Background
KRAS genotype, wild-type or mutant, addresses the medical treatment of metastatic colorectal cancer (MCRC) patients (pts), consisting of triplet regimens combining chemotherapeutic drugs, or doublets plus targeted agents [
1]. The addition of anti-epidermal growth factor receptor (anti-EGFR) treatment is not effective in
KRAS mutant patients [
2,
3]; anti-vascular endothelial growth factor (anti-VEGF) treatment added to doublet chemotherapy was effective in
KRAS wild-type and mutant pts [
4,
5]. In liver limited (L-L) MCRC, these first line options, integrated with secondary resection of liver metastases, may significantly increase survival [
6‐
13].
The prognostic relevance of the
KRAS genotype can be assessed by evaluation of clinical outcome (progression-free survival (PFS), overall survival (OS)) in wild-type and mutant pts, depending on differential tumor biological aggressiveness and predictive effectiveness of treatment strategies. The median OS of
KRAS wild-type and mutant MCRC pts treated with irinotecan, 5-fluorouracil and leucovorin (IFL) plus bevacizumab (BEV) was 27.7 and 19.9 months, respectively [
4,
5]. The hazard ratio (HR) for risk of death was 0.51 and statistically significant only when
KRAS and
BRAF wild-type pts were compared with pts harboring mutations in one gene. In
KRAS wild-type pts and in
BRAF wild-type pts compared to mutant, HR was 0.64 and 0.38, respectively, but did not reach statistical significance [
4]. Recently, phase II studies proposed by Masi
et al. [
8], and by our group [
7], showed that intensive medical treatment consisting of triplet chemotherapy plus BEV, according to FOLFOXIRI plus BEV and FIr-B/FOx schedules, respectively, may increase the activity and efficacy of the treatment in MCRC pts with the
KRAS wild-type and mutant genotypes [
8,
13]. Median OS of pts treated with FIr-B/FOx was different in
KRAS wild-type and mutant pts (38 months and 21 months, respectively), but not significantly [
13]. L-L pts compared to other/multiple metastatic (O/MM) pts achieve significantly increased PFS and OS; in addition,
KRAS wild-type pts with L-L disease may achieve a significantly greater benefit from integration with liver metastasectomies, with respect to
KRAS mutant patients [
11,
13].
The
KRAS wild-type genotype predicts favorable clinical outcomes when anti-EGFR or anti-VEGF molecules are added to doublet chemotherapy [
2,
5]. The
KRAS mutant genotype significantly predicts prolonged PFS up to 9.3 months, while there was no increase in OS and activity [
14,
5], in pts treated with BEV added to IFL compared to IFL.
KRAS mutations occur in 35% to 45% of colorectal cancer (CRC), mostly in codon 12 (80%), c.35 G > A (G12D) and c.35 G > T (G12V) transversions, representing 32.5% [
14,
15] and 22.5% [
14,
16], respectively, and codon 13, predominantly c.38 G > A (G13D) mutations [
17]. These mutations impair the intrinsic GTPase activity of KRAS, thus leading to constitutive, growth-factor-receptor independent activation of downstream signalling [
18]. In the
in vitro model proposed by Guerrero
et al. [
19], codon 12 mutations increase aggressiveness by the differential regulation of KRAS downstream pathways that lead to inhibition of apoptosis, enhanced loss of contact inhibition and increased predisposition to anchorage-independent growth [
19]. Codon 13 mutations showed reduced transforming capacity compared to codon 12 mutations [
20].
The biological aggressiveness of codon 12
KRAS mutant tumors seems to confer worse clinical behavior. A multivariate analysis suggested that the presence of
KRAS mutation significantly increased the risk of recurrence and death; the codon 12 c.35 G > T (G12V) mutation retained an independent increased risk of recurrence and death [
21], and significantly reduced disease-free survival and OS of Dukes C pts [
16]. The poorer prognosis conferred by codon 12
KRAS mutations was not confirmed in other studies [
22,
23].
We report a retrospective exploratory analysis evaluating the prognostic value of the prevalent codon 12 c.35 G > A (G12D)
KRAS mutation in MCRC pts enrolled in a previously reported phase II study [
7] and in an expanded clinical program proposing FIr-B/FOx intensive regimen as first line treatment.
Discussion
The prognostic relevance of
KRAS status, wild-type or mutant, is not significantly different in MCRC pts treated with BEV-containing chemotherapy. Reported median OS ranges from 29.9 to 38 months in
KRAS wild-type and 19.9 to 21 months in
KRAS mutant pts [
4,
5,
8,
13]. The addition of anti-EGFR or anti-VEGF molecules to doublet chemotherapy predicts a favorable clinical outcome in
KRAS wild-type pts [
2,
5]. BEV addition to IFL compared to IFL significantly predicts prolonged PFS up to 9.3 months, but not increased OS and activity, in
KRAS mutant pts [
5,
14]. BEV addition to triplet chemotherapy, according to FIr-B/FOx or FOLFOXIRI/BEV schedules, resulted in high activity and efficacy in
KRAS wild-type and mutant MCRC pts [
8,
13]. In particular,
KRAS mutant pts had an ORR of 67% and 71%, median PFS of 11 and 12.6 months, and median OS 20 months, respectively [
8,
13]. We recently reported a significantly favorable prognosis (PFS and OS) in
KRAS wild-type L-L compared to O/MM pts [
11,
13]. Conversely, in
KRAS mutant MCRC pts, median PFS and OS were not significantly affected by the extension of metastatic disease (L-L compared to O/MM) [
11,
13].
