AZD5438, an Inhibitor of Cdk1, 2, and 9, Enhances the
Radiosensitivity of Non-Small Cell Lung Carcinoma Cells
Pavithra Raghavan, MS,*,1Vasu Tumati, BS,*,1Lan Yu, PhD,* Norman Chan, MD, PhD,y
Nozomi Tomimatsu, PhD,* Sandeep Burma, PhD,*,zRobert G. Bristow, MD, PhD,y
and Debabrata Saha, PhD*,z
*Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, Texas;yDepartments
of Medical Biophysics and Radiation Oncology, Princess Margaret Hospital, University Health Network, University
of Toronto, Ontario, Canada; andzSimmons Comprehensive Cancer Center, Dallas, Texas
Received Feb 24, 2012, and in revised form Apr 25, 2012. Accepted for publication May 3, 2012
This preclinical study evalu-
ated the efficacy of new
therapies in the treatment of
lung cancer. The combina-
tion of a new generation Cdk
inhibitor, AZD5438, and
therapy was tested in several
non-small-cell lung cancer
(NSCLC) models. Treatment
with AZD5438 significantly
enhanced the response of
NSCLC cells to radiation,
both in vitro and in vivo.
These findings indicate that
Cdk inhibitors are promising
candidates for clinical eval-
uation as adjuvant therapy
Purpose: Radiation therapy (RT) is one of the primary modalities for treatment of non-small
cell lung cancer (NSCLC). However, due to the intrinsic radiation resistance of these tumors,
many patients experience RT failure, which leads to considerable tumor progression including
regional lymph node and distant metastasis. This preclinical study evaluated the efficacy of
a new-generation cyclin-dependent kinase (Cdk) inhibitor, AZD5438, as a radiosensitizer in
several NSCLC models that are specifically resistant to conventional fractionated RT.
Methods and Materials: The combined effect of ionizing radiation and AZD5438, a highly
specific inhibitor of Cdk1, 2, and 9, was determined in vitro by surviving fraction, cell cycle
distribution, apoptosis, DNA double-strand break (DSB) repair, and homologous recombination
(HR) assays in 3 NSCLC cell lines (A549, H1299, and H460). For in vivo studies, human xeno-
graft animal models in athymic nude mice were used.
Results: Treatment of NSCLC cells with AZD5438 significantly augmented cellular radiosen-
sitivity (dose enhancement ratio rangeing from 1.4 to 1.75). The degree of radiosensitization by
AZD5438 was greater in radioresistant cell lines (A549 and H1299). Radiosensitivity was
enhanced specifically through inhibition of Cdk1, prolonged G2-M arrest, inhibition of HR, de-
layed DNA DSB repair, and increased apoptosis. Combined treatment with AZD5438 and irra-
diation also enhanced tumor growth delay, with an enhancement factor ranging from 1.2-1.7.
Conclusions: This study supports the evaluation of newer generation Cdk inhibitors, such as
AZD5438, as potent radiosensitizers in NSCLC models, especially in tumors that demonstrate
variable intrinsic radiation responses. ? 2012 Elsevier Inc.
Reprint requests to: Debabrata Saha, PhD, Department of Radiation
Oncology, Division of Molecular Radiation Biology, University of Texas
Southwestern Medical Center, 2201 Inwood Rd, Dallas, TX 75390-9187.
Tel: (214) 648-7750; Fax: (214) 648-5995; E-mail: debabrata.saha@
This work was supported by Flight Attendant Medical Research
Institute grants, W81XWH-11-1-0270, R01CA149461, NNX10AE08G,
and RP100644 and by a clinical research fellowship from the Doris Duke
Conflict of interest: none.
1Pavithra Raghavan and Vasu Tumati contributed equally to this
this articlecan be found at
Int J Radiation Oncol Biol Phys, Vol. 84, No. 4, pp. e507ee514, 2012
0360-3016/$ - see front matter ? 2012 Elsevier Inc. All rights reserved.
