Granulocyte-Macrophage Colony-Stimulating Factor as ...

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Granulocyte-Macrophage Colony-Stimulating Factor as Immunomodulating Factor Together with Influenza Vaccination in Stem Cell Transplant Patients Karlis Pauksen,1 Annika Linde,2 Viera Hammarstro¨m,3 Jan Sjo¨lin,1 Jan Carneskog,4,a Gunilla Jonsson,2 ¨ berga,1 Hans Engelmann,5 and Per Ljungman3 Gunnar O

From the 1Department of Medical Sciences, Sections of Infectious Diseases and Medicine, Uppsala University Hospital, Uppsala, 2 Swedish Institute for Infectious Disease Control, Solna, Department of Laboratory Medicine, Karolinska Hospital, Stockholm, 3 Department of Hematology, Huddinge University Hospital, Karolinska Institute, Huddinge, 4Department of Medicine, Section of Hematology, Sahlgrenska University Hospital, Go¨teborg, Sweden; and 5Sa¨chsisches Serumwerk, Dresden, Germany

The effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) on the serological response at influenza vaccination was studied in 117 patients who had undergone stem cell transplantation (SCT). The vaccine response was evaluated as significant increases in levels of influenza hemagglutination-inhibition (HAI) antibodies and of IgG antibodies measured by enzyme-linked immunosorbent assay (ELISA). There was no difference in antibody response to either influenza A or B in 64 patients who received GM-CSF at vaccination, compared with the 53 who did not. In the subgroup of allogeneic SCT patients, HAI showed that the response rate to the influenza B vaccine was significantly higher in the treatment group (P ! .05). ELISA showed that autologous SCT patients with breast cancer who received GM-CSF had a better response to influenza A (P ! .05 ) and B (P ! .01). At early vaccination, 4–12 months after stem cell transplantation, these responses were more pronounced. GMCSF appears to improve the response to influenza vaccination in some groups of SCT patients, but only to a limited extent.

Among stem cell transplantation (SCT) patients, influenza virus causes increased morbidity and mortality [1–3]. After SCT, many patients lose the immunity established by previous immunization or natural infection and need to be revaccinated to obtain protective antibody levels [4–7]. In most stem cell transplantation centers, revaccination of SCT patients usually begins >1 year after transplantation, when the immune system has recovered and matured [8]. However, studies of tetanus vaccination administered sooner after transplantation have shown favorable results [9, 10]. Previous studies of influenza vaccination have shown a low response rate in SCT patients vaccinated during the first 2 years following transplantation, and when a 2-dose regimen has been used [11]. Because of the high morbidity and mortality associated with influenza in these

Received 2 June 1999; revised 30 September 1999; electronically published 7 February 2000. Informed consent was obtained from the patients or their guardians, and study was approved by the ethical committees at the participating hospitals and by the Swedish Medical Products Agency. Grant support: The study was supported by grants from the Swedish Cancer Foundation and the Swedish Schering-Plough Company. a Deceased. Reprints or correspondence: Dr. Karlis Pauksen, Department of Infectious Diseases, Uppsala University Hospital, Akademiska Sjukhuset, SE 751 85 Uppsala, Sweden ([email protected]) Clinical Infectious Diseases 2000; 30:342–8 q 2000 by the Infectious Diseases Society of America. All rights reserved. 1058-4838/2000/3002-0018$03.00

patients, especially soon after SCT, it is crucial to improve the response to influenza vaccine [1, 2]. The hematopoietic growth factor granulocyte-macrophage colony-stimulating factor (GM-CSF) induces enhanced proliferation of progenitor cells and improves the function of antigenpresenting cells [12–15]. Studies in animals have shown increased antibody titer responses after vaccination in combination with GM-CSF [12, 13]. In humans, GM-CSF administered at vaccination has been shown to result in an improved response to hepatitis B vaccine in both healthy and immunocompromised patients [16, 17]. In studies of vaccination against malignancies, GM-CSF has been shown to induce long-lasting antibody responses [13, 18, 19]. Because of the immunostimulating effect and activation of antigen-presenting cells, administration of GM-CSF may be a way to improve the response rate at vaccination in SCT patients. In this study of SCT patients, we attempted to improve the response rate at influenza vaccination, especially soon after stem cell transplantation, by also administering GM-CSF.

