SEARCH

SEARCH BY CITATION

Keywords:

  • Src oncogene;
  • tyrosine kinases;
  • solid and hematologic malignancies;
  • Src inhibitors

Abstract

  1. Top of page
  2. Abstract
  3. Signaling Regulation by Protein Tyrosine Kinases
  4. Structure and Function of the c-Src Family of Kinases
  5. c-Src Regulation
  6. SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases
  7. Involvement of c-Src in Human Cancers
  8. c-Src Kinase Inhibitors
  9. CONCLUSION
  10. Acknowledgements
  11. REFERENCES

c-Src was the first protooncogene described and was among the first molecules in which tyrosine kinase activity was documented. c-Src has been defined as a common modular structure that participates in much of the crosstalk between the cytoplasmic protein tyrosine kinases and tyrosine kinase receptors. Understanding the structure and function of this important class of protein kinases and elucidating the molecular signaling events mediated by c-Src are important not only for identifying the critical pathways but also for designing new strategies to block or inhibit the action of these kinases. Despite the large amount of information available on c-Src, its precise functions in cancer remain to be elucidated. Recently, there has been renewed interest in c-Src as a molecular target for cancer therapy, and multiple c-Src inhibitors are entering clinical trials. In this review, the authors describe the function and expression of c-Src in human malignancies and the novel c-Src inhibitors and their potential applications for cancer treatment. Cancer 2006. © 2006 American Cancer Society.

In 1911, a transmissible virus was identified that appeared to cause tumor growth in chickens, and some investigators suggested that such an agent may induce solid tumors.1 More than 40 years later, the Rous sarcoma virus was discovered; in fact, it produced infected tumor cells.2 The viral Src gene (v-Src) finally was identified in 19703; and, since then, a great deal has been learned about it, including its mutations and how they affect its transformation potential.

The results from numerous studies indicate that introducing c-Src into cells was sufficient to elicit the properties of transformed cells, including growth factor-independent and anchorage-independent growth, motility, and invasiveness. Better understanding of the functions of c-Src has led to the development of c-Src and Src family of kinases (SFK) inhibitors with significant potential as anticancer drugs or osteoclast-targeted drugs for treating osteoporosis or osteolytic metastases.

Signaling Regulation by Protein Tyrosine Kinases

  1. Top of page
  2. Abstract
  3. Signaling Regulation by Protein Tyrosine Kinases
  4. Structure and Function of the c-Src Family of Kinases
  5. c-Src Regulation
  6. SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases
  7. Involvement of c-Src in Human Cancers
  8. c-Src Kinase Inhibitors
  9. CONCLUSION
  10. Acknowledgements
  11. REFERENCES

The principal function of the diverse protein tyrosine kinase (PTK) family of proteins is the regulation of cell-to-cell signals. PTKs are important regulators of intracellular signal-transduction pathways that mediate cellular development and multicellular communications. PTKs catalyze the transfer of phosphate in adenosine 5′ triphosphate (ATP) to tyrosine residues in cellular substrates. This activity is tightly regulated; signaling perturbations caused by mutations and other genetic alterations result in deregulated kinase activity and may lead to malignant transformation. c-Src is a nonreceptor PTK and member of a large family of structurally related kinases, many of which are expressed predominantly in highly differentiated cell types.4 Studies investigating the location of c-Src have provide considerable information regarding its functions. At the plasma membrane, it is believed that c-Src is involved in signal-transduction events that regulate cell growth and proliferation through activated growth factor receptors.5

Structure and Function of the c-Src Family of Kinases

  1. Top of page
  2. Abstract
  3. Signaling Regulation by Protein Tyrosine Kinases
  4. Structure and Function of the c-Src Family of Kinases
  5. c-Src Regulation
  6. SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases
  7. Involvement of c-Src in Human Cancers
  8. c-Src Kinase Inhibitors
  9. CONCLUSION
  10. Acknowledgements
  11. REFERENCES

The SFKs comprise at least 9 proteins (all approximately 60 kD in molecular weight)—c-Src, Fyn, c-Yes, Fgr, Lyn, Hck, Lck, Blk, and Yrk—with similar structures,4 and their domain structures have considerable homology.6 Some of these proteins are expressed ubiquitously, whereas others are expressed only in specific cell types.6 Among all family members, c-Src is implicated most often in cancer.

c-Src has 7 domains: an N-terminal membrane-association domain (the SH4 domain), a unique domain, the SH3 and SH2 domains, an SH2-kinase linker domain, a catalytic domain (the SH1 domain), and a negative regulatory domain (Fig. 1).6 The functions of these domains are as follows.

thumbnail image

Figure 1. This is an illustration of the linear structure of c-Src. Numbers above the linear molecule denote amino acid residues in sequence from the N terminus (N) to the C terminus (C). c-Src has 7 domains (SH1-SH7), which are depicted in different colors and are described in the text. Adapted with permission from Haskell et al.6

Download figure to PowerPoint

1) The N-terminal or SH4 domain is tethered through myristylation to membranes by the action of an N terminus covalently linked to a myristate moiety. 2) The function of the unique domain is not well understood; however, this area contains many sites of serine-threoninie that are mediated by cyclin-dependent kinase 2 (cdc2). 3) The SH3 domain comprises approximately 60 amino acids and is found in many signaling molecules. This domain is noncatalytic and mediates protein-protein interactions by binding to proline-rich sequences. 4) The SH2 domain comprises approximately 100 amino acids that bind phosphotyrosine (pTyr) residues to specific proteins. The affinity of this domain depends on the amino acid sequence from the C terminus to the pTyr motif.7 Inhibitors of specific SH2 domains are important in preventing and treating several diseases, including cancer, cardiovascular disease, and osteoporosis. 5) The SH2-kinase linker domain is a loop that functions as a pseudo-SH3-binding site. This domain contains Tyr416, which is activated by autophosphorylation and is required for optimal activity.8 6) The catalytic or SH1 domain is well conserved in all tyrosine kinases. This domain, which is responsible for the catalytic activity of the molecule, contains many subdomains that are closely related to the serine-threonine kinases, including the ATP-binding pocket. 7) The negative regulatory domain harbors a tyrosine residue (Tyr530) that becomes phosphorylated by another tyrosine kinase, C-terminal Src kinase (CSK). Phosphorylated Tyr527/Tyr530 can bind to its own SH2 domain in such a way that it inhibits kinase activity without physically blocking the catalytic site.

c-Src Regulation

  1. Top of page
  2. Abstract
  3. Signaling Regulation by Protein Tyrosine Kinases
  4. Structure and Function of the c-Src Family of Kinases
  5. c-Src Regulation
  6. SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases
  7. Involvement of c-Src in Human Cancers
  8. c-Src Kinase Inhibitors
  9. CONCLUSION
  10. Acknowledgements
  11. REFERENCES

There is increasing evidence that c-Src plays an important role in regulating mitotic events. Changes in activation and localization of c-Src during mitosis have been described, suggesting a role in G2/M transition.9 In addition, activated Src may abrogate the Myc requirement for G0/G1 transition but not for G1/S transition.10 Multiple processes can alter c-Src activity and, in general, all SFKs. Briefly, 3 events are capable of activating or inactivating c-Src: phosphorylation of specific sites on the c-Src molecule, interaction with binding proteins and other kinases that phosphorylate Tyr527/530, and mutations. Each of these potentially may activate Src, or they may act in concert with others.

