International Journal of
Molecular Sciences
Article
Right- vs. Left-Sided Metastatic Colorectal Cancer:
Differences in Tumor Biology and
Bevacizumab Efficacy
Paola Ulivi 1 , Emanuela Scarpi 2, *, Elisa Chiadini 1 , Giorgia Marisi 1 , Martina Valgiusti 3 ,
Laura Capelli 1 , Andrea Casadei Gardini 3 , Manlio Monti 3 , Silvia Ruscelli 3 ,
Giovanni Luca Frassineti 3 , Daniele Calistri 1 , Dino Amadori 3 and Alessandro Passardi 3
1
2
3
*
Biosciences Laboratory, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS,
via P. Maroncelli 40, 47014 Meldola, Italy; paola.ulivi@irst.emr.it (P.U.); chiadini.elisa@irst.emr.it (E.C.);
giorgia.marisi@irst.emr.it (G.M.); laura.capelli@irst.emr.it (L.C.); daniele.calistri@irst.emr.it (D.C.)
Unit of Biostatistics and Clinical Trials, Istituto Scientifico Romagnolo per lo Studio e la Cura dei
Tumori (IRST) IRCCS, via P. Maroncelli 40, 47014 Meldola, Italy
Department of Medical Oncology, Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST)
IRCCS, via P. Maroncelli 40, 47014 Meldola, Italy; martina.valgiusti@irst.emr.it (M.V.);
andrea.casadei@irst.emr.it (A.C.G.); manlio.monti@irst.emr.it (M.M.); silvia.ruscelli@irst.emr.it (S.R.);
giovanni.frassineti@irst.emr.it (G.L.F.); dino.amadori@irst.emr.it (D.A.);
alessandro.passardi@irst.emr.it (A.P.)
Correspondence: emanuela.scarpi@irst.emr.it; Tel.: +39-0543-739-262; Fax: +39 0543-739-290
Academic Editor: Peter J. K. Kuppen
Received: 28 April 2017; Accepted: 29 May 2017; Published: 9 June 2017
Abstract: There is evidence of a different response to treatment with regard to the primary
tumor localization (right-sided or left-sided) in patients with metastatic colorectal cancer (mCRC).
We analyzed the different outcomes and biomolecular characteristics in relation to tumor localization
in 122 of the 370 patients with metastatic colorectal cancer enrolled onto the phase III prospective
multicenter “Italian Trial in Advanced Colorectal Cancer (ITACa)”, randomized to receive first-line
chemotherapy (CT) or CT plus bevacizumab (CT + B). RAS and BRAF mutations; baseline expression
levels of circulating vascular endothelial growth factor (VEGF), endothelial nitric oxide synthase
(eNOS), cyclooxygenase-2 (COX2), ephrin type-B receptor 4 (EPHB4), hypoxia-inducible factor
1-alpha (HIF-1α), lactate dehydrogenase (LDH), and high-sensitivity C reactive protein (hs-CRP);
and inflammatory indexes such as the neutrophil-to-lymphocyte ratio, platelet-lymphocyte rate and
systemic immune-inflammation index were evaluated. Patients with right-sided tumors showed a
longer median progression-free survival in the CT + B arm than in the CT group (12.6 vs. 9.0 months,
respectively, p = 0.017). Baseline inflammatory indexes were significantly higher in left-sided tumors,
whereas eNOS and EPHB4 expression was significantly higher and BRAF mutation more frequent
in right-sided tumors. Our data suggest a greater efficacy of the CT + B combination in right-sided
mCRC, which might be attributable to the lower inflammatory status and higher expression of
pro-angiogenic factors that appear to characterize these tumors.
Keywords: metastatic colorectal cancer; bevacizumab; right-sided colon; left-sided colon
Int. J. Mol. Sci. 2017, 18, 1240; doi:10.3390/ijms18061240
www.mdpi.com/journal/ijms
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1. Introduction
Colorectal cancer is a heterogeneous disease that can develop in different parts of the colon,
with consequent differences in terms of risk factor, histological grade, tumor size and metastatic
features [1,2]. Left-sided tumors (originating in the splenic flexure, descending colon, sigmoid colon,
rectum, or one-third of the transverse colon) derive from the embryonic hindgut, whereas right-sided
tumors (originating in the appendix, cecum, ascending colon, hepatic flexure, or two-thirds of the
transverse colon) derive from the embryonic midgut [3].
