Pediatr Blood Cancer 2008;50:822–825
Safety, Efficacy, and Immune Reconstitution After Rituximab Therapy in Pediatric
Patients with Chronic or Refractory Hematologic Autoimmune Cytopenias
Aarati Rao, MD,1Michael Kelly, MD, PhD,2,5Mark Musselman, MD,3Jagadeesh Ramadas, MD,4
David Wilson, MD, PhD,1William Grossman, MD, PhD,1,5and Shalini Shenoy, MD1*
Autoreactive antibodies against hematopoietic cells result in
destruction of platelets, neutrophils or red blood cells resulting
in bleeding, anemia, or infectious complications that are
concerning especially in children. Traditional treatment strategies
for immune thrombocytopenic purpura (ITP) or autoimmune
hemolytic anemia (AIHA) [glucocorticoids, intravenous immune
globulin (IVIG), anti-D] inhibit uptake of antibody coated
hematopoietic cells by phagocytes or decrease antibody production
[1–3]. Chronicity of autoantibody-mediated destruction is rare in
children but poses a therapeutic challenge if it occurs. Alternate
therapeutic options are often invasive (splenectomy) or toxic
(chemotherapy, immunosuppressive agents) and carry a finite risk
of death from toxicity, necessitating surveillance for complications
Targeteddepletion ofB-lymphocytesis an alternate approach to
treatment of autoimmune disorders [8–11]. Rituximab, a huma-
nized chimeric monoclonal antibody directed against the CD20
antigen expressed on pre-B and B-lymphocytes rapidly depletes
target cells by complement-mediated cytotoxicity, antibody-
dependent cytotoxicity, induction of apoptosis and inhibition of
B-cell proliferation . Rituximab has been used to treat immune
cytopenias often with additional safety measures such as replace-
ment immunoglobulin therapy . There are few prospective
clinical trials documenting safety and efficacy of rituximab in
children with hematologic cytopenias . We conducted a
prospective multicenter trial of rituximab therapy for pediatric
hematologic cytopenias that was designed to evaluate safety,
efficacy, and effect of dose escalation, immune reconstitution, and
safe and useful for the treatment of children with autoimmune
The study was approved by Institutional Review Boards at
participating institutions. Informed consent was obtained from
were less than 21 years old, and had ‘refractory’ disease (failed
immunoglobulin and steroid therapy) or were ‘steroid dependent’
which was defined as the inability towean oral methylprednisolone
to less than 1 mg/kg/day after 4 weeks of higher dose therapy.
Patients with active infections, allergy to drug components, prior
therapy with monoclonal antibodies, pregnant, or lactating were
excluded. Splenectomy was not an exclusion criterion. Physical
examination and blood counts were performed at each visit.
Immunologic laboratory tests included absolute numbers of
peripheral blood T- and B-cells (by flow cytometry), serum
immunoglobulin (G, M, and A) levels, and tetanus toxoid antibody
prior to rituximab administration.
Background. Autoimmune hematologic cytopenias in children
often require therapeutic intervention. We report a prospective
pediatric multicenter trial of rituximab for refractory or steroid-
dependent patients. Methods. Four doses of rituximab (375 mg/m2/
dose) were administered weekly. Patients without response after
three doses were offered dose escalation to 750 mg/m2/dose/
week?3. Safety, efficacy, and immunologic tests were evaluated
after therapy. Results. Twenty-nine of 30 children (2–18 years) with
thrombocytopenia (21), hemolytic anemia (6), Evans syndrome (2),
and neutropenia (1) received at least four doses of rituximab. One
developed anaphylaxis with the first dose. One patient was
subsequently diagnosed with monosomy 7 myelodysplasia. Of
28 remaining patients, 9 received dose escalation. Responders
discontinued other therapy following rituximab. The overall
response rate was 72% with median follow-up of 18 months.
Complete remission was observed in 14 (50%); all received four
doses of rituximab. Partial remission (PR) was observed in six (22%);
five had received dose escalation. Of four relapses, 4–24 months
after therapy, two were retreated with rituximab and achieved
second remission. No major infections were encountered. Circulat-
ing B-cells weredepleted by1 monthand normalizedby 1 year. IgM,
Ig A, and IgG levels decreased 6, 9, and 12 months after therapy,
respectively, but remained near normal range. Tetanus toxoid
antibody titers remained detectable. Conclusions. Rituximab was
well tolerated, and induced sustained remissions in children with
refractory immune cytopenias. Dose escalation and re-treatment
after relapse elicited additional responses. Rituximab therapy should
be considered prior to potential interventions with higher toxicity.
Pediatr Blood Cancer 2008;50:822–825.