Regulation of c-Src by phosphorylation

One important mechanism of c-Src regulation is the control of its phosphorylation status. c-Src normally is maintained in an inactive (close) conformation (Fig. 2) with the SH2 domain engaged with the Tyr530, the SH3 domain engaged with the SH2-kinase linker, and Tyr419 dephosphorylated. Two major phosphorylation sites are present in human c-Src: Tyr(527/530) and Tyr(416/419). Phosphorylating the carboxyl-terminal residue Tyr(527/530) is critical in the inactivation of c-Src.11

thumbnail image

Figure 2. This is an illustration of c-Src regulation. Src normally is maintained in an inactive conformation (center) with the SH2 domain engaged with the tyrosine (Tyr) residue Tyr530, the SH3 domain engaged with the SH2-kinase linker, and Tyr419 dephosphorylated. Src activity is regulated by a variety of mechanisms, some of which are illustrated here. Dephosphorylation of Tyr530 by protein tyrosine phosphatase (PTP) action (right) disrupts intramolecular interaction between the SH2 domain and Tyr530; then, Tyr419 can become autophosphorylated, resulting in Src activation. Phosphorylation of Tyr530 by c-Src kinase (CSK) allows this interaction to reform, resulting in Src inactivation. Binding of Src to tyrosine phosphorylated growth factor receptors through its SH2 domain (left) displaces Tyr530, allowing Tyr419 phosphorylation and Src activation. Dissociation of Src from the growth factor receptor allows the intramolecular interactions to reform, inactivating Src. ATP indicates 5′-adenosine triphosphate; Y, tyrosine; P, phosphorylated; Mg, magnesium. Adapted by permission from Macmillan Publishers Ltd: Oncogene, Bjorge et al., 2000.5

Download figure to PowerPoint

A second phosphorylation site, Tyr(416/419), is present within the activation loop. When this site becomes phosphorylated, it is displaced from the substrate-binding pocket, giving the kinase access to substrates and, thus, becoming a positive regulator of c-Src.5 When it is dephosphorylated, Tyr(416/419) is a negative regulator.5, 12

Two other identified tyrosine kinases, c-Src kinase (CSK) and CSK homologous kinase (CHK), are capable of phosphorylating Tyr(527/530), also resulting in c-Src inactivation. The overexpression of CSK suppressed colon cancer metastasis in an animal model,13 suggesting that CSK has tumor-suppressor activity and that its reduced expression may result in the activation of c-Src, facilitating malignant cell behavior promoted by c-Src.

Despite structural similarities, CHK and CSK show striking differences. Whereas CSK is expressed ubiquitously, CHK expression in normal tissue is limited to brain and hematopoietic cells. CHK may play a specific role as a novel negative growth regulator in human cancer. For example, in breast cancer, studies have shown that 1) unlike CSK, CHK is expressed specifically in primary breast cancer specimens but not in normal breast tissues; 2) unlike CSK, which cannot associate with ErbB-2, CHK binds directly to phospho-Tyr1248 of ErbB-2/neu on heregulin stimulation and inhibits c-Src kinase activity; and 3) overexpression of CHK in breast cancer cells markedly inhibits cell growth, transformation, and invasion induced by heregulin and delays cell entry into mitosis.14 Thus, CHK not only inhibits breast cancer cell proliferation and transformation but may inhibit tumor cell invasion, suggesting a role in cell motility and metastasis in breast cancer.

Regulation of c-Src activity through interaction with binding proteins

The association of c-Src with various proteins has 2 important consequences: activation of c-Src and relocation of c-Src to sites of action.5 c-Src is activated after binding with platelet-derived growth factor receptor (PDGF-R) and focal adhesion kinases (FAKs) through SH2. These molecules compete for binding sites, disturb intramolecular interactions, and activate c-Src.15 The association of c-Src with binding proteins permits the relocation of c-Src within the cell close to substrates. FAKs are associated with c-Src in focal adhesions, which results in the phosphorylation of FAK followed by downstream activation of the Ras, mitogen-activated kinase (MEK), and extracellular signal-related (ERK) pathways.16

Regulation of c-Src activity through mutations

Disruption of the c-Src regulatory intramolecular site by directed mutagenesis leads to deregulation of the kinase activity. These intramolecular interactions may limit accessibility of the kinase domain and the binding surfaces of the SH3 and SH2 domains, further limiting the potential for these proteins to participate in signal transduction.17 Mutation of c-Src Y527 to phenylalanine results in an active, transformation-competent protein.18 However, genomic mutations are not the predominant mechanism of SFK activation in human cancers.

SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases

  1. Top of page
  2. Abstract
  3. Signaling Regulation by Protein Tyrosine Kinases
  4. Structure and Function of the c-Src Family of Kinases
  5. c-Src Regulation
  6. SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases
  7. Involvement of c-Src in Human Cancers
  8. c-Src Kinase Inhibitors
  9. CONCLUSION
  10. Acknowledgements
  11. REFERENCES

Recent studies have established that SFKs are involved in signaling from many receptor tyrosine kinases (RTKs), including PDGF-R, epidermal growth factor receptor (EGF-R), fibroblast growth factor receptor (FGF-R), insulin-like growth factor-1 (IGF-1) receptor, hepatocyte growth factor/scatter factor receptor, colony-stimulating factor 1 receptor, stem cell factor receptor, muscle-specific kinase, and others.19 SFKs can promote signaling from growth factor receptors in a number of ways, such as initiating the signaling pathways required for DNA synthesis, modulating RTKs, and controlling receptor turnover, actin cytoskeleton rearrangements, and FAKs.

Initiation of signal pathways required for DNA synthesis

SFKs appear to be required for growth factor-stimulated DNA synthesis, particularly those growth factors with receptors that are tyrosine kinases. Although an absolute requirement for SFKs may depend on the cell type and cellular context, available data suggest that SFKs promote mitogenesis from a variety of RTKs. Whether SFKs are required for mitogenesis in response to PDGF is a subject of debate. Considerable evidence has demonstrated that SFKs are required for mitogenesis in response to PDGF. This hypothesis is favored by study of SFK-neutralizing antibodies20 and the SFK inhibitor SU6656,21 both of which inhibit PDGF-stimulated DNA synthesis. However, other investigators suggest that SFKs are not required for PDGF-induced mitogenesis.22 Preclinical data have suggested that SFKs are indispensable for EGF-R-induced mitogenesis in fibroblasts.23

Amplification and overexpression of the neu (c-erbB2) protooncogene has been implicated in the pathogenesis of 20% to 30% of human breast cancers.24 In breast cancer, the interaction between the SFKs and the EGF-R is crucial, because both kinases are overexpressed in these tumors. Phosphorylation of EGF-R by c-Src is critical for mitogenic signaling initiated by the EGF-R itself and as by several G protein-coupled receptors (GPCRs). Thus, c-Src appears to have pleiotropic effects on cancer cells by modulating the action of multiple growth-promoting receptors.25 Certain GPCRs can trigger phosphorylation of EGF-R and activation of its downstream effectors Shc and microtubule-associated protein kinase, and this activation is dependent on c-Src kinase activity.26 These observations suggested that activation of the SFKs during mammary tumorigenesis may occur through direct interaction with activated Neu.

SFKs modulate RTKs

The relation between the SFKs and RTKs is complex, with a bidirectional interaction. SFKs modulate the activity and signaling of RTKs, particularly EGF-R, PDGF-R, and IGF-R.4 This regulation is particularly important for the function of EGF-R.6 Furthermore, RTKs undergo endocytosis after their engagement with their ligands. Cells that overexpressed c-Src had an increased rate of EGF-R internalization.27 That observation suggests that c-Src participates in receptor internalization, which, in turn, may alter some aspects of EGF-R signaling related to mitogenesis and tumorigenesis.

Regulation of cytoskeletal events

Growth factors induce changes in the actin cytoskeleton that affect cell migration. These changes in actin stress fibers occur with the involvement of the Rho family and GTPases.28 The integrity of the actin cytoskeleton is needed for progression into S phase of the cell cycle. c-Src regulates the EGF-dependent reorganization of the actin cytoskeleton through phosphorylation of p190.29

In epithelial cancer models, c-Src activation promotes a more migratory and invasive phenotype, inducing morphologic and biochemical changes more commonly associated with fibroblast and cells of mesenchymal origin (i.e., epidermal to mesenchymal-like transition).30 This phenotypic change is characterized by the breakdown of cadherin-mediated cell-cell junctions, which increases cell invasiveness. In endothelial and tumor cells, c-Src activation increases the migratory potential.31 Two main types of junction mediate adhesion in epithelial cells: adherens junctions and focal adhesions. Adherens junctions facilitate cell-cell adhesion through intercellular proteins, including E-catherin. Activated c-Src promotes disruption of adherens junction. Focal adhesions include more than 50 different proteins, including talin, vinculin, α-actinin, and paxillin. These assemble into supramolecular structures that allow cell adherence to extracellular matrix proteins.32 This process is regulated by crosstalk between integrins and other cell-surface molecules, such as cadherins and selectins. c-Src contributes to the disassembly of these focal adhesions when cells move away from the extracellular matrix and disrupts adherens junctions by inducing phosphorylation and ubiquitination of E-cadherin.33 Therefore, the interaction of c-Src with these molecular interactions may disrupt cellular adhesions, resulting in cell detachment from the extracellular matrix.

