Nutrition and Cancer

ISSN: 0163-5581 (Print) 1532-7914 (Online) Journal homepage: https://www.tandfonline.com/loi/hnuc20

Are Ergothioneine Levels in Blood Associated

with Chronic Peripheral Neuropathy in Colorectal
Cancer Patients Who Underwent Chemotherapy?

Renate M. Winkels, Lieve van Brakel, Harm van Baar, Robert B. Beelman,
Fränzel J. B. van Duijnhoven, Anne Geijsen, Henk K. van Halteren, Bibi M.
E. Hansson, John P. Richie, Dongxiao Sun, Evertine Wesselink, Moniek van
Zutphen, Ellen Kampman & Dieuwertje E. Kok

To cite this article: Renate M. Winkels, Lieve van Brakel, Harm van Baar, Robert B. Beelman,
Fränzel J. B. van Duijnhoven, Anne Geijsen, Henk K. van Halteren, Bibi M. E. Hansson, John P.
Richie, Dongxiao Sun, Evertine Wesselink, Moniek van Zutphen, Ellen Kampman & Dieuwertje
E. Kok (2020) Are Ergothioneine Levels in Blood Associated with Chronic Peripheral Neuropathy
in Colorectal Cancer Patients Who Underwent Chemotherapy?, Nutrition and Cancer, 72:3,
451-459, DOI: 10.1080/01635581.2019.1637005

To link to this article: https://doi.org/10.1080/01635581.2019.1637005

© 2018 The Author(s). Published with Published online: 12 Jul 2019.

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NUTRITION AND CANCER
2020, VOL. 72, NO. 3, 451–459
https://doi.org/10.1080/01635581.2019.1637005

Are Ergothioneine Levels in Blood Associated with Chronic Peripheral

Neuropathy in Colorectal Cancer Patients Who Underwent Chemotherapy?
Renate M. Winkelsa , Lieve van Brakela,b, Harm van Baarb, Robert B. Beelmanc, Fr€anzel J. B. van
Duijnhovenb, Anne Geijsenb, Henk K. van Halterend, Bibi M. E. Hanssone, John P. Richiea, Dongxiao Sunf,
Evertine Wesselinkb, Moniek van Zutphenb, Ellen Kampmanb , and Dieuwertje E. Kokb
a
Department of Public Health Sciences, Penn State College of Medicine, Hershey, PA, USA; bDivision of Human Nutrition and Health,
Wageningen University and Research, Wageningen, The Netherlands; cDepartment of Food Science, Penn State College of Agricultural
Sciences, University Park, PA, USA; dInternal Medicine, ADRZ Medical Center, Goes, The Netherlands; eDepartment of Surgery, Canisius
Wilhelmina Ziekenhuis, Nijmegen, The Netherlands; fDepartment of Pharmacology, Penn State College of Medicine, Hershey, PA, USA

ABSTRACT ARTICLE HISTORY

Objective: Chronic Chemotherapy-Induced Peripheral Neuropathy (CIPN) is highly prevalent Received 22 February 2019
among colorectal cancer (CRC) patients. Ergothioneine (ET) – a dietary antioxidant -pro- Accepted 16 June 2019
tected against CIPN in experimental models, but human studies are lacking. We explored
whether whole blood ET levels were associated with chronic peripheral neuropathy among
CRC patients who had completed chemotherapy.
Methods: At diagnosis, median ET-concentration in whole blood of 159 CRC patients was
10.2 lg/ml (7.2–15.8). Patients completed questionnaires on peripheral neuropathy 6 months
after completion of chemotherapy. We calculated prevalence ratios (PR) to assess
associations of ET-concentrations and prevalence of peripheral neuropathy and used linear
regression to assess associations with severity of peripheral neuropathy.
Results: Prevalence of total and sensory peripheral neuropathy were both 81%. Higher
ET-concentrations tended to be associated with lower prevalence of total and sensory
peripheral neuropathy, but not statistically significant (highest versus lowest tertile of ET: PR
¼ 0.93(0.78, 1.11) for total neuropathy, and PR ¼ 0.84(0.70, 1.02) for sensory neuropathy).
ET-concentrations were not associated with severity of neuropathy.
Conclusion: Statistically significant associations were not observed, possibly because of lim-
ited sample size. Although data may putatively suggest higher levels of ET to be associated
with a lower prevalence of neuropathy, analyses should be repeated in larger populations
with larger variability in ET-concentrations.