Patients and Methods Patients. We studied 117 patients aged 116 years who had undergone allogeneic or autologous stem cell transplantation at the university hospitals of Uppsala, Huddinge, or Gothenburg. Eightythree patients were vaccinated between 4 and 12 months and 34 patients between 12 and 24 months after SCT (interval was calculated according to the transplantation day). Patient character-

CID 2000;30 (February)

GM-CSF and Influenza Vaccine

istics are shown in table 1. Exclusion criteria were as follows: (1) immunoglobulin therapy given within 3 months preceding study entry; (2) ongoing infections; (3) known or suspected allergy to components of the vaccines, to GM-CSF, or other recombinant drugs; (4) ongoing immunosuppressive therapy for chronic graftversus-host disease (GVHD) with the exceptions of cyclosporin A or corticosteroids (20 was regarded as seroconversion. ELISA-antibody testing. We also used IgG antibody ELISA to determine the antibody responses to influenza HA, targeting recombinant (r) antigens representing influenza type (Texas 36/ 91)A/H1/N1 (rA/H1N1-ag), (Beijing 32/92)A/H3/N2 (rA/H3N2ag), and (Panama 45/90)B (rB-ag) HA (MicroGene Systems, Meriden, CT). IgG ELISA antibody testing was also performed targeting the whole virus influenza antigens A/H3N2-ag (virA/ H3N2-ag) and B/Harbin (virB-ag). The strains A/PR8 and B/Harbin produced from egg-grown virus were used. The sera were diluted 1 : 1000 for both types of ELISAs. We tested 20 acute and convalescent serum pairs from patients known not to have influenza in both the recombinant and whole virus IgG ELISA, and found the mean optical density (OD) difference (53 SD) between the pairs was 0.18. An OD of 0.2 could therefore be safely used as a lower limit for significant increase of specific antibodies in paired samples examined in the same assay. The antibody titers were calculated as OD 3 1000 (dilution factor). The ELISA antigen rep-

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Table 3. Rate of response to vaccination in groups of patients who were or were not given granulocyte-macrophage colony-stimulating factor (GM-CSF), according to type of transplant and time after stem cell transplantation, as measured with ELISA as an increase in IgG antibodies (10.2 OD).

resents various HA epitopes and is not subtype specific. It cannot therefore be used for evaluation of protective antibody levels. There were no epidemics of the influenza type A/H1N1 during the study years. During the 1995–1996 season, influenza type A/ H3N2 dominated, and during the 1996–1997 season, there was a cocirculation of the influenza type A/H3N2 and influenza B. For both seasons, the vaccine strains were well matched to the influenza strains circulating in the community. Statistical analyses. The largest increase of antibody titers were used from the HAI and the ELISA tests for each patient, as measured at 3 or 6 weeks after vaccination. For presentation, antibody were logarithmically transformed; an HAI antibody titer !10 was set to 5 in the calculations. The Mann-Whitney U test or the Wilcoxon paired test were used to compare increases in antibody titers between different patient groups and control subjects. The x2 test or Fisher’s exact test were used, where appropriate, to compare the response rates between the different patient groups and control subjects. The null hypothesis was defined as the increase in response with GM-CSF !25%. To test this with a power of 80% and a significance level of 5%, 53 patients were needed in each group for the primary analysis.