FAK as a substrate for c-Src

FAK was among the first c-Src substrates to be identified, and it is believed that FAK regulates growth factor-mediated and integrin-mediated cellular motility, adhesion, and invasion as well as cell proliferation and survival.34 In multiple tumors, overexpression and activation of both c-Src and FAK have been demonstrated leading to increased invasive and metastatic potential.35

Involvement of c-Src in Human Cancers

  1. Top of page
  2. Abstract
  3. Signaling Regulation by Protein Tyrosine Kinases
  4. Structure and Function of the c-Src Family of Kinases
  5. c-Src Regulation
  6. SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases
  7. Involvement of c-Src in Human Cancers
  8. c-Src Kinase Inhibitors
  9. CONCLUSION
  10. Acknowledgements
  11. REFERENCES

Several studies suggest that c-Src plays an important role in the genesis and progression of multiple human cancer types, including carcinomas of the breast, gastrointestinal tract, lung, and ovary, and myeloproliferative disorders.36, 17 The promotion of events, such as alterations in adhesion, angiogenesis, tumor cell invasiveness, tumor growth, and apoptosis, plays a significant role in the development of the metastatic phenotype.36 Studies using human neoplastic cell lines and in vivo murine models have documented the complex network of c-Src-interacting proteins that affect numerous signal-transduction pathways (Fig. 3). However, high cytoplasmic levels of c-Src protein in tumor cells may not accurately reflect the protein kinase activity.37 Overexpression of c-Src is associated with the development of some epithelial cancers,14 but it may have different effects on the development of other types of cancers.

thumbnail image

Figure 3. This is an illustration of the role of c-Src expression and activation in human cancer. EGFR indicates epidermal growth factor receptor; PDGFR, platelet-derived growth factor receptor; FGFR, fibroblast growth factor receptor, ECM, extracellular matrix, PI3K, phosphoinositide-3 kinase; Stat3, signal transducer and activator of transcription type 3; MMPs, matrix metalloproteinases; VEGF, vascular endothelial growth factor. Adapted by permission from Macmillan Publishers Ltd: Oncogene, Irby and Yeatman.36

Download figure to PowerPoint

SFKs in colon cancer

The activation and functions of SFKs have been studied most extensively in colon cancer. Increased c-Src kinase activity has been identified in the adjacent normal mucosa in colon cancer38 and in colonic polyps, particularly in malignant polyps and in benign polyps with villous structure and severe dysplasia.39 The degree of c-Src activation may be linked to the site of metastatic growth, suggesting that the c-Src activity either influences the site of ectopic growth or is influenced itself by the ectopic environment.40

Irby et al.41 identified a truncating mutation in c-Src at amino acid 531 in 12% of patients with advanced colon cancer and demonstrated that the mutation was activating, transforming, tumorigenic, and metastasis-promoting. However, this mutation was not detected in other studies.42, 43 Other investigators suggested that c-Src activity was an independent indicator of prognosis at all stages of colonic carcinoma.44 It also has been observed that c-Yes is activated in colon metastases, and activation is associated with a worse clinical outcome.45 Thus, in colon cancer, c-Src activation may contribute to both early stages and later stages of tumor progression.

SFKs in breast cancer

Several studies have suggested a role for SFKs in the progression of breast cancer. In 1 study, it was observed that c-Src kinase activity was 4-fold to 20-fold greater in human mammary carcinomas than in normal tissues.46 In addition, tyrosine kinase activity was increased in all 72 patients with breast cancer studied, and 70% of that increase was attributed to c-Src kinases.46

Muthuswamy et al.47 observed that mice with overexpression of the neu oncogene also developed mammary tumors that had 6-fold to 8-fold greater c-Src kinase activity than adjacent normal tissue. In other preclinical models, elevated levels or activities of c-Src and HER-1 also occurred in a subset of later stage breast cancers, and the investigators suggested that interactions between these 2 molecules may contribute to a more aggressive clinical course.48 In addition, Src may play a critical role in ErbB2-mediated breast cancer invasion and metastasis. Tan et al.49 observed that ErbB2 activation up-regulates Src protein levels and kinase activity. Increased Src protein levels resulted from 2 mechanisms: 1) ErbB2 promoted Src protein synthesis by activating the Akt/mammalian target of rapamycin/4E-BP1 protein-translation pathway, and 2) ErbB2 stabilized Src by inhibiting calpain-mediated Src protein degradation.

SFKs in leukemias, lymphomas, and myelomas

Most SFKs are expressed primarily in hematopoietic cells and are involved in signaling pathways that regulate cell growth and proliferation. Several laboratory reports have indicated that myeloid cells primarily express Lyn, Hck, and Fgr.50 However, relatively little evidence has implicated SFK activity definitively in these processes.17

Lyn may be involved in the growth, survival, and motility of various hematologic malignancies. Interleukin 3 (IL-3)-induced up-regulation of Lyn kinase activity may be mediated by the 120-kD common subunit of human IL-3 and granulocyte-macrophage–colony stimulating factor receptors. This evidence suggests that Lyn kinase participates in early IL-3-initiated signaling events.

Danhauser-Riedl et al.51 first reported that the activity of Lyn and Hck was increased in hematopoietic cells that expressed Bcr-Abl. This interaction of Bcr-Abl with Src-kinases may be independent of Src and Abl kinase activity.52 In addition, expression of a kinase-defective mutant of Hck in a cytokine-dependent myeloid cell line significantly suppressed Bcr-Abl-induced, cytokine-independent outgrowth of these cells.53 Ptasznik et al.54 reported evidence that Lyn contributes to Bcr-Abl cell motility through phosphatidylinositol 3-kinase pathway signaling. Hu et al.,55 using BCR-ABL1 retrovirus-transduced bone marrow from mice that lacked all 3 Src kinases, efficiently induced chronic myeloid leukemia (CML) but not B-cell acute lymphoblastic leukemia (B-ALL) in recipients. In that study, the kinase inhibitor CGP76030 impaired the in vitro proliferation of Bcr-Abl-positive B-lymphoid cells and prolonged the survival of mice with B-ALL but not mice with CML. Those results suggested that SFKs were required for Bcr-Abl-positive B-lymphoblastic leukemia but not for CML.56 This may be different in the setting of failure to respond to imatinib, in which resistance to imatinib may be caused by the overexpression of Lyn in some patients.56

Another member of the SFKs, Lck, encodes a lymphocyte-specific protein tyrosine kinase, p56lck, which has been implicated in the pathogenesis of lymphoid neoplasia.57 Lck is expressed primarily in T lymphocytes, in which it probably plays an role during lymphopoiesis, proliferation, and receptor signaling.50 Jayaraman and Marks,58 using human T-cells that were deficient of inositoltriphosphate receptor 1 (a functional downstream target of Lck), demonstrated that those cells were highly resistant to ionizing radiation. Their report suggests that the modulation of calcium signals by Lck may be involved in the initial apoptotic phase.