Introduction
The treatment of choice for colorectal cancer (CRC)
is mostly surgery, combined with (neo) adjuvant
chemotherapy and/or radiotherapy depending on the
age of the patient, disease stage, and tumor location
(1). Chemotherapy can have severe direct and con-
tinued adverse effects (2). One common and severe
adverse effect reported by CRC patients is
Chemotherapy-Induced Peripheral Neuropathy
(CIPN). During treatment, CIPN is a major reason
to modify treatment intensity (3). CIPN regularly
continues after discontinuation of chemotherapy, and
can become chronic and irreversible (4). Previous
studies support that about one-third of patients
suffer from chronic and persistent CIPN 6 months
after the end of chemotherapy (5). CIPN is predom-
inantly characterized by sensory symptoms such as
numbness and tingling of hands and feet, pain, and
aberrant temperature sensation (3,6). Platinum-
derived chemotherapeutic agents used in CRC treat-
ment, such as oxaliplatin, are frequently causing
CIPN (4).
Strategies to prevent or reduce chronic CIPN are
needed, since CIPN strongly affects quality of life
(5,7). Ergothioneine (ET) has been suggested as a
potential dietary factor that may impact CIPN based
on experimental studies (8–10). ET is a sulfur-con-
taining amino acid (11), and a potent antioxidant

CONTACT Renate M. Winkels rwinkels@phs.psu.edu Department of Public Health Sciences, Penn State College of Medicine, 400 University Drive,
Hershey, PA 17033, USA.
ß 2018 The Author(s). Published with license by Taylor & Francis Group, LLC.
This is an Open Access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives License (http://creativecommons.org/licenses/by-
nc-nd/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited, and is not altered, transformed,
or built upon in any way
452 R. M. WINKELS ET AL.
produced only by fungi and some cyanobacteria (12).
Only some foods contain ET, of which mushrooms
are the most predominant source (12,13); other foods
containing ET are organ meat as kidney and liver, red
and black beans, and oat bran (13). ET is found in
the human body with relatively high concentrations in
red blood cells (11). The specific role of ET for
human health has yet to be fully elucidated, but it has
been proposed that ET may serve as an adaptive anti-
oxidant to protect injured tissues (14).
ET has been suggested to have neuroprotective
effects against toxicity of platinum-derived chemother-
apeutics in several experimental models (8–10), but
this has not been explored in humans. Based on
rodent studies, it has been suggested that ET may
inhibit accumulation of oxaliplatin in dorsal root gan-
glion neurons (10). The uptake of ET is thought to be
regulated through the Organic Cation/Carnitine
Transporter (OCTN1)) (15); interestingly, through
rodent studies it has been suggested that the uptake of
oxaliplatin by dorsal root ganglion neurons may occur
via this same transporter OCTN1 (8,16). As accumu-
lation of platinum in dorsal root ganglia has been
reported as a major determinant of neurotoxicity,
inhibition of oxaliplatin accumulation by ET may par-
tially prevent CIPN-related symptoms (10,17).
Moreover, ET acts as a potent antioxidant (11) and
hence, it has been suggested that ET mediates its
potential preventive effects on CIPN through reduc-
tion of oxidative stress in dorsal root ganglion
neurons (10).
The aim of this study was to assess whether ET lev-
els in blood were associated with chronic peripheral
neuropathy in CRC patients who had been treated
with chemotherapy.
Methods
Design and Population
We used data of the ongoing COLON study for the
current analysis (18). Briefly, for the current study we
included newly diagnosed CRC patients who were
recruited in 11 hospitals in The Netherlands between
2010 and 2017. Participants had to be fluent in Dutch,
could not have a history of CRC, could not have had
a partial bowel resection, hereditary CRC, chronic
inflammatory bowel disease, and should be able to fill
out surveys to be eligible for the study. Eligible
patients received an information leaflet from their
physician or nurse practitioner during a routine visit
shortly after being diagnosed with CRC. All partici-
pants provided written informed consent. The study
was approved by a medical ethical review board. For
the current study, we selected those participants of the
COLON study who planned to receive chemotherapy
(n ¼ 441). Of this group: n ¼ 196 participants had to
be excluded because ‘Quality of Life Questionnaire to
assess chronic Chemotherapy-induced Peripheral
Neuropathy’ (QLQ-CIPN) data were missing (mainly
because this survey was not administered in the
COLON study from the start); n ¼ 61 participants had
to be excluded because blood samples were missing;
n ¼ 25 participants had to be excluded because they
did not receive chemotherapy. This resulted in a total
study population of n ¼ 159 participants (see
Figure 1).
Measurements
Nonfasted whole blood samples were drawn in EDTA
tubes from all participants shortly after diagnosis, but
before the start of any chemotherapy, and stored at
80  C until further processing.
Twelve months after diagnosis, participants com-
pleted the QLQ-CIPN20 (19). We used 16 items of
this 20-item survey, as previous studies have shown
that those 16 items accurately address the prevalence
and severity of peripheral neuropathy and are most
clinically relevant (19). Of those 16 items, eight items
address sensory symptoms, and eight items address
motor symptoms. Validity and clinical relevance of
these 16 items of the QLQ-CIPN20 have been
reported previously (19). In the survey, participants
were asked to rate each item on a 4-point Likert scale
summing up to a total score of 8–32 for the eight
items on sensory neuropathy and to a total score of
16–64 for all 16 items. As recommended for this sur-
vey (19), both scores were then linearly transformed
to a 0–100 scale. A higher score reflects more severe
symptoms. We used the data of total peripheral neur-
opathy based on the 16 items, and the data of the sen-
sory subscore based on the eight items on sensory
neuropathy, since CIPN is mainly characterized by
sensory symptoms (20–22). Cutoff points based on
mean scores of the Dutch population were used to
determine which participants suffered from chronic
neuropathy: >3.6 for the total score and >3.2 for the
sensory subscore (23).
ET was analyzed in whole blood samples at the
Core lab of the Pennsylvania State University, College
of Medicine in Hershey, using the following method.
Labeled ET (ET-d9) was spiked into whole blood sam-
ples as internal standard. Acetonitrile was added to
the samples to precipitate proteins; all blood samples

