Significance of KRAS/PAK1/Crk pathway in non-small cell lung cancer oncogenesis.

BackgroundKey effector(s) of mutated KRAS in lung cancer progression and metastasis are unknown. Here we investigated the role of PAK1/Crk axis in transduction of the oncogenic KRAS signal in non-small cell lung cancer (NSCLC).MethodsWe used NSCLC clinical specimens to examine the correlation among KRAS mutations (codon 12, 13 and 61); PAK1/Crk axis activation [p-PAK1(Thr423), p-Crk(Ser41)]; and adhesion molecules expression by immunohistochemistry. For assessing the role of proto-oncogene c-Crk as a KRAS effector, we inhibited KRAS in NSCLC cells by a combination of farnesyltransferase inhibitor (FTI) and geranylgeranyltransferase inhibitor (GGTI) and measured p-Crk-II(Ser41) by western blotting. Finally, we disrupted the signaling network downstream of KRAS by blocking KRAS/PAK1/Crk axis with PAK1 inhibitors (i.e., IPA-3, FRAX597 or FRAX1036) along with partial inhibition of all other KRAS effectors by prenylation inhibitors (FTI + GGTI) and examined the motility, morphology and proliferation of the NSCLC cells.ResultsImmunohistochemical analysis demonstrated an inverse correlation between PAK1/Crk phosphorylation and E-cadherin/p120-catenin expression. Furthermore, KRAS mutant tumors expressed higher p-PAK1(Thr423) compared to KRAS wild type. KRAS prenylation inhibition by (FTI + GGTI) completely dephosphorylated proto-oncogene c-Crk on Serine 41 while Crk phosphorylation did not change by individual prenylation inhibitors or diluent. Combination of PAK1 inhibition and partial inhibition of all other KRAS effectors by (FTI + GGTI) dramatically altered morphology, motility and proliferation of H157 and A549 cells.ConclusionsOur data provide evidence that proto-oncogene c-Crk is operative downstream of KRAS in NSCLC. Previously we demonstrated that Crk receives oncogenic signals from PAK1. These data in conjunction with the work of others that have specified the role of PAK1 in transduction of KRAS signal bring forward the importance of KRAS/PAK1/Crk axis as a prominent pathway in the oncogenesis of KRAS mutant lung cancer

An integrative approach unveils FOSL1 as an oncogene vulnerability in KRAS-driven lung and pancreatic cancer

KRAS mutated tumours represent a large fraction of human cancers, but the vast majority remains refractory to current clinical therapies. Thus, a deeper understanding of the molecular mechanisms triggered by KRAS oncogene may yield alternative therapeutic strategies. Here we report the identification of a common transcriptional signature across mutant KRAS cancers of distinct tissue origin that includes the transcription factor FOSL1. High FOSL1 expression identifies mutant KRAS lung and pancreatic cancer patients with the worst survival outcome. Furthermore, FOSL1 genetic inhibition is detrimental to both KRAS-driven tumour types. Mechanistically, FOSL1 links the KRAS oncogene to components of the mitotic machinery, a pathway previously postulated to function orthogonally to oncogenic KRAS. FOSL1 targets include AURKA, whose inhibition impairs viability of mutant KRAS cells. Lastly, combination of AURKA and MEK inhibitors induces a deleterious effect on mutant KRAS cells. Our findings unveil KRAS downstream effectors that provide opportunities to treat KRAS-driven cancers

Impact of the Specific Mutation in KRAS Codon 12 Mutated Tumors on Treatment Efficacy in Patients with Metastatic Colorectal Cancer Receiving Cetuximab-Based First-Line Therapy: A Pooled Analysis of Three Trials

Purpose: This study investigated the impact of specific mutations in codon 12 of the Kirsten-ras (KRAS) gene on treatment efficacy in patients with metastatic colorectal cancer (mCRC). Patients: Overall, 119 patients bearing a KRAS mutation in codon 12 were evaluated. All patients received cetuximab-based first-line chemotherapy within the Central European Cooperative Oncology Group (CECOG), AIO KRK-0104 or AIO KRK-0306 trials. Results: Patients with KRAS codon 12 mutant mCRC showed a broad range of outcome when treated with cetuximab-based first-line regimens. Patients with tumors bearing a KRAS p.G12D mutation showed a strong trend to a more favorable outcome compared to other mutations (overall survival 23.3 vs. 14-18 months; hazard ratio 0.66, range 0.43-1.03). An interaction model illustrated that KRAS p.G12C was associated with unfavorable outcome when treated with oxaliplatin plus cetuximab. Conclusion: The present analysis suggests that KRAS codon 12 mutation may not represent a homogeneous entity in mCRC when treated with cetuximab-based first-line therapy. Copyright (C) 2012 S. Karger AG, Base

RelA regulates CXCL1/CXCR2-dependent oncogene-induced senescence in murine Kras-driven pancreatic carcinogenesis

Tumor suppression that is mediated by oncogene-induced senescence (OIS) is considered to function as a safeguard during development of pancreatic ductal adenocarcinoma (PDAC). However, the mechanisms that regulate OIS in PDAC are poorly understood. Here, we have determined that nuclear RelA reinforces OIS to inhibit carcinogenesis in the Kras mouse model of PDAC. Inactivation of RelA accelerated pancreatic lesion formation in Kras mice by abrogating the senescence-associated secretory phenotype (SASP) gene transcription signature. Using genetic and pharmacological tools, we determined that RelA activation promotes OIS via elevation of the SASP factor CXCL1 (also known as KC), which activates CXCR2, during pancreatic carcinogenesis. In Kras mice, pancreas-specific inactivation of CXCR2 prevented OIS and was correlated with increased tumor proliferation and decreased survival. Moreover, reductions in CXCR2 levels were associated with advanced neoplastic lesions in tissue from human pancreatic specimens. Genetically disabling OIS in Kras mice caused RelA to promote tumor proliferation, suggesting a dual role for RelA signaling in pancreatic carcinogenesis. Taken together, our data suggest a pivotal role for RelA in regulating OIS in preneoplastic lesions and implicate the RelA/CXCL1/CXCR2 axis as an essential mechanism of tumor surveillance in PDAC