Type of Antigen rA/H1N1-ag Patient group

GM-CSF

All Allogeneic Autologous !12 Allogeneic !12 Autologous !12 112

20/63 8/28 12/35 15/47 5/19 10/28 5/16

virB-ag

No adjuvant

(32) (29) (34) (32) (26) (36) (31)

19/53 7/22 12/31 9/35 4/16 5/19 10/18

(36) (32) (39) (26) (25) (26) (56)

GM-CSF 25/64 9/28 16/36 20/48 6/19 14/29 5/16

Table 2. Rate of response to vaccination in groups of patients who were or were not given granulocyte-macrophage colony-stimulating factor (GM-CSF), according to type of transplant and time following stem cell transplantation, as measured with hemagglutination inhibition (HAI) as a 4-fold antibody titer increase. Types of influenza A/H1N1

All Allogeneic Autologous !12 Allogeneic !12 Autologous !12 112

18/63 7/28 11/35 14/47 6/19 8/28 4/16

(29) (28) (31) (30) (32) (29) (25)

A/H3N2

No adjuvant 16/53 6/22 10/31 11/35 5/16 6/19 5/18

(30) (27) (32) (31) (31) (32) (28)

12/53 3/22 9/31 4/35 0/16 4/19 8/18

(23) (14) (29) (11) (0) (21) (44)

virus antigen virB-ag was chosen for presentation of antibody reactivity to influenza B. Antibody responses in healthy control subjects and patients. The frequency of protective HAI antibodies was low in patients (12%–16%) and control subjects (0–15%) before vaccination. We evaluated the response rate as a 4-fold HAI titer increase after vaccination in both healthy control subjects (n = 13 ) and patients (n = 117). Results were as follows: response to A/H1N1 was 46% in control subjects versus 29% in patients, (P not significant [NS]); response to A/H3N2, 54% versus 25% (P ! .05, x2 test); to B, 62% versus 34% (NS). For IgG antibodies measured with ELISA, the corresponding responses to rA/ H1N1-ag were 77% versus 34 % (P ! .05); to virB-ag, 69% versus 32% (P ! .05). Antibody responses in patients given and not given GM-CSF. When we compared all 64 patients who received GM-CSF at

Expression of ELISA results. For the 3 influenza A antigens used in ELISA (rA/H1N1-ag, rA/H3N2-ag and virA/H3N2ag), patients who did and those who did not receive GM-CSF showed similar response rates at vaccination. There was a statistically significant correlation between responses to the rA/ H3N2-ag and the virA/H3N2-ag. Because there was no epidemic of the influenza type A/H1N1 during the study years in Sweden, the results from the rA/H1N1-ag ELISA were chosen for further presentation and analysis of antibody reactivity to influenza A after vaccination. The response to the rB-ag after vaccination was very low in healthy control subjects. Therefore, the response to the whole

GM-CSF

No adjuvant

NOTE. Data are no. responded to vaccination/total no. vaccinated in group (%). !12 and 112, months between transplantation and vaccination. OD, optical density; rA/H1N1-ag, recombinent antigen (Texas 36/91) A/H1/N1; virB-ag, whole virus influenza antigen B/Harbin. a P < .01 (x2 test). b P < .05 (x2 test).

Results

Patient group

(39) (32) (44) a (42) b (32) (48) (31)

GM-CSF 15/63 6/28 9/36 8/48 2/19 6/28 7/15

(24) (21) (25) (17) (11) (21) (47)

B

No adjuvant 14/53 3/22 11/31 8/35 1/16 7/19 6/18

(26) (14) (35) (23) (6) (37) (33)

GM-CSF 26/64 10/28 16/36 20/48 7/19 13/29 6/16

(40) a (36) (44) b (42) (37) (45) (38)

No adjuvant 14/53 2/22 12/31 5/35 0/16 5/19 8/18

(26) (9) (39) (14) (0) (26) (44)

NOTE. Data are no. responded to vaccination/total no. vaccinated in group (%). !12 and 112, patients vaccinated !12 mo after and 112 mo after transplantation, respectively. A/H1N1, A/Singapore/6/86(H1N1)–like strain; A/H3N2, A/Johannesburg/33/94-(H3N2)–like strain; B, influenza B. a P < .05 (x2 test). b P < .01 (x2 test).