Figure 1. Flow chart of colorectal cancer patients participating in a prospective study on ergothioneine levels in blood and
chemotherapy-induced peripheral neuropathy.
were diluted 100 times. Then, samples were mixed
thoroughly and centrifuged at 4  C for 10 min at
14,000 rpm. After centrifuging, the supernatant was
loaded onto the ultra-performance liquid chromatog-
raphy tandem mass spectrometer (UPLC/MS/MS) sys-
tem. A standard curve for ET was created, which had
a linear range of 5–1,000 ng/ml. An ABSciex 4000 Q
Trap mass spectrometer coupled with a Waters
Acquity UPLC separation system was used to analyze
ET. ET was separated from impurities on a 1.7 mm
Acquity UPLC ethylene bridged hybrid C18 analytical
column (2.1  100 mm, Waters, Ireland). Mobile phase
A consisted of 0.1% formic acid in water and mobile
phase B consisted of acetonitrile. The ABSciex 4000 Q
Trap mass spectrometer was equipped with an electro-
spray ionization probe operated in positive mode. The
multiple reaction monitoring mode was used to ana-
lyze and quantify ET and ET-d9, with transitions of a
mass over charge (m/z) of 230.3 to 127.3 for ET and
m/z of 239.3 to 127.3 for ET-d9. All peaks were inte-
grated and quantified by ABSciex Multiquan
3.0 software.
In addition, participants completed surveys to
report personal and demographic factors around time
of diagnosis, including a 204-item semi-quantitative
food frequency questionnaire (FFQ) developed by the
NUTRITION AND CANCER 453
Division of Human Nutrition and Health of
Wageningen University & Research, The Netherlands,
to assess habitual dietary intake during the previous
month. This FFQ has been validated for several
nutrients (24,25), and has been shown to capture
energy intake reasonably well (26).
Statistics
The association of ET-concentration in blood and
peripheral neuropathy prevalence (yes/no) was
assessed in two ways: (1) with ET as a continuous
variable, and (2) with ET categorized into tertiles. In
the first analysis, we assessed associations between
ET-concentrations and the prevalence of total or
sensory peripheral neuropathy; ET-concentrations
were natural log-transformed to approach normal
distributions. In the second analysis, we created tertiles
of ET-concentrations in blood, and assessed whether
there was an association between tertile of ET and
prevalence of total or sensory peripheral neuropathy.
We used Cox regression models with a constant time
period to determine prevalence ratios (PRs), and used
robust variance estimation in the estimation confidence
intervals. PRs were used and not odds ratios, since
454 R. M. WINKELS ET AL.

