Gene Summary

Gene:CRKL; CRK like proto-oncogene, adaptor protein
Summary:This gene encodes a protein kinase containing SH2 and SH3 (src homology) domains which has been shown to activate the RAS and JUN kinase signaling pathways and transform fibroblasts in a RAS-dependent fashion. It is a substrate of the BCR-ABL tyrosine kinase, plays a role in fibroblast transformation by BCR-ABL, and may be oncogenic.[provided by RefSeq, Jan 2009]
Databases:OMIM, HGNC, Ensembl, GeneCard, Gene
Protein:crk-like protein
Source:NCBIAccessed: 29 August, 2019


What does this gene/protein do?
Show (13)
Pathways:What pathways are this gene/protein implicaed in?
Show (9)

Cancer Overview

Research Indicators

Publications Per Year (1994-2019)
Graph generated 29 August 2019 using data from PubMed using criteria.

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

  • CRKL
  • Antineoplastic Agents
  • K562 Cells
  • Neoplastic Cell Transformation
  • Transfection
  • Drug Resistance
  • Benzamides
  • Protein Kinase Inhibitors
  • Protein Binding
  • Cancer Gene Expression Regulation
  • Oncogene Fusion Proteins
  • Phosphoproteins
  • Cell Movement
  • Signal Transducing Adaptor Proteins
  • Cell Proliferation
  • Young Adult
  • Protein-Tyrosine Kinases
  • Mutation
  • Recombinant Fusion Proteins
  • Nuclear Proteins
  • Piperazines
  • TOR Serine-Threonine Kinases
  • alpha-Fetoproteins
  • Fusion Proteins, bcr-abl
  • Trans-Activators
  • Proto-Oncogene Proteins c-crk
  • Signal Transduction
  • Phosphorylation
  • Apoptosis
  • MicroRNAs
  • Tyrosine
  • src Homology Domains
  • Stromal Cells
  • Thiazoles
  • Leukemic Gene Expression Regulation
  • Chromosome 22
  • Chronic Myelogenous Leukemia
  • Transcription
  • Lung Cancer
  • Imatinib Mesylate
  • Pyrimidines
Tag cloud generated 29 August, 2019 using data from PubMed, MeSH and CancerIndex

Specific Cancers (2)

Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications: CRKL (cancer-related)

Wen DY, Pan DH, Lin P, et al.
Downregulation of miR‑486‑5p in papillary thyroid carcinoma tissue: A study based on microarray and miRNA sequencing.
Mol Med Rep. 2018; 18(3):2631-2642 [PubMed] Free Access to Full Article Related Publications
Abnormal expression of microRNA (miR) is associated with the occurrence and progression of various types of cancers, including papillary thyroid carcinoma (PTC). In the present study, the aim was to explore miR‑486‑5p expression and its role in PTC, as well as to investigate the biological function of its potential target genes. The expression levels of miR‑486‑5p and its clinicopathological significance were examined in 507 PTC and 59 normal thyroid samples via The Cancer Genome Atlas (TCGA). Subsequently, the results were validated using data from Gene Expression Omnibus (GEO) and ArrayExpress. Receiver operating characteristic and summary receiver operating characteristic curves were used to assess the ability of miR‑486‑5p in distinguishing PTC from normal tissue. Furthermore, potential miR‑486‑5p mRNA targets were identified using 12 prediction tools and enrichment analysis was performed on the encoding genes using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes. The expression levels of miR‑486‑5p were consistently downregulated in PTC compared with in normal tissue across datasets from TCGA, GEO (GSE40807, GSE62054 and GSE73182) and ArrayExpress (E‑MTAB‑736). The results also demonstrated that miR‑486‑5p expression was associated with cancer stage (P=0.003), pathologic lymph node (P=0.047), metastasis (P=0.042), neoplasm (P=0.012) and recurrence (P=0.016) in patients with PTC. In addition, low expression of miR‑486‑5p in patients with PTC was associated with a worse overall survival. A total of 80 miR‑486‑5p‑related genes were observed from at least 9 of 12 prediction platforms, and these were involved in 'hsa05200: Pathways in cancer' and 'hsa05206: MicroRNAs in cancer'. Finally, three hub genes, CRK like proto‑oncogene, phosphatase and tensin homolog and tropomyosin 3, were identified as important candidates in tumorigenesis and progression of PTC. In conclusion, it may be hypothesized that miR‑486‑5p contributes towards PTC onset and progression, and may act as a clinical target. However, in vitro and in vivo experiments are required to validate the findings of the present study.

Huang N, Huang Z, Gao M, et al.
Induction of apoptosis in imatinib sensitive and resistant chronic myeloid leukemia cells by efficient disruption of bcr-abl oncogene with zinc finger nucleases.
J Exp Clin Cancer Res. 2018; 37(1):62 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: The bcr-abl fusion gene is the pathological origin of chronic myeloid leukemia (CML) and plays a critical role in the resistance of imatinib. Thus, bcr-abl disruption-based novel therapeutic strategy may warrant exploration. In our study, we were surprised to find that the characteristics of bcr-abl sequences met the design requirements of zinc finger nucleases (ZFNs).
METHODS: We constructed the ZFNs targeting bcr-abl with high specificity through simple modular assembly approach. Western blotting was conducted to detect the expression of BCR-ABL and phosphorylation of its downstream STAT5, ERK and CRKL in CML cells. CCK8 assay, colony-forming assay and flow cytometry (FCM) were used to evaluate the effect of the ZFNs on the viablity and apoptosis of CML cells and CML CD34
RESULTS: The ZFNs skillfully mediated 8-base NotI enzyme cutting site addition in bcr-abl gene of imatinib sensitive and resistant CML cells by homology-directed repair (HDR), which led to a stop codon and terminated the translation of BCR-ABL protein. As expected, the disruption of bcr-abl gene induced cell apoptosis and inhibited cell proliferation. Notably, we obtained similar result in CD34
CONCLUSION: These results reveal that the bcr-abl gene disruption based on ZFNs may provide a treatment choice for imatinib resistant or intolerant CML patients.