Miscellaneous cancers

Increased levels of c-Src protein and/or kinase activity have been reported in other tumors, including lung carcinoma (50–80% of patients),59 ovarian carcinoma,60 esophageal carcinoma (3-fold to 4-fold increased activity in Barrett syndrome and 6-fold elevation in adenocarcinoma),61 gastric carcinoma,62 and pancreatic carcinoma.63

c-Src Kinase Inhibitors

  1. Top of page
  2. Abstract
  3. Signaling Regulation by Protein Tyrosine Kinases
  4. Structure and Function of the c-Src Family of Kinases
  5. c-Src Regulation
  6. SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases
  7. Involvement of c-Src in Human Cancers
  8. c-Src Kinase Inhibitors
  9. CONCLUSION
  10. Acknowledgements
  11. REFERENCES

The important role of c-Src in cancer and other diseases triggered the interest in developing c-Src inhibitors (Fig. 4). c-Src inhibitors can be classified as pyrazolo[3,4-d]pyrimidines (e.g., PP1 and PP2); pyrrolo[2,3-d]pyrimidines (e.g., CGP76030 and CGP77675); pyrido-[2,3-d]pyrimidines (e.g., PD166585, PD173955, and PD180970); quinolines (SKI606); indolinones (SU6656); dual c-Src-Abl inhibitors (e.g., dasatinib [N-(2-chloro-6-methylphenyl)-2-(6-[4-(2-hydroxyethyl)piperazin-1-yl]-2-methylpyrimidin-4-ylamino)thiazole-5-carboxamide]); or dual Lyn-Abl inhibitor (NS-187).

thumbnail image

Figure 4. These drawings illustrate the chemical structure Src inhibitors.

Download figure to PowerPoint

Pyrazolo[3,4-d]pyrimidines (PP1 and PP2)

First, it was shown that PP1 inhibited Lck and FynT, anti-CD3-induced protein tyrosine phosphorylation, and subsequent IL-2 gene activation in T lymphocytes.64 PP1 also showed selectivity for SFKs over other tyrosine kinase families, including ZAP-70 and Janus kinase 2, and over EGF-R.64

The urokinase-type plasminogen activator receptor (u-PAR) is a cell surface receptor that promotes cell migration and adhesion, extracellular matrix degradation, and EGF signaling. High u-PAR protein levels in patients with resected colon cancer was predictive of shortened survival.65 PP2 reduced u-PAR expression in colon cancer,66 and a combination of PP2 with a pharmacologic antagonist of u-PAR was superior to the individual agents in diminishing in vitro invasion.

Expression of the M2 subunit of ribonucleotide reductase (RRM2) is recognized as a promoter of chemoresistance.67 PP2 suppressed RRM2 expression, and incubation of pancreatic cell lines with escalating doses of PP2 and gemcitabine resulted in augmented, gemcitabine-induced, caspase-mediated apoptosis and suppressed RRM2 expression with decreased tumor growth and inhibition of metastasis.68

The E-cadherin cell adhesion system often is down-regulated in epithelial tumors. Nam et al.69 demonstrated that PP2 inhibited E-cadherin-mediated cell-cell adhesion and metastatic potential in colon, liver, and breast cancer cell lines. That study provided evidence that PP2 can activate the E-cadherin-mediated cell adhesion system, which is associated with the suppression of metastasis.

FGF-2 induced angiogenesis in vivo and endothelial cell differentiation in vitro.70 PP2 inhibited the formation of lamellopodia, and the expression of kinase-inactive c-Src reduced the phosphorylation of cortactin and paxillin, suggesting a model in which Src kinases are involved in organization of the actin cytoskeleton. These data suggest that pharmacologic inactivation of the SFKs inhibits FGF-2-stimulated angiogenesis by interfering with organization of the actin cytoskeleton in smooth muscle cells, affecting cell migration. In addition, PP2 inhibited the stimulation of angiogenesis by E-selectin.71

Pyrrolo[2,3-d]pyrimidines (CGP77675 and CGP76030)

The demonstration that mutant mice with a disrupted Src gene have functionally inactive osteoclasts suggests that c-Src plays a crucial role in osteoclastic bone resorption. c-Src is an essential enzyme for osteoclastic bone resorption, and it also has been demonstrated that synthetic c-Src inhibitors interfere with this process.72 CGP77675 reduced bone resorption in organ cultures and prevented in vivo IL-1β-induced hypercalcemia and bone loss in ovariectomized rats.73 The mechanism of action of CGP77675 is inhibiting phosphorylation of peptide substrates and c-Src Y416 autophosphorylation. CGP77675 inhibits phosphorylation of peptide substrates and autophosphorylation of purified c-Src. Although CGP77675 inhibited other kinases, the 50% inhibitory concentration (IC50) of Src was lower than the IC50 for Cdc2 (>500-fold), EGF-R (7.5-fold), vascular endothelial growth factor receptor (>50-fold), v-Abl (15-fold), and FAK (>25-fold).73

Similar results were obtained with another related inhibitor, CGP76030.74 Src inhibition resulted in impaired osteoclast function and cell damage suggestive of apoptosis probably through selective, sustained ERK1/ERK2 phosphorylation.74 In addition, CGP76030 inhibited the growth and survival of imatinib-resistant Bcr-Abl-expressing cells.75 CGP76030 and PP1 inhibited Bcr-Abl in a concentration-dependent manner by overlapping binding modes. In cells that expressed imatinib-resistant Bcr-Abl mutants, CGP76030, such as PP1, inhibited the activity of SFKs and Akt but not the activity of signal transducer and activator of transcription 5 or JUN kinase.75 These results suggested a possible role of c-Src inhibitors in overcoming imatinib resistance in CML.

Pyrido[2,3-d]-pyrimidines (PD 166285, PD173955, and PD180970)

Pyrido-pyrimidines are small molecules that can act as antagonists of c-Src. Pyrido-pyrimidines bind near the ATP-binding site, which is a highly conserved nucleotide-binding pocket within the kinase domain of protein tyrosine kinases, thereby blocking access of ATP. PD166285 has unique structural and biologic features, including 1) a novel pyrido[2,3-d]pyrimidine bicyclic structure; 2) ATP competitiveness for PDGF-R, EGF-R, and FGF-R tyrosine kinases and c-Src kinase; 3) a highly potent inhibitor of tyrosine kinase-mediated cellular growth, adhesion, migration and matrix metalloproteinase production, and 4) long-lasting inhibition of growth factor-mediated cellular functions.76 PD166285 inhibited the growth of human leukemic cells lines and leukemic blasts; and, when combined with doxorubicin, it had an additive effect on the inhibition of leukemic cell growth.77

PD173955 is another inhibitor of the same class that blocks cell proliferation and mitotic progression through inhibition of the SFKs.78 PD173955 also inhibits Bcr-Abl (IC50, 1–2 nM) and c-Kit (IC50, ≈ 50 nM).79 PD173955 is at least 10-fold more potent than imatinib and is able to target both the active and inactive configurations of Abl whereas, imatinib is effective only against the inactive conformation.80

PD180970 is the most recent member of this class. PD180970 induces apoptosis of K562 cells,81, 82 whereas it has no detectable effect on Philadelphia chromosome-negative HL60 human leukemic cells,81 and it induced apoptosis of CD34-positive leukemic cells from patients with imatinib-resistant CML in blastic phase. Gab2 is a multisite docking protein that was cloned from Bcr-Abl-transformed hematopoietic cells.83 Gab2 is tyrosine phosphorylated constitutively and is associated with the protein tyrosine phosphatase SHP2 in Bcr-Abl-transformed hematopoietic cells. PD180970 potently inhibits Gab2 tyrosine phosphorylation in K562 cells.84 These findings provide rationale to test the in vivo efficacy of pyrido-pyrimidines against imatinib-resistant Bcr-Abl-positive acute leukemias.85, 86

Quinolines (SKI-606)

SKI-606 is a 4-anilino-3-quinolinecarbonitrile inhibitor of Src with the following properties: 1) It inhibits Src in an enzyme assay with an IC50 of 1.2 nM, 2) it inhibits anchorage-independent growth of Src-transformed fibroblasts with an IC50 of 100 nM, and 3) it inhibits Src-dependent protein tyrosine phosphorylation at comparable or lower concentrations.87 SKI-606 shares multiple similar inhibitory properties with imatinib; however, SKI-606 exerts inhibition in multiple SFKs and does not affect other growth factor RTKs, such us PDGF-R, I-LGF I, EGF-R, FGF-R, Akt, and Cdk4. In the culture cell lines KU812, K562, and MEG-01, SKI-606 showed that it is a potent inhibitor of CML cell proliferation and survival (IC50, 5–20 nM).88 SKI-606 has a pattern inhibiting protein tyrosine phosphorylation similar to that of imatinib but with increased potency and is able to inhibit several mutants of the Abl kinase domain. In a K562 model xenografts, SKI-606 administered daily for 5 days resulted in complete regression of the tumor for up to 64 days.88 Studies of SKI-606 in solid tumors and CML are ongoing.