The different origins consequently lead to tumors with a different gene expression and mutation
profile. In particular, right-sided tumors show a higher frequency of BRAF mutation and microsatellite
instability and more often occur in patients with a genetic predisposition to colorectal cancer (e.g.,
Lynch syndrome). Conversely, left-sided tumors are characterized by chromosomal instability and
a gene expression profile involving the activation of the epidermal growth factor receptor (EGFR)
pathway [2,4]. These differences result in different prognoses for the two tumor types, with right-sided
tumors associated with poorer patient outcome [2–4].
In addition to its prognostic relevance, there is evidence to suggest that tumor localization may
be predictive of treatment efficacy with targeted agents, especially those directed against EGFR and
vascular endothelial growth factor (VEGF) pathways [3,5–10]. Although data on this specific topic are
discordant due to the heterogeneity of the studies carried out, left-sided RAS wild type (wt) tumors
appear to be more responsive to EGFR inhibitors, possibly due to the higher frequency of BRAF
mutations in right-sided disease [2,7,9,11]. Results on the efficacy of bevacizumab (B) are even more
conflicting, some studies finding no correlation with respect to tumor position [12,13] and others,
conversely, reporting a link between the effectiveness of the monoclonal antibody and the side of the
colon affected [13,14]. Among the latter, some authors found that left-sided or rectal tumors benefited
more from B-based treatment [14], whereas others observed that the drug prolonged progression-free
(PFS) and overall survival (OS) in right- rather than left-sided tumors [13].
Our extensive research into colorectal cancer revealed a correlation between different biomarkers
involved in angiogenic and inflammatory processes and B efficacy [15–20], but we have never focused
on these different markers in relation to tumor localization. We thus decided to investigate B efficacy,
the distribution of a series of parameters involved in angiogenesis and inflammatory processes, and
RAS and BRAF mutations in relation to tumor localization in a case series of metastatic colorectal cancer
patients enrolled in the phase III multicenter, prospective, randomized “Italian Trial in Advanced
Colorectal Cancer (ITACa)” trial [21] The ITACa trial is registered on ClinicalTrials.gov (NCT01878422).
2. Results
2.1. Patient Characteristics
The clinical pathological characteristics of patients are reported in Table 1. Sixty patients were
randomized to receive chemotherapy (CT) + B and 62 to receive CT alone. In the overall case series
51 and 71 patients had right- and left-sided tumors, respectively. Patient characteristics were well
balanced between right- and left-sided tumors. However, a significantly higher percentage of G3
tumors was observed in right-sided disease (p = 0.044).
2.2. Clinical Outcome in Relation to Tumor Localization
In the overall case series, no significant differences were observed in terms of progression free
survival (PFS) or overall survival (OS) in either treatment group (Figure 1a,b and Table 2). Conversely,
among right-sided tumors, a better outcome was observed in CT + B patients compared to the CT-only
group. In particular, median PFS was 12.6 (95% CI 8.6–16.0) and 9.0 (95% CI 6.5–10.3) months in
right-sided CT + B and CT-only patients, respectively (p = 0.017) (Figure 1c and Table 2). Significance
was maintained after adjusting for CT (FOLFOX4/FOLFIRI), gender, age, ECOG performance status
and KRAS status (p = 0.049). This difference was not observed in patients with left-sided tumors
Int. J. Mol. Sci. 2017, 18, 1240
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(Figure 1e and Table 2). Similar, albeit not significant, results were observed with regard to median
OS: 27.5 (95% CI 15.9–35.7) months for CT + B and 20.4 (95% CI 13.8–26.4) months for CT only in the
group with right-sided tumors (p = 0.380) (Figure 1d and Table 2). An inverse trend was observed
for patients with left-sided tumors (median OS 19.7 (95% confidence interval (CI) 12.7–27.1) and 27.1
(18.2–36.6) months in CT + B and CT-only patients, respectively, p = 0.194) (Figure 1f and Table 2).