Shi X, Xiao X, Yuan N, et al.
MicroRNA-379 Suppresses Cervical Cancer Cell Proliferation and Invasion by Directly Targeting V-crk Avian Sarcoma Virus CT10 Oncogene Homolog-Like (CRKL).
Oncol Res. 2018; 26(7):987-996 [PubMed] Related Publications
Cervical cancer is the fourth most common malignancy among females worldwide. MicroRNA-379 (miR-379) is aberrantly expressed in multiple human cancer types. However, the expression pattern, roles, and detailed regulatory mechanisms of miR-379 in cervical cancer remain unknown. In this study, we found that miR-379 expression was downregulated in cervical cancer tissues and cell lines. Low miR-379 expression was correlated with International Federation of Gynecology and Obstetrics (FIGO) stage, lymph node metastasis, and distant metastasis. Additionally, miR-379 overexpression suppressed the proliferation and invasion of cervical cancer cells. Furthermore, V-crk avian sarcoma virus CT10 oncogene homolog-like (CRKL) was identified as a direct target of miR-379 in cervical cancer. CRKL was upregulated in cancer tissues and negatively correlated with miR-379 expression. Moreover, restored CRKL expression rescued the inhibitory effects of miR-379 overexpression on cell proliferation and invasion. In conclusion, miR-379 may serve as a tumor suppressor in cervical cancer by directly targeting CRKL. Restoring miR-379 expression may be an effective strategy for the treatment of cervical cancer.

Bian X, Liang Z, Feng A, et al.
HDAC inhibitor suppresses proliferation and invasion of breast cancer cells through regulation of miR-200c targeting CRKL.
Biochem Pharmacol. 2018; 147:30-37 [PubMed] Free Access to Full Article Related Publications
Although histone deacetylase (HDAC) inhibitors have been shown to effectively induce the inhibition of proliferation and migration in breast cancer, the anticancer mechanism remains poorly understood. Our studies show that miR-200c was significantly downregulated in breast cancer cell lines compared to normal cell lines and inversely correlated with the levels of class IIa HDACs and CRKL. HDAC inhibitors and the ectopic expression of miR-200c as tumor suppressors inhibited the proliferation, invasion, and migration of breast cancer cells by downregulating CRKL. These results indicate that the anticancer mechanism of HDAC inhibitor was realized partially by regulating miR-200c via CRKL targeting. Our findings suggest that the HDAC-miR200c-CRKL signaling axis could be a novel diagnostic marker and potential therapeutic target in breast cancer.

Fadare O, Roma AA, Parkash V, et al.
Does a p53 "Wild-type" Immunophenotype Exclude a Diagnosis of Endometrial Serous Carcinoma?
Adv Anat Pathol. 2018; 25(1):61-70 [PubMed] Related Publications
An aberrant p53 immunophenotype may be identified in several histotypes of endometrial carcinoma, and is accordingly recognized to lack diagnostic specificity in and of itself. However, based on the high frequency with which p53 aberrations have historically been identified in endometrial serous carcinoma, a mutation-type immunophenotype is considered to be highly sensitive for the histotype. Using an illustrative case study and a review of the literature, we explore a relatively routine diagnostic question: whether the negative predictive value of a wild-type p53 immunophenotype for serous carcinoma is absolute, that is, whether a p53-wild type immunophenotype is absolutely incompatible with a diagnosis of serous carcinoma. The case is an advanced stage endometrial carcinoma that was reproducibly classified by pathologists from 3 institutions as serous carcinoma based on its morphologic features. By immunohistochemistry, the tumor was p53-wild type (DO-7 clone), diffusely positive for p16 (block positivity), and showed retained expression of PTEN, MSH2, MSH6, MLH1, and PMS2. Next generation sequencing showed that there indeed was an underlying mutation in TP53 (D393fs*78, R213*). The tumor was microsatellite stable, had a low mutational burden (4 mutations per MB), and displayed no mutations in the exonuclease domain of DNA polymerase epsilon (POLE) gene. Other genomic alterations included RB1 mutation (R46fs*19), amplifications in MYST3 and CRKL, and ARID1A deletion (splice site 5125-94_5138del108). A review of the recent literature identified 5 studies in which a total of 259 cases of serous carcinoma were whole-exome sequenced. The average TP53 mutational rate in endometrial serous carcinoma was only 75% (range, 60 to 88). A total of 12 (33%) of 36 immunohistochemical studies reported a p53-aberrant rate of <80% in endometrial serous carcinoma. We discuss in detail several potential explanations that may underlie the scenario of serous carcinoma-like morphology combined with p53-wild-type immunophenotype, including analytic limitations, a nonserous histotype displaying morphologic mimicry of serous carcinoma, and true biological phenomena (including the possibility of a TP53-independent pathway of endometrial serous carcinogenesis). Ultimately, our central thematic question is provisionally answered in the negative. At present, the available data would not support a categorical conclusion that a p53 alteration is a necessary and obligate component in the genesis and/or diagnosis of endometrial serous carcinoma. On the basis of their collective experience, the authors proffer some recommendations on the use of p53 immunohistochemistry in the histotyping of endometrial carcinomas.

Ren Y, Shang J, Li J, et al.
The long noncoding RNA
J Biol Chem. 2017; 292(43):17939-17949 [PubMed] Free Access to Full Article Related Publications
The long non-coding RNA (lncRNA)

Mazzu YZ, Hu Y, Soni RK, et al.
miR-193b-Regulated Signaling Networks Serve as Tumor Suppressors in Liposarcoma and Promote Adipogenesis in Adipose-Derived Stem Cells.
Cancer Res. 2017; 77(21):5728-5740 [PubMed] Related Publications
Well-differentiated and dedifferentiated liposarcomas (WDLS/DDLS) account for approximately 13% of all soft tissue sarcoma in adults and cause substantial morbidity or mortality in the majority of patients. In this study, we evaluated the functions of miRNA (miR-193b) in liposarcoma

