Supplementation with Folic Acid during Methotrexate
Therapy for Rheumatoid Arthritis
A Double-Blind, Placebo-controlled Trial
Sarah L. Morgan, MD, RD; Joseph E. Baggott, PhD; William H. Vaughn, BS; Janet S. Austin, MA;
Tonya A. Veitch, BS; Jeannette Y. Lee, PhD; William J. Koopman, MD;
Carlos L. Krumdieck, MD, PhD; and Graciela S. Alarcon, MD, MPH
• Objective: To determine the effect of two different
weekly doses of folic acid on the toxicity and efficacy of
low-dose methotrexate therapy for rheumatoid arthritis.
• Design: Randomized, double-blind, placebo-
controlled study.
• Patients: 79 persons between 19 and 78 years of
age who fulfilled the American Rheumatism Associa-
tion's criteria for rheumatoid arthritis.
• Intervention: Participants were randomly assigned
to visually identical placebo or to 5 mg or 27.5 mg of
folic acid each week.
• Measurements: Duration, intensity, and clinical se-
verity of toxic events; efficacy (indices of joint tender-
ness and swelling and grip strength); plasma and eryth-
rocyte folate levels; and other laboratory variables.
• Results: Folic acid supplementation at either dose
did not affect the efficacy of methotrexate therapy as
judged by joint indices and patient and physician as-
sessments of disease. Patients given folic acid supple-
ments had lower toxicity scores than did participants
given placebo (P < 0.001). Low blood folate levels and
increased mean corpuscular volumes were associated
with substantial methotrexate toxicity, whereas daily
dietary intakes of more than 900 nmol (400 fig) of folic
acid were associated with little methotrexate toxicity.
• Conclusions: Folic acid, an inexpensive vitamin, is
safe in a broad range of doses and protects patients
with rheumatoid arthritis who are taking methotrexate
from toxicity while preserving the efficacy of methotrex-
ate.
Ann Intern Med. 1994;121:833-841.
From the Birmingham Veteran's Administration Hospital and
the University of Alabama at Birmingham, Birmingham, Ala-
bama. For current author addresses, see end of text.
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1 he folic acid antagonist methotrexate (N-10-methyl-ami-
nopterin) is useful in low doses (2.5 to 20 mg/wk) for
treating chronic inflammatory diseases (1-7). Many trials
have established the efficacy of methotrexate in rheuma-
toid arthritis (7-13). Compared with other disease-modi-
fying drugs, methotrexate has the highest probability of
drug continuation at 10 years. Dose response-related
toxic effects have been reported in 30% to 90% of pa-
tients given methotrexate (13). Toxic effects include gas-
trointestinal intolerance, hematologic abnormalities, alo-
pecia, hepatotoxicity, and pulmonary toxicity (14-22).
Some side effects of methotrexate administration, such
as gastrointestinal intolerance, mimic complicated folate
deficiency (23). Folate deficiency occurs frequently in pa-
tients with rheumatoid arthritis; further, folate stores are
decreased in patients with rheumatoid arthritis who take
methotrexate, suggesting that impaired folate status is
related to toxicity (24-26).
Folic acid supplementation has been reported anecdot-
ally to lessen toxicity in patients receiving methotrexate
treatment (27, 28). In a 6-month, double-blind, placebo-
controlled trial, 7 mg of folic acid weekly (1 mg/d or 2265
nmol/d) decreased methotrexate toxicity without affecting
efficacy (29). This was confirmed by Stewart and col-
leagues (30) in a retrospective chart review.
Folinic acid (leucovorin, citrovorum factor) is a one-
carbon-substituted, fully reduced folate that has also been
administered during methotrexate therapy (31-36). Low
doses of the vitamin (1 to 7 mg/wk) have decreased
methotrexate toxicity (35, 36). Higher doses negate effi-
cacy and lessen toxicity (31, 32). Thus, the folinic acid
dose may critically affect the efficacy of methotrexate ther-
apy.
The influence of the folic acid dose on methotrexate
toxicity and efficacy remains controversial, and the effects
of different doses of folic acid are not known (37, 38).
Some investigators argue that if toxic effects occur, the
most rational approach is to reduce the dose of metho-
trexate rather than to provide folic acid supplements (37).
We designed a larger and longer study to evaluate differ-
ent doses of folic acid, assuming that toxicity could be
reduced without changing the efficacy of methotrexate.
Methods
Participants
Patients aged 19 to 78 years who fulfilled the American Col-
lege of Rheumatology's revised criteria for rheumatoid arthritis
© 1994 American College of Physicians 833
 consented to participate in the trial (39). Enrollment criteria
included rheumatoid arthritis diagnosed more than 6 months
previously, onset after the age of 16 years, and at least three of
the following signs or symptoms: 3 or more swollen joints, 6 or
more tender joints, at least 45 minutes of morning stiffness, and
a Westergren erythrocyte sedimentation rate greater than or
equal to 28 mm/h.
Referring rheumatologists and the principal investigator did
the screening. Exclusion criteria included serious concomitant
medical illnesses, liver enzyme levels twice the upper limit of
normal, leukocyte counts less than 3.5 X 109/L or platelet counts
less than 150 X 109/L, and use of methotrexate within the past 6
months. Gold salts were stopped for at least 10 days before the
trial. This short washout period mirrors actual rheumatology
practice. Patients remained under the care of their rheumatolo-