High-Resolution Three-Dimensional NMR Structure Of The KRAS Proto-Oncogene Promoter Reveals Key Features Of A G-Quadruplex Involved In Transcriptional Regulation

Non-canonical base pairing within guanine-rich DNA and RNA sequences can produce G-quartets, whose stacking leads to the formation of a G-quadruplex (G4). G4s can coexist with canonical duplex DNA in the human genome and have been suggested to suppress gene transcription, and much attention has therefore focused on studying G4s in promotor regions of disease-related genes. For example, the human KRAS proto-oncogene contains a nuclease-hypersensitive element located upstream of the major transcription start site. The KRAS nuclease-hypersensitive element (NHE) region contains a G-rich element (22RT; 5′-AGGGCGGTGTGGGAATAGGGAA-3′) and encompasses a Myc-associated zinc finger-binding site that regulates KRAS transcription. The NEH region therefore has been proposed as a target for new drugs that control KRAS transcription, which requires detailed knowledge of the NHE structure. In this study, we report a high-resolution NMR structure of the G-rich element within the KRAS NHE. We found that the G-rich element forms a parallel structure with three G-quartets connected by a four-nucleotide loop and two short one-nucleotide double-chain reversal loops. In addition, a thymine bulge is found between G8 and G9. The loops of different lengths and the presence of a bulge between the G-quartets are structural elements that potentially can be targeted by small chemical ligands that would further stabilize the structure and interfere or block transcriptional regulators such as Myc-associated zinc finger from accessing their binding sites on the KRAS promoter. In conclusion, our work suggests a possible new route for the development of anticancer agents that could suppress KRAS expression

Precision delivery of RAS-inhibiting siRNA to KRAS driven cancer via peptide-based nanoparticles

Over 95% of pancreatic adenocarcinomas (PDACs), as well as a large fraction of other tumor types, such as colorectal adenocarcinoma, are driven by KRAS activation. However, no direct RAS inhibitors exist for cancer therapy. Furthermore, the delivery of therapeutic agents of any kind to PDAC in particular has been hindered by the extensive desmoplasia and resultant drug delivery challenges that accompanies these tumors. Small interfering RNA (siRNA) is a promising modality for anti-neoplastic therapy due to its precision and wide range of potential therapeutic targets. Unfortunately, siRNA therapy is limited by low serum half-life, vulnerability to intracellular digestion, and transient therapeutic effect. We assessed the ability of a peptide based, oligonucleotide condensing, endosomolytic nanoparticle (NP) system to deliver siRNA to KRAS-driven cancers. We show that this peptide-based NP is avidly taken up by cancer cell

A polymorphism in a let-7 microRNA binding site of KRAS in women with endometriosis.

Endometriosis is found in 5-15% of women of reproductive age and is more frequent in relatives of women with the disease. Activation of KRAS results in de novo endometriosis in mice, however, activating KRAS mutations have not been identified in women. We screened 150 women with endometriosis for a polymorphism in a let-7 microRNA (miRNA) binding site in the 3'-UTR of KRAS and detected a KRAS variant allele in 31% of women with endometriosis as opposed to 5% of a large diverse control population. KRAS mRNA and protein expression were increased in cultured endometrial stromal cells of women with the KRAS variant. Increased KRAS protein was due to altered miRNA binding as demonstrated in reporter assays. Endometrial stromal cells from women with the KRAS variant showed increased proliferation and invasion. In a murine model, endometrial xenografts containing the KRAS variant demonstrated increased proliferation and decreased progesterone receptor levels. These findings suggest that an inherited polymorphism of a let-7 miRNA binding site in KRAS leads to abnormal endometrial growth and endometriosis. The LCS6 polymorphism is the first described genetic marker of endometriosis risk

Characterizing genomic alterations in cancer by complementary functional associations.

Systematic efforts to sequence the cancer genome have identified large numbers of mutations and copy number alterations in human cancers. However, elucidating the functional consequences of these variants, and their interactions to drive or maintain oncogenic states, remains a challenge in cancer research. We developed REVEALER, a computational method that identifies combinations of mutually exclusive genomic alterations correlated with functional phenotypes, such as the activation or gene dependency of oncogenic pathways or sensitivity to a drug treatment. We used REVEALER to uncover complementary genomic alterations associated with the transcriptional activation of β-catenin and NRF2, MEK-inhibitor sensitivity, and KRAS dependency. REVEALER successfully identified both known and new associations, demonstrating the power of combining functional profiles with extensive characterization of genomic alterations in cancer genomes

EGFR inhibitor as second-line therapy in a patient with mutant RAS metastatic colorectal cancer: circulating tumor DNA to personalize treatment

A 47-year-old male patient presented in March 2016 to our unit with a palpable painless left supraclavicular mass. A whole-body contrastenhanced computed tomography (CT) scan revealed a left supraclavicular lymphadenopathy, transverse colon thickening (3 cm), multiple chest and abdominal lymphadenopathies, and peritoneal carcinomatosis. Colonoscopy revealed a bleeding area at 15 cm from the anal verge; biopsy was performed, and the result was negative for a primary cancer