International Journal of
Non-small cell lung cancer (NSCLC) is both the most prevalent
type of lung cancer and the leading cause of cancer death
worldwide. Up to 40% of NSCLC patients present with locally
advanced and mostly inoperable disease (1). For patients who
present withadvanced disease, concurrent
therapy remains the only effective treatment; combined therapy
results in 2-year survival rates of between 8% and 43% (2). Poor
overall survival rates in NSCLC patients may be attributed to the
intrinsic radiation resistance of many tumors. Solid tumors,
including NSCLC, are heterogeneous and contain subpopulations
of cells with divergent levels of sensitivity to established cancer
therapy including radiation therapy (RT). Perturbation of cell
cycle regulation is a key factor in the development of most
cancers (3). The regulatory proteins that control cell cycle
progression are the cyclins, cyclin-dependent kinases (Cdks), and
their substrate proteins Cdk inhibitors, tumor suppressor gene
products, p53 and pRb. Several Cdk inhibitors including fla-
vopiridol, indisulam, AZD5438, P276-00, EM-1421, seliciclib,
PD0332991, and SCH727965 have entered clinical trials (4, 5)
and have demonstrated promising outcomes especially in
combination with other chemotherapeutic agents (4). Cdk inhib-
itors preferentially target proliferating cells, but these inhibitors
can also induce cell death in noncycling radioresistant tumor
In this study, we tested the efficacy of AZD5438 (9), a new-
generation inhibitor of Cdk 1, 2, and 9 in combination with
fractionated RT in NSCLC cell lines (A549, H1299, and H460)
and in animal models. AZD5438 significantly enhanced the effect
of radiation in NSCLC cells. This enhanced radiosensitivity was
due mostly to Cdk1 inhibition and was partially attributed to
persistent DNA double-strand breaks (DSB) and the inhibition of
DNA homologous recombination (HR) repair.
Methods and Materials
Cell culture and reagents
The human NSCLC cell lines H460, A549, and H1299 were
kindly provided by Dr John D. Minna at University of Texas
Southwestern Medical Center, Dallas, TX, and maintained in
RPMI 1640 medium with 10% fetal bovine serum and
50 units/mL penicillin and 50 mg/mL streptomycin in 5% CO2
at 37?C. AZD5438 (molecular weight, 471.36) was obtained
from AstraZeneca (London, UK). Cells were irradiated using
ciates, San Fernando, CA) at a dose rate of 3.47 Gy/min (8).
137Cs source (Mark 1-68 irradiator; JL Shepherd and Asso-
Clonogenic survival assay
Cells were treated with AZD5438 for 24 h and then treated
with increasing doses of IR (0, 2, 4, 6, and 8 Gy). Colony
formation assay (CFA) and determination of dose enhancement
ratio (DER) were performed as described previously (7, 8).
CFA was also performed using short interfering RNA (siR-
NAs) against Cdk1 and Cdk9 (Life technologies Grand Island,
NY) and Cdk2 (Dharmacon, Inc Chicago, IL). Cells were
transiently transfected witheitherindividual siRNAs or
scrambled siRNAs. After 48 h, cells were plated for CFA and
Western blot analysis.
Western blot assay
Cell lysates were prepared from each sample, and total protein
(20 mg) was subjected to immunoblot analysis and probed with
antibodies as indicated. b-Actin was used for loading control.
Double-strand break repair assay
DSB repair assay was performed as described previously (10).
The number of phospho-gH2AX foci (green) was determined
at each time point (average of 50 nuclei), and the percentage
of foci remaining was plotted against time to obtain DSB
repair kinetics (10). Data is represented as mean ? SEM.
Immunoblot analysis of Cdk1, Cdk2, and Cdk9 in NSCLC cells.
(B) Clonogenic survival assay for drug sensitivity determination.
NSCLC cells were plated with or without AZD5438 (5 nM, 50
nM, 500 nM, 5 mM, and 50 mM). Colonies were counted and
percentage of survival was plotted against log [Dose]. (C)
AZD5438 prevents Cdk2 activity. H460 cells were treated with
nocodazole (50 ng/mL) for 16 h to synchronize cells in M phase.
