The impact of KRAS mutations on prognosis in surgically resected colorectal cancer patients with liver and lung metastases: a retrospective analysis
Kim et al. BMC Cancer
The impact of KRAS mutations on prognosis in surgically resected colorectal cancer patients with liver and lung metastases: a retrospective analysis
Hae Su Kim 0 2 3 5
Jin Seok Heo 0 1 3
Jeeyun Lee 2 3
Ji Yun Lee 2 3
Min-Young Lee 2 3
Sung Hee Lim 2 3
Woo Yong Lee 1 3
Seok Hyung Kim 3 4
Yoon Ah Park 1 3
Yong Beom Cho 1 3
Seong Hyeon Yun 1 3
Seung Tae Kim 2 3
Joon Oh Park 2 3
Ho Yeong Lim 2 3
Yong Soo Choi 3 6
Woo Il Kwon 1 3
Hee Cheol Kim 1 3
Young Suk Park 2 3
0 Equal contributors
1 Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine , Seoul , Korea
2 Division of Hematology-Oncology, Department of Medicine, Samsung Medical Center, Sungkyunkwan University School of Medicine , Seoul , Korea
3 Hae Su Kim and Jin Seok Heo are co-first author
4 Pathology, Samsung Medical Center, Sungkyunkwan University School of Medicine , Seoul , Korea
5 Division of Hematology-Oncology, Department of Medicine, Veterans Health Service Medical Center , Seoul , Korea
6 Thoracic Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine , Seoul , Korea
Background: KRAS mutations are common in colorectal cancer (CRC). The role of KRAS mutation status as a prognostic factor remains controversial, and most large population-based cohorts usually consist of patients with non-metastatic CRC. We evaluated the impact of KRAS mutations on the time to recurrence (TTR) and overall survival (OS) in patients with metastatic CRC who underwent curative surgery with perioperative chemotherapy. Methods: Patients who underwent curative resection for primary and synchronous metastases were retrospectively collected in a single institution during a 6 year period between January 2008 and June 2014. Patients with positive surgical margins, those with known BRAF mutation, or those with an unknown KRAS mutation status were excluded, and a total of 82 cases were identified. The pathological and clinical features were evaluated. Patients' outcome with KRAS mutation status for TTR and OS were investigated by univariate and multivariate analysis. Results: KRAS mutations were identified in 37.8 % of the patients and not associated with TTR or OS between KRAS wild type and KRAS mutation cohorts (log-rank p = 0.425 for TTR; log-rank p = 0.137 for OS). When patients were further subdivided into three groups according to mutation subtype (wild-type vs. KRAS codon 12 mutation vs. KRAS codon 13 mutation) or amino acid missense mutation type (G > A vs. G > T vs. G > C), there were no significant differences in TTR or OS. Mutational frequencies were significantly higher in patients with lung metastases compared with those with liver and ovary/bladder metastases (p = 0.039), however, KRAS mutation status was not associated with an increased risk of relapsed in the lung. Conclusions: KRAS mutation was not associated with TTR or OS in patients with metastatic CRC who underwent curative surgery with perioperative chemotherapy.
Colorectal cancer; KRAS mutation; Prognosis; Metastases
Colorectal cancer (CRC) is the fourth leading cause of
cancer-related death worldwide [
]. Although the
development of molecular-targeted therapy has improved the
survival of patients with metastatic CRC [
majority of patients with stage IV CRC who undergo
complete resection die from metastatic disease.
Nevertheless, a good proportion of patients demonstrate good
recurrence-free survival. CRC tumorigenesis is
characterized by the accumulation of genetic alterations, and
V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog
(KRAS) mutations are an early event in tumorigenesis
. KRAS mutations occur in approximately 30 to 40 %
of patients with CRC, and 90 % of KRAS mutations
occur in codon 12 or 13 [
2, 5, 6
]. KRAS mutations lead
to constitutive activation of downstream pathways,
including the Ras/Raf/MAP/MEK/ERK and/or PTEN/
PI3K/Akt pathways [
]. KRAS mutations are
established biomarkers for predicting the poor efficacy of
anti-epidermal growth factor receptor (EGFR)
monoclonal antibodies in patients with stage IV CRC [
2, 5, 11
but the prognostic relevance of KRAS mutations remains
]. Recent studies, in patients with
resected stage II and/or III CRC, have highlighted the
prognostic value of KRAS codon12 and 13 mutations,
showing correlations between mutation subtype, cancer
recurrence, and poor overall survival [
Large population-based cohorts usually consist of
patients with non-metastatic CRC [
12, 14, 16, 17
prognostic impact of KRAS mutation in patients with
synchronous metastatic CRC who undergo curative
resection with perioperative chemotherapy is unknown.