vaccination with those 53 who did not, we found no significant differences in increases in antibody titers or in the response rates to influenza A(H1N1, H3N2) or B, measured either with HAI or with ELISA (tables 2 and 3). In the subgroup of allogeneic SCT patients, which consisted mainly of those with hematologic malignancies, HAI showed that the response rate to influenza B was significantly higher in those patients who received GM-CSF than in those who did not (10 of 28 vs. 2 of 22; P ! 0.05, x2 test; table 2). The group of autologous SCT patients consisted of patients with different hematologic malignancies, lymphomas, and solid tumors (mainly breast cancer; table 1). In the subgroup of patients with breast cancer, IgG ELISA showed that patients who received GM-CSF had a significantly higher antibody reactivity to rA/H1N1-ag (P ! .05) and to virB-ag (P ! .01 ) than did those who did not receive GM-CSF. No significant differences were found in the subgroups of lymphoma and hematologic malignancies (table 4). When the response was measured with HAI, no significant differences were found in any of the subgroups (table 5). At early vaccination (4–12 months after transplantation), HAI showed that the response rate to influenza B was significantly higher in those 48 patients who received GM-CSF than in those 35 patients who did not (20 of 48 vs. 5 of 35; P ! .01); IgG ELISA showed similar results (20 of 48 vs. 4 of 35; P ! .01; tables 2 and 3). This improved response was also seen in the allogeneic SCT patients, as measured by both HAI (7 of 19 vs. 0 of 16, P ! .01, Fisher’s exact test) and IgG ELISA (6 of 19 vs. 0 of 16, P ! .05). Among autologous SCT patients, an improved antibody reactivity with the rA/H1N1-ag (P ! .05) and the virB-ag vaccines (P ! .05) was seen only in the subgroup of patients with breast cancer (table 4). Neither HAI nor IgG ELISA showed any significant differences in the response rates to influenza A in the whole group of patients vaccinated early. GVHD, TBI and T cell depletion. No significant differences in antibody responses were found between patients who had received TBI (n = 58) and those who did not (n = 59), or among allogeneic SCT patients who had chronic GVHD treatment (n = 13) at vaccination and those who did not (results not shown). Neither TBI nor chronic GVHD treatment significantly influenced the response rates, irrespective of whether patients were given GM-CSF at the time of influenza vaccination. Antibody responses in revaccinated patients. Eleven patients were vaccinated in the autumn of 1995 and revaccinated in the autumn of 1996. HAI showed that antibody titers to A/H3N2 increased significantly more at revaccination, compared with the first dose of vaccine given (median, 150; range, 0–1240 vs. median, 0; range, 0–19; P ! .01, Wilcoxon’s paired test), and, for influenza B, the corresponding increase in antibody titers were seen (median, 240; range, 0–2500 vs. median; range, 0–150; P ! .05, Wilcoxon’s paired test). ELISA showed that antibody titers to rA/H1N1-ag also increased more at revaccination (me-

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Table 4. Rate of response in the largest diagnosis groups of patients who underwent autologous stem cell transplantation and who were or were not given granulocyte-macrophage colony-stimulating factor (GM-CSF), as measured with ELISA as an increase in IgG antibodies (optical density 10.2). Type of Antigen rA/H1N1-ag Diagnosis group a

Hematalogic diseases All lymphoma All breast cancer Breast cancer !12

virB-ag

GM-CSF

No adjuvant

GM-CSF

No adjuvant

2/7 (29) 3/10 (30) b 9/16 (56) c 8/13 (62)

5/8 (63) 2/6 (33) 3/15 (20) 1/10 (10)

3/8 (38) 3/10 (30) c 10/16 (63) b 8/13 (62)

3 /8(38) 2/6 (33) 2/15 (13) 2/10 (20)

NOTE. Data are no. responded to vaccination/total no. vaccinated in group (%). !12, months between transplantation and vaccination. rA/H1N1-ag, recombinant antigen (Texas 36/91) A/H1/N1; virB-ag, whole virus influenza antigen B/ Harbin. a Hematologic malignancies: acute lymphoblastic leukemia, acute myeloblastic leukemia, chronic lymphogenous leukemia, chronic myelogenous leukemia, and multiple myeloma). b P < .05 (x2 test). c P < .01 (x2 test).