? 2007 Wiley-Liss, Inc.
autoimmune hemolytic anemia; immune thrombocytopenia; rituximab
? 2007 Wiley-Liss, Inc.
This article contains Supplementary Material available at http://
1Washington University School of Medicine, St. Louis Children’s
Hospital, St. Louis, Missouri;2St. Louis University, Cardinal Glennon
Children’s Hospital, St. Louis, Missouri;3University of Illinois, Carle
Clinic, Champagne, Illinois;
Pennsylvania;5Medical College of Wisconsin, Children’s Hospital of
Wisconsin, Milwaukee, Wisconsin
4Geisinger Medical Center, Danville,
*Correspondence to: Shalini Shenoy, Box 8116, One Children’s Place,
St. Louis, MO 63110. E-mail: firstname.lastname@example.org
Received 12 February 2007; Accepted 16 April 2007
Rituximab was administered at a dose of 375 mg/m2weekly for
4 weeks. Dose escalation of rituximab was offered to patients that
had no improvement in affected blood counts after the first
three doses. The dose was increased to 750 mg/m2weekly?3 to
total six doses if parents/guardians and the medical care team were
in agreement. Premedication prior to each dose consisted of oral
acetaminophen and IV diphenhydramine and hydrocortisone. All
infusions were administered in the outpatient setting over
approximately 4 hours and monitored by medical staff. Side effects
during infusion were documented and treated symptomatically. In
the event of improvement in blood counts, steroid therapy was
tapered over 2–4 weeks following response.
Patients were evaluated for response if they completed 4 or
6 weekly infusions. Response rates were determined based on a
other therapy. Respondents were classified as complete responders
(CR), partial responders (PR), or nonresponders (NR). CR was
defined as platelet counts maintained ?75?109/L; hemoglobin
stable without transfusion ?10 gm/dl; absolute neutrophil count
30–75?109/L; hemoglobin 7.5–10 gm/dl without transfusion;
absolute neutrophil count ?0.5?109/L. Partial response criteria
were chosen arbitrarily based on the premise that cytopenia related
complications were unlikely if PR was achieved.
age were enrolled between January 2001 and July 2005. Twenty-
nine children received at least four doses of rituximab. One patient
discontinued therapy when the first dose resulted in transient
hypotension and bronchoconstriction that responded to withdrawal
incidence of infusion-related events with first and subsequent doses
in other patients were minor and transient. Cough (14%), rash
(10%), muscle cramps (10%), headache (7%), and angioedema
(7%) were most frequent. Shortness of breath, hypertension, chills,
and fever were noticed in 3% of patients. Symptoms subsided after
completion of infusion.
Of twenty-nine children completed at least four doses of
rituximab, one child with severe thrombocytopenia was subse-
quently diagnosed to have monosomy 7 induced myelodysplastic
syndrome that later evolved into acute myeloid leukemia and was
withdrawn from the study. Of the remaining 28, 19 received four
doses of rituximab and 9 received dose escalation to total six doses.
28 patients. Further details on individual patients are available in
Supplemental Table I online.
The overall response rate was 72%. Following four doses
(n¼19), 14 (74%) had CR, 1 had PR, and 4 (21%) were NR. Of the
nine that received six doses, five (55%) achieved PR and four were
received rituximab after failing splenectomy but did not respond.
Three NR with ITP, AIHA, and Evans syndrome proceeded to
splenectomy; all achieved PR.
Median follow-up period was 18 months (range 3–30 months).
Three children who achieved CR (ITP-2; AIHA-1) and one in PR
(ITP) recurred 4, 6, 12, and 24 months after rituximab. Two (ITP-1;
AIHA-1) were retreated with four doses of rituximab and both
responded, achieving either a second CR (1) or PR (1).
Immune reconstitution is described in Table II. Prior to therapy,
peripheral blood B-cells ranged between 6% and 53% (median
16%); absolute number of circulating B-cells were 73–7,350/
cu mm (median 251/cu mm). B-cells were undetectable in
peripheral blood 1 and 3 months after commencing therapy.
at 1 year. Recovery was similar in patients that received four or six
6, and 9 months, respectively; tetanus toxoid antibody titers were
detectable throughout the follow-up period. CD3þ, CD4þ, and
CD8þT-cell numbers remained within normal limits during the
study period (data not shown). No major infectious complications
occurred in any patient. No hospital admissions were necessary
during the period of B-cell depletion.