gists, abstained from alcohol use, did not become pregnant, and
received stable doses of aspirin and nonsteroidal anti-inflamma-
tory drugs. If prednisone was taken at entry, the dose could not
exceed 10 mg/d. Hydroxychloroquine therapy was allowed during
the study.
Study Design
Figure 1 shows the trial design. To maintain the double-blind
status of the trial, the statistician carried out the randomization
using a computer program in which the algorithm was transpar-
ent and a coded vial number represented the treatment assign-
ment. Patients were assigned to treatment groups by a sequential
treatment assignment process designed to balance the sample
with respect to baseline features, including age, sex, folate-con-
taining vitamin use, rheumatoid factor status, and prednisone use
(40). Patients agreed to discontinue therapy with folate-contain-
ing vitamins during the trial. Rheumatoid factor was considered
positive if the level was more than 30 IU/mL or if the titer was
more than 1:160. Patients received either visually identical pla-
cebo or 5 mg (low-dose folic acid group) or 27.5 mg folic acid
(high-dose folic acid group) each week, prepared by the Hospital
Investigational Drug Service. Spectrophotometric analysis indi-
cated that the mean ± SD folic acid content was 1 ±0.15 mg
(2.3 ± 0.3 jLtmol) and 5.5 ± 0.3 mg (12.5 ± 0.7 /Ltmol) per capsule
in the low-dose and high-dose folic acid groups, respectively.
Lederle Laboratories provided the methotrexate (Rheumatrex;
Pearl River, New York), which was started in a median oral dose
of 16.5 jLtmol (7.5 mg) per week and increased in 5.5-/xmol (2.5
mg) increments at the rheumatologist's discretion. Methotrexate
was taken either in an undivided or a divided dose (that is, every
12 hours for three doses). The methotrexate dosing regimens
were identical among the study groups. Folic acid supplements
were given 5 days per week when methotrexate was not ingested.
Compliance with the regimen was reinforced using a digital re-
minder cap (Counter Cap; Senetics, Boulder, Colorado). All
participants and investigators were blinded to vitamin capsule
content until the study was complete.
Clinical Assessment
Patients were evaluated immediately before methotrexate ini-
tiation at a mean of 13, 26, 39, and 53 weeks (Figure 1). Each
patient was assessed by the same physician-nutritionist (SLM).
Two research assistants (JSA or WHV) did the joint evaluations.
In most cases, patients were examined by the same observer
throughout the study. The joint counts for tenderness and swell-
ing were not significantly different between the two observers
(P = 0.6 for tenderness; P = 0.9 for swelling). The following vari-
ables were evaluated at each visit: 1) number of the 58 diarth-
rodial joints with swelling; 2) number of the 60 joints with
tenderness on pressure or with pain on passive motion (or both);
3) joint swelling and tenderness indices, expressed as a sum, with
each joint graded for swelling as 0 (none), 1 (mild), 2 (moder-
ate), or 3 (severe); 4) mean grip strength (three measurements)
for both hands expressed in mm Hg; 5) duration of morning
stiffness expressed in hours; 6) patient's and physician's global
assessments of disease activity graded as 0 (asymptomatic), 1
(mild), 2 (moderate), 3 (severe), or 4 (very severe); 7) current
medications and doses; 8) a 1-day dietary recall using the Min-
nesota Nutrition Data System software, Food Database version
6A, Nutrient Database version F21 (41); 9) eight activities of
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daily living assessed and averaged at baseline and at 12 months
using the modified Health Assessment Questionnaire and scored
on a scale of 1 (no difficulty) to 4 (unable to perform) (42); and
10) presence, duration, intensity, and severity of toxic effects at
every follow-up visit and every 3 weeks by telephone interview
(done by SLM).
Laboratory Assessments
At the initial visit, complete blood cell count, Westergren
erythrocyte sedimentation rate, liver enzyme (aspartate amino
transferase and alkaline phosphatase), rheumatoid factor by
nephelometry, and serum creatinine values were obtained. The
complete blood cell count, creatinine, and liver function tests
were repeated at all visits. If laboratory values were obtained
more frequently than stipulated in the protocol, they were re-
corded and became part of the toxicity score.
Blood for plasma and erythrocyte folate levels, using a meth-
otrexate-resistant Lactobacillus casei microbiological assay, was
drawn at all visits (43). At baseline, blood was drawn for a
vitamin panel (plasma and erythrocyte folate, vitamins B 12 , A, C,
and E, carotene, riboflavin, thiamin, pyridoxine) (44-51). If pa-
tients had abnormal values for any of the vitamins, other than
folate, the abnormality was treated with single vitamin supple-
ments.
Radiographic Assessment
Hand and wrist radiographs taken within 6 months of entering
the trial were assessed by one of the rheumatologists (GSA)
without knowledge of study status. Joint erosions and space
narrowing were determined by the modified method of Sharp
(52). Results were expressed as the mean raw scores for joint
erosion and joint space narrowing.
Toxicity: Frequency and Severity
Toxic effects and discontinuation of therapy with study medi-
cations due to toxicity were considered primary outcomes. We
determined a toxicity score, modified from the one previously
used, for each patient at each visit or until methotrexate therapy