dian, 670; range, 0–2200 vs. median, 35; range, 0–630; P ! .05, Wilcoxon’s paired test). The response rates at revaccination compared with the first dose of vaccine given within 12 months after transplantation, as measured by HAI, were as follows: response to A/H1N1, 46% versus 18% (NS); to A/H3N2, 73% versus 0% (P ! .05, Fisher’s exact test); and to B, 64% versus 27% (NS). The corresponding rates measured with ELISA were as follows: response to rA/H1N1-ag, 73% versus 27% (P ! .05); and to virB-ag 64% versus 55% (NS). Adverse events. Severe adverse events were reported in 5 patients (8%) who received GM-CSF and in none for those who did not. Three patients had marked inflammatory reactions with muscle pain and/or pelvic aches, 2 with CML and 1 with AML who had undergone allogeneic SCT. The fourth patient, who had testicular cancer and had undergone autologous SCT, had a severe myalgic reaction with chest pain and was observed overnight at the hospital. These reactions started within an hour after injection and resolved without treatment after ∼3–4 h. One woman with lymphoma, who had undergone autologous SCT and received GM-CSF, had a possible reactivation of her autoimmune thyroiditis. Because most patients lived far from the university hospitals, they were questioned mainly by telephone. Thus local reactions at the vaccination site could not be registered objectively. Patients who received GM-CSF seemed to have had a stronger local inflammatory reaction than did patients who did not receive GM-CSF.

Discussion In this study, the population consisted of a heterogeneous group of patients with many different underlying diseases. In the group of patients who underwent allogeneic SCT, almost all had a hematologic malignancy with different diagnosis. The

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Table 5. Rate of response to vaccination in the 2 largest diagnosis groups of patients who underwent autologous stem cell transplantation and who were or were not given granulocyte-macrophage colony-stimulating factor (GM-CSF), as measured with hemagglutination inhibition (HAI) as a 4fold antibody titer increase. Types of influenza A/H1N1 Diagnosis group a

Hematalogic diseases All lymphoma All breast cancer Breast cancer !12

A/H3N2

B

GM-CSF

No adjuvant

GM-CSF

No adjuvant

GM-CSF

No adjuvant

3/8 (38) 3/10 (30) 4/15 (27) 4/12 (25)

3/8 (38) 1/6 (17) 5/15 (33) 3/10 (30)

1/8 (13) 1/10 (10) 7/16 (44) 4/13 (31)

2/8 (25) 0/6 (0) 8/15 (53) 5/10 (50)

2/8 (25) 5/10 (50) 9/16 (56) 7/13 (54)

1/8 (13) 3/6 (50) 5/15 (33) 3/10 (30)

NOTE. Data are no. responded to vaccination/total no. vaccinated in group (%). !12, months between transplantation and vaccination. A/H1N1, A/Singapore/6/86-(H1N1)–like strain; A/H3N2, A/Johannesburg/33/ 94-(H3N2)–like strain. a Hematological malignancies: acute lymphoblastic leukemia, acute myeloblastic leukemia, chronic lymphogenous leukemia, chronic myelogenous leukemia, and multiple myeloma).

group of autologous SCT patients consisted of patients with different hematologic malignancies, lymphomas, and solid tumors, mainly breast cancer. However, in the case of allogeneic SCT, previous studies have shown that the most important factors for the response to vaccination are the recovery from the procedure itself and GVHD treatment, not the underlying disease [5]. Therefore, the allogeneic SCT patients could be regarded as a single group. Although the immunosuppressive treatment for GVHD might affect the results, only those who had received a low dose of immunosuppressive treatment were included. Furthermore, only a small number of the patients in the study received immunosuppressive therapy for GVHD, and these patients were evenly distributed among the different groups. Although most studies have not found that different patient groups who underwent autologous SCT differed in response to vaccination or loss of immunity to previous vaccinations, there still might be a difference that was not detected because of the low number of patients in each patient group [4, 6]. A difference in retainment of tetanus immunity was found in a study on patients with hematologic malignancies. This study has shown that patients with advanced disease stage or lymphoid malignancy more frequently lost antibodies to tetanus [25]. Therefore, for the group of autologous SCT patients, we have shown the results from the major diagnosis groups and the entire group of SCT patients separately. Before SCT, no routine vaccination against influenza was performed, and most of the patients were seronegative at vaccination. This suggests that a booster response at vaccination could not be expected, as has been shown for SCT patients who were seronegative for polio virus [6]. The aim of this study was to try to improve the antibody response to influenza vaccination in SCT patients by administering GM-CSF. We used conventional methods to determine the HAI antibody responses to the vaccine strains, and we also measured the IgG antibody reactivity to various influenza A and B antigens by use of ELISA. As determined by HAI, response rates after influenza vaccination (25%–34%) were similar