Cells were washed and treated with AZD5438 (435 nM) for 4 h,
followed by immunoblot analysis of pRb (Ser780). Total Rb
protein was used for loading control. (D) AZD5438 does not
abrogate Chk1/2 activation. Both A549 (250 nM) and H460 (435
nM) cells were treated with AZD5438 for 24 h and treated with IR
(5 Gy) for 30 min. Immunoblot analysis using pChk1(Ser317) and
pChk2 (T68). Total Chk1 and Chk2 proteins and actin were used
for loading control.
Specificity of AZD5438 in human NSCLC cells. (A)
Raghavan et al. International Journal of Radiation Oncology ? Biology ? Physics
Direct repeat-green fluorescent protein HR assay
The direct repeat-green fluorescent protein (DR-GFP) assay was
performed as described by Chan et al (11). Transient expression of
the I-SceI endonuclease generates a DNA DSB at the integrated
GFP gene sequences and stimulates HR. For each experiment,
50,000 cells were scored per treatment group, and the frequency of
recombination events was calculated from the number of GFP-
positive cells divided by the number of cells analyzed after
correction for transfection efficiency.
Cell cycle analysis
Cells cycle assays were performed with propidium iodide (PI, 100
mg/mL) as previously described (10, 12). At least 20,000 cells were
counted, and the proportions of cells at different phases were gated
and calculated using Flowjo version 8.7.1 software (Tree Star, Inc).
Tumor growth delay
Female athymic nude mice (nu/nu, 5-6 weeks old) were injected
(1 ? 106cells in 100 mL) subcutaneously into the right posterior
flanks. Tumors were treated when they reached 2-3 mm in
diameter. Treatment groups (5 animals per group) included
untreated control (treated with 0.9% saline), those treated with
AZD5438 (25 mg/kg/day for 5 days, by mouth [po]), with radi-
ation (2 Gy/day, 5 days), and those that received combined
treatment with AZD5438 and IR. AZD5438 was administered 1 h
before radiation. Tumor growth delay and the dose enhancement
factor were then determined (7). All experiments were conducted
AZD5438. A549 (75 nM), H1299 (50 nM), and H460 (200 nM) cells were treated with AZD5438 for 24 h and treated with IR as indicated.
Cells were trypsinized immediately and colony formation was counted. (A-C, right panels) AZD5438 in combination with IR enhanced the
tumor growth delay in NSCLC cells: subcutaneous tumors were treated, as indicated, with IR (2 Gy per day, 5 days), AZD5438 (25 mg/kg/
day for 5 days), and IR plus AZD5438 (25 mg/kg/day plus 2 Gy per day for 5 days). AZD5438 was administrated po 1 h before IR. Tumor
volume (mm3) was measured twice per week and was plotted against days.
AZD5438 increased the sensitivity of NSCLC cells to IR. (A-C, left panels) Clonogenic survival of NSCLC cells with or without
Volume 84 ? Number 4 ? 2012 Lung cancer and radiotherapy e509
under Institutional Animal Care and Use Committee of UTSW
approved guidelines for animal welfare.
Statistical analysis of DSB repair and HR assays were done using
1-sided unpaired t-tests. Clonogenic survival curves were modeled
with the linear quadratic equation (S Z e?[aD þ bD2]) for radiation
treatment and a four-parameter variable slope regression for drug
Specificity of AZD5438 in NSCLC cells
AZD5438 inhibits Cdk 1, 2, and 9, therefore, the levels of Cdk
protein in 3 NSCLC cell lines were determined. The relative
expression levels of these 3 proteins were similar in all 3 cell lines
highly sensitive, while H460 was the most resistant (435.8 nM) to
AZD5438(Fig. 1B). AZD5438specificallyinhibited the
phosphorylation of Rb (pSer780) by inhibiting Cdk2 activity
(Fig. 1C), whereas the activity levels of other cell cycle regulatory
proteins such as Chk1 (pSer137) and Chk2 (pThr68) were not per-
turbed. These results show that AZD5438 is highly specific to Cdks
and demonstrates differential sensitivity in NSCLC cells.