Indolinones (SU6656)

SU6656 and SU6657 are similar structurally c-Src inhibitors, which differ only in the substitution of the sulfonamide group at position 5 of the oxindole core (Fig. 4), but they have remarkably different selectivity profiles.21 SU6657 is a potent inhibitor of PDGF-R, FGF-R1 and Met, whereas SU6656 did not inhibit PDGF-R and exhibited greater than 6.5-fold selectivity for Src relative to other kinases. The IC50 of SU6656 for c-Src was ≈ 10-fold lower than that for Lck. In addition, SU6656 inhibits not only c-Src but other SFKs, such as Fyn and Yes, although it differs in the level of inhibition. For example, SU6656 was a potent inhibitor of Lyn but a rather poor inhibitor of Lck.21

Dasatinib-BMS354825

Recent structural analyses of Abl in an autoinhibited conformation revealed that its SH3, SH2, and kinase domains adopt an assembled conformation similar to that of Src kinases,89 despite the fact that Abl lacks the autoinhibitory tail phosphorylation site that is crucial for Src inhibition. Thus, several compounds designed as Src kinase inhibitors also are effective against Bcr-Abl.

Dasatinib (N-[2-chloro-6-methylphenyl]-2-[6-(4-[2-hydroxyethyl)piperazin-1-yl]-2-methylpyrimidin-4-ylamino]thiazole-5-carboxamide) is a synthetic, small-molecule, ATP-competitive inhibitor of SFKs with broad-spectrum antiproliferative activity against hematologic and solid tumor cell lines. Dasatinib is a dual-specific c-Src and Abl kinase inhibitor that can bind Bcr-Abl in both the active and inactive conformations.90 Compared with other c-Src inhibitors, dasatinib has less stringent conformational requirements for the Abl kinase inhibition. Dasatinib is >300-fold more potent than imatinib against the wild-type Bcr-Abl, binds ABL in both the active and inactive conformations, and has little if any inhibitory effect against normal hematopoietic progenitors.91,92 Dasatinib demonstrated in vitro and in vivo activity against 14 of 15 Bcr-Abl mutants against which it was tested: The only exception was T315I.92 In vitro studies have shown that the combination of dasatinib and imatinib may be synergistic against Bcr-Abl-positive cells, greatly reducing the recovery of mutated clones.93

A Phase I study of dasatinib in Philadelphia chromosome-positive CML after failure or intolerance to imatinib recently was reported.94 The doses of dasatinib were from 15 mg to 240 mg orally daily 5 to 7 days weekly in patients with chronic-phase CML and from 35 mg to 90 mg orally twice daily in patients with advanced-stage disease. Eighty-four patients were treated (Table 1).94 In chronic phase, 93% of patients achieved a complete histologic response, and 63% of patients achieved a cytogenetic response, which was complete 35% of those patients. In accelerated phase, a hematologic response was achieved in 9 of 11 patients, and a cytogenetic response was achieved in 40% of patients. In blastic phase, a hematologic response was achieved in 61%, and a cytogenetic response was achieved in 52% of patients. Seventeen different imatinib-resistant point mutations were identified in 67% of 63 patients prior to therapy. Clinical activity of dasatinib was observed in all patients except those with T315I.95 The dose-limiting toxicities were myelosuppression and occasional pleural effusion, mostly in the patients with advanced-stage disease. Dasatinib was absorbed rapidly with peak concentration in 2 hours and a terminal phase half-life of approximately 5 hours. Pharmacodynamic studies of dasatinib demonstrated > 50% inhibition of CRKL and Lyn phosphorylation, consistent with the serum concentrations. Phase II studies have initiated in the use of dasatinib all phases of CML after imatinib failure, and the results after a minimum of 6 months of follow-up are summarized on Table 1. These clinical data demonstrate the clinical efficacy of dasatinib and provide compelling evidence to support the further development of these agents.

Table 1. Use of Dasatinib in Chronic Myeloid Leukemia
PhaseNo. of patientsResponse rate (%)
HematologicCytogenetic
OverallCompleteOverallComplete
  • CP indicates chronic phase; AP, accelerated phase; MyBP, myeloid blastic phase; LyBP, lymphoid blastic phase; Ph+ ALL, Philadelphia chromosome-positive acute lymphoblastic.

  • *

    See Talpaz et al., 2005.94

  • See Hochhaus et al., 2006103; Talpaz et al., 2006104; Coutre et al., 2006105; Cortes et al., 2006106 (minimum follow-up, 6 months).

Phase I*
 CP4093936335
 AP1181453618
 MyBP2361355226
 LyBP1080709030
Phase II
 CP18690904533
 AP10759333621
 MyBP7451244227
 LyBP-Ph+ ALL7840265446

Another second-generation inhibitor, nilotinib (AMN107), recently has been introduced into clinical trials.96 This is a selective Abl inhibitor without Src-inhibitory activity. Significant activity was also achieved with nilotinib in CML after failure on imatinib. Although it is difficult to compare results with dasatinib, it is noteworthy that there was a high response rate to dasatinib in Philadelphia chromosome-positive ALL, suggesting that Src may play a particularly prominent role in this disease.55 Pleural effusions seldom occur with nilotinib or imatinib, but the role of Src inhibition in this event is unclear.

Other dual c-Src/Abl kinase inhibitors (AP23464, AP22408, AP23236, and NS-187)

AP23464 is a potent ATP-based inhibitor of Src and Abl that has antiproliferative activity against human CML cell lines and Bcr-Abl-transduced Ba/F3 cells (IC50, 14 nM).97 AP23464 blocks cell cycle progression and promotes apoptosis of Bcr-Abl-expressing cells. AP23464 also inhibits many Bcr-Abl mutants, but it does not inhibit T315I. In preclinical studies, AP23464 was more potent than PP1 in causing growth inhibition and Src kinase inhibition in the human myeloid cell line MV4-11.98 It is noteworthy that MV4-11 cells do not express Abl, indicating that Abl is not the drug target. In addition, AP23464 inhibits the D816 mutant of c-KIT, which is resistant to imatinib and is found frequently in systemic mastocytosis.99 Two additional compounds, AP22408 and AP23236, differ mechanistically by blocking c-Src-dependent, noncatalytic or catalytic activities in osteoclasts. These novel agents are bone-targeted compounds that were active in the treatment of osteoporosis and osteolytic bone metastasis in preclinical studies.100

NS-187 (INNO 406) is a dual Lyn-Abl inhibitor that is from 25 to 55 times more potent than imatinib in suppressing the growth of Bcr-Abl-bearing tumors.101 NS-187 inhibits Lyn without affecting the phosphorylation of Src, Blk, or Yes and is effective in inhibiting Bcr-Abl with most Abl kinase domain mutations except T315I.102 NS-187 inhibits in vitro growth of cells that express mutant Bcr-Abl proteins and prolonged the survival of mice engrafted with BaF3/E255K cells. Studies with NS-187 (INNO 406) in imatinib-resistant CML are ongoing.

CONCLUSION

  1. Top of page
  2. Abstract
  3. Signaling Regulation by Protein Tyrosine Kinases
  4. Structure and Function of the c-Src Family of Kinases
  5. c-Src Regulation
  6. SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases
  7. Involvement of c-Src in Human Cancers
  8. c-Src Kinase Inhibitors
  9. CONCLUSION
  10. Acknowledgements
  11. REFERENCES

During the last 20 years, studies on c-Src have provided a wealth of information on signal transduction; however, much regarding its intracellular functions remains unclear. Understanding the structure and function of this important class of protein kinases and elucidating the molecular signaling events mediated by them are important not only for deciphering the critical pathways that they regulate but also for designing new strategies to inhibit their action in abnormal or pathologic situations. Selective Src inhibitors are entering the clinic and offer hope for a rich family of anticancer agents.