The current study investigated the impact of KRAS
mutations on the time to recurrence (TTR) and overall
survival (OS) in patients with stage IV CRC who underwent
curative surgery with perioperative chemotherapy. In
addition, the recurrence pattern according to KRAS
mutation status after complete resection was evaluated.
In this retrospective study, patients who underwent
curative resection for primary and synchronous
metastases at our institution between January 2008 and June
2014 were identified from the hospital records. Patients
who underwent separate colorectal resection and
metastasectomy were excluded if the duration between the
two procedures exceeded 2 months. Patients with
positive surgical margins, those with known v-Raf murine
sarcoma viral oncogene homolog B (BRAF) mutations,
or those with an unknown KRAS mutation status were
also excluded. All patients included in the study were
administered 5-FU with/without oxaliplatin or
irinotecanbased chemotherapy. Clinical and pathological data
including sex, patient age, tumor location, resection site,
staging at surgery (performed in accordance with the
classification of the 6th Edition of the American Joint
Committee on Cancer guidelines), BRAF mutation
status, perioperative chemotherapy regimens, use of
molecular targeting agents including cetuximab and
bevacizumab, were collected. The study protocol was
reviewed and approved by the SMC institutional review
Perioperative chemotherapy regimens
Oxaliplatin based chemotherapy was FOLFOX
(oxaliplatin 85 mg/m2 on day 1, infused during 2 h; LV 200 mg/
m2, infused during 2 h, followed by 5-FU as a 400 mg/
m2 intravenous bolus then a 1200 mg/m2 infusion
during 22 h on days 1 and 2) in 2 week treatment cycles or
XELOX(oxaliplatin 130 mg/m2 on day 1 followed by oral
capecitabine 1000 mg/m2 twice daily (day 1 to 14) in
3 week treatment cycles. Irinotecan based chemotherapy
was FORFIRI (irinotecan 180 mg/m2 on day 1, infused
during 2 h; LV 200 mg/m2, infused during 2 h, followed
by 5-FU as a 400 mg/m2 intravenous bolus then a
1200 mg/m2 infusion during 22 h on days 1 and 2) in
2 week treatment cycles or XELIRI (irinotecan 250 mg/
m2 on day 1 followed by oral capecitabine 1000 mg/m2
twice daily (day 1 to 14) in 3 week treatment cycles. If
bevacizumab or cetuximab was used, patients received
cetuximab (initial dose 400 mg/m2 infused during 2 h,
and 250 mg/m2 weekly) or bevacizumab (5 mg/kg)
followed by FOLFOX or FOLFIRI.
DNA extraction and mutation analysis
DNA was isolated from 10-μm formalin-fixed,
paraffinembedded tumor specimens using FFPE-DNA isolation
kit (Qiagen, Hilden, Germany). A Qiagen the rascreen
KRAS mutation kit was used to detect the seven most
common KRAS codon 12 and 13 mutations. Specifically,
the mutation was detected by real-time polymerase
chain reaction based on amplification-refractory
mutation system and Scorpion probes (Gly12Asp [GGT >
GAT] G12D, Gly12Val [GGT > GAC] G12V, Gly12Cys
[GGT > TGT] G12C, Gly12Ser [GGT > AGT] G12S,
Gly12Ala [GGT > GCT] G12A, Gly12Arg [GGT > CGT]
G12R, Gly13Asp [GGC > GAC] G13D).
Patients were subdivided into wild-type KRAS and
mutant KRAS cohorts. The primary objective was to
investigate the effect of KRAS mutation on the TTR. TTR was
defined as the time from the date of operation to the
date of local or metastatic recurrence. As of November
2014, overall survival data are not yet available for the
mutant KRAS group. Data from recurrence-free patients
were censored at the date of the last follow-up.
Fig. 1 Time to recurrence (a) and overall survival (b) according to KRAS status. KRAS mutation status had no impact on time to recurrence (p = 0.425)
and overall survival (p = 0.137)
To compare baseline characteristics, categorical
outcomes were analyzed using the chi-square test or Fisher’s
exact test. Continuous variables are presented as medians
and ranges. TTR and OS were calculated using the
Kaplan-Meier method, and data was compared using the
log-rank test. The Cox proportional hazard model was
used to assess hazard ratios (HRs) of prognostic factor. All
factors of statistical significance (p < 0.10) in univariate
analysis were included in the multivariate analysis.