Kong Y, Wu YL, Song Y, et al.
Ruxolitinib/nilotinib cotreatment inhibits leukemia-propagating cells in Philadelphia chromosome-positive ALL.
J Transl Med. 2017; 15(1):184 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: As one of the major treatment obstacles in Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph
METHODS: RNA sequencing (RNA-Seq) and real-time quantitative polymerase chain reaction (qRT-PCR) were performed to analyze the gene expression profiles of the sorted LPCs and other cell fractions from patients with de novo Ph
RESULTS: RNA-Seq and qRT-PCR demonstrated that JAK2 was more highly expressed in the sorted LPCs than in the other cell fractions in de novo Ph
CONCLUSIONS: In summary, this pre-clinical study provides a scientific rationale for simultaneously targeting BCR-ABL and JAK2 activities as a promising anti-LPCs therapeutic approach for patients with de novo Ph

Fujioka M, Asano Y, Nakada S, Ohba Y
SH2 Domain-Based FRET Biosensor for Measuring BCR-ABL Activity in Living CML Cells.
Methods Mol Biol. 2017; 1555:513-534 [PubMed] Related Publications
Fluorescent proteins (FPs) displaying distinct spectra have shed their light on a wide range of biological functions. Moreover, sophisticated biosensors engineered to contain single or multiple FPs, including Förster resonance energy transfer (FRET)-based biosensors, spatiotemporally reveal the molecular mechanisms underlying a variety of pathophysiological processes. However, their usefulness for applied life sciences has yet to be fully explored. Recently, our research group has begun to expand the potential of FPs from basic biological research to the clinic. Here, we describe a method to evaluate the responsiveness of leukemia cells from patients to tyrosine kinase inhibitors using a biosensor based on FP technology and the principle of FRET. Upon phosphorylation of the tyrosine residue of the biosensor, binding of the SH2 domain to phosphotyrosine induces conformational change of the biosensor and brings the donor and acceptor FPs into close proximity. Therefore, kinase activity and response to kinase inhibitors can be monitored by an increase and a decrease in FRET efficiency, respectively. As in basic research, this biosensor resolves hitherto arduous tasks and may provide innovative technological advances in clinical laboratory examinations. State-of-the-art detection devices that enable such innovation are also introduced.

Wang J, Yunyun Z, Wang L, et al.
ABCG2 confers promotion in gastric cancer through modulating downstream CRKL in vitro combining with biostatistics mining.
Oncotarget. 2017; 8(3):5256-5267 [PubMed] Free Access to Full Article Related Publications
ABCG2, member of ATP-binding cassette (ABC) transporter family, is known as crucial regulator related to multi-drug resistance in human tumors and has recently been putatively studied as human carcinoma cell biomarker. While, effects of ABCG2 on human gastric cancer (GC) has not been illustrated thoroughly. In this study, by applying biostatistics mining methods, we observed that ABCG2 is frequently aberrantly expressed in GC patients through exploring dataset of GSE19826 in NCBI GEO database. Contemporary, extreme up-regulation of ABCG2 was discovered in both GC specimens and cell lines of our center, from which we observed high level of ABCG2 associated with GC clinicopathologic features and poor outcomes. Depletion of ABCG2 in MKN-45 GC cells, the cell proliferation was significantly impacted along with cell cycle arrest, and cell apoptosis was induced. Interestingly, combined with data mining of NCBI database, CRKL, a pivotal GC promoter, presents a significant positive correlation with ABCG2. And the expression of CRKL in GC cells was obviously affected through ABCG2 depletion. Simultaneously, over-expression of CRKL in MKN-45 cells significantly rescued most of the phenotypes induced by ABCG2 depletion. Thus, we suggest that ABCG2 is a potential biomarker and target upstream CRKL, which could be further studied for GC diagnosis and therapeutic treatment.

Horiguchi M, Fujioka M, Kondo T, et al.
Improved FRET Biosensor for the Measurement of BCR-ABL Activity in Chronic Myeloid Leukemia Cells.
Cell Struct Funct. 2017; 42(1):15-26 [PubMed] Related Publications
Although the co-development of companion diagnostics with molecular targeted drugs is desirable, truly efficient diagnostics are limited to diseases in which chromosomal translocations or overt mutations are clearly correlated with drug efficacy. Moreover, even for such diseases, few methods are available to predict whether drug administration is effective for each individual patient whose disease is expected to respond to the drug(s). We have previously developed a biosensor based on the principle of Förster resonance energy transfer to measure the activity of the tyrosine kinase BCR-ABL and its response to drug treatment in patient-derived chronic myeloid leukemia cells. The biosensor harbors CrkL, one of the major substrates of BCR-ABL, and is therefore named Pickles after phosphorylation indicator of CrkL en substrate. The efficacy of this technique as a clinical test has been demonstrated, but the number of cells available for analysis is limited in a case-dependent manner, owing to the cleavage of the biosensor in patient-derived leukemia cells. Here, we describe an improved biosensor with an amino acid substitution and a nuclear export signal being introduced. Of the two predicted cleavage positions in CrkL, the mutations inhibited one cleavage completely and the other cleavage partially, thus collectively increasing the number of cells available for drug evaluation. This improved version of the biosensor holds promise in the future development of companion diagnostics to predict responses to tyrosine kinase inhibitors in patients with chronic myeloid leukemia.

Gao M, Huang ZL, Tao K, et al.
Depression of oncogenecity by dephosphorylating and degrading BCR-ABL.
Oncotarget. 2017; 8(2):3304-3314 [PubMed] Free Access to Full Article Related Publications
Aberrant phosphorylation and overexpression of BCR-ABL fusion protein are responsible for the main pathogenesis in chronic myeloid leukemia (CML). Phosphorylated BCR-ABL Y177 recruits GRB2 adaptor and triggers leukemic RAS-MAPK and PI3K-AKT signals. In this study, we engineered a SPOA system to dephosphorylate and degrade BCR-ABL by targeting BCR-ABL Y177. We tested its effect on BCR-ABL phosphorylation and expression, as well as cell proliferation and apoptosis in CML cells. We found that SPOA remarkably dephosphorylated BCR-ABL Y177, prevented GRB2 recruitment, and uncoupled RAS-MAPK and PI3K-AKT signals. Meanwhile, SPOA degraded BCR-ABL oncoprotein in ubiquitin-independent manner and depressed the signal transduction of STAT5 and CRKL by BCR-ABL. Furthermore, SPOA inhibited proliferation and induced apoptosis in CML cells and depressed the oncogenecity of K562 cells in mice. These results provide evidence that dephosphorylating and degrading oncogenic BCR-ABL offer an alternative CML therapy.