REFERENCES

  1. Top of page
  2. Abstract
  3. Signaling Regulation by Protein Tyrosine Kinases
  4. Structure and Function of the c-Src Family of Kinases
  5. c-Src Regulation
  6. SFKs Are Pathways Downstream of Several Receptor Tyrosine Kinases
  7. Involvement of c-Src in Human Cancers
  8. c-Src Kinase Inhibitors
  9. CONCLUSION
  10. Acknowledgements
  11. REFERENCES
  • 1
    Rous P. A sarcoma of the fowl transmissible by an agent separable for the tumor cells. J Exp Med. 1911; 13: 397411.
  • 2
    Rubin H. Quantitative relations between causative virus and cell in the Rous no. 1 chicken sarcoma. Virology. 1956; 6: 669688.
  • 3
    Martin GS. Rous sarcoma virus: a function required for the maintenance of the transformed state. Nature. 1970; 227: 10211023.
  • 4
    Thomas SM, Brugge JS. Cellular functions regulated by Src family kinases. Annu Rev Cell Dev Biol. 1997; 13: 513609.
  • 5
    Bjorge JD, Jakymiw A, Fujita DJ. Selected glimpses into the activation and function of Src kinase. Oncogene. 2000; 19: 56205635.
  • 6
    Haskell MD, Slack JK, Parsons JT, et al. c-Src tyrosine phosphorylation of epidermal growth factor receptor, P190 RhoGAP, and focal adhesion kinase regulates diverse cellular processes. Chem Rev. 2001; 101: 24252440.
  • 7
    Sawyer TK. Src homology-2 domains: structure, mechanism and drug discovery. Biopolymers. 1998; 47: 243261.
  • 8
    Johnson LN, Noble MLM, Owen DJ. Active and inactive protein kinases: structural basis for regulation. Cell. 1996; 85: 149158.
  • 9
    Roche S, Fumagalli S, Courtneidge SA. Requirement for Src family protein tyrosine kinases in G2 for fibroblast cell division. Science. 1995; 269: 15671569.
  • 10
    Prathapam T, Tegen S, Oskarsson T, Trumpp A, Martin GS. Activated Src abrogates the Myc requirement for the G0/G1 transition but not for the G1/S transition. Proc Natl Acad Sci USA. 2006; 103: 2695700.
  • 11
    Cooper JA, Gould KL, Cartwright CA, et al. Tyr527 is phosphorylated in pp60c-src: implications for regulation. Science. 1986; 231: 14311434.
  • 12
    Roskoski RJr. Src kinase regulation by phosphorylation and dephosphorylation. Biochem Biophys Res Commun. 2005; 331: 114.
  • 13
    Nakagawa S, Tanaka H, Suzuki H, et al. Overexpression of the CSK gene suppresses tumor metastasis in vivo. Int J Cancer. 2000; 88: 384391.
  • 14
    Lee BC, Lee TH, Zagozdzon R, Avraham S, Usheva A, Avraham HK. Carboxyl-terminal Src kinase homologous kinase negatively regulates the chemokine receptor CXCR4 through YY1 and impairs CXCR4/CXCL12 (SDF-1α)-mediated breast cancer cell migration. Cancer Res. 2005; 65: 28402845.
  • 15
    Alonso G, Koegl M, Mazurenko N, et al. Sequence requirements for binding of Src family tyrosine kinases to activated growth factor receptors. J Biol Chem. 1995; 270: 98409848.
  • 16
    Schlaepfer DD, Jones KC, Hunter T, et al. Multiple Grb2-mediated integrin-stimulated signaling pathways to ERK2/mitogen-activated protein kinase: summation of both c-Src- and focal adhesion kinase initiated tyrosine phosphorylation events. Mol Cell Biol. 1998; 18: 25712585.
  • 17
    Summy JM, Gallick GE. Src family kinases in tumor progression and metastasis. Cancer Metastasis Rev. 2003; 22: 337358.
  • 18
    Cartwright CA, Eckhart W, Simon S, et al. Cell transformation by pp60c-src mutated in the carboxyterminal regulatory domain. Cell. 1987; 49: 8391.
  • 19
    Parsons JT, Parsons SJ. Src family protein tyrosine kinases: cooperating with growth factor and adhesion signaling pathways. Curr Opin Cell Biol. 1997; 9: 187192.
  • 20
    Twamley-Stein GM, Pepperkok R, Ansorge W, et al. The Src family tyrosine kinases are required for platelet-derived growth factor-mediated signal transduction in NIH 3T3 cells. Proc Natl Acad Sci USA. 1993; 90: 76967700.
  • 21
    Blake RA, Broome MA, Liu X, et al. SU6656, a selective Src family kinase inhibitor, used to probe growth factor signaling. Mol Cell Biol. 2000; 20: 90189027.
  • 22
    DeMali KA, Kazlauskas A. Activation of Src family members is not required for the platelet-derived growth factor beta receptor to initiate mitogenesis. Mol Cell Biol. 1998; 18: 20142022.
  • 23
    Roche S, Koegl M, Barone MV, et al. DNA synthesis induced by some but not all growth factors requires Src family protein tyrosine kinases. Mol Cell Biol. 1995; 15: 11021109.
  • 24
    Gullick WJ, Love SB, Writh C, et al. c-erbB-2 protein overexpression in breast cancer is a risk factor in patients with involved and uninvolved lymph nodes. Br J Cancer. 1992; 63: 954961.
  • 25
    Biscardi JS, Ishizawar RC, Silva CM, et al. Tyrosine kinase signaling in breast cancer: Epidermal growth factor receptor and c-Src interactions in breast cancer. Breast Cancer Res. 2000; 2: 203210.
  • 26
    Daub H, Wallasch C, Lankenau A, et al. Signal characteristics of G protein-transactivated EGF receptor. EMBO J. 1997; 16: 70327044.
  • 27
    Ware MF, Tice DA, Parsons SJ, et al. Overexpression of cellular Src in fibroblasts enhances endocytic internalization of epidermal growth factor receptor. J Biol Chem. 1997; 272: 3018530190.
  • 28
    Hall A. Rho GTPases and the actin cytoskeleton. Science. 1998; 279: 509514.
  • 29
    Chang JH, Gill S, Settleman J, et al. c-Src regulates the simultaneous rearrangement of actin cytoskeleton, p190RhoGAP, and p120RasGAP following epidermal growth factor stimulation. J Cell Biol. 1995; 130: 355368.
  • 30
    Avizienyte E, Frame MC. Src and FAK signaling controls adhesion fate and the epithelial-to-mesenchymal transition. Curr Opin Cell Biol. 2005; 17: 547547.
  • 31
    Potter MD, Barbero S, Cheresh DA. Tyrosine phosphorylation of VE-cadherin prevents binding of p120-and-beta catenin and maintains cellular mesenchymal state. J Biol Chem. 2005; 280: 3190631912.
  • 32
    Zamir E, Geiger B. Molecular complexity and dynamics of cell-matrix adhesions. J Cell Sci. 2001; 114: 35833590.
  • 33
    Sastry SK, Burridge K. Focal adhesions: a nexus for intracellular signaling and cytoskeletal dynamics. Exp Cell Res. 2000; 261: 2536.
  • 34
    Schaller MD. Biochemical signals and biological responses elicited by the focal adhesion kinase. Biochim Biophys Acta. 2001; 1540: 121.
  • 35
    Owens LV, Xu L, Craven RJ, et al. Overexpression of the focal adhesion kinase (p125FAK) in invasive human tumors. Cancer Res. 1995; 55: 27522755.
  • 36
    Irby RB, Yeatman TJ. Role of Src expression and activation in human cancer. Oncogene. 2000; 19: 56365642.
  • 37
    Brown MT, Cooper JA. Regulation, substrates and functions of Src. Biochim Biophys Acta. 1996; 1287(2–3): 121149.
  • 38
    Bolen JB, Veillette A, Schwartz AM, et al. Activation of pp60 (c-Src) protein kinase activity in human colon carcinoma. Proc Natl Acad Sci USA. 1987; 84: 22512255.
  • 39