Overall, no differences in terms of PFS or OS were observed between right-sided and left-sided tumors,
whereas, within the CT + B group, right-sided tumors were associated with higher, albeit not significant,
median PFS and OS (12.6 (8.6–16.0) and 27.5 (15.9–37.7) months, respectively) than left-sided disease
(9.1 (6.8–10.9) and 19.7 (12.7–27.1) months, respectively) (p = 0.069 and p = 0.270, respectively).
Table 1. Patient characteristics.
Total (n = 122)
Patient characteristics
Right-sided (n = 51) No. (%)
Left-sided (n = 71) No. (%)
Median age, years (range)
68 (37–83)
63 (34–82)
Gender
Male
Female
28 (54.9)
23 (45.1)
42 (59.1)
29 (40.9)
Performance Status (ECOG)
0
1+2
43 (84.3)
8 (15.7)
57 (80.3)
14 (19.7)
Stage at diagnosis
I–III
IV
8 (15.7)
43 (84.3)
20 (28.2)
51 (71.8)
Grading
1+2
3
22 (48.9)
23 (51.1)
43 (68.2)
20 (31.7)
51 (100.0)
0
68 (95.8)
3 (4.2)
31 (60.8)
20 (39.2)
44 (62.0)
27 (38.0)
Prior cancer therapy
Surgery
Radiotherapy
Adjuvant CT
45 (88.2)
0
10 (19.6)
50 (70.4)
12 (16.9)
8 (11.3)
Treatment group
CT + B
CT
26 (51.0)
25 (49.0)
34 (47.9)
37 (52.1)
Histological type
Adenocarcinoma NOS
Mucinous cancer
CT regimen
FOLFOX4
FOLFIRI
CT, chemotherapy; B, bevacizumab; NOS, not otherwise specified.
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Overall
a
1.00
1.00
0.80
0.80
p = 0.435
p = 0.601
CT
0.60
OS
PFS
0.60
b
CT + B
0.40
0.40
CT + B
CT
0.20
0.20
0.00
0.00
0
6
12
18
24
0
30
6
12
18
24
months
months
Months
c
1.00
30
36
42
48
Months
Right-sided tumors
d
1.00
0.80
0.80
p = 0.017
p = 0.380
0.60
OS
PFS
0.60
CT + B
0.40
CT + B
CT
0.40
0.20
CT
0.20
0.00
0.00
0
6
12
18
24
30
0
Months
6
12
18
e
30
36
42
48
42
48
Months
Left-sided tumors
f
1.00
24
1.00
0.80
0.80
0.60
p = 0.194
0.60
OS
PFS
p = 0.458
0.40
CT
CT + B
0.40
CT
0.20
0.20
CT + B
0.00
0.00
0
6
12
18
Months
24
30
0
6
12
18
24
30
36
Months
Figure 1. Progression-free (PFS) and overall survival (OS) of mCRC patients treated with CT + B or
CT alone: in the overall population (a,b); in those with right-sided tumors (c,d); and in those with
left-sided tumors (e,f).
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Table 2. PFS and OS in relation to tumor localization in the two treatment groups (CT + B and CT).
PFS
OS
No.
Events
Median OS
(months)
(95% CI)
No.
Patients
No.
Events
Median PFS
(months)
(95% CI)
Overall
CT + B
CT
60
62
53
56
9.6 (8.3–12.4)
9.1 (8.3–10.0)
0.435
49
49
21.4 (14.4–28.8)
23.2 (18.2–28.0)
0.601
Right-sided
CT + B
CT
26
25
23
24
12.6 (8.6–16.0)
9.0 (6.5–10.3)
0.017
21
20
27.5 (15.9–35.7)
20.4 (13.8–26.4)
0.380
Left-sided
CT + B
CT
34
37
30
32
9.1 (6.8–10.9)
9.1 (7.2–13.0)
0.458
28
29
19.7 (12.7–27.1)
27.1 (18.2–36.6)
0.194
p
p
PFS, progression-free survival; OS, overall survival; CT, chemotherapy; B, bevacizumab.