Wang J, Fei X, Wu W, et al.
SLC7A5 Functions as a Downstream Target Modulated by CRKL in Metastasis Process of Gastric Cancer SGC-7901 Cells.
PLoS One. 2016; 11(11):e0166147 [PubMed] Free Access to Full Article Related Publications
SLC7A5, who is also named LAT-1, has been validated as a promoter regulated by miRNA-126 in our previous research for gastric cancer cells. However, the mechanisms driving SLC7A5 to affect the bio-function of gastric cancer cells are unclear, remaining us lots of to elucidate. The aim of this study is to investigate the regulating effect of CRKL, one of the critical genes involving with gastric cancer progression, on SLC7A5 expression. By studying the gastric cancer cell lines and clinical pathological specimens, we found that the expression of SLC7A5 was significantly correlated to CRKL. By depleting CRKL in gastric cancer SGC-7901 cells, the SLC7A5 expression was impaired, and the invasion and migration of SGC-7901 cells were suppressed. Ectopic expression of SLC7A5 could drastically rescue the phenotypes induced by CRKL depletion in this study. Accordingly, we conclude that SLC7A5 functions as a promoter in gastric cancer metastasis, and CRKL could be one of its regulators modulating the expression of SLC7A5 and consequentially affect the metastatic feature of SGC-7901 cells. The findings in this study indicate a regulation relationship between CRKL and SLC7A5, and provide useful evidence for gastric cancer therapeutic strategies.

Koczkowska M, Wierzba J, Śmigiel R, et al.
Genomic findings in patients with clinical suspicion of 22q11.2 deletion syndrome.
J Appl Genet. 2017; 58(1):93-98 [PubMed] Free Access to Full Article Related Publications
Chromosome 22q11.2 deletion syndrome, one of the most common human genomic syndromes, has highly heterogeneous clinical presentation. Patients usually harbor a 1.5 to 3 Mb hemizygous deletion at chromosome 22q11.2, resulting in pathognomic TBX1, CRKL and/or MAPK1 haploinsufficiency. However, there are some individuals with clinical features resembling the syndrome who are eventually diagnosed with genomic disorders affecting other chromosomal regions. The objective of this study was to evaluate the additive value of high-resolution array-CGH testing in the cohort of 41 patients with clinical features of 22q11.2 deletion syndrome and negative results of standard cytogenetic diagnostic testing (karyotype and FISH for 22q11.2 locus). Array-CGH analysis revealed no aberrations at chromosomes 22 or 10 allegedly related to the syndrome. Five (12.2 %) patients were found to have other genomic imbalances, namely 17q21.31 microdeletion syndrome (MIM#610443), 1p36 deletion syndrome (MIM#607872), NF1 microduplication syndrome (MIM#613675), chromosome 6pter-p24 deletion syndrome (MIM#612582) and a novel interstitial deletion at 3q26.31 of 0.65 Mb encompassing a dosage-dependent gene NAALADL2. Our study demonstrates that the implementation of array-CGH into the panel of classic diagnostic procedures adds significantly to their efficacy. It allows for detection of constitutional genomic imbalances in 12 % of subjects with negative result of karyotype and FISH targeted for 22q11.2 region. Moreover, if used as first-tier genetic test, the method would provide immediate diagnosis in ∼40 % phenotypic 22q11.2 deletion subjects.

Agatheeswaran S, Pattnayak NC, Chakraborty S
Identification and functional characterization of the miRNA-gene regulatory network in chronic myeloid leukemia lineage negative cells.
Sci Rep. 2016; 6:32493 [PubMed] Free Access to Full Article Related Publications
Chronic myeloid leukemia (CML) is maintained by leukemic stem cells (LSCs) which are resistant to the existing TKI therapy. Hence a better understanding of the CML LSCs is necessary to eradicate these cells and achieve complete cure. Using the miRNA-gene interaction networks from the CML lin(-) cells we identified a set of up/down-regulated miRNAs and corresponding target genes. Association studies (Pearson correlation) from the miRNA and gene expression data showed that miR-1469 and miR-1972 have significantly higher number of target genes, 75 and 50 respectively. We observed that miR-1972 induces G2-M cell cycle arrest and miR-1469 moderately arrested G1 cell cycle when overexpressed in KCL22 cells. We have earlier shown that a combination of imatinib and JAK inhibitor I can significantly bring down the proliferation of CML lineage negative cells. Here we observed that imatinib and JAK inhibitor I combination restored the expression pattern of the down-regulated miRNAs in primary CML lin(-) cells. Thus effective manipulation of the deregulated miRNAs can restore the miRNA-mRNA networks that can efficiently inhibit CML stem and progenitor cells and alleviate the disease.

Wang Y, Dong X, Hu B, et al.
The effects of Micro-429 on inhibition of cervical cancer cells through targeting ZEB1 and CRKL.
Biomed Pharmacother. 2016; 80:311-321 [PubMed] Related Publications
MicroRNA-429 (miR-429) has been suggested to inhibit epithelial-mesenchymal transition (EMT), mainly due to targeting of ZEB1 and ZEB2, which are repressors of the cell to cell contact protein, E-cadherin. In this study, we indicated that regulation of miR-429 in cervical cancer cells modulates cell migration, elongation, as well as transforming growth factor β (TGF-β)-induced stress fiber formation through regulating the cytoskeleton reorganization which is likely independent of the zinc finger E-box binding homeobox (ZEB)/E-cadherin axis. ZEB1 and Crk-like adapter protein (CRKL), as novel targets of miR-429 and direct regulators of the actin cytoskeleton were identified. Remarkably, expression levels of ZEB1 and CRKL were inversely associated with the level of miR-429 in cervical cancer cell lines. In addition, individual knockdown and over-expression of these targeting genes phenocopied the roles of miR-429 over-expression and inhibition on cell elongation, migration, stress fiber formation, and invasion. Targeting of ZEB1 by miR-429 led to a decreased expression and transcriptional activity of CRB3, regulated by interference with the translocation of the CRB3. This finally led to decreasing of the expression of Crumbs 3 (CRB3), which is needed for the formation of stress fiber and contractility. Therefore, miR-429 affects cervical cancer by modulating some EMT-related processes. And in this study, evidences were provided to support a role for miR-429 as a novel target suppressing invasion and migration of human cervical cancer cells through modulation of its targeting genes ZEB1 and CRKL. Taken together, our data indicate that miR-429 plays a pivotal role in cervical cancer progression, which is a potential therapeutic target for patients.

