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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

PAK1 activation is correlated with tumor stage at presentation. A-Dot plot demonstrating the expression of E-cadherin in the examined tumors in relation to the surgical stage of each tumor. The correlation between variables were examined by Spearman Rank Correlation analysis. B- Dot plot demonstrating the expression of p-PAK1(Thr423) in stage I and stage II/III tumors. The mean between groups was compared by student T-test. C- Dot plot demonstrating the expression of p-Crk-II(Ser41) in stage I and stage II/III tumors. The mean between groups was compared by student T-test.
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Fig4: PAK1 activation is correlated with tumor stage at presentation. A-Dot plot demonstrating the expression of E-cadherin in the examined tumors in relation to the surgical stage of each tumor. The correlation between variables were examined by Spearman Rank Correlation analysis. B- Dot plot demonstrating the expression of p-PAK1(Thr423) in stage I and stage II/III tumors. The mean between groups was compared by student T-test. C- Dot plot demonstrating the expression of p-Crk-II(Ser41) in stage I and stage II/III tumors. The mean between groups was compared by student T-test.

Mentions: Considering we identified that PAK1 activation is closely correlated with E-cadherin and p120-catenin expression in our clinical samples, we were facing the question whether PAK1 activation establishes any association with tumor stage at presentation. In order to answer this question, we reviewed the medical records for each case and extracted relevant clinical parameters to surgical staging (i.e., tumor size and mediastinal nodal involvement) and correlated the results with E-cadherin, p-PAK1(Thr423) and p-Crk-II(Ser41) expression in our samples. All examined tumors were surgically removed so none of the cases had distant metastasis therefore our cases ranged from Stage I to III. Of note, majority of resected lung tumors including cases in our repository are from stage I disease with a limited number of stage II-III cases. As expected the intensity of E-cadherin expression established a close correlation with surgical stage of tumors (Figure 4A). Furthermore, we observed that the mean of p-PAK1(Thr423) expression was significantly higher in stage II/III vs. stage I tumors (Figure 4B). However, the difference between mean of p-Crk-II(Ser41) expression in stage I vs stage II/III tumors did not reach the statistical threshold (Figure 4C). We should mention that the tumor stage (i.e., tumor size, nodal involvement and distant metastasis) is a function of (i) tumor biology, and (ii) the time interval that tumor has been growing. In other words, aggressive tumors generally present at higher stages (as we see in case of most tumors with activated PAK1) however we also see one case with no PAK1 activation and high stage. The high stage of this case can be explained by the long period of time this tumor has been growing prior to being detected and surgically removed.Figure 4


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)

PAK1 activation is correlated with tumor stage at presentation. A-Dot plot demonstrating the expression of E-cadherin in the examined tumors in relation to the surgical stage of each tumor. The correlation between variables were examined by Spearman Rank Correlation analysis. B- Dot plot demonstrating the expression of p-PAK1(Thr423) in stage I and stage II/III tumors. The mean between groups was compared by student T-test. C- Dot plot demonstrating the expression of p-Crk-II(Ser41) in stage I and stage II/III tumors. The mean between groups was compared by student T-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Fig4: PAK1 activation is correlated with tumor stage at presentation. A-Dot plot demonstrating the expression of E-cadherin in the examined tumors in relation to the surgical stage of each tumor. The correlation between variables were examined by Spearman Rank Correlation analysis. B- Dot plot demonstrating the expression of p-PAK1(Thr423) in stage I and stage II/III tumors. The mean between groups was compared by student T-test. C- Dot plot demonstrating the expression of p-Crk-II(Ser41) in stage I and stage II/III tumors. The mean between groups was compared by student T-test.
Mentions: Considering we identified that PAK1 activation is closely correlated with E-cadherin and p120-catenin expression in our clinical samples, we were facing the question whether PAK1 activation establishes any association with tumor stage at presentation. In order to answer this question, we reviewed the medical records for each case and extracted relevant clinical parameters to surgical staging (i.e., tumor size and mediastinal nodal involvement) and correlated the results with E-cadherin, p-PAK1(Thr423) and p-Crk-II(Ser41) expression in our samples. All examined tumors were surgically removed so none of the cases had distant metastasis therefore our cases ranged from Stage I to III. Of note, majority of resected lung tumors including cases in our repository are from stage I disease with a limited number of stage II-III cases. As expected the intensity of E-cadherin expression established a close correlation with surgical stage of tumors (Figure 4A). Furthermore, we observed that the mean of p-PAK1(Thr423) expression was significantly higher in stage II/III vs. stage I tumors (Figure 4B). However, the difference between mean of p-Crk-II(Ser41) expression in stage I vs stage II/III tumors did not reach the statistical threshold (Figure 4C). We should mention that the tumor stage (i.e., tumor size, nodal involvement and distant metastasis) is a function of (i) tumor biology, and (ii) the time interval that tumor has been growing. In other words, aggressive tumors generally present at higher stages (as we see in case of most tumors with activated PAK1) however we also see one case with no PAK1 activation and high stage. The high stage of this case can be explained by the long period of time this tumor has been growing prior to being detected and surgically removed.Figure 4

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