Neonatal Lethality in Knockout Mice Expressing the Kinase-Dead Form of the Gefitinib Target GAK Is Caused by Pulmonary Dysfunction
et al. (2011) Neonatal Lethality in Knockout Mice Expressing the Kinase-Dead Form of the Gefitinib
Target GAK Is Caused by Pulmonary Dysfunction. PLoS ONE 6(10): e26034. doi:10.1371/journal.pone.0026034
Neonatal Lethality in Knockout Mice Expressing the Kinase-Dead Form of the Gefitinib Target GAK Is Caused by Pulmonary Dysfunction
Hiroe Tabara 0
Yoko Naito 0
Akihiko Ito 0
Asako Katsuma 0
Minami A. Sakurai 0
Shouichi Ohno 0
Hiroyuki Shimizu 0
Norikazu Yabuta 0
Hiroshi Nojima 0
Hiroaki Matsunami, Duke University, United States of America
0 1 Department of Molecular Genetics, Research Institute for Microbial Diseases, Osaka University , Osaka , Japan , 2 Department of Pathology, Kinki University Faculty of Medicine , Osaka , Japan
Gefitinib (Iressa) is an inhibitor of the epidermal growth factor receptor (EGFR) that has shown promising activity in the treatment of patients with non-small cell lung cancer (NSCLC). However, adverse side effects of gefitinib treatment, such as respiratory dysfunction, have limited the therapeutic benefit of this targeting strategy. The present results show that this adverse effect can be attributed to the inhibition of the novel gefitinib target GAK (Cyclin G-associated kinase), which is as potently inhibited by the drug as the tyrosine kinase activity of EGFR. Knockout mice expressing the kinase-dead form of GAK (GAK-kd) died within 30 min after birth primarily due to respiratory dysfunction. Immunohistochemical analysis revealed that surfactant protein A (SP-A) was abundant within alveolar spaces in GAK-kd+/+ mice but not in GAK-kd-/- pups. E-cadherin and phosphorylated EGFR signals were also abnormal, suggesting the presence of flat alveolar cells with thin junctions. These results suggest that inhibition of GAK by gefitinib may cause pulmonary alveolar dysfunction, and the present study may help prevent side effects associated with gefitinib therapy in NSCLC patients.
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Funding: This work was supported in part by grants-in-aid for Scientific Research S (No. 15101006), Scientific Research B (No. 20370081) and Exploratory
Research (No. 21651085) from the Ministry of Education, Culture, Sports, Science and Technology of Japan (http://www.mext.go.jp/english/) to Hiroshi Nojima. No
additional external funding was received for this study. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of
the manuscript.
Competing Interests: The authors have declared that no competing interests exist.
. These authors contributed equally to this work.
EGFR is a membrane receptor tyrosine kinase that is activated
by ligand binding and dimerization, resulting in the activation of a
signaling pathway that controls cell proliferation, differentiation,
and survival [1]. Constitutively active EGF-EGFR signaling due to
overexpression of mutated or wild-type EGFR is found in a broad
range of human carcinomas, leading to the activation of
antiapoptotic pathways and uncontrolled cell proliferation [2], [3].
EGFR selective tyrosine kinase inhibitors (TKIs) such as gefitinib
(Iressa) and erlotinib (Tarceva) that bind to the adenosine
triphosphate (ATP)-binding site of the enzyme have been used
as successful treatments for NSCLC patients, particularly in the
presence of activating mutations within the EGFR gene [4], [5].
Although occurring at low frequency, progressive respiratory
dysfunction, including acute interstitial pneumonia (IP) is the most
severe adverse effect of gefitinib [6], which has limited the
therapeutic benefit of this drug. Tumor regression in gefitinib
treated NSCLC patients is at least partly due to apoptotic death of
tumor cells. Shutdown of the EGFR-MEK-ERK signaling cascade
induces activation of the proapoptotic BH3-only protein BIM,
causing gefitinib-induced tumor cell apoptosis [7]. Moreover,
induction of another BH3-only protein, p53 up-regulated
modulator of apoptosis (PUMA), by p73, is also involved in
EGFR inhibitor-induced apoptosis [8], [9]. However, the
molecular mechanisms underlying the development of IP in
response to gefitinib treatment and the selectivity of the drug for its
cellular targets are not fully understood.
Two protein kinases were identified by liquid chromatography
(LC)-MS/MS as novel gefitinib targets [10], namely a negative
regulator of EGFR signaling, GAK [11] and Rip2/RICK
(receptor-interacting caspase-like apoptosis-regulatory kinase), a
signal transducer and integrator of signals for both the innate and
adaptive immune systems that functions through the promotion of
nuclear factor kappa B and caspase activation [12], [13]. Both
targets are affected by gefitinib as potently as the tyrosine kinase
activity of wild-type EGFR in vitro [10]. Although the physiological
significance of these phenomena needs to be elucidated for the
selection of EGFR-directed drugs with minimal side effects, there
is little data presently available.
The ubiquitously expressed kinase GAK was first identified as a
(...truncated)