to those found in previous studies of SCT patients and solid organ transplant patients [11, 26–29]. The response rate at the time of influenza vaccination among patients who had undergone SCT during the preceding 2 years also remained lower than the rate in healthy control subjects. Among healthy subjects, the response rate at influenza vaccination is usually 70%–80%, but in our study it was between 46% and 62% when measured as a fourfold increase in HAI-antibody titers. The main reason for this low frequency was probably the low number of healthy subjects tested. However, in healthy people the response rate measured with HAI might also be lower, as shown by the immunogenicity data given above in Patients and Methods. The HAI results for the sera included were similar at parallel testing from two laboratories. The prevaccination titers to the influenza strains used were low in study patients and control patients; thus high titers prior to vaccination could not interfere with the response rate. When the antigen reactivity was measured in IgG ELISAs, the response rates were higher both in control subjects (69%–77%) and in patients (34%–43%) than they were when measured with HAI. This is not surprising since, unlike HAI, the antibodies measured by ELISA are not epitope-specific and can detect all kinds of influenza-reactive antibodies. The response rate to influenza B was higher than the response rate to A among both study patients and control subjects, when measured by HAIantibody titers and IgG ELISA with 3 virus antigens. Either the B vaccine was more immunogenic or the assays for influenza B were more efficient than those for the A types. Overall, there were no statistically significant differences in the response rates between patients who received GM-CSF and those who did not. However, at vaccination !1 year after SCT, patients who received GM-CSF had a higher response rate to the influenza B strain. The group of allogeneic SCT patients who received GM-CSF had a higher response rate to the influenza B antigen than those who did not, and the difference was more pronounced at early vaccination when measured with both HAI and ELISA. In autologous SCT patients with breast cancer who received GM-CSF, IgG ELISA showed an in-



creased antibody reactivity both to influenza A and B. This increase was also most pronounced after early vaccination. The other diagnosis groups were too small and too heterogenous to allow any reliable conclusions. Thus the most pronounced effect of GM-CSF as adjuvant at vaccination was seen shortly after transplantation. At later vaccination, the difference diminished. This may be explained by the gradual maturation of the immune system, which results in a better response to vaccination with time after SCT. This improving response has also been shown in a previous influenza vaccination study [11]. A similar tendency was also seen in this study, but since the number of patients who were vaccinated late in this study was small, no firm conclusions could be drawn as to whether GM-CSF had any effect at late vaccination. However, it is soon after SCT that patients contract the most severe influenza infections, and thereby most urgently need vaccine protection. GM-CSF has been shown to increase the expression of major histocompatibility complex (MHC) class I and II molecules on antigen-presenting cells, such as macrophages and dendritic cells, and has been used as an immunomodulator in cancer vaccination [12, 14, 18, 19]. Some studies with cancer vaccines have shown promising results with long-lasting antibody responses [18, 19]. In many of these studies, GM-CSF was given more frequently than in ours. Repeated doses or a slow-release preparation of GM-CSF may be required to obtain a better vaccine response. There is also the possibility that the immune systems of patients who did not respond to vaccination were not mature enough to be stimulated by GM-CSF, and, since quite severe side-effects were related to the agent, the risk/benefit ratio must be much better established before the preparation can be used on a larger scale. However, the adverse events seem to be dose-dependent. The dose of GM-CSF given in this study, 2.5 mg/kg body weight, was chosen from results of a study that showed the best response at influenza vaccination in healthy elderly individuals and the lowest frequency of adverse events [30]. A further increase of the dose showed both an increased frequency of severe adverse events and an impaired response to the vaccine. Later studies of hepatitis B vaccination in healthy individuals showed an increased response at much lower doses of GM-CSF (20–40mg) and a tendency toward impaired responses when the dose was increased [16]. Also, in the cancer vaccination studies, a lower dose has been given, producing fewer side-effects [18]. Because of these results, a bimodal dose response to GM-CSF at vaccination has been suggested. Thus, it is possible that in SCT patients a lower dose of GM-CSF would have been both more efficacious and associated with fewer side-effects. We conclude that GM-CSF at a dose of 2.5 mg/kg body weight appears to improve the response to influenza vaccination in SCT patients, but only to a limited extent. The best effect of GM-CSF as an immunomodulator at vaccination was seen in allogeneic SCT patients in the response to the influenza B strain, and in autologous SCT patients among those who had