was discontinued (29) (Appendix).
Efficacy
We determined patient response to treatment using a modifi-
cation of the criteria used in our previous folic acid supplemen-
tation trial and by others (8, 29, 53). We defined marked im-
provement in the joint swelling index and the joint tenderness
and pain index as a net decrease of 50% or more in value at any
Figure 1. Study design for the double-blind, placebo-controlled
trial. The randomization was done by the statistician using a
sequential treatment assignment program in which the algorithm
was transparent and a coded vial number was used as the treat-
ment assignment.
 follow-up visit compared with visit 1. We defined moderate im-
provement as a 30% to 49% net decrease in the index, and no
change as the index value remaining within 30% of the original
value. We defined worsening as an increase in the index value of
30% or more.
We examined the effects of folic acid supplementation on
changes in physician and patient global assessments, grip
strength, morning stiffness, erythrocyte sedimentation rate, find-
ings of the modified Health Assessment Questionnaire, complete
blood cell counts, blood folate levels, and dietary folate and
vitamin B 12 intake.
Statistical Analysis
Sample Size and Analysis
In our previous study, the mean toxicity scores for patients
receiving low-dose folic acid and placebo recipients were 0.21
and 1.06, respectively, for a difference of 0.85. The sample size
tested the null hypothesis for no difference between placebo and
low- and high-dose folic acid treatments, with respect to the
mean toxicity score. The alternative hypothesis was that the
difference in the score between at least one pair of treatments
(placebo compared with low-dose folic acid, placebo compared
with high-dose folic acid, or low-dose folic acid compared with
high-dose folic acid) was greater than or equal to the difference
observed previously. As described by Cohen (54) and imple-
mented by Borenstein and Cohen (55), 25 patients per group was
a sufficient number to detect a difference in the mean toxicity
score of 0.85 between any pair of treatments at a significance
level of 0.05 and a power of 0.80.
An underlying assumption in sample size estimates was that
the scores are normally distributed. To address the potential
non-normal character of the toxicity scores, we increased the
sample size estimates by 4% to maintain the significance level
and power if nonparametric analyses were used. To allow for a
20% attrition rate, the enrollment goal was 31 patients per
group.
Patients vvere withdrawn if more than 3 weeks of methotrexate
treatment was missed for any reason or if noncompliance was
documented. Patients who withdrew from the trial because of
toxic effects did so after consulting their rheumatologists. Pa-
tients contributed to data analyses until therapy with methotrex-
ate was discontinued. For patients who withdrew before receiving
12 months of therapy, we computed toxicity and efficacy data
based on data collected until drug therapy was discontinued.
Comparison of Data among Groups
We tested the baseline values of all outcome measures for
normality using the Shepin-Wilk statistic. For normally distrib-
uted data, we used chi-square analyses to compare proportions
and analysis of variance to evaluate the effects of treatment,
time, and their interaction.
Non-normal distributions occurred in joint indices for pain and
tenderness, physician and patient assessment of disease, grip
strength, answers to the modified Health Assessment Question-
naire, and plasma and erythrocyte folate levels. We used the
Wilcoxon rank-sum test to compare the three groups and Wil-
coxon rank-sum tests for pair-wise comparison, if appropriate.
Comparison of Toxicity Scores
We used the Fisher exact test to compare the incidence of
toxic effects in the folic acid and placebo groups. Because the
primary outcome measure was the toxicity score, we included
data until the study was complete or the participant withdrew. To
evaluate the effects of time, treatment group, and their interac-
tion on the toxicity score, we used analysis of variance on the
rank of the toxicity scores. We did two-way analyses of variance
(or on the ranks for data that were non-normally distributed) to
evaluate the effects of time, treatment, and their interaction.
Results
Patients
W e enrolled 94 patients (age range, 19 to 78 years) into
the trial. Subsequently, we withdrew 15 patients because
of noncompliance (numbers were equally distributed
among the study groups). O u r results are from 79 patients
who completed the trial: 25 in the low-dose folic acid
group, 26 in the high-dose folic acid group, and 28 in the
placebo group. T h e demographic and selected clinical
features of the patients in the three study groups were
similar (Table 1). N o patients were taking sulfasalazine
before enrollment. W e found no statistical difference
a m o n g the groups in previous use of gold salts (P = 0.92).
Four patients in the placebo group (23%), 4 in the low-
dose folic acid group (14.3%), and 6 in the high-dose folic
acid group (23%) previously had therapy with methotrex-
ate discontinued because of toxic effects (P = 0.76). Pa-
tients who previously discontinued methotrexate therapy
because of toxic effects were randomly distributed in the
three groups. O t h e r initial variables such as hemoglobin
level, hematocrit, leukocyte count, m e a n corpuscular vol-
u m e , creatinine level, aspartate aminotransferase value,