AZD5438 increases the sensitivity of NSCLC cells to
IR in vitro and in vivo
Radiation caused a dose-dependent reduction in clonogenic
survival in all NSCLC lines, and we found significant variation in
intrinsic radiosensitivity (Fig. 2A-2C, left panels). The SF
(surviving Fraction) at 2 Gy (SF2) of the A549, H460, and H1299
cells were 0.84, 0.44, and 0.72, respectively. However, these SF2
values were significantly decreased to 0.44, 0.23, and 0.36 for the
A549, H460, and H1299 cells, respectively, upon treatment with
AZD5438 and IR. The DER at 10% survival was 1.5 for A549, 1.3
for H460, and 1.3 for H1299 cells. SF at different IR doses and the
corresponding DERs are listed in Supplementary Table ES1. To
further analyze the efficacy of AZD5438 as a radiosensitizer
in vivo, tumor growth delay assays were performed in athymic
nude mice (Fig. 2A-2C, right panels). Tumors were treated with
AZD5438 (po) 1 h prior to IR. For A549 xenografts (Fig. 2A),
siRNAs (200 pmol) (A-E) or scrambled siRNAs (200 pmol) using Lipofectamine 2000. After 48 h, cells were trypsinized, counted, and
plated for CFA. Remaining cells were lysed and subjected to immunoblot assay for detecting Cdks (insets). Three hours after plating, cells
were irradiated, and colonies were counted after 10 days. (A) A549 cells with Cdk1 siRNA; (B) A549 cells with Cdk2 siRNA; (C) A549
cells with Cdk9 siRNA; (D) H460 cells with Cdk1siRNA; (E) H1299 cells with Cdk1siRNA.
IR sensitization of NSCLC cells transfected with Cdk siRNAs. All 3 NSCLC cells were transfected with either Cdk-specific
Raghavan et al. International Journal of Radiation Oncology ? Biology ? Physics
a tumor volume of 800 mm3was reached in 41, 43, 46, and 55
days for the control, AZD5438 alone, IR alone, and IR plus
AZD5438 groups, respectively, which resulted in a dose
enhancement factor of 2.4. Whereas the dose enhancement factors
for H1299 and H460 xenografts were 1.8 and 0.6, respectively
(Fig. 2C and 2D, right panel). These results demonstrated that
A549 and H1299 xenografts are highly responsive to the
combined treatment of AZD5438 plus IR.
Cdk1 siRNA significantly enhanced IR sensitization
Because AZD5438 inhibits Cdk 1, 2, and 9, further analysis of the
role of each individual Cdk in IR sensitization was performed
using siRNAs. A significant decrease in the expression level of
targeted Cdks was achieved in all cell lines (Fig. 3A-3E, inset).
Cdk1 inhibition particularly caused IR sensitization in A549
(Fig. 3A) and H1299 cells (Fig. 3E). Modest sensitization was
observed in A549 cells when Cdk2 was knocked down (Fig. 3B),
whereas no effect was observed when Cdk9 was completely
ablated in A549 cells (Fig. 3C). SF and DER values upon Cdk1
inhibition are listed in Supplementary Table ES2. Interestingly,
Cdk1 knockdown did not cause IR sensitization in H460 cells
(Fig. 3D). Cdk2 and Cdk9 inhibition did not cause IR enhance-
ment in H1299 and H460 cells (data not shown). These results
indicate that the inhibition of the Cdk1 pathway by AZD5438 may
be associated with IR sensitization in A549 and H1299 cells.