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breast cancer, especially during the first year after SCT. Side effects were not negligible. Further studies with varying dosages of GM-CSF are warranted before large-scale use should be considered.

References 1. Ljungman P, Andersson J, Aschan J, et al. Influenza A in immunocompromised patients. Clin Infect Dis 1993; 17:244–7. 2. Whimbey E, Elting LS, Couch RB, et al. Influenza A virus infections among hospitalized adult bone marrow transplant recipients. Bone Marrow Transplant 1994; 13:437–40. 3. Aschan J, Ringde´n O, Ljungman P, Andersson J, Lewensohn FI, Forsgren M. Influenza B in transplant patients. Scand J Infect Dis 1989; 21:349–50. 4. Pauksen K, Duraj V, Ljungman P, et al. Immunity to and immunization against measles, rubella and mumps in patients after autologous bone marrow transplantation. Bone Marrow Transplant 1992; 9:427–32. 5. Ljungman P, Lewensohn-Fuchs I, Hammarsto¨m V, et al. Long-term immunity to measles, mumps, and rubella after allogeneic bone marrow transplantation. Blood 1994; 84:657–63. 6. Pauksen K, Hammarstro¨m V, Ljungman P, et al. Immunity to poliovirus and immunization with inactivated polio virus vaccine after bone marrow transplantation. Clin Infect Dis 1994; 18:547–52 7. Ljungman P, Wiklund HM, Duraj V, et al. Response to tetanus toxoid immunization after allogeneic bone marrow transplantation. J Infect Dis 1990; 162:496–500. 8. Ljungman P, Cordonnier C, de Bock R, et al. Immunisations after bone marrow transplantation: results of a European survey and recommendations from the infectious diseases working party of the European Group for Blood and Marrow Transplantation. Bone Marrow Transplant 1995; 15:455–60. ¨ lander R-M, Ruutu T, et al. A randomized comparison between 9. Parkkali T, O early and late vaccination with tetanus toxoid vaccine after allogeneic BMT. Bone Marrow Transplant 1997; 19:933–8. 10. Parkkali T, Stenvik M, Ruutu T, Hovi T, Volin L, Ruutu P. Randomized comparison of early and late vaccination with inactivated poliovirus vaccine after allogeneic BMT. Bone Marrow Transplant 1997; 20:663–8. 11. Engelhard D, Nagler A, Hardan I, et al. Antibody response to a two-dose regimen of influenza vaccine in allogeneic T cell–depleted and autologous BMT recipients. Bone Marrow Transplant 1993; 11:1–5. 12. Morrissey PJ, Bressler L, Park LS, Alpert A, Gillis S. Granulocyte-macrophage colony-stimulating factor augments the primary antibody response by enhancing the function of antigen-presenting cells. J Immunol 1987; 139:1113–9. 13. Liehl E, Hildebrandt J, Lam C, Mayer P. Predication of the role of granulocyte-macrophage colony-stimulating factor in animals and man from in vitro results. Eur J Clin Microbiol Infect Dis 1994; 13(Suppl 2):9–17. 14. Fischer H-G, Frosch S, Reske K, Reske-Kunz AB. Granulocyte-macrophage colony-stimulating factor activates macrophages derived from bone marrow cultures to synthesis of MHC class II molecules and to augmented antigen presentation function. J Immunol 1988; 141:3882–8. 15. Bender A, Bui LK, Feldman MAV, Larsson M, Bhardwaj N. Inactivated influenza virus, when presented on dendritic cells, elicits human CD81 cytolytic T cell responses. J Exp Med 1995; 182:1663–71. 16. Tarr PE, Lin R, Mueller EA, Kovarik JM, Guillaume M, Jones TC. Evaluation of tolerability and antibody response after recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) and a single dose of recombinant hepatitis B vaccine. Vaccine 1996; 14: 1199–1204. 17. Hess G, Kreiter F, Ko¨ster W, Deusch K. The effect of granulocyte-macrophage colony-stimulating factor (GM-CSF) on hepatitis B vaccination in hemodialysis patients. J Viral Hepat 1996; 3:149–53.