Table 1. Demographic and Selected Clinical Characteristics of 79 Patients with Rheumatoid Arthritis*
Characteristics Study Group
Placebo Low-Dose Folic Acid High-Dose Folic Acid
(n = 28) (n = 25) (n = 26)

Mean age, y 52.2 ± 13.0 54.4 ± 14.0 53.2 ± 14.3

Sex, % women 82 76 65
Race, % white 85.7 68 73
Mean disease duration, y 8.5 ± 8.2 7.4 ± 10 11.6 ± 9.5
Previous use of folate-containing vitamins, % 32 28 19
IgM rheumatoid factor positivity (>30 IU/mL or 1:160 titer), % 71 80 85
Concurrent use of aspirin or nonsteroidal anti-inflammatory
drugs, % 86 80 88
Concurrent prednisone use, % 79 80 88
Concurrent hydroxychloroquine use, n 1 1 4
Patients previously receiving gold salts, n 16 17 17
Patients previously receiving methotrexate, n 4 6 4
Previous disease-modifying antirheumatic drug use, % 64 68 77
Mean joint erosion score 9.2 ± 9.5 15 ± 16.7 17.7 ± 17.8
Mean joint space narrowing 20.2 ± 20.1 29.1 ± 25.1 35.3 ± 31.1

* All comparisons between groups, P > 0.05. Means are expressed ± SD.

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 Methotrexate Dose: Folate and B 1 2 Intake
We found no statistical differences in mean cumulative
methotrexate dose among the three groups. The mean
cumulative methotrexate doses at 53 weeks in the pla-
cebo, low-dose folic acid, and high-dose folic acid groups
were 206 ± 88 mg, 217 ± 113 mg, and 284 ± 187 mg,
respectively (P = 0.24). The average weekly methotrexate
dose was 8.5 ± 1 . 5 mg, 9.4 ± 2.4 mg, and 9.6 ± 2.6 mg in
the placebo, low-dose folic acid, and high-dose folic acid
groups, respectively (P = 0.27).
The mean dietary folate and vitamin B 1 2 intakes among
the groups were never statistically different. Median di-
etary folate intakes were 204 /xg/d in the placebo group,
195 jmg/d in the low-dose folic acid group, and 230 ju,g/d in
the high-dose folic acid group (P = 0.37 for all compar-
isons). The median daily intakes in all study groups did
not substantially differ from the median values of 150 to
250 jmg/d reported in the Second National Health and
Nutrition Examination series (56).

Primary Outcome: Toxic Effects

Figure 2. Median toxicity scores in the three study groups. The
placebo group had a statistically higher toxicity score (P = 0.001)
than the folic acid supplementation groups. The toxicity score
was not significantly different between the low-dose folic acid and
high-dose folic acid groups (P = 0.71). A = patients withdrawn
from the trial because of toxic effects; O = patients who com-
pleted the trial.
alkaline phosphatase level, and erythrocyte sedimentation
rate were also similar among the groups. The erosion and
joint space narrowing scores were similar at baseline
among the three groups (P = 0.16 for joint space narrow-
ing and P = 0.17 for erosions; Table 1).
Fifty-four patients (68%) experienced some form of
toxicity: 25 (89%) in the placebo group, 12 (48%) in the
low-dose folic acid group (P < 0.002 compared with pla-
cebo), and 17 (65%) in the high-dose folic acid group.
Figure 2 shows the median toxicity score for the three
groups (0.685, 0.016, and 0.031 in the placebo, low-dose
folic acid, and high-dose folic acid groups, respectively).
The toxicity score for the placebo group was greater than
those for the two folic acid-supplemented groups (P =
0.001 for both comparisons for low-dose folic acid and
high-dose folic acid compared with placebo; P - 0.71 for
low-dose folic acid compared with high-dose folic acid).
The most frequently reported toxicities were nausea and
indigestion (31 patients), diarrhea (11 patients), stomatitis
(9 patients), and rash (9 patients). When the patients
receiving hydroxychloroquine were omitted from the anal-
yses, the results were identical.
Other Outcomes
Efficacy
As noted in Table 2, the percentages of patients who