    Cartwright CA, Meisler AI, Eckhart W. Activation of the pp60 (c-Src) protein kinase is an early event in colonic carcinogenesis. Proc Natl Acad Sci USA. 1990; 87: 558562.
  • 40
    Termuhlen PM, Curley SA, Talamonty MS, et al. Site-specific differences in pp60 (c-Src) activity in human colorectal metastases. J Surg Res. 1993; 54: 293298.
  • 41
    Irby RB, Mao W, Coppola D, et al. Activating SRC mutation in a subset of advanced human colon cancer. Nat Genet. 1999; 21: 187190.
  • 42
    Daigo Y, Furukawa Y, Kawasoe T, et al. Absence of genetic alteration at codon 531 of the human c-Src gene in 479 advanced colorectal cancers from Japanese and Caucasian patients. Cancer Res. 1999; 59: 42224224.
  • 43
    Wang NM, Yeh KT, Tsai CH, Chen SJ, Chang JG. No evidence of correlation between mutation at codon 531 of src and the risk of colon cancer in Chinese. Cancer Lett. 2000; 150: 201204.
  • 44
    Allgayer H, Boyd DD, Heiss MM, et al. Activation of Src kinase in primary colorectal carcinoma: an indicator of poor clinical prognosis. Cancer. 2002; 94: 344351.
  • 45
    Han NM, Curley SA, Gallick GE, et al. Differential activation of pp60(c-Src) and pp62(c-Yes) in human colorectal carcinoma and liver metastasis. Clin Cancer Res. 1996; 2: 13971404.
  • 46
    Ottenhoff-Kalff AE, Rijksen G, van Beurden EA, et al. Characterization of protein tyrosine kinases from human breast cancer: involvement of the c-Src oncogene product. Cancer Res. 1992; 52: 47734778.
  • 47
    Muthuswamy SK, Siegel PM, Dankort DL, et al. Mammary tumors expressing the neu proto-oncogene possesses elevated c-Src tyrosine kinase activity. Mol Cell Biol. 1994; 14: 735743.
  • 48
    Biscardi JS, Belsches AP, Parsons SJ, et al. Characterization of human epidermal growth factor receptor and c-Src interaction in human breast tumor cells. Mol Carcinog. 1998; 21: 261272.
  • 49
    Tan M, Li P, Klos KS, et al. ErbB2 promotes Src synthesis and stability: novel mechanisms of Src activation that confer breast cancer metastasis. Cancer Res. 2005; 65: 18581867.
  • 50
    Palacios EH, Weiss A. Function of the Src-family kinases, Lck and Fyn, in T-cell development and activation. Oncogene. 2004; 23: 79908000.
  • 51
    Danhauser-Riedl S, Warmurth M, Druker BJ, et al. Activation of Src kinases p53/56lyn and p59hck by p210bcr-abl in myeloid cells. Cancer Res. 1996; 56: 35893596.
  • 52
    Warmuth M, Bergmann M, Priess A, et al. The Src family kinase Hck interacts with Bcr-Abl by a kinase-independent mechanism and phosphorylates the Grb2-binding site of Bcr. J Biol Chem. 1997; 272: 3326033270.
  • 53
    Lionberger JM, Wilson MB, Smithgall TE. Transformation of myeloid leukemia cells to cytokine independence by Bcr-Abl is suppressed by kinase-defective Hck. J Biol Chem. 2000; 275: 1858118585.
  • 54
    Ptasznik A, Urbanowska E, Chinta S, et al. Crosstalk between BCR/ABL oncoprotein and CXCR4 signaling through a Src family kinase in human leukemia cells. J Exp Med. 2002; 196: 667678.
  • 55
    Hu Y, Liu Y, Pelletier S, et al. Requirement of Src kinases Lyn, Hck and Fgr for BCR-ABL1-induced B-lymphoblastic leukemia but not chronic myeloid leukemia. Nat Genet. 2004, 36: 453461.
  • 56
    Donato NJ, Wu JY, Stapley J, et al. BCR-ABL independence and LYN kinase overexpression in chronic myelogenous leukemia cells selected for resistance to STI571. Blood. 2003; 101: 690698.
  • 57
    Marth JD, Overell RW, Meier KE, et al. Translational activation of the lck proto-oncogene. Nature. 1988; 332: 171173.
  • 58
    Jayaraman T, Marks AR. T cells deficient in inositol 1,4,5-trisphosphate receptor are resistant to apoptosis. Mol Cell Biol. 1997; 17: 30053012.
  • 59
    Mazurenko NN, Kogan EA, Zborovskaya IB, et al. Expression of pp60c-src in human small cell and non-small cell lung carcinomas. Eur J Cancer. 1992; 28: 372377.
  • 60
    Wiener JR, Nakano K, Kuzrock RP, et al. Decreased Src tyrosine kinase activity inhibits malignant human ovarian cancer tumor growth in a nude mouse model. Clin Cancer Res. 1999; 5: 21642170.
  • 61
    Kumble S, Omary MB, Cartwright CA, et al. Src activation in malignant and premalignant epithelia of Barrett's esophagus. Gastroenterology. 1997; 112: 348356.
  • 62
    Takeshima E, Hamaguchi M, Watanabe T, et al. Aberrant elevation of tyrosine-specific phosphorylation in human gastric cancer cells. Jpn J Cancer Res. 1991; 82: 14281435.
  • 63
    Lutz MP, Esser IB, Flossmann-Kast BB, et al. Overexpression and activation of the tyrosine kinase Src in human pancreatic carcinoma. Biochem Biophys Res Commun. 1998; 243: 503508.
  • 64
    Hanke JH, Gardner JP, Dow RL, et al. Discovery of a novel, potent, and Src family-selective tyrosine kinase inhibitor. J Biol Chem. 1996; 271: 695701.
  • 65
    Suzuki S, Hayashi Y, Wang Y, et al. Urokinase type plasminogen activator receptor expression in colorectal neoplasias. Gut. 1998; 43: 798805.
  • 66
    Boyd DD, Wang H, Avila H, et al. Combination of a Src kinase inhibitor with a novel pharmacological antagonist of the urokinase receptor diminished in vitro colon cancer invasiveness. Clin Cancer Res. 2004; 10: 15451555.
  • 67
    Goan YG, Zhou B, Hu E, et al. Overexpression of ribonucleotide reductase as a mechanism of resistance to 2,2-difluorodeoxycytidine in the human KB cancer cell line. Cancer Res. 1999; 59: 42044207.
  • 68
    Duxbury MS, Ito H, Zinner MJ, et al. Inhibition of Src tyrosine kinase impairs inherent and acquired gemcitabine resistance in human pancreatic adenocarcinoma cells. Clin Cancer Res. 2004; 10: 23072318.
  • 69
    Nam JS, Ino Y, Sakamoto M, et al. Src family kinase inhibitor PP2 restores the E-cadherin/catenin cell adhesion system in human cancer cells and reduces cancer metastasis. Clin Cancer Res. 2002; 8: 24302436.
  • 70
    Javerzat S, Auguste P, Bikfalvi A. The role of fibroblast growth factors in vascular development. Trends Mol Med. 2002; 8: 483489.
  • 71
    Kumar P, Amin MA, Harlow LA, et al. Src and phosphatidylinositol 3-kinase mediate soluble E-selectin-induced angiogenesis. Blood. 2003; 101: 39603968.
  • 72
    Altmann E, Missbach M, Green J, et al. Pyrrolidinyl- and 7-piperidinyl-5-aryl-pyrrolo[2,3-d]pyrimidines—potent inhibitors of the tyrosine kinase c-Src. Bioorg Med Chem Lett. 2001; 11: 853856.
  • 73
    Missbach M, Jeschke M, Feyen J, et al. Novel inhibitor of the tyrosine kinase Src suppresses phosphorylation of its major cellular substrates and reduces bone resorption in vitro and in rodent models in vivo. Bone. 1999; 24: 437449.
  • 74
    Recchia I, Rucci N, Funari A, et al. Reduction of c-Src activity by substituted 5,7-diphenyl-pyrrolo[2,3-d]-pyrimidines induces osteoclast apoptosis in vivo and in vitro. Involvement of ERK1/2 pathway. Bone. 2004; 34: 6579.
  • 75