Two-sided p values of <0.05 were considered as
statistically significant. All statistical analyses were performed
using the SPSS statistical software version 21 (IBM,
Armonk, NY. USA).
Between January 2008 and June 2014, 82 patients who
were diagnosed with synchronous metastatic CRC and
underwent curative resection of primary and metastatic
lesions with perioperative chemotherapy were included
in the analyses. Table 1 summarizes the patient
characteristics according to KRAS mutation status.
There was no significant difference in
clinicopathologic features between the two groups. Baseline
characteristics including age, sex, tumor location, tumor
grade, T stage, N stage, synchronous metastasectomy
site, and recurrence site were similar between the
KRAS wild type and KRAS mutation cohorts.
Regarding BRAF mutation status, all of the tested cases
(76.8 %) were BRAF wild type.
Subtype of KRAS mutations
Of 82 patients, KRAS mutations were detected in 31
(37.8 %) patients. Eighteen (58 %) patients harbored
codon 12 mutations including 9 with c.35G > A (p.G12D,
codon 12 GGT > GAT), 5 with c.35G > T (pG12V, codon
12 GGT > GTT), 2 with c.35G > C (p.G12A, codon 12
GGT > GCT), and 2 with c.34G > A (p.G12S, codon 12
GGT > AGT). For the 13 (42 %) patients with codon 13
mutations, all had the c.38G > A (p.G13D, codon 13
GGC > GAC) mutation. KRAS amino acid mutations were
also analyzed. The G > A missense mutation was the most
frequently observed mutation, followed by the G > T and
G > C mutations.
The impact of KRAS mutations on TTR and OS
The median follow-up durations were 25 months (range,
4–74) and 34 months (range, 9–63) for patients with KRAS
wild type and KRAS mutation status, respectively. During
follow-up in surviving participants, there were 57 events
for TTR analysis and 25 events for OS analysis. There were
no significant differences in survival time distributions
according to KRAS wild type and KRAS mutation status
(logrank p = 0.425 for TTR; log-rank p = 0.137 for OS, Fig. 1).
In univariate and multivariate analyses, there were no
significant differences in TTR or OS between KRAS wild type
and KRAS mutation cohorts (Tables 2, 3 and 4). When
patients were further subdivided into three groups according
to mutation subtype (wild-type vs. KRAS codon 12
mutation vs. KRAS codon 13 mutation) or amino acid missense
mutation type (G > A vs. G > T vs. G > C), there were no
significant differences in TTR or OS.
The effect of KRAS mutation status on the recurrence site
Mutational frequencies were significantly higher in
patients with lung metastases compared with those with
liver and ovary/bladder metastases (KRAS mutant: lung
9/13 [69 %], liver 18/57 [31 %], ovary/bladder 4/12
[33 %]; p = 0.039). However, KRAS mutation status was
not associated with an increased risk of relapse in the
lung, and the majority of recurrence occurred at the
previous metastasectomy sites (15/33 vs. 24/31 for KRAS
wild type vs. KRAS mutation, respectively).
The majority of studies evaluating the prognostic impact
of KRAS mutational status in CRC have been conducted
in patients with stage II/III disease. The QUASAR trial,
which mainly evaluated patients with stage II CRC,
revealed that KRAS mutations had a detrimental effect on
recurrence and OS, despite adjuvant chemotherapy [
In contrast, the CALGB 89803 and PETACC-3 trials
demonstrated that KRAS mutation status had no
significant effect on recurrence or OS in patients with stage II/
III colon cancer or CRC treated with adjuvant
]. However, conflicting findings were
reported simultaneously in two large studies conducted by
The Kirsten ras in-colorectal-cancer collaborative group,
the RASCAL and RASCAL II trials, which were
comprised of 2721 and 4268 patients, respectively [
Although the first RASCAL study reported an
association of KRAS mutations with an increased risk of
recurrence and death for patients with all stages of CRC,
recurrence in patients with Dukes’ D tumors was less
than might be expected. The RASCAL II study
concluded that there was a significant prognostic value in
failure-free survival alone in patients with Dukes’ C
cancer harboring a KRAS G12V mutation.