Table 2. Joint Swelling and Tenderness at 6 and 12 Months in the Three Treatment Groups
Outcome 6 Months 12 Months
Placebo Low-Dose Folic High-Dose Folic Placebo Low-Dose Folic High-Dose Folic
(n = 23) Acid (" = 24) Acic (" = 23) (n = 19) Acid (" = 23) Acid (n = 18)
n?_
< /u

Marked improvement
Swelling 35 38 61 68 78 78
Tenderness 30 29 39 53 43 61
Moderate improvement
Swelling 26 29 26 16 13 5
Tenderness 35 8 18 16 13 16
No change
Swelling 35 33 9 11 5 17
Tenderness 26 55 39 21 35 17
Worsening
Swelling 4 0 4 5 4 0
Tenderness 9 8 4 10 9 6

* P > 0.5 (chi-square) for comparisons among groups at 6 months and 12 months.

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 Table 3. Data for Efficacy and Other Outcomes at Baseline, 6 Months, and 12 Months*
Outcome Placebo Group Low-Dose Folic Acid Group High-Dose Folic Ac id Group
Baseline 6 Months 12 months Baseline 6 Months 12 months Baseline 6 Months 12 months

Joint indices for tendernesst 34 21 18 32 20 21 34 26 14

(2, 99) (0, 61) (4, 62) (6, 112) (2, 99) (0, 90) (2, 105) (0, 66) (2, 41)
Joint indices for swelling! 45 25 12 51 28 14 43 20 13
(6, 85) (3, 51) (0, 51) (14, 85) (2, 48) (2, 40) (18, 103) (4, 62) (1, 58)
Patient assessment of disease
activity! 3 2 2 3 2 2 3 2 2
(2,5) (1,4) (2,4) (2,5) (1,5) (1,5) (2,5) (1,4) (1,4)
Physician assessment of disease
activity! 4 3 2 4 3 2 4 3 2
(2,4) (2,4) (2,4) (2,5) (2,4) (2,3) (2,5) (2,5) (2,4)
Grip strength in right hand,
mm Hg"\ 56.5 66.7 61.7 40.5 45 48.3 35 53.3 46.7
(5, 100) (10, 205) (9.3, 167) (0, 173) (13, 127) (18.3, 133) (5, 132) (5, 180) (8.3, 118)
Grip strength in left hand,
mm Hgf 46.7 56.7 56.7 40 34.2 48.3 32.5 30 48.3
(4.3, 208) (10, 223) (10.7, 150) (5, 243) (5, 115) (10, 152) (1.3, 183) (6.7, 190) (10, 157)
Modified Health Assessment
Questionnaire 1.8 1.5 2 1.2 2 1.2
(1, 3.4) (1, 2.8) (1, 3.8) (1, 2.8) (1.1, 3.4) (1, 2.6)

* Median (minimum, maximum).

! P < 0.05 for time effects for the three treatment groups.

had marked improvement, moderate improvement, no im-
provement, or worsening were similar among the three
groups (P > 0.5). We observed marked improvement in
the pain and tenderness index after 12 months in 53%,
43%, and 61% of patients receiving placebo, low doses of
folic acid, and high doses of folic acid, respectively. We
found marked improvement in the swelling index in 68%,
78%, and 78% of patients receiving placebo, low doses of
folic acid, and high doses of folic acid, respectively.
Patient Withdrawal and Dropout
We withdrew 13 patients because of noncompliance.
One patient was withdrawn for taking additional folate
supplements and one because diagnosis was reconsidered.
Seven patients in the placebo group, 2 in the low-dose
folic acid group, and 2 in the high-dose folic acid group
discontinued methotrexate treatment because of toxic ef-
fects. More dropouts occurred in the placebo group
(25%) than in the folic acid groups (8%) (P = 0.08).
volume of 100 fL or more at one or more follow-up visits
compared with only one and none in the low-dose or
high-dose folic acid groups, respectively (P = 0.02).
Mean baseline plasma folate levels were 15, 10, and
13.5 nmol/mL (P > 0.1), and mean baseline erythrocyte
folate levels were 662, 591, and 624 nmol (P > 0.1) in the
placebo, low-dose folic acid, and high-dose folic acid
groups, respectively. After 1 year, mean plasma folate
levels in the placebo group decreased to 4.8 nmol/mL
(P < 0.01) and mean plasma folate levels increased four
to five times from baseline values in the folic acid groups
(P < 0.001). In contrast, mean erythrocyte folate levels in
the low-dose and high-dose folic acid groups were little
changed (± 5%) from baseline values after 1 year,
whereas levels in the placebo group decreased by approx-
imately 50% (349 nmol/mL; P < 0.001 for baseline values
compared with values at 1 year). Twelve (43%), 5 (20%),
and 4 (15%) patients in the placebo, low-dose folic acid,

