Disruption of Src function potentiates Chk1-inhibitor–induced apoptosis in human multiple myeloma cells in vitro and in vivo

Abstract

IntroductionMultiple myeloma (MM) is a neoplastic disorder of mature, differentiated B lymphocytes. Whereas recent insights into MM molecular pathogenesis prompted the introduction of effective new agents, including the proteasome inhibitor bortezomib and the immunomodulatory agents thalidomide and lenalidomide, MM remains largely incurable 1 and new strategies are clearly needed.DNA-damage checkpoints halt cell-cycle progression after extrinsic DNA damage (eg, by genotoxic agents or radiation) or intrinsic DNA-replication stress during the undisturbed cell cycle, permitting DNA-repair machinery initiation or DNA-replication block circumvention. 2 Checkpoint responses are initiated by ATM (Ataxia telangiectasia mutated) and ATR (Ataxia telangiectasia and Rad3-related), which induce checkpoint kinases (Chk1 and Chk2), thus disabling Cdk1/p34 cdc2 or Cdk2 by preventing dephosphorylation at inhibitory sites (T14/Y15) via inhibition/degradation of Cdc25 phosphatases, resulting in cell-cycle arrest. Genomic instability and defective DNA-damage checkpoints are characteristic of diverse human cancers, including MM. 3 Chk1 has a critical role in the DNA-damage-response network. 2 Moreover, novel Chk1 functions in the DNA-replication checkpoint, the mitoticspindle checkpoint, and DNA repair have been identified, 2,4 stimulating clinical development of multiple Chk1 inhibitors, including UCN-01 (Kyowa), AZD7762 (AstraZeneca), LY2603618 (Lilly), SCH900776 (Schering-Plough), and PF-00477736 (Pfizer). Whereas these efforts have focused on chemotherapy or radiation sensitization, 2,5,6 recent evidence implicating Chk1 in normal cell-cycle checkpoints (eg, the DNA replication checkpoint) suggests alternative therapeutic strategies.We previously reported that Chk1 inhibitors (eg, UCN-01 or more specific Chk1 inhibitors) activate extracellular signalregulated kinase 1/2 (ERK1/2) in human MM and leukemia cells, while blockade of this event by MEK1/2 (mitogen-activated protein kinase [MAPK]/ERK kinase 1/2) inhibitor dramatically induces apoptosis. 7,8 Furthermore, interruption of Ras function by farnesyltransferase inhibitors 9,10 or statins 11 acted similarly. Because Src plays an important role in Ras 3 ERK1/2 signaling activation, 12 the possibility that Src may be involved in Chk1-inhibitor-mediated ERK1/2 activation arose. Src family kinases (SFKs) are up-regulated/activated in multiple human tumors. 13 Src itself has been implicated in transformation, survival, proliferation, adhesion, migration, invasion, 12,13 and angiogenesis. 14 Src is generally activated by receptor tyrosine kinases or integrin-related kinases (eg, focal adhesion kinase [FAK] 16 Recently, Src inhibitors (eg, BMS354825) were shown to inhibit angiogenesis and the proliferative/survival effects of growth factors, including vascular endothelial growth factor (VEGF) and The online version of this article contains a data supplement.The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact,...

Dasatinib reverses the multidrug resistance of breast cancer MCF-7 cells to doxorubicin by downregulating P-gp expression via inhibiting the activation of ERK signaling pathway

“…44,45 The quantification of P-ERK expression was normalized to the total content of ERK. Each column shows the mean § SD of 3 independent experiments, performed in triplicate.…”

Enhancing CHK1 inhibitor lethality in glioblastoma

“…The results are similar to what have been observed in non-CNS tumor cell types. [12][13][14] Medulloblastoma is the most common cause of CNS tumor in children and although disease free response rates now approach ~80% they do so with considerable morbidity to the child. Clearly, better therapeutic approaches are required to treat this CNS tumor type.…”

“…Our studies focused in particular detail on are essentially in G 0 phase of the cell cycle, CHK1 and MEK1/2 inhibitors synergized to kill arguing against the lethal effect being dependent on changes in cell cycle progression. 12 The present studies were performed to define whether inhibitors of the SRC-MEK1/2-ERK1/2 pathway interacted with dephosphorylation of S6 protein and with p38 MAPK and/or JNK1-3 activation.…”

“…8,9 We have published several studies combining SRC/MEK1/2 inhibitors with CHK1 inhibitors in established tumor cell types, notably breast cancer cells, but not in CNS tumor cells that are generally thought to be much more resistant to many therapeutic modalities, and demonstrated that the lethality of CHK1 inhibitors is enhanced by their combination with inhibitors of the SRC-RAS-MEK1/2-ERK1/2 pathway. [9][10][11][12][13][14] Treatment of tumor cells with CHK1 inhibitors enhances signaling through the SRC-RAS-MEK1/2-ERK1/2 pathway and expression of a dominant negative CHK1 protein elevates basal ERK1/2 activity and blocks CHK1 inhibitor-induced ERK1/2 activation. [9][10][11][12][13][14] Interruption of MEK1/2-ERK1/2 pathway by MEK1/2 inhibitors dramatically potentiated UCN-01-induced cell death.…”

“…[9][10][11][12][13][14] Treatment of tumor cells with CHK1 inhibitors enhances signaling through the SRC-RAS-MEK1/2-ERK1/2 pathway and expression of a dominant negative CHK1 protein elevates basal ERK1/2 activity and blocks CHK1 inhibitor-induced ERK1/2 activation. [9][10][11][12][13][14] Interruption of MEK1/2-ERK1/2 pathway by MEK1/2 inhibitors dramatically potentiated UCN-01-induced cell death. 15,16 What is perhaps more of note are findings that non-transformed blood cells, e.g., peripheral mononuclear cells, CD34 + stem cells or primary cultures of human mammary epithelial cells or of rodent hepatocytes were resistant to the CHK1 inhibitor based drug combination toxicity.…”