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Pauksen et al.

18. Osterborg A, Yi Q, Henriksson L, et al. Idiotype immunization combined with granulocyte-macrophage colony-stimulating factor in myeloma patients induced type I, major histocompatibility complex–restricted, CD8and CD4-specific T-cell responses. Blood 1998; 91:2459–66. 19. Samanci A, Yi Q, Fagerberg J, et al. Pharmacological administration of granulocyte/macrophage-colony-stimulating factor is of significant importance for the induction of a strong humoral and cellular response in patients immunized with recombinant carcinoembryonic antigen. Cancer Immunol Immunother 1998; 47:131–42. 20. Ringde´n O, Pihlstedt P, Markling L, et al. Prevention of graft-versus-host disease with T-cell depletion or cyclosporin and methotrexate: a randomized trial in adult leukemic marrow recipients. Bone Marrow Transplant 1991; 7:221–6. 21. Lo¨nnerholm G, Carlsson K, Bratteby LE, et al. Renal function after autologous bone marrow transplantation. Bone Marrow Transplant 1991; 8: 129–34. 22. Simonsson B, Burnett AK, Prentice HG, et al. Autologous bone marrow transplantation with monoclonal antibody purged marrow for high risk acute lymphoblastic leukemia. Leukemia 1989; 3:631–6. 23. Ljungman P, Bjo¨rkstrand B, Fornander T, et al. High-dose chemotherapy with autologous stem cell support in patients with responding stage IV breast cancer. Bone Marrow Transplant 1998; 22:445–8.


24. Lennette EH, Lennette DA, Lennette ET. Diagnostic procedures for viral, rickettsial and chlamydial infections. 7th ed. Washington, DC: American Public Health Association, 1995:441–5. ¨ berg G, Paul C, Ljungman P. 25. Hammarstro¨m V, Pauksen K, Svensson H, O Tetanus immunity in patients with hematological malignancies. Support Care Cancer 1998; 6:469–72. 26. Blumberg EA, Albano C, Pruett T, et al. The immunogenicity of influenza virus vaccine in solid organ transplant recipients. Clin Infect Dis 1996; 22:295–302. 27. Grekas D, Alivanis P, Kiriazopoulou V, et al. Influenza vaccination on renal transplant patients is safe and serologically effective. Int J Clin Pharmacol Ther Toxicol 1993; 31:553–6. 28. Edvardsson VO, Flynn JT, Deforest A, et al. Effective immunization against influenza in pediatric renal transplant recipients. Clin Transplant 1996; 10:556–60. 29. Beyer WE, Diepersloot Rj, Masurel N, Simoons ML, Weimar W. Double failure of influenza vaccination in a heart transplant patient [letter]. Transplantation 1987; 43:319. 30. Taglietti M. Vaccine adjuvance: a new potential area of development for GM-CSF [abstract 00]. 3d Dendritic Cells Symposium (Annecy, France), 1994:34.