AZD5438 inhibits HR repair
Several reports describe the role of Cdk1 in DNA HR repair
processes (13, 14), and a recent study with a dual kinase inhibitor,
NU6027 (ATR: ATM and Rad3-related and Cdk2), inhibited
Rad51 foci formation and blocked HR repair (15). The effect of
AZD5438 on HR repair was then studied using H1299 cells
containing an integrated DR-GFP HR reporter in which functional
GFP can only be restored by HR repair (Fig. 4A) (11). H1299 cells
were transfected with vector encoding I-SceI endonuclease to
generate a DSB, with pGFP for transfection efficiency control, and
with phCMV-1 I-Seaford negative control in the presence or
absence of AZD5438 (75 nM). Results clearly showed that
AZD5438 reduced the frequency of HR by almost 50% (Fig. 4B
and 4C). There was also a noticeable decrease in Rad51 expres-
sion after treatment with AZD5438 (Fig. 4C, right panel).
IR sensitization by AZD5438 is associated with
delayed DSB repair
To measure the induction and repair of IR-induced DSBs, the
3 NSCLC cells were exposed to AZD5438 for 24 h, followed by IR
(Fig. 5A-5C). The effect of AZD5438 treatment on gH2AX foci
(a surrogate for DNA DSB repair) formation is shown in Fig. 5A,
indicating no induction of foci formation. In all 3 cell lines, almost
complete (>95%) repair occurred within 8 h post-IR (Fig. 5B and
5C) but nearly 30%-40% of foci were retained at 8 h when treated
H460 cells, while in A549 and H1299 cells, a small number of foci
(5%-10%) remained. These results indicate that AZD5438 modu-
lates the repair kinetics of radiation-induced gH2AX foci.
AZD5438 treatment selectively induced G2-M
checkpoint arrest in NSCLC cells
The NSCLC cells showed G2 arrest at 8 h post-IR (2 Gy).
Results shown in Figure 6 indicate that approximately 50% of
were transfected with phCMV-1-I-SceI (negative control), pCMV3xnls-I-SceI (functional endonuclease), or pGFP (transfection efficiency
control), and 75 nM of AZD5438 was added as indicated. GFP signal was assayed at 3 days post-transfection with a FACSCalibur flow
cytometer. A total of 50,000 cells were scored per treatment group. (C, left and middle panels) The frequency of recombination events was
calculated from the number of GFP-positive cells divided by the number of cells analyzed following correction for transfection efficiency.
(Right panel) Immunoblot analysis of Rad51expression in H1299 cells treated with AZD5438 (75 nM) for 72 h, lysed for immunoblotting.
Volume 84 ? Number 4 ? 2012 Lung cancer and radiotherapy e511
A549 cells were blocked at G2-M after 24-h treatment of
AZD5438, whereas 32% of H1299 and 11% of H460 cells were
arrested at the same time. When these cells were treated with
AZD5438 (after 24 h) and IR (at 8 h), 68% of A549 and 49% of
H1299 cells were blocked at the G2-M checkpoint compared to
only 25% of H460 cells under similar conditions. In an addi-
tional study, apoptosis was measured at 24 and 48 h after
treatment with AZD5438 and radiation. Combined treatment
enhanced apoptosis specifically in A549 (w5-fold) and H1299
(w3-fold) cells at 48 h after treatment (see Supplemental Table
ES3).Taken together, these results clearly indicate that the
combined treatment of IR plus AZD5438 causes significant
delay in DSB repair, prolonged G2-M blockage, and enhanced
apoptosis, which may contribute to less survival.
Many lung cancers, especially NSCLC, display intrinsic radiation
resistance. Previous studies with older generation Cdk inhibitors
have shown increased radiation sensitivity (7, 8). SF and DER of
the three NSCLC cell lines used in this study changed signifi-
cantly when combined with AZD5438 treatment. Several mech-
anisms could contribute to AZD5438-mediated radiosensitizing
effects in NSCLC cells. Cdk2, it has been suggested, compensates
for lack of Cdk1 (14); however, an important finding of this study
(Fig. 3) indicates that AZD5438-mediated radiosensitivity is
induced mostly through the inhibition of Cdk1 alone and only
modestly through the Cdk2 pathway. These results are also in
3 NSCLC cell lines. (B) DNA DSB repair kinetics in NSCLC cell lines. Cells were incubated with or without AZD5438 for 24 h (210 nM,
90 nM, and 435 nM for A549, H1299 and H460, respectively), irradiated (2 Gy), immunostained for gH2AX foci, and counted for each
time point (average, 50 nuclei). (C) Repair kinetics of NSCLC cells was obtained by plotting the percentage of remaining foci against time;
D, AZD5438; R, radiation.