Table 1. Baseline description of the colorectal cancer patients in a study on ergothioneine (ET) levels and peripheral neuropathy.
Total Tertile 1 of ET- Tertile 2 of ET- Tertile 3 of ET-
population (n ¼ 159) concentration (n ¼ 53) concentration (n ¼ 53) concentration (n ¼ 53)
ET-concentration in whole 10.2 (7.6–15.8) 5.8 (4.5–7.6) 10.2 (9.4–11.1) 19.4 (16.0–23.0)
blood (lg/ml)
Gender: female 54 (34%) 19 (36%) 16 (30%) 19 (36%)
Age (years) 64.3 (60.0–68.4) 63.7 (59.7–66.7) 64.3 (60.1–68.4) 65.3 (60.2–69.1)
Tumor Stage
Stage II 10 (6%) 3 (6%) 5 (9%) 2 (4%)
Stage III 113 (71%) 40 (76%) 35 (66%) 38 (72%)
Stage IV 20 (13%) 6 (11%) 8 (15%) 6 (11%)
Missing 16 (10%) 4 (8%) 5 (9%) 7 (13%)
Cancer Site
Colon 139 (89%) 46 (87%) 46 (87%) 47 (89%)
Rectal 17 (11%) 7 (13%) 6 (12%) 4 (8%)
missing 3 (2%) 0 (0%) 1 (2%) 2 (4%)
Chemotherapy Regimen
a
CAPOX 119 (75%) 39 (74%) 42 (79%) 38 (72%)
FOLFOXb 4 (3%) 2 (4%) 2 (4%) 0 (0%)
Capecitabine 14 (9%) 3 (6%) 5 (9%) 6 (11%)
Other 4 (3%) 1 (2%) 0 (0%) 3 (6%)
Missing 18 (11%) 8 (15%) 4 (8%) 6 (11%)
Diabetes mellitus: yes 10 (7%) 4 (8%) 5 (10%) 1 (2%)
Smoking
Current smoker 14 (9%) 7 (13%) 4 (8%) 3 (6%)
Former smoker 98 (62%) 30 (57%) 37 (70%) 31 (58%)
Never smoker 47 (30%) 16 (30%) 12 (23%) 19 (36%)
Alcohol consumption (g/day) 8.2 (1.6–18.0) 4.5 (0.9–15.0) 7.9 (1.0–17.1) 10.3 (4.5–23.3)
2
Body mass index (BMI) (kg/m ) 26.2 (24.0–29.3) 25.9 (23.7–28.4) 26.2 (24.3–29.3) 26.3 (24.1–29.4)
Total energy intake (kcal/day) 1850 (1521–2254) 1696 (1464–2383) 1938 (1460–2511) 1872 (1615–2132)
Total fruit and vegetable 213 (120–307) 171 (120–243) 239 (121–312) 250 (123–392)
consumption (g/day)
Chemotherapy-Induced Peripheral Neuropathy (CIPN)
Total score for peripheral 14.6 (6.3–27.1) 18.7 (6.3–29.2) 14.6 (6.3–27.1) 14.6 (6.3–22.9)
neuropathyc
Sensory score for peripheral 20.8 (4.2–37.5) 25.0 (8.3–37.5) 16.7 (8.3–37.5) 16.7 (2.1–33.3)
neuropathyc
Prevalence of total CIPN (cutoff 129 (81%) 45 (85%) 42 (79%) 42 (79%)
based on scores in
general population)
Prevalence of sensory CIPN 129 (81%) 46 (87%) 44 (83%) 39 (75%)
(cutoff based on scores in
general population)
Prevalence of total CIPN 49 (31%)d 21 (40%) 15 (28%) 13 (25%)
(upper tertile cutoff)
Note. Values are expressed as median (interquartile range), or count (percentage).
a
CAPOX: combination of capecitabine and oxaliplatin.
b
FOLFOX: combination of 5-fluorouracil, leucovorin, and oxaliplatin.
c
All scores are on a 0–100 scale.
d
Number of participants with scores in the highest tertile of total CIPN, see methods for further explanation.

