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

Mortazavi F, Lu J, Phan R, Lewis M, Trinidad K, Aljilani A, Pezeshkpour G, Tamanoi F - BMC Cancer (2015)

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Division of Hematology/Oncology, West Los Angeles VA, Los Angeles, CA, USA. fredmortazavi@ucla.edu.

ABSTRACT

Background: Key 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).

Methods: We 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.

Results: Immunohistochemical 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.

Conclusions: Our 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.

No MeSH data available.


Related in: MedlinePlus

Addition of PAK1 inhibitor to KRAS prenylation inhibitors dramatically alters cell morphology and motility. Photomicrographs of standard wound healing assays in H157 and A549 cells (KRAS mutant). Cells were exposed to (i) farnesyltransferase inhibitor (BMS-225975 at 2 μM) and geranylgeranyltransferase inhibitor (P61A6 at 2 μM) [F + G]; (ii) PAK1 inhibitor (IPA-3 at 5 μM); (iii) combination of the three inhibitors or (iv) the inhibitor vehicles for 24-48 hours.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4477307&req=5

Fig7: Addition of PAK1 inhibitor to KRAS prenylation inhibitors dramatically alters cell morphology and motility. Photomicrographs of standard wound healing assays in H157 and A549 cells (KRAS mutant). Cells were exposed to (i) farnesyltransferase inhibitor (BMS-225975 at 2 μM) and geranylgeranyltransferase inhibitor (P61A6 at 2 μM) [F + G]; (ii) PAK1 inhibitor (IPA-3 at 5 μM); (iii) combination of the three inhibitors or (iv) the inhibitor vehicles for 24-48 hours.

Mentions: In fact, addition of PAK1 inhibitor (IPA-3) to (FTI + GGTI) and exposure of H157 cells to these inhibitors resulted in a prominent loss of cellular motility and a clear change in cellular morphology in a standard wound healing assay (Figure 7). H157 cells that have a mesenchymal morphology, obtained an epithelial phenotype with a smooth border when exposed to IPA-3 and (FTI + GGTI) combination for 48 hours. Of note, exposure of H157 cells to (FTI + GGTI) or IPA-3 alone did not demonstrate a noticeable effect in the wound healing assay. A similar but less pronounced effect was observed in A549 cells at 24 hours.Figure 7


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

Mortazavi F, Lu J, Phan R, Lewis M, Trinidad K, Aljilani A, Pezeshkpour G, Tamanoi F - BMC Cancer (2015)

Addition of PAK1 inhibitor to KRAS prenylation inhibitors dramatically alters cell morphology and motility. Photomicrographs of standard wound healing assays in H157 and A549 cells (KRAS mutant). Cells were exposed to (i) farnesyltransferase inhibitor (BMS-225975 at 2 μM) and geranylgeranyltransferase inhibitor (P61A6 at 2 μM) [F + G]; (ii) PAK1 inhibitor (IPA-3 at 5 μM); (iii) combination of the three inhibitors or (iv) the inhibitor vehicles for 24-48 hours.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4477307&req=5

Fig7: Addition of PAK1 inhibitor to KRAS prenylation inhibitors dramatically alters cell morphology and motility. Photomicrographs of standard wound healing assays in H157 and A549 cells (KRAS mutant). Cells were exposed to (i) farnesyltransferase inhibitor (BMS-225975 at 2 μM) and geranylgeranyltransferase inhibitor (P61A6 at 2 μM) [F + G]; (ii) PAK1 inhibitor (IPA-3 at 5 μM); (iii) combination of the three inhibitors or (iv) the inhibitor vehicles for 24-48 hours.
Mentions: In fact, addition of PAK1 inhibitor (IPA-3) to (FTI + GGTI) and exposure of H157 cells to these inhibitors resulted in a prominent loss of cellular motility and a clear change in cellular morphology in a standard wound healing assay (Figure 7). H157 cells that have a mesenchymal morphology, obtained an epithelial phenotype with a smooth border when exposed to IPA-3 and (FTI + GGTI) combination for 48 hours. Of note, exposure of H157 cells to (FTI + GGTI) or IPA-3 alone did not demonstrate a noticeable effect in the wound healing assay. A similar but less pronounced effect was observed in A549 cells at 24 hours.Figure 7

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Division of Hematology/Oncology, West Los Angeles VA, Los Angeles, CA, USA. fredmortazavi@ucla.edu.

ABSTRACT

Background: Key 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).

Methods: We 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.

Results: Immunohistochemical 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.

Conclusions: Our 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.

No MeSH data available.


Related in: MedlinePlus