IR sensitization by AZD5438 was associated with delayed DSB repair in NSCLC cells. (A) Effect of AZD5438 on DNA DSB in
Raghavan et al. International Journal of Radiation Oncology ? Biology ? Physics
agreement with the important physiological roles of Cdk1 such as
(1) promotion of mitotic entry, (2) inhibition that causes G2arrest,
and (3) involvement in DNA repair through HR (5, 13). Results
shown in Figure 4 indicate that AZD5438 causes significant
reduction (50%) in HR repair in H1299 cells. Thus, the blockage
of Cdk1 may be the mechanism by which NSCLC cells are
sensitized to IR.
Generally, cells show significant variation in their radiation
sensitivity based on the cell cycle, with G2-M being the most
radiation-sensitive phase (16). Radiation-induced arrest at G2-M is
critical in preventing cell death. This study shows that in combi-
nation with radiation, AZD5438 treatment causes a greater
number of cells to be arrested at G2-M, at least in the A549 and
H1299 cell populations, which implies a larger proportion of
cancer cells are disrupted by combination therapy.
Cell cycle inhibitors, such as flavopiridol, have been shown to
induce apoptosis in a variety of cell lines (17). AZD5438 alone
enhances apoptosis nearly 3.0-fold in A549 cells, whereas
AZD5438 combined with IR increases the number of apoptotic
cells approximately fivefold in A549 cells and threefold in H1299
cells after 48 h. The smaller population of cells undergoing
apoptosis after combined treatment with AZD5438 plus IR may
indicate that other modes of cell death are involved; the exact
mechanism is currently under investigation. There are reports
demonstrating that Cdk1 is involved in several modes of cell death
including apoptosis and mitotic catastrophe (5, 18).
Although all 3 cell lines tested in this study had similar levels
of Cdk1, -2, and -9 as well as similar levels of all the other cell
cycle regulatory proteins (see Supplementary Table ES4), they
displayed divergent levels of sensitivity to AZD5438. In addition,
all 3 cell lines carried a variety of mutations (see Supplementary
Table ES5), but none was exclusive to the responding cell lines or
explained this divergence. This is also an important finding of this
study, indicating that the patient population with the H460
phenotype (metastatic large-cell carcinoma) will not receive
significant therapeutic advantage from treatment with combined
radiation and this class of cell cycle inhibitor. While Cdk inhibi-
tion may be an effective strategy for inducing radiation sensitivity,
it is clear that the differential patterns of tumor growth and
response to therapy (radiation, drug, and both) lie in the genetic
background of each cell line, which ultimately regulates thera-
peutic outcome. Therefore, patient selection is extremely impor-
tant in ensuring the highest efficacy is received when combined
treatments of radiation and Cdk inhibitors are administered.
In conclusion, AZD5438 enhances radiation-induced cell death by
blocking Cdk1 in A549 and H1299 cells. This enhanced radio-
sensitivity is associated with inhibition of DNA DSB repair
processes through HR repair. While clinical development of
AZD5438 has been discontinued due to low tolerability in phase II
studies (19), several other Cdk inhibitors such as SCH727965,
P276-00, and EM-1421, which belong to the same new generation
of inhibitors, are currently under investigation in phase I/II trials
for treatment of both solid tumors and chronic lymphocytic
leukemia (5). This preclinical study confirms that Cdk inhibitors
are potent radiation-sensitizing agents and are promising candi-
dates for clinical evaluation as part of a combined regimen.
1. Jemal A, Siegel R, Ward E, et al. Cancer statistics, 2008. CA Cancer
J Clin 2008;58:71-96.
collected 8 h post-IR. (A) Cells were stained with PI to detect distribution of cell cycle after the treatments. (B) Quantitation of the
perecentage of cells at different phases of the cell cycle in response to drug, IR, and IR plus drug.