An R, Wang Y, Voeller D, et al.
CRKL mediates EML4-ALK signaling and is a potential therapeutic target for ALK-rearranged lung adenocarcinoma.
Oncotarget. 2016; 7(20):29199-210 [PubMed] Free Access to Full Article Related Publications
Anaplastic lymphoma kinase (ALK) gene rearrangements are oncogenic drivers in a small subset of patients with non-small-cell lung cancer (NSCLC). The ALK inhibitors are highly effective in NSCLC patients harboring ALK rearrangements; however, most patients acquire resistance to the therapy following an initial response. Mechanisms of acquired resistance are complex. We used LC-MS/MS-based phosphotyrosine-peptide profiling in the EML4-ALK rearranged H3122 and H2228 cells treated with ALK inhibitors, to identify downstream effectors of ALK. We then used Western blot, siRNA experiments, cell proliferation, viability and migration assays to validate our findings. We identified CRKL as a novel downstream effector of ALK signaling. We demonstrated that CRKL tyrosine phosphorylation was repressed by pharmacological inhibition or small interfering RNA (siRNA) knockdown of ALK in the ALK-rearranged cells. More importantly, CRKL knockdown attenuated their cell proliferation, viability, and migration, but it had no effect on ALK phosphorylation and expression in these cells. Furthermore, CRKL tyrosine phosphorylation was inhibited by dasatinib (an inhibitor of ABL and SRC kinases), which in combination with the ALK inhibitor crizotinib displayed a synergistic inhibitory effect in vitro. In conclusion, our study suggests that CRKL is a key downstream effector of ALK, and combined inhibition of ALK and CRKL may represent an effective strategy for treating ALK-rearranged NSCLC patients.

Wang F, Jiang C, Sun Q, et al.
Downregulation of miR‑429 and inhibition of cell migration and invasion in nasopharyngeal carcinoma.
Mol Med Rep. 2016; 13(4):3236-42 [PubMed] Related Publications
Viral, dietary and genetic factors have been implicated in nasopharyngeal carcinoma (NPC), however, the molecular mechanism underlying its pathogenesis remains to be fully elucidated. MicroRNAs (miRNAs) have been reported to be important in NPC tumorigenesis, with a previous miRNA microarray study showing the downregulation of miRNA (miR)‑429 in NPC cells. However, the possible mechanisms of action of miR‑429 have not been examined. In the present study, the expression profiles of miR‑429 were detected using reverse transcription‑quantitative polymerase chain reaction analysis in CNE‑1 and CNE‑2 cells, which are two generally used NPC cells with different degrees of differentiation. Subsequently, cell proliferation, invasion and migration were analyzed in miR‑429‑overexpressing CNE‑2 cells, and the modulatory function of miR‑429 was also investigated using two target genes, zinc finger E‑Box‑binding homeobox 1 (ZEB1) and CRK‑like (CRKL), by transfection with miR‑429 mimic or anti‑miR‑429. Significant changes in the expression of miR‑429 were detected, particularly in low‑differentiated CNE‑2 cells, with higher levels of epidemicity and malignancy. Additional results revealed that miR‑429 inhibited the invasion and migration of the CNE‑2 cells, whereas no significant effect on cell growth was observed. In addition, the mRNA and protein expression levels of the two target genes, ZEB1 and CRKL, were negatively regulated by miR‑429, demonstrated through gain‑of‑function and loss‑of‑function investigations, indicating that these two functional downstream targets may be involved in the inhibitory effects of miR‑429 on NPC migration and invasion. miR‑429 may act as a negative regulatory factor of NPC tumorigenesis, involving the functions of its downstream targets, ZEB1 and CRKL. The results suggested miR‑429 as a potential candidate for miRNA‑based prognosis or therapy against NPC.

Ghosh A, Ghosh A, Datta S, et al.
Hepatic miR-126 is a potential plasma biomarker for detection of hepatitis B virus infected hepatocellular carcinoma.
Int J Cancer. 2016; 138(11):2732-44 [PubMed] Related Publications
Controversies about the origin of circulating miRNAs have encouraged us to identify organ specific circulating miRNAs as disease biomarkers. To identify liver-specific miRNAs for hepatocellular carcinoma (HCC), global expression profiling of miRNAs in liver tissue of HBV-HCC and HBV-control with no or mild fibrosis was evaluated. A total of 40 differentially expressed miRNAs were identified in HCC. Among ten highly altered miRNAs, six miRNAs were successfully validated in tissues, whereas only two miRNAs, miR-126 and miR-142-3p showed increased expression in plasma of HBV-HCC compared to HBV-non-HCC patients. Subsequently, ROC curve analysis revealed that neither miR-126 nor miR-142-3p performed better than AFP in discriminating HCC from non-HCC while combination of each with AFP showed significantly higher efficiency rather than AFP alone (AUC: 0.922, 0.908 vs. 0.88; sensitivity: 0.84, 0.86 vs. 0.82 and specificity: 0.92, 0.94 vs. 0.86 respectively). Interestingly, triple combination of markers (miR-126 + miR-142-3p + AFP) showed no additive effect on efficiency (AUC: 0.925) over the dual combination. Again, the expression of only miR-126 was noticed significantly higher in HBV-HCC patients with low-AFP [<250 ng/ml] compared to either non-HCC or liver cirrhosis (AUC: 0.77, 0.64, respectively). Furthermore, no alteration in expression of mir-126 in HCV-HCC or non-viral-HCC revealed that miR-126 + AFP might be specific to HBV-HCC. To understand the physiological role of these two miRNAs in hepato-carcinogenesis, target genes related to cancer pathways (APAF1, APC2, CDKN2A, IRS1, CRKL, LIFR, EGR2) were verified. Thus, combination of circulating miR-126 + AFP is a promising noninvasive diagnostic biomarker for HBV-HCC and may be useful in the management of HCC patients.

