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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 alters the proliferation of NSCLC cells. Line charts demonstrating mean cell count of H157 and A549 cells upon exposure to prenylation inhibitors [FTI (BMS-225975) and GGTI (P61A6) at 500 nM each], PAK1 inhibitor (FRAX1036 at 10 μM) or combination in comparison to cells exposed to inhibitors’ diluent (DMSO).
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Fig9: Addition of PAK1 inhibitor to KRAS prenylation inhibitors alters the proliferation of NSCLC cells. Line charts demonstrating mean cell count of H157 and A549 cells upon exposure to prenylation inhibitors [FTI (BMS-225975) and GGTI (P61A6) at 500 nM each], PAK1 inhibitor (FRAX1036 at 10 μM) or combination in comparison to cells exposed to inhibitors’ diluent (DMSO).

Mentions: Considering downstream effectors of both PAK1 and KRAS include modulators of cell proliferation, we investigated whether inhibition of PAK1 along with KRAS prenylation inhibition may alter proliferation of NSCLC cells. Towards this end, we exposed A549 and H157 cells to (i) prenylation inhibitors [i.e., combination of FTI (BMS-225975) and GGTI (P61A6) at 500 nM each], (ii) PAK1 inhibitor (FRAX1036 at 10 μM, Genentech), (iii) prenylation inhibitors and PAK1 inhibitor, and (iv) inhibitors’ vehicle (Figure 9). H157 cells continued to grow in the presence of either prenylation inhibitors or PAK1 inhibitor at the above mentioned concentrations however combination of prenylation inhibitors and PAK1 inhibitor synergistically reduced the proliferation of these cells. On the other hand, proliferation of A549 cells were affected by PAK1 or prenylation inhibitors to some extent nevertheless combination of PAK1 and prenylation inhibitors showed a much stronger effect in halting the proliferation of A549 cells.Figure 9


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 alters the proliferation of NSCLC cells. Line charts demonstrating mean cell count of H157 and A549 cells upon exposure to prenylation inhibitors [FTI (BMS-225975) and GGTI (P61A6) at 500 nM each], PAK1 inhibitor (FRAX1036 at 10 μM) or combination in comparison to cells exposed to inhibitors’ diluent (DMSO).
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Fig9: Addition of PAK1 inhibitor to KRAS prenylation inhibitors alters the proliferation of NSCLC cells. Line charts demonstrating mean cell count of H157 and A549 cells upon exposure to prenylation inhibitors [FTI (BMS-225975) and GGTI (P61A6) at 500 nM each], PAK1 inhibitor (FRAX1036 at 10 μM) or combination in comparison to cells exposed to inhibitors’ diluent (DMSO).
Mentions: Considering downstream effectors of both PAK1 and KRAS include modulators of cell proliferation, we investigated whether inhibition of PAK1 along with KRAS prenylation inhibition may alter proliferation of NSCLC cells. Towards this end, we exposed A549 and H157 cells to (i) prenylation inhibitors [i.e., combination of FTI (BMS-225975) and GGTI (P61A6) at 500 nM each], (ii) PAK1 inhibitor (FRAX1036 at 10 μM, Genentech), (iii) prenylation inhibitors and PAK1 inhibitor, and (iv) inhibitors’ vehicle (Figure 9). H157 cells continued to grow in the presence of either prenylation inhibitors or PAK1 inhibitor at the above mentioned concentrations however combination of prenylation inhibitors and PAK1 inhibitor synergistically reduced the proliferation of these cells. On the other hand, proliferation of A549 cells were affected by PAK1 or prenylation inhibitors to some extent nevertheless combination of PAK1 and prenylation inhibitors showed a much stronger effect in halting the proliferation of A549 cells.Figure 9

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