odds ratios tend to overestimate the size of the associ-
ation when the outcome is common (27).
In the tertile analysis, the lowest tertile served
as reference category. Based on the literature, age,
gender, physical activity, energy intake, diabetes (28),
and smoking (5) were considered as possible con-
founding variables. Variables were added to the model
one by one. We adjusted for factors that changed PRs
with at least 10%. In test for trend analyses (p for
trend), we assigned the median score of each tertile of
ET to every individual in this tertile. This new variable
was included in the Cox model as a continuous variable.
In addition, we assessed the association between
ET-concentrations in blood and the severity of total
CIPN score and severity of sensory symptoms. For
these analyses, we used multiple linear regression and
adjusted for possible confounding variables as
described above. In the linear regression analyses, we
included ET and severity of total peripheral neur-
opathy as continuous variables: both peripheral neur-
opathy severity scores and ET-concentrations were
natural log-transformed to approach normal
distributions.
As an additional analysis, we repeated all analyses
with a different cutoff point for prevalence of total
peripheral neuropathy and sensory peripheral neur-
opathy. We included this analysis, because the preva-
lence of CIPN in our study based on cutoff points
published in the literature, was much higher than the
prevalence found in other studies (5). In this
 NUTRITION AND CANCER 455

additional analysis, we created tertiles based on the

p for trend

0.14

0.59
peripheral neuropathy scores, and defined peripheral
neuropathy ‘yes’ as a score in the upper tertile.
In a sensitivity analysis, all analyses were repeated

0.74 (0.50, 1.10)

0.73 (0.42, 1.26)

0.63 (0.35, 1.13)

0.83 (0.51, 1.34)

0.62 (0.30, 1.26)

0.78 (0.41, 1.49)
excluding participants who did not receive oxaliplatin
as part of their chemotherapy regimen (n ¼ 14) or for
PR (95%CI)b
Total CIPN, upper tertilea

Reference

Reference
whom type of chemotherapy was unknown (n ¼ 22),
as oxaliplatin is particularly known to cause neur-
opathy (4), and we excluded patients with stage IV
disease (n ¼ 20) as chemotherapy treatment for those
n cases
49

21
15
13

33

14
8
11
patients may be very different than for patients with
an earlier stage of disease, e.g. it could be a lower
number of cycles or in combination with bevacizu-
159

53
53
53

103

35
34
34
n

mab. This dataset for the sensitivity analyses consisted

of n ¼ 103 patients. All analyses were performed using
p for trend

0.09

0.09

StataSE 14 software (Statacorp, TX, USA).

Results
0.91 (0.79, 1.03)

0.95 (0.80, 1.11)

0.84 (0.70, 1.02)

0.90 (0.79, 1.03)

0.91 (0.79, 1.06)

0.83 (0.68, 1.01)
Sensory CIPN, cutoff based on scores
CIPN prevalence

Median age of the participants was 64.3 years; most

PR (95%CI)b

PR (95%CI)b
Reference

Reference

participants were diagnosed with stage III cancer

in general population

(71%), with the majority of participants having a

tumor in the colon (89%). Of all participants, 78% of
participants were treated with CAPOX (also known as
n cases

n cases
129

46
44
39

89

33
30
26

Xelox) or FOLFOX, which are both oxaliplatin-con-

taining regimens. The median age of participants who
Table 2. Association of ET-concentration in blood and prevalence of CIPN in CRC patients.

had to be excluded of the dataset (see Figure 1) was

158

53
53
52

103

35
34
34
n

64.6 years, 69% had stage III disease, and 84% with a
tumor in the colon. Of these excluded participants,
p for trend

82% were treated with CAPOX or FOLFOX.