2.3. Circulating Biomarkers in Right-Aided and Left-Sided Tumors
Overall, higher baseline circulating expression levels of ephrin type-B receptor 4 (EPHB4) and
endothelial nitric oxide synthase (eNOS) were observed in right-sided tumors than in left-sided ones.
In particular, median relative expression levels of EPHB4 and eNOS were 3.57 (range 0.68–69.55)
and 7.14 (range 0.59–123.16), respectively, in right-sided tumors and 2.67 (range 0.21–129.51) and
5.09 (0.41–118.15), respectively, in left-sided lesions (p = 0.027 and p = 0.036, respectively) (Table 3).
No significant differences were observed for the other biomarkers (Table 3).
Table 3. Median baseline values of circulating biomarkers in relation to tumor localization.
Biomarker
VEGF
COX
HIF1-α
EPHB4
eNOS
Right-Sided
Left-Sided
Median Value (Range)
Median Value (Range)
2.36 (0.68–37.69)
1.37 (0.34–6.07)
1.17 (0.28–4.23)
3.57 (0.68–69.55)
7.14 (0.59–123.16)
2.22 (0.54–50.80)
1.12 (0.37–4.78)
1.07 (0.34–5.38)
2.67 (0.21–129.51)
5.09 (0.41–118.15)
p
0.194
0.067
0.358
0.027
0.036
Significantly lower levels of inflammatory indexes were observed in right-sided compared
to left-sided tumors (Table 4). Applying the chosen cut-off for each inflammatory index,
neutrophil-to-lymphocyte ratio (NLR) <3 and systemic immune-inflammation index (SII) <730 were
more frequently observed in patients with right- vs. left- sided colon cancers (66.7% vs. 44.3%,
and 58.8% vs. 40.0%, respectively) (p = 0.015 and 0.041, respectively). Platelet-to-lymphocyte ratio
(PLR) < 169 was 52.9% and 37.1% in right- and left-sided tumors, respectively (p = 0.085). NLR,
PLR and SII median values were 2.37 (range 0.90–10.73), 161.76 (range 64.73–310.45) and 641.47
(range 175.42–3614.67), respectively, in right-sided tumors, and 3.19 (range 0.78–12.32), 192.07 (range
38.11–909.72) and 876.77 (range 140.36–8069.24), respectively, in left-sided lesions (p = 0.003, p = 0.020
and p = 0.005, respectively). No differences were seen in high-sensitivity C-reactive protein (hs-CRP)
or lactate dehydrogenase (LDH) levels between right- and left-sided tumors (Table 4).
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Table 4. Systemic inflammatory biomarkers in relation to tumor localization.
Total (n = 122)
Biomarker
p
Right-Sided (n = 51) No. (%)
Left-Sided (n = 71) No. (%)
NLR
<3
≥3
34 (66.7)
17 (33.3)
31 (44.3)
39 (55.7)
0.015
PLR
<169
≥169
27 (52.9)
24 (47.1)
26 (37.1)
44 (62.9)
0.085
SII
<730
≥730
30 (58.8)
21 (41.2)
28 (40.0)
42 (60.0)
0.041
hs-PCR
<13.1
≥13.1
29 (58.0)
21 (42.0)
39 (60.0)
26 (40.0)
0.829
LDH
≤UNL
>UNL
16 (31.4)
35 (68.6)
21 (29.6)
50 (70.4)
0.832
NLR, neutrophil-to-lymphocyte ratio; PLR, platelet-lymphocyte ratio; SII, systemic immune-inflammation index;
hs-PCR, high-sensitivity C-reactive protein; LDH, lactate dehydrogenase; UNL, upper normal limit.
2.4. eNOS and VEGF Polymorphism Distribution in Relation to Tumor Localization
No differences were seen in eNOS and VEGF polymorphism distribution with respect to rightand left-sided tumor localization. With regard to eNOS haplotype analysis, the haplotype combination
Haplo1/Haplo1 + Haplo2/Haplo2 was found to be more highly represented in right-sided lesions
(48.1% vs. 31.0% in left-sided tumors (p = 0.050) (data not shown).