Other Indices

As noted in Table 3, patient and physician assessment,

joint indices for swelling and tenderness, and grip
strength improved with time. We found significant time
effects with respect to joint tenderness indices (P =
0.025), joint swelling indices (P = 0.027), physician assess-
ment of disease (P = 0.011), patient assessment of disease
(P= 0.008), right-hand grip strength (P = 0.032), and
left-hand grip strength (P = 0.047). We observed no sig-
nificant time-treatment interactions.
At the follow-up visits, changes in the median or mean
values for the following variables were not statistically
different among the groups: modified Health Assessment
Questionnaire, erythrocyte sedimentation rate, hemoglo-
bin level, hematocrit, leukocyte count, mean corpuscular
Figure 3. Toxicity score as a function of dietary folate intake in
volume, creatinine concentration, and aspartate amino- the placebo group. An exponential decay curve was fitted to the
transferase or alkaline phosphatase levels. However, six of data. To convert micrograms to nanomoles, divide micrograms by
the patients in the placebo group had a mean corpuscular 0.441 (the formula weight of folic acid is 441.4).

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 and high-dose folic acid groups, respectively, had eryth-
rocyte folate levels less than 320 nmol/L (that is, <140
ng/mL) at one or more of the follow-up visits (P = 0.02).
Figure 3 shows the toxicity score in the placebo group
plotted as a function of mean daily dietary folate intake.
Negligible toxic effects occurred as the dietary folate in-
take was increased to more than 900 nmol/d (400 jLtg/d).
We used a multivariate general linear model to evaluate
the effects of the following factors on the toxicity score:
sex, race, disease duration, previous folate-containing vi-
tamin use, rheumatoid factor status, concurrent nonste-
roidal anti-inflammatory drug use, concurrent prednisone
use, previous disease-modifying antirheumatic drug use,
dietary folate intake, and supplemental folate intake (pla-
cebo; 5 mg/wk; and 27.5 mg/wk). Because the toxicity
score and dietary folate intake measurements were not
normally distributed, we used their ranks in the analysis.
The analysis indicated that dietary folate intake and sup-
plemental folate intake were related to toxicity (P = 0.016
for dietary intake; P = 0.016 for supplemental folate in-
take). Dietary folate was negatively correlated with toxic-
ity score, indicating that higher dietary folate intake re-
duced toxicity. With regard to supplemental folate intake,
both the 5 mg and 27.5 mg/wk doses reduced toxicity
compared with placebo, suggesting an ameliorative effect.
No interaction occurred between the dietary and supple-
mental folate intakes (P = 0.39).
Discussion
This controlled trial shows that folic acid supplementa-
tion of 11 327 nmol (5 mg) or 62 302 nmol (27.5 mg) per
week decreases methotrexate toxicity without compromis-
ing efficacy. The toxicity of methotrexate in both folic acid
groups was low and nearly identical. The data suggest that
dietary folate also helps protect against methotrexate tox-
icity. This finding suggests that the intake of one multiple-
vitamin pill containing 900 nmol of folic acid (400 jLig/d)
may also modulate methotrexate toxicity in patients with
other micronutrient deficiencies (57).
We designed this trial using completers analysis. One
year into the study, we considered modifying the trial
toward intention-to-treat analysis. However, we could not
have obtained complete data on the participants who had
already withdrawn in the first year. We wanted to com-
pare our study with other methotrexate clinical trials that
used completers analyses and thus chose not to analyze
the data by intent to treat. The overall results of the study
were not greatly influenced by the study design. By using
intent-to-treat analysis, we might have observed a lesser
degree of difference for efficacy, but we would have ob-
served the same toxic effects and resulting discontinuation
of drug therapy.
We also re-examined the data on the basis of the most
recent recommendations from the American College of
Rheumatology for monitoring hepatic conditions in pa-
tients with rheumatoid arthritis who are receiving meth-
otrexate, but this did not change the toxicity scores (58).
We previously showed that low baseline plasma and
erythrocyte folate levels can predict future toxicity (29).
Because large doses of folic acid can mask and exacerbate
vitamin B 1 2 deficiency, adequate vitamin B 1 2 status should
be assured before folic acid supplementation (59). Based
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on the data presented, baseline mean corpuscular volume,
blood folate, and vitamin B, 2 values should be deter-
mined in patients in whom methotrexate will be initiated.
Because anemia is a late finding in nutritional deficien-
cies, and patients may have dimorphic anemia (iron plus
B 12 or folate deficiency), yielding a normal mean corpus-
cular volume, we think vitamin levels are useful. There is
no contraindication to starting folic acid at a dose of 5 to
7 mg/wk during methotrexate therapy initiation. Supple-
mentation with higher doses may be indicated if an in-
creasing mean corpuscular volume is observed or if sus-
pected methotrexate toxic effects occur. The dose may be
adjusted to levels as high as 27.5 mg/wk to control or
reverse both of these clinical findings. Serial evaluation of
the mean corpuscular volume has been suggested as an
inexpensive means to monitor incipient toxicity (60).
However, other investigators have found that the mean
corpuscular volume does not predict toxicity in patients
given folic acid (30). Our data suggest that a high mean
corpuscular volume and a low blood folate level are often
associated with methotrexate toxicity.
The mechanism or mechanisms by which folate pro-
duces the clinical effects on toxicity are not clear. Folinic
acid, 5-formyl-tetrahydrofolic acid, also has been used to
reduce side effects of methotrexate (31-36). Perhaps folic
acid and folinic acid correct a state of folate deficiency, or
perhaps folinic acid, by circumventing dihydrofolate re-
ductase inhibition, relieves toxicity. However, these two
mechanisms are not mutually exclusive.
The vitamin form and ratio of folate to methotrexate
are important factors in determining whether efficacy will
be lost with combination vitamin and antivitamin therapy.
Because folinic acid is a fully reduced and one-carbon-
substituted folate, it can bypass folate-metabolizing steps
that reduce the pteridine moiety and add a one-carbon
fragment. Table 4 shows the ratios of folate to metho-
trexate and clinical outcomes in previous studies. Folinic
acid in relatively high doses negates the effectiveness of
methotrexate therapy for rheumatoid arthritis (31). Our
previous trial used a folic acid-to-methotrexate ratio of
0.93 to 1, causing decreased toxicity but not decreased
efficacy (29). Similar results are now reported with a ratio
as high as 2.85 to 1. The participants who received 27.5
mg/wk had a folate-to-methotrexate ratio that was much
higher than in the folinic acid studies by Joyce and col-
leagues and Tishler and colleagues (31, 32), in which
efficacy was negated. A study by Duhra (61) using 35 mg
of folic acid per week during methotrexate therapy for
psoriasis showed no loss of efficacy and supports a wide
margin of safety for folic acid. Although folinic acid can
lessen methotrexate toxicity, the dose level is more critical
than that for folic acid.
The optimal dosing schedule of folate supplements in
relation to methotrexate is not known. The half-life for
oral, intramuscular, and intravenous low-dose methotrex-
ate is about 4.5 to 13 hours (62-65). Table 4 shows the
timing of the folate to methotrexate in previously pub-
lished reports. In the studies by Joyce and associates (31),
Tishler and coworkers (32), and Buckley and colleagues
(34), folinic acid was given within the first half-life of the
methotrexate dose. The ratio of folate to methotrexate
used by these three investigators was 2.3, 0.95, and 0.5,
respectively, with the lowest ratio causing no flare in
 Table 4. Ratios of Folic Acid to Methotrexate and Folinic Acid to Methotrexate and Effects on Clinical Efficacy*