0.52

0.45

The median concentration of ET in whole blood

was 10.2 mg/ml, see Table 1 for a description of base-
line characteristics. Total median peripheral neur-
0.93 (0.81, 1.07)

0.93 (0.77, 1.11)

0.93 (0.78, 1.11)

0.93 (0.80, 1.08)

0.94 (0.78, 1.13)

0.92 (0.75, 1.13)
Total CIPN, cutoff based on scores in

opathy score was 14.6; for the sensory domain median

PR (95%CI)b

Reference

Reference

score was 20.8. Based on cutoff points derived from

general population

scores in the general population (23), the prevalence

of peripheral neuropathy in our study population was
81% (n ¼ 129 participants), and 81% (n ¼ 129 partici-
n cases
129

45
42
42

87

31
29
27

pants) for sensory peripheral neuropathy. Total per-

ipheral neuropathy scores and sensory peripheral
neuropathy scores were highest among participants in
159

53
53
53

103

35
34
34
N

the lowest tertile of ET. Prevalence of total peripheral

in the upper tertile of CIPN total score.

neuropathy or sensory peripheral neuropathy were

Sensitivity analysis, excluding stage IV
patients and patients not receiving

highest among participants in the lowest tertiles of ET

as compared with the higher tertiles.
c

ET-concentration in ln(mg/ml)c
ET-concentration in ln(mg/ml)

ET levels in blood were not associated with preva-

lence of total peripheral neuropathy (PR ¼ 0.93 per
for age and gender.

ln(mg/ml) ET (0.1; 1.07)), nor with prevalence of sen-

sory peripheral neuropathy (PR ¼ 0.91 per ln(mg/ml)
Medium (T2)

Medium (T2)
Main analysis

oxaliplatin

ln transformed.
High (T3)

High (T3)
Low (T1)

Low (T1)

ET (0.79, 1.03)), see Table 2. In the tertile analysis,

ET tertiles

ET tertiles

aParticipants

bAdjusted

being in the highest tertile of ET tended to be associ-

ated with a lower prevalence of (total) peripheral
c
456 R. M. WINKELS ET AL.

Table 3. Association of ET-concentration in blood and severity of peripheral neuropathy in CRC patients.

Total score (ln transformed) Sensory score (ln transformed)

Peripheral neuropathy scores a
n b (95%CI) b
n b (95%CI)b
ET blood concentration in ln(mg/ml) 129 0.11 (0.33, 0.11) 129 0.14 (0.37, 0.10)
Sensitivity Analysis, Excluding Stage IV Patients and Patients not Receiving Oxaliplatin
ET blood concentration in ln(mg/ml) 87 0.07 (0.33, 0.20) 89 0.06 (0.34, 0.22)
Note. Severity scores of neuropathy and ET-concentrations were both natural log-transformed to approach normal distributions.
a
Included in this analysis are the participants with CIPN according to cutoff values based on scores in the general Dutch population.
b
Adjusted for age and gender.