2.5. RAS and BRAF Mutation Profile
A higher percentage of BRAF mutations were observed in right-sided than left-sided tumors
(15.7% and 2.8%, respectively) (p = 0.017). No significant differences were observed with regard to
KRAS or NRAS mutations in relation to tumor localization (Table 5).
Table 5. Gene mutation in relation to tumor localization.
Gene
Total (n = 122)
p
Patient Characteristics
Right-Sided (n = 51) No. (%)
Left-Sided (n = 71) No. (%)
KRAS
Wild type
Mutated
29 (56.9)
22 (43.1)
47 (66.2)
24 (33.8)
0.296
BRAF
Wild type
Mutated
43 (84.3)
8 (15.7)
69 (97.2)
2 (2.8)
0.017
NRAS
Wild type
Mutated
48 (94.1)
3 (5.9)
70 (98.6)
1 (1.4)
0.307
3. Discussion
The ITACa trial, a prospective multicenter randomized phase III study whose aim was to evaluate
the improvement in PFS obtained by adding B to CT with respect to CT alone, reported similar PFS and
OS in the CT and CT + B treatment arms [21]. In the present study, we confirmed the lack of difference
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in survival between the CT + B and CT-only groups in a subgroup of ITACa patients. Of note, however,
a benefit from the addition of B to CT was only seen in right-sided tumors, in agreement with results
from the CALGB/SWOG 80405 trial in which treatment with cetuximab led to a clear improvement
in PFS and OS in patients with left-sided KRAS wt tumors, while B significantly improved survival
parameters in those with right-sided lesions [13]. In our study the benefit from B in right-sided tumors
was evident for both PFS and OS but was only significant for PFS. However, given that ITACa trial
patients who received first-line CT alone went on to receive second-line CT + B, it can be hypothesized
that OS may have been impacted by the subsequent lines of therapy.
The distinct molecular features characterizing right- and left-sided tumors account for their
different sensitivity to targeted drugs. First, the higher frequency of BRAF and RAS mutations
found in right-sided tumors may explain the higher activity of EGFR inhibitors in left-sided lesions
as these alterations represent well known mechanisms of resistance to the drug. However, EGFR
inhibitor activity has been shown to be higher in left-sided tumors even when all RAS wt patients are
considered [3,13], suggesting that other molecular mechanisms such as higher EGFR ligand expression
(epiregulin and amphiregulin) might be involved in left-sided lesions [2,22]. The reported higher
frequency of BRAF mutations in right-sided compared to left-sided tumors, also confirmed in this study,
may partially explain the lower responsiveness of right-sided lesions to EGFR inhibitors. Another
important factor is histological type. Mucinous cancer is a CRC subtype more frequently found
in females and in the right colon. Although this tumor has been associated with poor outcome,
its real clinical importance, especially with regard to its response to targeted agents, needs further
investigation. There were only three cases of mucinous cancer in our patient population, surprisingly
all left-sided. We do not believe this aspect could have influenced clinical results.
Our previous studies focusing on identifying biomarkers predictive of B efficacy indicated that a
series of parameters might be associated with different activity of the drug. In particular, we found
that specific eNOS polymorphisms were correlated with significantly higher PFS and OS in the CT
+ B group [15]. Interestingly, the present study revealed that eNOS polymorphisms associated with
higher B efficacy were more frequent in right-sided tumors, reflecting their higher sensitivity to
the drug. However, this result was of borderline significance and the biological reason why these
polymorphisms were more frequently associated with right-sided tumors remains to be clarified in
larger patient population.
We observed a significant difference between right- and left-sided tumors in terms of baseline
inflammatory index values. In particular, higher levels of SII, NLR and PLR were found in patients
left-sided tumors with respect to right-sided ones. Our previous results showed that, in the overall
ITACa case series, patients with low systemic inflammatory indexes (especially low NLR) benefited the
most from the addition of B to CT in terms of PFS, suggesting that low systemic inflammatory indexes
are associated with an increase in B activity [18]. These results may help to explain the higher activity
of B in right-sided tumors in which lower values of SII, NLR and PLR were present. Conversely, we
found that circulating levels of biomarkers associated with angiogenesis, i.e., eNOS and EPHB4, were
higher in right-sided tumors with respect to left-sided ones, reflecting more marked angiogenesis that
may correlate with a greater B efficacy.