    Warmuth M, Simon N, Mitina O, et al. Dual-specific Src and Abl kinase inhibitors, PP1 and CGP76030, inhibit growth and survival of cells expressing imatinib mesylate-resistant Bcr-Abl kinases. Blood. 2003; 101: 664672.
  • 76
    Panek RL, Lu GH, Klutchko SR, et al. In vitro pharmacological characterization of PD 166285, a new nanomolar potent and broadly active protein tyrosine kinase inhibitor. J Pharmacol Exp Ther. 1997; 283: 14331444.
  • 77
    Roginskaya V, Zuo S, Caudell E, et al. Therapeutic targeting of Src-kinase Lyn in myeloid leukemic cell growth. Leukemia. 1999; 13: 855861.
  • 78
    Moasser MM, Srethapakdi M, Sachar KS, et al. Inhibition of Src kinases by a selective tyrosine kinase inhibitor causes mitotic arrest. Cancer Res. 1999; 59: 61456152.
  • 79
    Wisniewski D, Lambek CL, Liu C, et al. Characterization of potent inhibitors of the Bcr-Abl and the c-Kit receptor tyrosine kinases. Cancer Res. 2002; 62: 42444255.
  • 80
    Nagar B, Bornmann WG, Pellicena P, et al. Crystal structures of the kinase domain of c-Abl in complex with the small molecule inhibitors PD173955 and imatinib (STI-571). Cancer Res. 2002; 62: 42364243.
  • 81
    Dorsey JF, Jove R, Kraker AJ, et al. The pyrido[2,3-d]pyrimidine derivate PD180970 inhibits p210Bcr-Abl tyrosine kinase and induces apoptosis of K652 leukemic cells. Cancer Res. 2000; 60: 31273131.
  • 82
    La Rosse P, Corbin AS, Stoffregen EP, et al. Activity of the Bcr-Abl kinase inhibitor PD180970 against clinically relevant Bcr-Abl isoforms that cause resistance to imatinib mesylate (Gleevec, STI571). Cancer Res. 2002; 62: 71497153.
  • 83
    Gu H, Pratt JC, Burakoff SJ, et al. Cloning of p97/Gab2, the major SHP2-binding protein in hematopoietic cells, reveals a novel pathway for cytokine-induced gene activation. Mol Cell. 1999; 2: 729740.
  • 84
    Kraker AJ, Hartl BG, Amar A, et al. Biochemical and cellular effects of c-src selective pyrido[2,3-d]pyrimidine tyrosine kinase inhibitors. Biochem Pharmacol. 2000; 60: 885898.
  • 85
    Nimmanapally R, O'Bryan E, Huang M, et al. Molecular characterization and sensitivity of STI-571 (imatinib mesylate-Gleevec)-resistant, Brc-Abl-positive, human acute leukemia cells to SRC kinase inhibitor PD180970 and 17-allylamino-17-demethoxygeldanamycin. Cancer Res. 2002; 62: 57615769.
  • 86
    von Bubnoff N, Veach DR, Miller T, et al. Inhibition of wild-type and mutant Bcr-Abl by pyrimidine-type small molecule kinase inhibitors. Cancer Res. 2003; 63: 63956404.
  • 87
    Boschelli DH, Ye F, Wang YD, et al. Optimization of 4-phenylamino-3-quinolinecarbonitriles as potent inhibitors of Src kinase activity. J Med Chem. 2001; 44: 39653977.
  • 88
    Golas JM, Arndt K, Etienne C, et al. SKI-606, a 4-anilino-3-quinolinecarbonitrile dual inhibitor of Src and Abl kinases, is a potent antiproliferative agent against chronic myelogenous leukemia cells in culture and causes regression of K562 xenografts in nude mice. Cancer Res. 2003; 63: 375381.
  • 89
    Nagar B, Hantschel O, Young M, et al. Structural basis for the autoinhibition of c-Abl tyrosine kinase. Cell. 2003; 112: 859871.
  • 90
    Tokarshi JS, Newitt J, Lee FY, et al. The crystal structure of Abl kinase with BMS354825, a dual Src/Abl kinase inhibitor. Blood. 2004; 104: 160a. Abstract 553.
  • 91
    Burgess MR, Skaggs BJ, Shah NP, et al. Comparative analysis of two clinically active BCR-ABL kinase inhibitors reveals the role of conformation-specific binding in resistance. Proc Natl Acad Sci USA. 2005; 102: 33953400.
  • 92
    Shah NP, Tran C, Lee FY, et al. Overriding imatinib resistance with a novel ABL kinase inhibitor. Science. 2004; 305: 399401.
  • 93
    O'Hare T, Walters DK, Stoffregen EP, et al. In vitro activity of Bcr-Abl inhibitors AMN107 and BMS-354825 against clinically relevant imatinib-resistant Abl kinase domain mutants. Cancer Res. 2005; 65: 45004505.
  • 94
    Talpaz M, Shah NP, Kantarjian H, et al. Dasatinib in imatinib-resistant Philadelphia chromosome-positive leukemias. N Eng J Med. 2006; 354: 25312596.
  • 95
    Shah N, Sawyers CL, Kantarjian H, et al. Correlation of clinical response to BMS-354825 with BCR-ABL mutation status in imatinib-resistant patients with chronic myeloid leukemia (CML) and Philadelphia chromosome-associated acute lymphoblastic leukemia (Ph+ ALL). J Clin Oncol. 2005; 16S: 564s. Abstract 6521.
  • 96
    Kantarjian H, Giles F, Wunderle L, et al. Nilotinib in imatinib-resistant CML and Philadelphia chromosome-positive ALL. N Eng J Med. 2006; 354: 25422596.
  • 97
    O'Hare T, Pollock R, Stoffregen EP, et al. Inhibition of wild-type and mutant Bcr-Abl by AP23464, a potent ATP-based oncogenic protein kinase inhibitor: implications for CML. Blood. 2004; 104: 25322539.
  • 98
    Kanerva J, Nwawka O, Hwang K, et al. A potent Src/Abl kinase inhibitor (AP23464) potently inhibits the growth of myeloid leukemic cells characterized by Flt3-ITD expression, GM-CSF dependency, or G-CSF responsiveness via and Abl-independent mechanism. Blood. 2004; 104: 202b. Abstract 4479.
  • 99
    Corbin AS, Demehri S, Griswold IJ, et al. In vitro and in vivo activity of the Src/Abl kinase inhibitor AP23464 and analogs against activation loop mutants of KIT. Blood. 2004; 104: 227a. Abstract 793.
  • 100
    Shakespeare WC, Metcalf CAIII, Wang Y, et al. Novel bone-targeted Src tyrosine kinase inhibitor drug discovery. Curr Opin Drug Discov Dev. 2003; 6: 729741.
  • 101
    Kimura S, Naito H, Segawa H, et al. NS-187, a potent and selective dual Bcr-Abl/Lyn tyrosine kinase inhibitor, is a novel agent for imatinib-resistant leukemia. Blood. 2005; 106: 39483954.
  • 102
    Naito H, Kimura S, Nakaya Y, et al. In vivo antiproliferative effect of NS-187, a dual Bcr-Abl/Lyn tyrosine kinase inhibitor, on leukemic cells harbouring Abl kinase domain mutations. Leukemia Res. 2006 [Epub ahead of print].
  • 103
    Hochhaus A, Kantarjian H, Baccarani M, et al. Dasatinib in patients with chronic phase chronic myeloid leukemia who are resistant or intolerant to imatinib: results of the CA180013 “START-C” Study. J Clin Oncol. 2006; 24: 339a. Abstract 6508.
  • 104
    Talpaz M, Apperley JF, Kim DW, et al. Dasatinib in patients with accelerated phase chronic myeloid leukemia who are resistant or intolerant to imatinib: results of the CA180005 “START-A” Study. J Clin Oncol. 2006; 24: 343a. Abstract 6526.
  • 105
    Coutre S, Martinelli G, Dombret H, et al. Dasatinib in patients with chronic myeloid leukemia in lymphoid blast crisis or Philadelphia-chromosome positive acute lymphoblastic leukemia who are imatinib-resistant or intolerant: the CA180015 ‘START-L’ Study. J Clin Oncol. 2006; 24: 344a. Abstract 6528.
  • 106
    Cortes J, Kim DW, Rosti G, et al. Dasatinib in patients with chronic myeloid leukemia in myeloid blast crisis who are imatinib-resistant or intolerant: the CA180006 ‘START-B’ Study. J Clin Oncol. 2006; 24: 344a. Abstract 6529.