neuropathy, but this was not statistically significant Experimental rodent studies intervened with ET and
(PR for the highest vs. the lowest tertile 0.93 (0.78, assessed ET and oxaliplatin levels directly in dorsal
1.11). For sensory peripheral neuropathy, being in the root ganglions, while in our study we used blood
highest tertile of ET tended to be associated with a levels of ET as a biomarker of ET status. Although we
lower prevalence of sensory peripheral neuropathy, could not directly assess ET levels in neurons, we
but again this association was not statistically signifi- believe that our findings provide support for a
cant (PR for the highest vs. the lowest tertile 0.84 possible association between ET levels and CIPN in
(0.70, 1.02). humans. Importantly, we measured levels of ET in
When prevalence of peripheral neuropathy was blood and studied a possible association with CIPN;
defined as scores in the upper tertile of total periph- we did not intervene with ET in this study as
eral neuropathy scores, having a high blood concen- safety and relevance need to be established first.
tration in the highest tertile of ET-concentrations (T3) Observational studies like this are necessary before
was associated with a lower prevalence of total periph- any potential step toward intervening with ET in
eral neuropathy, but again not statistically significant cancer patients should be made.
(PR for T3 vs. T1 0.63 (0.35, 1.13). We did not have complete data on the cumulative
In the linear regression analysis presented in dose of oxaliplatin received during chemotherapy.
Table 3, ET levels in blood were not associated with CAPOX and FOLFOX both are combinations of
severity of total peripheral neuropathy (b¼ 0.11 chemotherapy including oxaliplatin, and specifically
(0.33; 0.11)), nor with severity of sensory peripheral oxaliplatin is known to be associated with acute and
neuropathy (b¼ 0.14 (0.37; 0.10)). with chronic CIPN (3,20). In an earlier observational
The results of the sensitivity analyses, excluding study among 207 CRC patients who underwent
participants who had not received oxaliplatin-based chemotherapy, chronic CIPN years after treatment
chemotherapy and excluding stage IV patients, did was associated with having received higher cumulative
not differ substantially from the main analysis as dose of oxaliplatin (3), but patients who had received
shown in Tables 2 and 3. a dose reduction were still at risk of developing
chronic CIPN (3). In our study, a higher dose of oxa-
liplatin may have resulted in a higher risk of chronic
Discussion
CIPN, but severe symptoms of acute CIPN may have
This study did not find statistically significant resulted in modification of treatment, resulting in a
associations between ET levels in blood and chronic lower cumulative dose of oxaliplatin. Yet, acute CIPN
peripheral neuropathy in CRC patients who had been is correlated with a higher risk of developing chronic
treated with chemotherapy. However, although none CIPN (3). This makes it challenging to predict how
of the associations was statistically significant, it including information on the total cumulative dose of
is noteworthy that the direction of the associations oxaliplatin would affect our results.
reported through PRs and betas in this paper were all Earlier studies reported that about one-third of
in agreement with our hypothesis. This may putatively patients suffer from chronic CIPN 6 months after
suggest that a higher ET-concentration in blood could chemotherapy (5), whereas peripheral neuropathy was
be associated with lower prevalence and severity of 81% in the current study. This difference in preva-
chronic peripheral neuropathy, but that there was lence compared with other studies likely resulted from
insufficient statistical power to detect this association. differences in methodology to determine chronic
To the best of our knowledge this is the first CIPN (29). Our study used a patient-reported ques-
human study to explore possible associations of circu- tionnaire, which is prone to overestimate the outcome
lating ET-concentrations and peripheral neuropathy. (22). Nevertheless, this questionnaire is shown to be