It has been demonstrated that tumors with microsatellite instability (MSI), known to be more
frequent in right-sided disease, are associated with a higher cytotoxic T-cell infiltration and higher
microvessel density, suggesting a higher angiogenic capacity of tumors with this localization [23].
MSI-high CRC with long interspersed nucleotide element-1 (LINE-1) hypomethylation has been seen
to have a poor prognosis [24,25], suggesting a complex biological interaction between MSI and LINE-1
hypomethylation. Moreover, differences in the mucosal microbiota of patients who develop rightor left-sided colorectal cancer may also help to explain the different angiogenic and inflammatory
properties of the two types of lesions [26]. These findings attest to the better outcome of patients with
right-sided tumors treated with B-based treatment.
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4. Materials and Methods
4.1. Case Series
The ITACa protocol was approved by the Local Ethics Committee (Comitato Etico Area Vasta
Romagna e I.R.S.T. no. 674) on 19 September 2007, and informed consent was obtained from all
patients before blood samples were obtained for genotype testing. Participation in the ITACa biological
study was not mandatory for those taking part in the clinical trial. Of the 376 patients with mCRC
enrolled onto the ITACa trial, 122 had sufficient archived biological material to be considered for this
secondary analysis. Patients were randomized to receive first-line CT (FOLFOX4 or FOLFIRI) only or
CT plus B (CT + B). FOLFOX4 consisted of oxaliplatin 85 mg/m2 as a two-hour infusion on day 1 and
leucovorin 100 mg/m2 as a 2-h infusion followed by bolus 5-FU 400 mg/m2 and a 22-h infusion of 5-FU
600 mg/m2 on days 1–2 every two weeks. FOLFIRI consisted of the same 5-FU + leucovorin regimen
with the addition of irinotecan 180 mg/m2 as a 90-min infusion on day 1. B was administered as a
30- to 90-min intravenous infusion at a dose of 5 mg/kg on day 1 of each two-week cycle. Treatment
was to be continued until progressive disease (PD), withdrawal of consent or unacceptable toxicity,
whichever came first. Tumor assessment tests were performed within 28 days of starting the study
treatment and repeated every eight weeks during treatment until PD. All patients were evaluated
for response (according to Response Evaluation Criteria in Solid Tumors (RECIST) guidelines), PFS
and OS.
The study was performed in accordance with the principles of Good Clinical Practice and the
ethical standards of the Declaration of Helsinki.
4.2. Biomarker Analysis
Laboratory staff blinded to patient outcome performed expression analyses of VEGF-A,
cyclooxygenase-2 (COX2), hypoxia inducible factor 1 α (HIF-1α), EPHB4 and eNOS on total RNA
extracted from blood collected in PAX-Gene blood RNA kit (PreAnalytix-Qiagen, Hilden, Germany)
before the start of therapy, as previously described [20]. hs-CRP and LDH levels were evaluated on
serum collected at baseline, as previously reported [16,17].
Information on neutrophil, lymphocyte and platelet counts from blood tests carried out at
baseline (before systemic treatment) was collected. SII was calculated as platelet count × neutrophil
count/lymphocyte count, NLR was obtained by dividing the absolute neutrophil count by the absolute
lymphocyte count, and PLR was calculated by dividing the absolute platelet count by the absolute
lymphocyte count [18]. Genotyping analysis of 5 single nucleotide polymorphisms (SNPs) (VEGF
−2578C>A, −1498C>T, −1154G>A, −634C>G, +936C>T) for VEGF and 2 SNPs (eNOS −786T>C,
+894G>T) and one variable number tandem repeat (VNTR) of 27 nucleotides for eNOS was performed
on peripheral blood samples, as previously reported [15]. Exons 2, 3 and 4 of KRAS and NRAS and
exon 15 of BRAF genes were also analyzed by pyrosequencing, as previously described [27,28].
