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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

KRAS mutant NSCLC specimens express higher activated PAK1 compared to KRAS wild type samples. A-Dot plot demonstrating immunohistochemical expression of p-PAK1(Thr423) in KRAS mutant and KRAS wild type NSCLC clinical specimens. p-PAK1(Thr423) is detectable in all KRAS mutant specimens while most KRAS wild type samples express lower p-PAK1(Thr423). B- Dot plot demonstrating immunohistochemical expression of E-cadherin in KRAS mutant vs. KRAS wild type NSCLC clinical specimens.
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Fig5: KRAS mutant NSCLC specimens express higher activated PAK1 compared to KRAS wild type samples. A-Dot plot demonstrating immunohistochemical expression of p-PAK1(Thr423) in KRAS mutant and KRAS wild type NSCLC clinical specimens. p-PAK1(Thr423) is detectable in all KRAS mutant specimens while most KRAS wild type samples express lower p-PAK1(Thr423). B- Dot plot demonstrating immunohistochemical expression of E-cadherin in KRAS mutant vs. KRAS wild type NSCLC clinical specimens.

Mentions: Mutated KRAS transduces its oncogenic signal through several downstream effectors. Recently, PAK1 has been identified as one of the prominent players that transduce the oncogenic KRAS signal [1-4]. Here we examined whether PAK1 is activated in KRAS mutant NSCLC specimens. Towards this end, we examined the presence of KRAS mutations in codon 12, 13 and 61 by examining KRAS exons 2 and 3 (as explained in the Methods) and compared PAK1 phosphorylation between KRAS mutant and wild type groups (Figure 5). We observed that KRAS mutant group demonstrated a statistically significant higher mean of p-PAK1(Thr423) expression as compared to samples harboring wild type KRAS (P < 0.0265) “see Additional file 1”. Furthermore, we did not observe any KRAS mutant sample that did not express p-PAK1(Thr423). We should mention that we noticed one case that carried wild type KRAS along with elevated p-PAK1(Thr423). This finding suggests that upstream oncogenes other than mutated KRAS also transduce their signal through PAK1 (e.g., PI3K) [13]. KRAS mutant samples also demonstrated lower mean of E-cadherin expression compared to wild type group however this difference was not statistically significant. These data further suggest that PAK1 is activated by mutated KRAS in lung cancer.Figure 5


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)

KRAS mutant NSCLC specimens express higher activated PAK1 compared to KRAS wild type samples. A-Dot plot demonstrating immunohistochemical expression of p-PAK1(Thr423) in KRAS mutant and KRAS wild type NSCLC clinical specimens. p-PAK1(Thr423) is detectable in all KRAS mutant specimens while most KRAS wild type samples express lower p-PAK1(Thr423). B- Dot plot demonstrating immunohistochemical expression of E-cadherin in KRAS mutant vs. KRAS wild type NSCLC clinical specimens.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4477307&req=5

Fig5: KRAS mutant NSCLC specimens express higher activated PAK1 compared to KRAS wild type samples. A-Dot plot demonstrating immunohistochemical expression of p-PAK1(Thr423) in KRAS mutant and KRAS wild type NSCLC clinical specimens. p-PAK1(Thr423) is detectable in all KRAS mutant specimens while most KRAS wild type samples express lower p-PAK1(Thr423). B- Dot plot demonstrating immunohistochemical expression of E-cadherin in KRAS mutant vs. KRAS wild type NSCLC clinical specimens.
Mentions: Mutated KRAS transduces its oncogenic signal through several downstream effectors. Recently, PAK1 has been identified as one of the prominent players that transduce the oncogenic KRAS signal [1-4]. Here we examined whether PAK1 is activated in KRAS mutant NSCLC specimens. Towards this end, we examined the presence of KRAS mutations in codon 12, 13 and 61 by examining KRAS exons 2 and 3 (as explained in the Methods) and compared PAK1 phosphorylation between KRAS mutant and wild type groups (Figure 5). We observed that KRAS mutant group demonstrated a statistically significant higher mean of p-PAK1(Thr423) expression as compared to samples harboring wild type KRAS (P < 0.0265) “see Additional file 1”. Furthermore, we did not observe any KRAS mutant sample that did not express p-PAK1(Thr423). We should mention that we noticed one case that carried wild type KRAS along with elevated p-PAK1(Thr423). This finding suggests that upstream oncogenes other than mutated KRAS also transduce their signal through PAK1 (e.g., PI3K) [13]. KRAS mutant samples also demonstrated lower mean of E-cadherin expression compared to wild type group however this difference was not statistically significant. These data further suggest that PAK1 is activated by mutated KRAS in lung cancer.Figure 5

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