Reference Weekly Folate-to- Effect on Efficacy in Timing of Folate Relative to

Methotrexate Ratio Rheumatoid Arthritis Methotrexate Dose

Folinic acidf
31 2.3 : 1 Flare in disease activity Oral folinic acid 2 hours after weekly oral
methotrexate dose
32 0.95 : 1 Flare in disease activity 15 mg oral folinic acid for 3 consecutive days,
starting 4 to 6 hours after intravenous meth-
otrexate dose
33 0.5:1 No adverse effect on disease activity Folinic acid orally for 2 days, beginning 4 days
before next intramuscular methotrexate dose
34 0.5 : 1 No adverse effect on disease activity Oral folinic acid 4 hours after oral methotrex-
ate dose
35 0.09 : 1 No change in efficacy—one patient in Oral folinic acid 24 hours after oral metho-
folinic acid arm for lack of efficacy trexate dose
36 0.05 : 1 No change in efficacy Oral folinic acid given simultaneously with oral
methotrexate dose
Folic acid
29 0.93 : 1 No change in efficacy Not specified
30 1: 1 No change in efficacy Not specified
This trial 0.53 : 1 and 2.85 : 1 No change in efficacy Folic acid administered on 2 days when oral
methotrexate not taken

* Folic acid = Pteroylglutamic acid; folinic acid = 5-Formyl-tetrahydro-pteroylglutamic acid.

t Only one half of the folinic acid dose is biologically active because of the presence of stereoisomers.