Abbreviations: CI confidence interval, A.A amino acid, HR hazard ratio
Factors of statistical significance (p < 0.10) in univariate analysis presented with boldface
Few studies have evaluated the relationship between
patients with stage IV disease at the time of diagnosis
and KRAS mutations [
]. Patients with metastatic
CRC with limited metastases undergo curative primary
resection with or without metastasectomy, anti-EGFR
antibody therapy, and heterogeneous chemotherapy
regimens, making it difficult to evaluate the precise
prognostic value of KRAS status in this setting. To overcome
this limitation, in this study, we included only patients
who underwent curative resection of the primary and
metastatic sites who received perioperative
chemotherapy. To our knowledge, this study is the first to report
TTR in such patients. In this homogenous cohort of
Korean patients with metastatic CRC, we observed that
KRAS mutation was not associated with TTR or OS,
which is congruent with previous studies [
Phipps et al., reported that KRAS mutations did not
differ by stage at diagnosis, and that the prognostic value
of KRAS mutations only became evident in patients with
stage I-III disease . Furthermore, Nash et al.,
reported that the prevalence of KRAS mutations did not
vary with stage, but that KRAS mutations were strong
independent predictors of survival for patients with stage
I-III CRC [
We also investigated the association KRAS mutations
with recurrence pattern in our cohort. KRAS mutations
were significantly more common in lung metastases
compared with liver and bladder/ovary metastases.
These finding were concordant with those of Tie et al.,
who observed a significantly higher prevalence of KRAS
mutations in patients with lung metastases compared
with those with liver metastases [
]. In addition, in
their study, KRAS mutations were associated with an
increased risk of lung relapse in patients with stage II/III
CRC who were enrolled on the VICTOR clinical trial
]. However, in the present study, we did not observe
recurrence-specific associations with KRAS mutation
status. The differential impact of KRAS mutations on
recurrence-specific sites according to disease stage
requires evaluation in further studies.
Limitations of the present study included the relatively
short follow-up, where the median OS was not reached
in the KRAS mutation group. Nevertheless, sufficient
TTR events occured enabling analysis of recurrence. In
addition, the BRAF mutation status was not determined
for 19 (33 %) patients, but BRAF mutations were only
detected in a small proportion of patient and were not
significantly different between KRAS wild type and
KRAS mutated patients. In addition, the small sample
size did not allow us to evaluate the impact of different
KRAS mutation subtypes.
In conclusion, KRAS mutation was not associated with
TTR or OS in curatively resected, metastatic CRC.
Further validation of these finding is needed in metastatic
CRC patients treated with curative resection in
prospective controlled trials.
The present study, to our knowledge, is the first report
on the effect of KRAS mutations on prognosis in
surgically treated CRC patients with synchronous metastases.
The most of previous studies evaluating the prognostic
impact of KRAS in CRC have been conducted in
patients with non-metastatic CRC, and the influence of
KRAS mutations on outcome is conflicting. In our study,
KRAS mutation was not associated with TTR or OS in
metastatic CRC patients who undergo curative surgery
and perioperative chemotherapy. KRAS mutation status
was also not linked to recurrence pattern. Prospective
studies will be necessary to evaluate the prognostic effect
of KRAS mutation in metastatic CRC patients.
This research is strictly retrospective and involving the
collection of existing data and records. The study
protocol was reviewed and approved consent exemptions by
the SMC institutional review board.
BRAF: v-Raf murine sarcoma viral oncogene homolog B; CIs: Confidence
intervals; CRC: Colorectal cancer; EGFR: Epidermal growth factor receptor;
KRAS: V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog; OS: Overall
survival; TTR: Time to recurrence.
The authors declare that they have no competing interests.
HSK, JL, JH conceived and designated the study; JYL, ML and SHL helped to
conceive the study and revised manuscript critically for important intellectually
content; SK reviewed the pathologic specimens; WYL, YAP, YBC and SY critically
revised the manuscript; STK, JOP and HYL helped acquisition and interpretation
of data; YSC and WIK participated in statistical analysis and interpretation of
data; HCK, YSP conceived the study, participated in the design of it and
coordination. All authors read and approved the final manuscript.
This work was supported by a grant of the Korea Health Technology R&D
Project through the Korea Health Industry Development Institute (KHIDI),
funded by the Ministry of Health & Welfare, Republic of Korea (grant number:
HI14C2750, and HI14C3418).
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