WT K-ras were selected. 26 They were grown in Advanced Eagle's minimum essential medium (Gibco) supplemented with 2% fetal bovine serum (Gibco), 2 mM L-glutamine, 1 mM sodium pyruvate and 350 mg l À1 gentamicin and 1 ml l À1 crystacillin. The control scrambled sequences were, by manufacturer's assurance, designed in a way that had no homology to any known gene in human genome.…”

Section: Colorectal Tumor Cell Linementioning

confidence: 99%

MicroRNAs targeting mutant K-ras by electrotransfer inhibit human colorectal adenocarcinoma cell growth in vitro and in vivo

et al. 2010

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Mutations of K-ras have been found in 30-60% of colorectal carcinomas and are believed to be associated with tumor initiation, tumor progression and metastasis formation. Therefore, silencing of mutant K-ras expression has become an attractive therapeutic strategy for colorectal cancer treatment. The aim of our study was to investigate the effect of microRNA (miRNA) molecules directed against K-ras (miRNA-K-ras) on K-ras expression level and the growth of colorectal carcinoma cell line LoVo in vitro and in vivo. In addition, we evaluated electroporation as a gene delivery method for transfection of LoVo cells and tumors with plasmid DNA encoding miRNA-K-ras (pmiRNA-K-ras). Results of our study indicated that miRNAs targeting K-ras efficiently reduced K-ras expression and cell survival after in vitro electrotransfection of LoVo cells with pmiRNA-K-ras. In vivo, electroporation has proven to be a simple and efficient delivery method for local administration of pmiRNA-K-ras molecules into LoVo tumors. This therapy shows pronounced antitumor effectiveness and has no side effects. The obtained results demonstrate that electrogene therapy with miRNA-K-ras molecules can be potential therapeutic strategy for treatment of colorectal cancers harboring K-ras mutations.

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Section: Colorectal Tumor Cell Linementioning

confidence: 99%

MicroRNAs targeting mutant K-ras by electrotransfer inhibit human colorectal adenocarcinoma cell growth in vitro and in vivo

et al. 2010

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“…Phenocopies might occur first with their eventual replacement by more stable or dramatic sequence-based modes of inheritance. It should be noted that the finding of sequence-based mutations in clinical material requires a level of sensitivity approximately 1% because the mutation is often present in only a minority of cells in a biopsy sample (Khanna et al 1999). This is interpreted to be a consequence of normal cells infiltrating the tumor.…”

Section: Differing Lineage Qualitiesmentioning

confidence: 99%

Hereditary stability and variation in evolution and development

Thaler

1999

Evolution and Development

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SUMMARY Evolution and development are both lineage processes but are often conceptualized as occurring by different and mutually exclusive mechanisms. It is conventionally asserted that evolution occurs via the random generation of diversity and the subsequent survival of those that pass selection. On the other hand, development is too often presented as proceeding via the unfolding of a deterministic program encoded in the DNA sequence. In biology, universal generalizations are rare and dogmas are often wrong for particular cases. Deterministic mechanisms contribute some of the new DNA sequences that subsequently become substrates for natural selection. Conversely, stochastic and selective mechanisms are intrinsic to development, and also to maintenance of the immune, and possibly, nervous systems. Cancer appears to be another process that straddles distinctions between evolutionary and developmental modes of hereditary change and stabilization. DNA sequence changes are an essential feature of many cancers, but there are also aspects of the disease similar to developmental lineage gone awry. The literature suggests that the cellular changes that give rise to cancer occur by mechanisms commonly associated with both evolutionary and developmental lineage pathways.

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“…Genomic DNA was extracted using a DNA isolation kit (BioFlux, Japan), and genotyping was performed using the PCR-LDR method. 26 The primers for amplification were derived from published genomic sequences (NCBI RefSeq: NT_009952) (Table 1), and the PCR reactions were carried out in an ABI 9600 (Applied Biosystems, USA) in a total volume of 15 μl including 10 ng genomic DNA, 1.5 μl 10*PCR Buffer, 0.3 mM dNTPs, 2 μM each primer and 1 U Taq DNA polymerase (TaKaRa, Japan). Cycling parameters were as follows: 94°C for 1 min; 35 cycles of 94°C for 10 s; 56°C for 20 s; 72°C for 40 s; and a final extension step at 72°C for 10 min.…”

Section: Patientmentioning

confidence: 99%

ERCC5 promoter polymorphisms at –763 and +25 predict the response to oxaliplatin-based chemotherapy in patients with advanced colorectal cancer

Chen

Xie²,

Wang³

et al. 2009

Cancer Biology & Therapy

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This study aimed to investigate whether single nucleotide polymorphisms (SNPs) in the promoter of the excision repair cross complementation group 5 (ERCC5) gene influences response to oxaliplatin-based chemotherapy. Eighty-three patients with cytologically or histologically confirmed advanced colorectal cancer (CRC), at least one measurable lesion and underwent oxaliplatin-based chemotherapy were studied. To this end, six polymorphisms (-1415C>T, -763A>G, -413C>T, +25A>G, +202C>T, +372C>T) in the ERCC5 promoter were selected for investigation. Genomic DNA was obtained from peripheral blood cells, and polymerase chain reaction-ligation detection reaction was used to analyze these SNPs. The χ 2 test or Fisher's exact test was then used to investigate the association between polymorphisms and chemotherapy response. Our results showed that the response rate among patients with the -763GG genotype (72.7%) was significantly higher than that of other genotypes (22.2% for AA genotype, p = 0.008 and 37.2% for AG genotype, p = 0.046 respectively). In addition, the response rate among patients with the +25AA genotype (75%) was significantly higher than that of other genotypes (24.1% for GG genotype, p = 0.004 and 35.7% for AG genotype, p = 0.022 respectively). Patients with the -763A/+25G haplotype had a higher risk of non-response to oxaliplatin chemotherapy compared to those carrying the -763G/+25A haplotype (OR 2.672, 95% CI 1.353-5.278, p = 0.004). However, no genetic variation was observed at site -413, and no significant association was found between the -1415C>T, +202C>T or +372C>T polymorphisms and chemotherapy response. Therefore, these data suggest that ERCC5 promoter polymorphisms at -763 and +25 may be important predictors of response to oxaliplatin chemotherapy.

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Multiplex PCR/LDR for detection of K-ras mutations in primary colon tumors

Cited by 158 publications

References 71 publications

MicroRNAs targeting mutant K-ras by electrotransfer inhibit human colorectal adenocarcinoma cell growth in vitro and in vivo

MicroRNAs targeting mutant K-ras by electrotransfer inhibit human colorectal adenocarcinoma cell growth in vitro and in vivo

Hereditary stability and variation in evolution and development

ERCC5 promoter polymorphisms at –763 and +25 predict the response to oxaliplatin-based chemotherapy in patients with advanced colorectal cancer

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