Ungewiss C, Rizvi ZH, Roybal JD, et al.
The microRNA-200/Zeb1 axis regulates ECM-dependent β1-integrin/FAK signaling, cancer cell invasion and metastasis through CRKL.
Sci Rep. 2016; 6:18652 [PubMed] Free Access to Full Article Related Publications
Tumor cell metastasis is a complex process that has been mechanistically linked to the epithelial-mesenchymal transition (EMT). The double-negative feedback loop between the microRNA-200 family and the Zeb1 transcriptional repressor is a master EMT regulator, but there is incomplete understanding of how miR-200 suppresses invasion. Our recent efforts have focused on the tumor cell-matrix interactions essential to tumor cell activation. Herein we utilized both our Kras/p53 mutant mouse model and human lung cancer cell lines to demonstrate that upon miR-200 loss integrin β1-collagen I interactions drive 3D in vitro migration/invasion and in vivo metastases. Zeb1-dependent EMT enhances tumor cell responsiveness to the ECM composition and activates FAK/Src pathway signaling by de-repression of the direct miR-200 target, CRKL. We demonstrate that CRKL serves as an adaptor molecule to facilitate focal adhesion formation, mediates outside-in signaling through Itgβ1 to drive cell invasion, and inside-out signaling that maintains tumor cell-matrix contacts required for cell invasion. Importantly, CRKL levels in pan-cancer TCGA analyses were predictive of survival and CRKL knockdown suppressed experimental metastases in vivo without affecting primary tumor growth. Our findings highlight the critical ECM-tumor cell interactions regulated by miR-200/Zeb1-dependent EMT that activate intracellular signaling pathways responsible for tumor cell invasion and metastasis.

Luo LY, Hahn WC
Oncogenic Signaling Adaptor Proteins.
J Genet Genomics. 2015; 42(10):521-529 [PubMed] Free Access to Full Article Related Publications
Signal transduction pathways activated by receptor tyrosine kinases (RTK) play a critical role in many aspects of cell function. Adaptor proteins serve an important scaffolding function that facilitates key signaling transduction events downstream of RTKs. Recent work integrating both structural and functional genomic approaches has identified several adaptor proteins as new oncogenes. In this review, we focus on the discovery, structure and function, and therapeutic implication of three of these adaptor oncogenes, CRKL, GAB2, and FRS2. Each of the three genes is recurrently amplified in lung adenocarcinoma or ovarian cancer, and is essential to cancer cell lines that harbor such amplification. Overexpression of each gene is able to transform immortalized human cell lines in in vitro or in vivo models. These observations identify adaptor protein as a distinct class of oncogenes and potential therapeutic targets.

Murali R, Chandramohan R, Möller I, et al.
Targeted massively parallel sequencing of angiosarcomas reveals frequent activation of the mitogen activated protein kinase pathway.
Oncotarget. 2015; 6(34):36041-52 [PubMed] Free Access to Full Article Related Publications
Angiosarcomas are rare malignant mesenchymal tumors of endothelial differentiation. The clinical behavior is usually aggressive and the prognosis for patients with advanced disease is poor with no effective therapies. The genetic bases of these tumors have been partially revealed in recent studies reporting genetic alterations such as amplifications of MYC (primarily in radiation-associated angiosarcomas), inactivating mutations in PTPRB and R707Q hotspot mutations of PLCG1. Here, we performed a comprehensive genomic analysis of 34 angiosarcomas using a clinically-approved, hybridization-based targeted next-generation sequencing assay for 341 well-established oncogenes and tumor suppressor genes. Over half of the angiosarcomas (n = 18, 53%) harbored genetic alterations affecting the MAPK pathway, involving mutations in KRAS, HRAS, NRAS, BRAF, MAPK1 and NF1, or amplifications in MAPK1/CRKL, CRAF or BRAF. The most frequently detected genetic aberrations were mutations in TP53 in 12 tumors(35%) and losses of CDKN2A in9 tumors (26%). MYC amplifications were generally mutually exclusive of TP53 alterations and CDKN2A loss and were identified in 8 tumors (24%), most of which (n = 7, 88%) arose post-irradiation. Previously reported mutations in PTPRB (n = 10, 29%) and one (3%) PLCG1 R707Q mutation were also identified. Our results demonstrate that angiosarcomas are a genetically heterogeneous group of tumors, harboring a wide range of genetic alterations. The high frequency of genetic events affecting the MAPK pathway suggests that targeted therapies inhibiting MAPK signaling may be promising therapeutic avenues in patients with advanced angiosarcomas.

Tamura M, Sasaki Y, Kobashi K, et al.
CRKL oncogene is downregulated by p53 through miR-200s.
Cancer Sci. 2015; 106(8):1033-40 [PubMed] Free Access to Full Article Related Publications
Tumor suppressive miRNAs that target oncogenes are frequently downregulated in cancers, and this downregulation leads to oncogene pathway activation. Thus, tumor suppressive miRNAs and their target oncogenes have been proposed as useful targets in cancer treatment. miR-200 family downregulation has been reported in cancer progression and metastasis. The miR-200 family consists of two gene clusters, miR-200b/200a/429 and miR-200c/141, which are located on human chromosomes 1 and 12, respectively. Here, we identified that p53 response elements are located around both clusters of the miR-200 family and confirmed that miR-200s are transcriptional targets of the p53 family. In silico analyses of miRNA targets established the CRKL oncogene as a potential target for miR-200b/200c/429. Moreover, miR-200b/200c/429 inhibited CRKL mRNA and protein expression by directly targeting its 3'-UTR region. Importantly, endogenous CRKL expression was decreased in cancer cells through the introduction of p53 family and endogenous p53 activation. Moreover, the downregulation of CRKL by siRNA inhibited cancer cell growth. The Oncomine database demonstrates that CRKL is overexpressed in a subset of cancer types. Furthermore, CRKL is significantly overexpressed in primary breast cancer tissues harboring mutant TP53. Our results demonstrate that the p53 target miR-200b/200c/429 miRNAs are negative regulators of the CRKL oncogene.