4.3. Statistical Analysis
The objectives of this secondary analysis were to examine the efficacy of B in the ITACa population
on the basis of tumor localization and to study the distribution of a series of parameters involved in
angiogenesis and in inflammatory processes, together with RAS and BRAF mutations. The data cut-off
for the analysis was 31 December 2013 when the median duration of follow-up was 36 months (range
1–65). The primary aim of the ITACa study was PFS and the secondary endpoint included OS. PFS
was defined as the time from random assignment to the first documentation of progressive disease or
death from any cause or last tumor evaluation. Patients undergoing curative metastasectomy were
censored at the time of surgery. OS was defined as the time interval between random assignment and
death or last follow-up visit. PFS and OS were estimated by the Kaplan–Meier method and curves
were compared by the logrank test (at a significance level of 5%). Nonparametric tests (Wilcoxon) were
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used to examine the potential correlation between median baseline circulating biomarker levels and
tumor localization.
Receiver Operating Characteristic (ROC) curve analysis was performed to determine the best
threshold of hs-PCR levels, with hs-PCR ≥ 13.1 considered as elevated [17]. We distinguished 2 patient
subgroups based on LDH levels at baseline: low LDH if within or below the normal range and high
LDH if above the upper limit of the normal range [16]. X-tile 3.6.1 software (Yale University, New
Haven, CT, USA) was used for bioinformatics analysis of baseline data to determine the cut-off value
for pre-treatment levels of each immune-inflammation index. SII ≥ 730, NLR ≥ 3 and PLR ≥ 169 were
considered elevated, as previously reported [18]. The association between baseline biomarker levels
and tumor localization was evaluated using the Chi-square test. All p values were based on two-sided
testing. Statistical analyses were performed using SAS statistical software version 9.4 (SAS Inc., Cary,
NC, USA).
5. Conclusions
Our data suggest a higher benefit of adding B to CT in right-sided mCRC compared to left-sided
disease. The efficacy of B may be attributable to a lower systemic inflammatory status and a higher
expression of pro-angiogenic factors, both of which appear to characterize patients with right-sided
tumors. A prospective validation of these data in a larger patient population is warranted.
Acknowledgments: The authors wish to thank Cristiano Verna for editorial assistance. This work was partially
supported by the Italian Medicines Agency (AIFA) (grant number FARM6FJJAY) and Roche S.p.A. The study
sponsors were not involved in the study design or in the collection, analysis, and interpretation of data. The study
sponsors did not provide writing support for the report. All authors had full access to all the data in the study.
The corresponding author had the final responsibility to submit for publication.
Author Contributions: Paola Ulivi and Alessandro Passardi conceived and designed the experiments; Alessandro
Passardi, Martina Valgiusti, Andrea Casadei Gardini, Manlio Monti, Silvia Ruscelli and Giovanni Luca Frassineti
were responsible for data collection; Elisa Chiadini, Laura Capelli and Giorgia Marisi performed the experiments;
Elisa Chiadini, Emanuela Scarpi, Paola Ulivi and Alessandro Passardi analyzed the data. Paola Ulivi, Alessandro
Passardi and Emanuela Scarpi wrote the manuscript; and Dino Amadori and Daniele Calistri revised the
manuscript critically for important intellectual content. All authors read and approved the final version
for submission.
Conflicts of Interest: The authors declare no conflict of interest.
Abbreviations
mCRC
CT
B
VEGF
eNOS
COX2
EPHB4
HIF-1
LDH
hs-CRP
NLR
PLR
SII
PFS
OS
Metastatic colorectal cancer
Chemotherapy
Bevacizumab
Vascular endothelial growth factor
Endothelial nitric oxide synthase
Cyclooxygenase-2
Ephrin type-B receptor 4
Hypoxia-inducible factor 1
Lactate dehydrogenase
High-sensitivity C-reactive protein
Neutrophil-lymphocyte ratio
Platelet-lymphocyte ratio
Systemic immune-inflammation index
Progression-free survival
Overall survival
Int. J. Mol. Sci. 2017, 18, 1240
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