AZD5438 exhibited G2-M checkpoint arrest in NSCLC cells. Cells were treated with AZD5438 followed by IR (2 Gy) and
Volume 84 ? Number 4 ? 2012 Lung cancer and radiotherapy e513
2. Jassem J. The role of radiotherapy in lung cancer: where is the Download full-text
evidence? Radiother Oncol 2007;83:203-213.
3. Sherr CJ. G1 phase progression: cycling on cue. Cell 1994;79:551-555.
4. Dickson MA, Schwartz GK. Development of cell-cycle inhibitors for
cancer therapy. Curr Oncol 2009;16:36-43.
5. Manchado E, Guillamot M, Malumbres M. Killing cells by targeting
mitosis. Cell Death Differ 2012;19:369-377.
6. Bible KC, Kaufmann SH. Flavopiridol: a cytotoxic flavone that
induces cell death in noncycling A549 human lung carcinoma cells.
Cancer Res 1996;56:4856-4861.
7. Kim JC, Saha D, Cao Q, et al. Enhancement of radiation effects by
combined docetaxel and flavopiridol treatment in lung cancer cells.
Radiother Oncol 2004;71:213-221.
8. Kodym E, Kodym R, Reis AE, et al. The small-molecule Cdk
inhibitor, SNS-032, enhances cellular radiosensitivity in quiescent
and hypoxic non-small-cell lung cancer cells. Lung Cancer 2009;
9. Byth KF, Thomas A, Hughes G, et al. AZD5438, a potent oral
inhibitor of cyclin-dependent kinases 1, 2, and 9, leads to pharmaco-
dynamic changes and potent antitumor effects in human tumor
xenografts. Mol Cancer Ther 2009;8:1856-1866.
10. Kong Z, Xie D, Boike T, et al. Downregulation of human DAB2IP
gene expression in prostate cancer cells results in resistance to
ionizing radiation. Cancer Res 2010;70:2829-2839.
11. Chan N, Koritzinsky M, Zhao H, et al. Chronic hypoxia decreases
synthesis of homologous recombination proteins to offset chemo-
resistance and radioresistance. Cancer Res 2008;68:605-614.
12. Kong Z, Raghavan P, Xie D, et al. Epothilone B confers radiation dose
enhancement in DAB2IP gene knock-down radioresistant prostate
cancer cells. Int J Radiat Oncol Biol Phys 2010;78:1210-1218.
13. Ira G, Pellicioli A, Balijja A, et al. DNA end resection, homologous
recombination and DNA damage checkpoint activation require CDK1.
14. Johnson N, Li YC, Walton ZE, et al. Compromised CDK1 activity
sensitizes BRCA-proficient cancers to PARP inhibition. Nat Med
15. Peasland A, Wang LZ, Rowling E, et al. Identification and evaluation
of a potent novel ATR inhibitor, NU6027, in breast and ovarian cancer
cell lines. Br J Cancer 2011;105:372-381.
16. Gorski DH, Beckett MA, Jaskowiak NT, et al. Blockage of the
vascular endothelial growth factor stress response increases the anti-
tumor effects of ionizing radiation. Cancer Res 1999;59:3374-3378.
17. Melillo G, Sausville EA, Cloud K, et al. Flavopiridol, a protein kinase
inhibitor, down-regulates hypoxic induction of vascular endothelial
growth factor expression in human monocytes. Cancer Res 1999;59:
18. Castedo M, Perfettini JL, Roumier T, et al. Cyclin-dependent kinase-1:
linking apoptosis to cell cycle and mitotic catastrophe. Cell Death
19. Boss DS, Schwartz GK, Middleton MR, et al. Safety, tolerability,
pharmacokinetics and pharmacodynamics of the oral cyclin-dependent
kinase inhibitor AZD5438 when administered at intermittent and
continuous dosing schedules in patients with advanced solid tumours.
Ann Oncol 2010;21:884-894.
Raghavan et al. International Journal of Radiation Oncology ? Biology ? Physics