This study prospectively evaluated the safety of administration
after therapy. There is only one other prospective trial of rituximab
therapy in children which included both untreated and refractory
patients with ITP and reported a 31% response rate . Our study
included children with any immune hematologic cytopenia but
only those patients that were refractory to immunoglobulin or
steroids, or had failed a steroid wean. In this group of patients,
(69.5%) and AIHA (62.5%) were comparable. With a median
follow-up period of 18 months, this high response rate is
encouraging and durable response persisted in 54%. The single
Pediatr Blood Cancer DOI 10.1002/pbc
TABLE I. Patient Demographics and Response to Therapy (n¼28)
Duration of disease prior to rituximab
median (range in months)
Number of doses
Number of responses
ITP, immune thrombocytopenic purpura; AIHA, autoimmune hemolytic anemia; AIN, autoimmune neutropenia; CR, complete response; PR,
partial response; NR, no response.
Rituximab in Pediatric Autoimmune Cytopenias 823
patient with immune neutropenia failed to respond to rituximab.
There is only one previous case report of autoimmune neutropenia
thatwas treated with rituximabsuccessfully . Response ratesof
54% have been described in adult patients with chronic thrombo-
cytopenia . Smaller, retrospective reports have described
response rates between 25% and 78% in adults and children with
ITP and excellent response (100%) in patients with AIHA [8–
10,13,16,17]. The variable response rates are likely attributable to
Only one patient in our series had undergone splenectomy prior
to rituximab. Patients enrolled on this trial would have received
alternate therapeutic interventions to avert complications of
Withdrawal of close monitoring, restricted activity, more therapy,
patients that had initial response support the need for intermittent
vigilance. Relapse occurred after both CR and PR, and at variable
intervals. There seemed to be no relation to B-cell recovery in
peripheral blood. We favor a second course of rituximab based on
the efficacy and low toxicity profile of the drug with repeated
Twelve patients had no response after three doses of rituximab
andninewenton toreceiveatotal ofsixdoseswithdoseescalation.
Responderstosixdoses ofrituximab (five ofnine achieved PR) had
blood counts that increased after completion of the sixth dose and
were either benefited by dose escalation and prolonged therapy or
were late responders. In a previous study, the median time to
response in children with ITP was 3.5 weeks and the majority
responded within 8 weeks after four doses . Pediatric
AIHA responded between 2–4 weeks . Patients with a ‘poor’
response (did not achieve even a PR after the initial three doses of
maybe made foratrialof doseescalation tofurther facilitate B-cell
depletion since there is considerable variability in the strength and
duration of response to rituximab and high dose schedules
have demonstrated longer duration of response without increased
toxicity in malignant disorders [18,19]. It should be noted that the
group of patients with ITP who received six doses had persistent
disease for a longer period (median duration 19 vs. 6 months). It is
encouraging to note that four of the six had partial responses to
therapy. It is also likely that some patients will respond to fewer
doses but developing clear response based treatment criteria with
dose escalation guidelines is difficult.
Rituximab is a murine-human protein conjugate that can cause
allergic reactions. Reversible serum sickness symptoms were
previously reported in 12% [8,14]. In our series, infusion reactions
40% and 48% with first and subsequent doses, respectively, after
premedication. One patient (3%) did not tolerate the drug. No
delayed infusion reactions were encountered.
Immune function analysis was undertaken for correlation with
infection risks. Infectious complications following rituximab are
more common in the setting of adjuvant therapy for malignancy or
systemic autoimmunity and rare with rituximab alone [14,20]. We
encountered no early or delayed infectious complications. B-cell
recovery took 6–12 months irrespective of the dose. Immunoglo-
period. Tetanus toxoid antibody levels were preserved. Routine use
of immunoglobulin prophylaxis is not recommended. However,
1 year until B-cell recovery is complete.
In conclusion, rituximab was well tolerated and provided
durable responses in a significant number of pediatric patients with
refractory or persistent autoimmune hematologic cytopenias. The
high response rate and low toxicity profile warrant a trial of
sion or splenectomy.
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TABLE II. B-Lymphocyte Dependent Immunologic Laboratory Tests Following Rituximab Therapy
Absolute number B-cells
Ig G (mg/dl) median
Tetanus toxoid Ab titer
(IU/ml) Median (SD)
Pretherapy 251 (1,827)1,020 (625)109 (56)100 (66)0.8 (0.8)
1 month0 (64)890 (540)122 (70) 93 (50)0.6 (1)
3 months 0 (52)899 (419)103 (62)44 (36)0.6 (0.5)
6 months17 (124)868 (330) 80 (70)41 (50)0.5 (0.4)
9 months98 (133)826 (375) 75 (78)46 (26) 0.5 (1)
1 year372 (95)738 (327) 87 (62)74 (114) 0.3 (0.2)
N, normal range; SD, standard deviation; Ab, antibody.
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Rituximab in Pediatric Autoimmune Cytopenias 825