Han G, Wu D, Yang Y, et al.
CrkL meditates CCL20/CCR6-induced EMT in gastric cancer.
Cytokine. 2015; 76(2):163-169 [PubMed] Related Publications
BACKGROUND: In recent years, Crk-like adapter protein (CrkL) has been identified as a key regulator in the epithelial-to-mesenchymal transition (EMT). However, the molecular mechanisms underlying the CC chemokine receptor 6 (CCR6) and chemokine (C-C motif) ligand 20 (CCL20)-induced EMT in gastric cancer are still unclear.
METHODS: We conducted the immunohistochemistry and immunoblotting to detect the expression of CCR6 and CrkL in 90 cases of gastric cancer tissues and five kinds of cell lines. And then, gastric cancer cells were subjected to small interfering RNA (siRNA) treatment and in vitro assay.
RESULTS: Both CCR6 and CrkL were aberrantly expressed in gastric cancer specimens and closely correlated with differentiation of cell lines. The expression of CCR6 and CrkL was also significantly associated with metastasis, stage, and poor prognosis of gastric cancer. In addition, we validated CCL20 activated the expression of p-CrkL, p-Erk1/2, p-Akt, vimentin, N-cadherin and MMP2 in MGC803 cells in a dose-dependent manner. However, si-CrkL abrogated the CCL20-induced p-Erk1/2, vimentin, N-cadherin and MMP2 expression. Most importantly, the knockdown of CrkL decreased migration and invasion of MGC803 cells.
CONCLUSIONS: CrkL mediates CCL20/CCR6-induced EMT via Akt pathway, instead of Erk1/2 pathway in development of gastric cancer, which indicated CCL20/CCR6-CrkL-Erk1/2-EMT pathway may be targeted to antagonize the progression of gastric cancer.

Shagisultanova E, Gaponova AV, Gabbasov R, et al.
Preclinical and clinical studies of the NEDD9 scaffold protein in cancer and other diseases.
Gene. 2015; 567(1):1-11 [PubMed] Free Access to Full Article Related Publications
Cancer progression requires a significant reprogramming of cellular signaling to support the essential tumor-specific processes that include hyperproliferation, invasion (for solid tumors) and survival of metastatic colonies. NEDD9 (also known as CasL and HEF1) encodes a multi-domain scaffolding protein that assembles signaling complexes regulating multiple cellular processes relevant to cancer. These include responsiveness to signals emanating from the T and B cell receptors, integrins, chemokine receptors, and receptor tyrosine kinases, as well as cytoplasmic oncogenes such as BCR-ABL and FAK- and SRC-family kinases. Downstream, NEDD9 regulation of partners including CRKL, WAVE, PI3K/AKT, ERK, E-cadherin, Aurora-A (AURKA), HDAC6, and others allow NEDD9 to influence functions as pleiotropic as migration, invasion, survival, ciliary resorption, and mitosis. In this review, we summarize a growing body of preclinical and clinical data that indicate that while NEDD9 is itself non-oncogenic, changes in expression of NEDD9 (most commonly elevation of expression) are common features of tumors, and directly impact tumor aggressiveness, metastasis, and response to at least some targeted agents inhibiting NEDD9-interacting proteins. These data strongly support the relevance of further development of NEDD9 as a biomarker for therapeutic resistance. Finally, we briefly discuss emerging evidence supporting involvement of NEDD9 in additional pathological conditions, including stroke and polycystic kidney disease.

Aljedai A, Buckle AM, Hiwarkar P, Syed F
Potential role of Notch signalling in CD34+ chronic myeloid leukaemia cells: cross-talk between Notch and BCR-ABL.
PLoS One. 2015; 10(4):e0123016 [PubMed] Free Access to Full Article Related Publications
Notch signalling is critical for haemopoietic stem cell (HSC) self-renewal and survival. The role of Notch signalling has been reported recently in chronic myeloid leukaemia (CML) - a stem cell disease characterized by BCR-ABL tyrosine kinase activation. Therefore, we studied the relationship between BCR-ABL and Notch signalling and assessed the expression patterns of Notch and its downstream target Hes1 in CD34+ stem and progenitor cells from chronic-phase CML patients and bone marrow (BM) from normal subjects (NBM). We found significant upregulation (p<0.05) of Notch1, Notch2 and Hes1 on the most primitive CD34+Thy+ subset of CML CD34+ cells suggesting that active Notch signalling in CML primitive progenitors. In addition, Notch1 was also expressed in distinct lymphoid and myeloid progenitors within the CD34+ population of primary CML cells. To further delineate the possible role and interactions of Notch with BCR-ABL in CD34+ primary cells from chronic-phase CML, we used P-crkl detection as a surrogate assay of BCR-ABL tyrosine kinase activity. Our data revealed that Imatinib (IM) induced BCR-ABL inhibition results in significant (p<0.05) upregulation of Notch activity, assessed by Hes1 expression. Similarly, inhibition of Notch leads to hyperactivation of BCR-ABL. This antagonistic relationship between Notch and BCR-ABL signalling was confirmed in K562 and ALL-SIL cell lines. In K562, we further validated this antagonistic relationship by inhibiting histone deacetylase (HDAC) - an effector pathway of Hes1, using valproic acid (VPA) - a HDAC inhibitor. Finally, we also confirmed the potential antagonism between Notch and BCR/ABL in In Vivo, using publically available GSE-database, by analysing gene expression profile of paired samples from chronic-phase CML patients pre- and post-Imatinib therapy. Thus, we have demonstrated an antagonistic relationship between Notch and BCR-ABL in CML. A combined inhibition of Notch and BCR-ABL may therefore provide superior clinical response over tyrosine-kinase inhibitor monotherapy by targeting both quiescent leukaemic stem cells and differentiated leukaemic cells and hence must be explored.