valid and reliable (19). Moreover, the total median
CIPN score we observed in our population (14.6) is
similar to the mean score of 13.0 as observed in survi-
vors of CRC who had received oxaliplatin as part of
their chemotherapy treatment (30). In addition, it is
important to realize that there is no clinically defined
cutoff for CIPN for this instrument. We therefore
used the reported mean score of a sample of respond-
ents from the general Dutch population, as the cutoff
for CIPN. Thus, given that we used the mean in that
standard population as cutoff for CIPN, the expect-
ation was that at least 50% of our study population
would have neuropathy. To mitigate this issue, we
decided to also report on an analysis of tertiles with
the upper tertile defined as ‘having peripheral neur-
opathy’. Some patients may have had neuropathy at
baseline, as a result of diabetes or other comorbidities,
but we did not assess neuropathy at baseline and
could therefore not adjust for that.
Although the associations found in our data were
not statistically significant, the directions of the associ-
ation were in agreement with our hypothesis. Thus,
further study into these associations using more com-
prehensive methods to assess CIPN throughout and
after the period of cancer treatment (29) is warranted.
It is important to acknowledge that our study is
observational, and that there could be residual con-
founding by other clinical/personal factors. Given the
paucity of data on lifestyle factors that may affect the
risk of CIPN (7), it is challenging to judge whether
we may have missed specific confounding variables.
As stated earlier, we may not have had sufficient stat-
istical power to detect associations. We calculated that
if we assume that the prevalence of neuropathy in the
upper tertile of ET is 25%, and 40% in the lowest ter-
tile (PR0.63), we would have needed a total sample
size of 465 participants to detect a statistically signifi-
cant association, assuming a power of 80% and an
alpha of 0.05. Therefore, our results should be inter-
preted as hypothesis-generating and future studies
require a larger sample size.
Mushrooms are the most important source of ET
in the human diet (13). In comparison with other
countries, especially Asian countries (31), mushroom
intake in The Netherlands is relatively low (32).
Additionally, in The Netherlands white buttons are
the most eaten variety of mushrooms, which are low
in ET compared with other mushroom varieties (12).
As comparison, in healthy volunteers in a study from
Japan, ET blood levels ranged from 20 to 70 mg/ml,
compared with a range of 4.5–23.0 mg/ml in our study.
The relatively low levels of ET, and lack of wide
NUTRITION AND CANCER 457
variability in ET levels in our study, may have further
reduced our ability to detect significant associations.
Thus, it would be interesting to repeat these analyses
in Japan or other countries where mushroom intake is
traditionally higher than in The Netherlands (33).
ET is a potent antioxidant. The effects of antioxi-
dants in general on CIPN have been studied, but
results are fairly mixed (7,34–36). For example, some
studies suggest that there could be a role for the anti-
oxidants vitamin E and acetyl-L-carnitine in protecting
against CIPN (37–40), but other studies could not
confirm this (41,42), or even suggested detrimental
effects on CIPN-related outcomes (35,41). As also
proposed by others (7), there could be a role for anti-
oxidants or for dietary strategies in general in the pre-
vention of CIPN, but further research is warranted to
draw firm conclusions.
To conclude, ET-concentration in blood was not asso-
ciated with prevalence or severity of peripheral neur-
opathy among CRC patients treated with chemotherapy.
Nevertheless, data may putatively suggest that a higher
ET-concentration in blood may be associated with a
lower prevalence of chronic peripheral neuropathy, espe-
cially for sensory peripheral neuropathy. Future studies
should consider different methodologies to assess chronic
CIPN at various time points before, during and after
cancer treatment, and should be performed in popula-
tions where intake of mushrooms, and therefore variabil-
ity in concentrations of ET in blood is larger.
Acknowledgments
The authors would like to thank the coworkers from
the following hospitals for their involvement in recruitment
for the COLON study: Hospital Gelderse Vallei, Ede;
RadboudUMC, Nijmegen; Slingeland Hospital,
Doetinchem,; Canisius Wilhelmina Hospital, Nijmegen;
Rijnstate Hospital, Arnhem; Gelre Hospitals, Apeldoorn/
Zutphen; Hospital Bernhoven, Uden; Isala, Zwolle; ZGT,
Almelo; Martini Hospital, Groningen; Admiraal de Ruyter
Hospital, Goes/Vlissingen, all in The Netherlands.
Disclosure Statement
The authors declare to have no conflicts of interest. The
funding agencies had no role in the design, data collection,
analysis or interpretation of the data, or in the writing of the
manuscript, or in the decision to submit for publication.
Author Contributions
Conceptualization, Renate Winkels, Lieve van Brakel,
Robert Beelman, John Richie, Ellen Kampman and
Dieuwertje Kok; Formal analysis, Renate Winkels and Lieve
van Brakel; Investigation, Harm van Baar, Franzel van
458 R. M. WINKELS ET AL.
Duijnhoven, Anne Geijsen, Henk van Halteren, Bibi
Hansson, Dongxiao Sun, Evertine Wesselink, Moniek van
Zutphen and Dieuwertje Kok; Supervision, Renate Winkels;
Writing – original draft, Renate Winkels and Lieve van
Brakel; Writing – review & editing, Renate Winkels, Lieve
van Brakel, Harm van Baar, Robert Beelman, Franzel van
Duijnhoven, Anne Geijsen, Henk van Halteren, Bibi
Hansson, John Richie, Dongxiao Sun, Evertine Wesselink,
Moniek van Zutphen, Ellen Kampman and Dieuwertje Kok.
All authors approved the final version of the manuscript
Funding
The COLON study was financially supported by Wereld
Kanker Onderzoek Fonds, including funds from grant 2014/
1179 as part of the World Cancer Research Fund
International Regular Grant Program; Alpe d’Huzes/Dutch
Cancer Society (UM 2012-5653, UW 2013-5927, UW 2015-
7946); and ERA-NET on Translational Cancer Research
(TRANSCAN/Dutch Cancer Society: UW2013-6397,
UW2014-6877 and The Netherlands Organization for
Health Research and Development (ZonMw, The
Netherlands).
ORCID
Renate M. Winkels http://orcid.org/0000-0002-0376-8811
Ellen Kampman http://orcid.org/0000-0002-8606-7075
Dieuwertje E. Kok http://orcid.org/0000-0001-7154-8207
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