disease activity. In contrast, when the two highest ratios
were administered, a flare in disease activity occurred,
which suggests that the ratio of folate to methotrexate,
not the dosing interval, was more important in determin-
ing whether efficacy is altered.
The cost of folinic acid compared with that of folic acid
therapy is also relevant (36). At our center, the price of a
dose of folinic acid is 43 times greater than the price of
an equivalent dose of folic acid; these figures are probably
similar to costs nationwide. Thus, safety and cost would
support folic acid as the better supplement. A controlled
trial of folic acid and folinic acid would answer these
questions most clearly.
The proposal to enrich flour with folic acid to prevent
neural tube defects has led to concern about the possi-
bility of negating the efficacy of methotrexate therapy in
autoimmune diseases (66, 67). Neither our data nor the
data of others support this concern (61). Enrichment of
flour to provide folic acid intakes of 900 to 2265 nmol
(400 jLtg to 1 mg/d) would not affect efficacy of low-dose
methotrexate therapy for rheumatic disease.
The enzyme or enzymes that must be inhibited to allow
for the efficacy of methotrexate therapy is not known. We
have postulated that inhibition of aminoimidazole carbox-
amide ribotide transformylase is necessary for the efficacy
of methotrexate therapy (68).
Widely varying dose levels of folic acid decrease the
toxicity of methotrexate without affecting its efficacy. The
flexibility in the dosing range, coupled with its low cost,
make folic acid the preferred vitamin form to treat pa-
tients with rheumatoid arthritis who are taking methotrex-
ate.
Appendix
Toxicity score = duration of toxic events [weeks] X (in-
tensity) X (clinical severity factor) + weeks in the proto-
col, where intensity is 1 (mild), 2 (moderate), or 3 (se-
vere), as gauged by patient report; and the clinical
severity factor is 1 (alopecia, malaise or fatigue, nausea,
pruritus, anorexia, or general gastrointestinal intolerance
[pyrosis, cramps, and so on]), 2 (vomiting, diarrhea, sto-
matitis, rash, headache, elevated liver enzyme levels, pe-
ripheral nodulosis), 3 (elevated serum creatinine level), or
4 (cytopenias, documented infections, or pulmonary tox-
icity, systemic nodulosis, profound lethargy). Any abnor-
mality in liver functions or complete blood cell count,
documented infection, or pulmonary toxicity was given an
intensity score of 3 (severe). Abnormal liver function tests
were defined as an aspartate aminotransferase or an al-
kaline phosphatase level greater than two times baseline;
cytopenias were defined as a leukocyte count less than 3.5
X 109/L or a platelet count less than 150 X 109/L; and
serum creatinine concentration was considered elevated if
more than 133 /xmol/L (1.5 mg/dL). Pulmonary toxicity
was defined as evidence of new interstitial pulmonary
infiltrates from baseline chest radiograph and evidence of
restrictive changes in pulmonary function tests, including
vital capacity less than 80% of predicted value, normal
expiratory flow rates, normal maximal voluntary ventila-
tion, and carbon monoxide diffusion capacity less than
70% of normal. Infection was defined as a documented
viral, bacterial, or fungal infection that compromised the
patient greatly and required hospitalization or administra-
tion of systemic antimicrobial agents, or both.
Acknowledgments: The authors thank their patient volunteers and their
referring rheumatologists (faculty and fellows) for participating in the
study and the staffs of the General Clinical Research Center, the Kirklin
Clinic, and the University of Alabama at Birmingham Clinical Vitaminol-
ogy Lab for assisting in the investigation.
Grant Support: In part by the National Institutes of Health Department of
Research Resources Clinical Research Center grant RR-32-31S1; United
States Public Health Service grants 5P01-CA-28103-10, 5P60-AR-20614,
and AR-03555; and Veteran's Administration Merit Review Award AR-
03555.
Requests for Reprints: Sarah L. Morgan, MD, RD, 212 Webb, Department
of Nutrition Sciences, U.A.B. Station, Birmingham, AL 35294-3360.
Current Author Addresses: Dr. Morgan: 212 Webb, Department of Nutri-
tion Sciences, U.A.B. Station, Birmingham, AL 35294-3360.

1 December 1994 • Annals of Internal Medicine • Volume 121 • Number 11 839

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 Dr. Baggott: 340 Webb, Department of Nutrition Sciences, U.A.B. Station,
Birmingham, AL 35294-3360.
Mr. Vaughan: 437 Webb, Department of Nutrition Sciences, U.A.B. Sta-
tion, Birmingham, AL 35294-3360.
Ms. Austin: CH 19 420, U.A.B. Station, Birmingham, AL 35294-2041.
Ms. Veitch: 210 Webb, Department of Nutrition Sciences, U.A.B. Station,
Birmingham, AL 35294-3360.
Dr. Lee: WTI 153, Cancer Center Biostatistics Unit, Division of Hema-
tology/Oncology, Department of Medicine, U.A.B. Station, Birmingham,
AL 35294-3300.
Dr. Koopman: 429A THT, Division of Clinical Immunology and Rheuma-
tology, Department of Medicine, U.A.B. Station, Birmingham, AL 35294-
0006.
Dr. Krumdieck: 326 Webb, Department of Nutrition Sciences, U.A.B.
Station, Birmingham, AL 35294-3360.
Dr. Alarcon: 301 MEB, Division of Clinical Immunology and Rheumatol-
ogy, Department of Medicine, U.A.B. Station, Birmingham, AL 35294-
3296.
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