The prevalent c.35 G > A (G12D)
KRAS mutation characterizes 10.3% of CRC and represents up to 30% of
KRAS mutations [
16]. In the present evaluation, 25.4% of MCRC pts harbored the c.35 G > A
KRAS mutation and exhibited a high activity of the FIr-B/FOx intensive regimen (ORR 71%). Liver metastasectomies were performed in 13% of pts (33% of L-L disease), median PFS and OS were 9 and 14 months, respectively. In pts with the
KRAS c.35 G > A mutation, activity and PFS were not significantly different, while OS was significantly worse compared to
KRAS wild-type,
KRAS/
BRAF wild-type, and other codon 12 and 13 mutant pts. Median OS was not significantly different in other
KRAS mutant compared to wild-type pts. This is the first report of a worse prognosis in
KRAS c.35 G > A (G12D) mutant MCRC pts, treated with intensive triplet chemotherapy plus BEV.
Codon 12
KRAS mutations may increase aggressiveness by the differential regulation of KRAS downstream pathways associated with higher AKT/protein kinase B activation, bcl-2, E-catherin, β-catenin, and focal adhesion kinase overexpression, and RhoA underexpression, whereas codon 13
KRAS mutant cells show increased sensitivity associated with increased activation of the c-Jun-NH
2-terminal kinase I pathway [
19]. Several studies compared the prognostic roles of
KRAS codon 12 with codon 13 mutations in CRC. RASCAL (Kirsten Ras in CRC) studies showed that the presence of the
KRAS mutation significantly increased the risk of death by 26% [
16,
21]; the c.35 G > T (G12V) mutation, but not c.35 G > A (G12D) or c.35 G > C (G12A), represented an independent risk factor for recurrence and death and significantly increased the risk of death by 44% [
21]. It also had a significantly worse impact on failure-free survival and OS, increasing the risk of recurrence or death by 30% [
16], and up to 50% in Dukes' C cancers [
16].
KRAS codon 12 mutations (in particular, c.35 G > T) were associated with inferior survival in patients with
KRAS-wild-type/
BRAF-wild-type cancers [
29].
In MCRC pts, specific
BRAF and
KRAS mutations can confer different biological aggressiveness and effectiveness of treatment strategies; the balance between aggressiveness and effectiveness can differentiate prognosis, that is, median OS. Comparison of median OS in pts with different genotypes can discriminate this net prognostic effect. Thus, specific mutations and treatment strategies (medical regimens and secondary liver surgery, further lines of treatment) could be major parameters determining different prognoses in MCRC. The prevalent
BRAF c.1799 T > A (V600E) mutation, characterizing 4.7% to 8.7% of CRC, demonstrated a negative prognostic effect compared to
BRAF wild-type pts in MCRC pts treated with doublet chemotherapy alone or added to cetuximab, BEV and cetuximab plus BEV, with a median PFS of 5.6 to 8 months and median OS of 10.3 to 15.9 months [
4,
30,
31]. The favorable predictive effect of cetuximab or BEV addition to chemotherapy was not significantly demonstrated in
BRAF mutant MCRC pts [
4,
31,
32]. Patients with tumors harbouring the
KRAS c.35 G > T mutation and other mutations were associated with a worse outcome when receiving chemotherapy plus cetuximab, compared with chemotherapy alone [
33].
In MCRC pts pre-treated with chemotherapy alone, the
KRAS c.38 G > A mutation (G13D) confers a significantly worse prognosis [
34]. Cetuximab or cetuximab plus chemotherapy significantly predicted increased OS (median 7.6 and 10.6 months, respectively) and PFS (median 4.0 and 4.1 months, respectively) compared to other
KRAS mutations [
34], and no different outcome was found compared to
KRAS wild-type pts [
34]. Recently, a retrospective pooled analysis confirmed the favorable predictive effect of c.38 G > A
KRAS mutation in first line cetuximab-containing chemotherapy [
33]: significantly improved PFS (median, 7.4 versus 6.0 months) and tumor response (40.5% versus 22.0) but not survival (median, 15.4 versus 14.7 months). Moreover, systematic reviews and meta analyses confirmed that
KRAS c.38 G > A (G13D) mutant pts demonstrated a significantly favorable predictive effect of cetuximab-containing associations compared to other
KRAS mutant MCRC, and significantly lower ORR, with no significantly different PFS and OS compared to
KRAS wild-type pts [
35,
36]. In patients with MCRC treated with panitumumab or control therapy in first-or second-line chemorefractory settings, no consistent associations were found between tumors with specific
KRAS mutations and patient outcome. Opposite findings were reported when panitumumab was combined with first line oxaliplatin, whereas similar data were reported when it was combined with second-line FOLFIRI [
37].
Prospective studies should be developed to confirm the differential prognosis and predictive effect of chemotherapeutics and/or targeted agents in MCRC pts harboring KRAS/BRAF mutations, specifically KRAS c.35 G > A (G12D), c.35 G > T (G12V), c.38 G > A (G13D) mutations and BRAF c.1799 T > A (V600E).
Competing interests
The authors declare that they have no competing interests.
Authors' contributions
GB contributed to the conception and design of the study, in the provision of study materials of patients, in data analysis and interpretation and in the manuscript writing. ER contributed to the conception and design of the study, in data analysis and interpretation and in the manuscript writing. KC, TF, MT, EA participated in data analysis and interpretation. GB, KC, CF and ER provided clinical management and data on patients. DDG, AL, JCS provided molecular genetic analysis. All authors participated in the collection and/or assembly of data. All authors read and approved the final manuscript.