Nizard P, Ezan F, Bonnier D, et al.
Integrative analysis of high-throughput RNAi screen data identifies the FER and CRKL tyrosine kinases as new regulators of the mitogenic ERK-dependent pathways in transformed cells.
BMC Genomics. 2014; 15:1169 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Cell proliferation is a hallmark of cancer and depends on complex signaling networks that are chiefly supported by protein kinase activities. Therapeutic strategies have been used to target specific kinases but new methods are required to identify combined targets and improve treatment. Here, we propose a small interfering RNA genetic screen and an integrative approach to identify kinase networks involved in the proliferation of cancer cells.
RESULTS: The functional siRNA screen of 714 kinases in HeLa cells identified 91 kinases implicated in the regulation of cell growth, most of them never being reported in previous whole-genome siRNA screens. Based on gene ontology annotations, we have further discriminated between two classes of kinases that, when suppressed, result in alterations of the mitotic index and provoke cell-cycle arrest. Extinguished kinases that lead to a low mitotic index mostly include kinases implicated in cytosolic signaling. In contrast, extinguished kinases that result in a high mitotic index mostly include kinases implicated in cell division. By mapping hit kinases in the PhosphPOINT phosphoprotein database, we generated scale-free networks consisting of 449 and 661 protein-protein interactions for kinases from low MI and high MI groups, respectively. Further analyses of the kinase interactomes revealed specific modules such as FER- and CRKL-containing modules that connect three members of the epidermal growth factor receptor (EGFR) family, suggesting a tight control of the mitogenic EGF-dependent pathway. Based on experimental studies, we confirm the involvement of these two kinases in the regulation of tumor cell growth.
CONCLUSION: Based on a combined approach of large kinome-wide siRNA screens and ontology annotations, our study identifies for the first time two kinase groups differentially implicated in the control of cell proliferation. We further demonstrate that integrative analysis of the kinase interactome provides key information which can be used to facilitate or optimize target design for new therapeutic strategies. The complete list of protein-protein interactions from the two functional kinase groups will provide a useful database for future investigations.

Lian X, Jiao Y, Yang Y, et al.
CrkL regulates SDF-1-induced breast cancer biology through balancing Erk1/2 and PI3K/Akt pathways.
Med Oncol. 2015; 32(1):411 [PubMed] Related Publications
The adapter protein CrkL is required for regulating the malignant potential of human cancers. However, the regulatory mechanisms of CrkL on the stromal cell-derived factor 1 (SDF-1)/CXCR4 signaling pathways in breast cancer are not well characterized. Here, CXCR4 and CrkL proteins were tested in breast cancer cell lines and 60 primary breast cancer tissues. In vitro, the roles of CrkL in SDF-1-induced MDA-MB-231 cell cycle, invasion and migration were investigated. In the present study, CXCR4 and CrkL were highly expressed in MCF-7, MDA-MB-231, MDA-MB-231HM MDA-MB-468 and tumor tissues (80 and 60 %, respectively) and closely correlated with lymph node metastasis. In vitro studies revealed that SDF-1 induced the activation of CrkL, Erk1/2, Akt and matrix metallopeptidase 9 (MMP9) in MDA-MB-231 cells. The si-CrkL treatment significantly down-regulated the phosphorylated Erk1/2 (p-Erk1/2) and MMP9, but up-regulated p-Akt, compared with control. Importantly, wound-healing and transwell invasion assays showed that si-CrkL significantly impaired the wound closure and inhibited the SDF-1-induced invasion; similarly, flow cytometry showed that si-CrkL affected cell cycle. In conclusion, these results suggest that CrkL plays a regulatory role in the SDF-1-induced Erk1/2 and PI3K/Akt pathways and further managed the invasion and migration of breast cancer cells. Thus, CrkL may be recommended as an interesting therapeutic target for breast cancer.

Ye ZB, Ma G, Zhao YH, et al.
miR-429 inhibits migration and invasion of breast cancer cells in vitro.
Int J Oncol. 2015; 46(2):531-8 [PubMed] Free Access to Full Article Related Publications
Accumulating evidence indicates that microRNAs (miRNAs) are involved in regulating cancer invasion and metastasis, and an increasing number of research demonstrates that miRNAs can promote or inhibit cell motility depending on genetic background of different cancers and the microenvironment. In the present study, we established an in vivo bone metastasis model of breast cancer by injecting MDA-MB-231 cells into the left ventricle of nude mice, and then screened the differentially expressed miRNAs between parental and bone-metastatic MDA-MB-231 cells using miRNA array. The results revealed that decreased expression of miR-429 was probably involved in negatively regulating bone metastasis of breast cancer cells. On the other hand, overexpression of miR-429 in MDA-MB-231 cells remarkably suppressed invasion in vitro. We identified ZEB1 and CRKL as potential targets of miR-429 by analyzing combined results from in silico search and global expression array of the same RNA samples. Immunoblot assay confirmed that miR-429 reduced their expression at protein level. Taken together, our results offer an opportunity for further understanding of the recondite mechanisms underlying the bone metastasis of breast cancer.

Fu L, Dong Q, Xie C, et al.
CRKL protein overexpression enhances cell proliferation and invasion in pancreatic cancer.
Tumour Biol. 2015; 36(2):1015-22 [PubMed] Related Publications
CRKL is an adapter protein which is overexpressed in many malignant tumors and plays crucial roles in tumor progression. However, expression pattern and biological roles of CRKL in pancreatic cancer have not been examined. In the present study, we found that CRKL expression in pancreatic cancer specimens was higher than that in normal pancreatic tissues. Colony formation assay and Matrigel invasion assay showed that the overexpression of CRKL in Bxpc3 and Capan2 cell lines with low endogenous expression increased cell proliferation and invasion. Flow cytometry showed that CRKL promoted cell proliferation by facilitating cell cycle. Further analysis of cell cycle- and invasion-related molecules showed that CRKL upregulated cyclin D1, cyclin A, matrix metalloproteinase 2 (MMP2) expression, and phosphorylated extracellular signal (ERK)-regulated kinase. In conclusion, our study demonstrated that CRKL was overexpressed in human pancreatic cancers and contributed to pancreatic cancer cell proliferation and invasion through ERK signaling.

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