Background Treatment with anti-B cell antibody rituximab may ameliorate the disease course in a subgroup of patients with polyneuropathy associated with IgM monoclonal gammopathy. Polymorphisms of leukocyte IgG receptors (FcγR) that influence efficiency of antibody-dependent cell-mediated cytotoxicity determine rituximab efficacy in patients with lymphoma and autoimmune disease.
Objective To investigate the association of FcγRIIA and FcγRIIIA polymorphisms with the response to rituximab treatment in a cohort of patients with polyneuropathy associated with IgM monoclonal gammopathy (PNP-IgM) with and without antimyelin-associated glycoprotein antibodies.
Methods We determined FcγRIIA-R/H131 and FcγRIIIA-V/F158 genotypes in 27 patients with PNP-IgM using allele-specific PCR and Sanger sequencing.
Results The FcγRIIIA-V/V158 genotype was associated with functional improvement (p=0.02) after 1 year.
Conclusions FcγRIIIA polymorphisms are potential biomarkers for response to rituximab treatment in polyneuropathy associated with IgM monoclonal gammopathy.
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Polyneuropathy associated with IgM monoclonal gammopathy (PNP-IgM) is characterised by progressive weakness, sensory ataxia and IgM antibodies against myelin-associated glycoprotein (MAG), sulfated glucuronyl paragloboside or gangliosides, such as GM1, GD1a, GD1b or GQ1b. IgM-PNP may lead to significant disability.1 Despite the presence of auto-antibodies, treatment strategies that target toxic B cell clones have been largely unsuccessful in patients with IgM-PNP.2 However, results from two recent randomised placebo-controlled clinical trials suggest that rituximab treatment may lead to improvement in function in a subgroup of patients.3 ,4 Rituximab is a chimeric monoclonal IgG antibody against the B cell surface protein CD20 and induces B cell depletion by several mechanisms including antibody-dependent cell-mediated cytotoxicity.5 Antibody-dependent cell-mediated cytotoxicity is mediated through IgG receptors (FcγR) on specific leukocytes such as natural killer cells.5 FcγR are classified in three classes and several subclasses.5 ,6 Two FcγR subclasses, FcγRIIA and FcγRIIIA, display single nucleotide polymorphisms that correlate with the efficacy of antibody-mediated leukocyte functions such as antibody-dependent cell-mediated cytotoxicity and are associated with the response to anti-CD20 monoclonal antibody treatment in patients with lymphoma and autoimmune disease.5 ,7 ,8 We therefore investigated whether the FcγRIIA-R/H131 and FcγR IIIA-F/V158 polymorphisms are associated with a favourable response to treatment with rituximab in Dutch patients with IgM-PNP.
Patients and methods
Thirty patients from a prospective cohort of 169 patients with IgM-PNP were treated with rituximab monotherapy between 2004 and 2011.1 Results of treatment in 17 patients were reported previously.9 DNA samples of 27 patients were available for this study. Informed consent was obtained from all patients. All patients were examined before and 1 year after initiation of rituximab treatment by a physician unaware of the results of FcγR genotyping. The treatment protocol consisted of 375 mg/m2 rituximab once weekly for four consecutive weeks.9 The level of functional impairments was assessed using the 12-point inflammatory neuropathy cause and treatment Overall Disability Sum Score (ODSS).9 ,10 Sensory deficits were scored with a 56-point sensory sum score to assess sensitivity to touch and perception of pain, vibration and passive movement. Motor deficits were assessed using an medical research council (MRC) distal sum score of four distal muscle groups in arms and legs on both sides (maximum total score 80 points).9 The primary outcome was defined as improvement of one or more than one point on the ODSS functional score after 1 year. Improvement of the sensory sum score and distal motor score of 5% at assessment 1 year after treatment were used as secondary outcome measures.9
Nerve conduction studies were performed using a standardised protocol and were used to classify the IgM-PNP as ‘demyelinating’ as outlined in the American Academy of Neurology (AAN) research criteria, or ‘axonal’ if these criteria were not fulfilled. The presence of anti-MAG antibodies was determined by means of ELISA (Bühlmann Laboratories, Switzerland).
FcγRIIA-R/H131 and FcγRIIIA-F/V158 polymorphisms were determined using genomic DNA and allele-specific PCR as described previously.5 Sanger sequencing was used to confirm FcγRIIIA genotyping results.5
Differences in outcome after rituximab treatment were assessed using the Fisher's exact test. For statistical analysis FcγRIIA and FcγRIIIA genotypes were divided in efficient (FcγRIIA-H/H131, FcγRIIIA-V/V158) and less efficient (FcγRIIA-R/H131 or R/R131 and FcγRIIIA-F/V158 or F/F158) genotypes for triggering antibody-dependent cell-mediated cytotoxicity.
Clinical characteristics and response to treatment
Twenty-seven patients were included in this study. Their median age was 60 (range 41–78); 17 patients were male. Twenty-six patients had a demyelinating polyneuropathy and 18 had antibodies against MAG. The median ODSS was 4, with a range from 2 to 7. Their median distal motor sum score was 71 (range 30–80), and their median sensory sum score 36 (range 7–50).
Twenty-five of 27 (93%) patients completed 4 weekly rituximab infusions of 375 mg/m2. Data from all 27 patients were used for final analysis. Three out of those 27 patients (11%) improved at least one point on the ODSS and another nine (33%) experienced improvement of sensory or motor sum scores: two patients improved on all three outcome measures, one patient on motor and sensory sum scores, but not on the ODSS, one patient improved only on the ODSS and eight patients improved on sensory sum scores alone. Two patients, both with anti-MAG antibodies, developed a paradoxical worsening of weakness between the second and third rituximab infusion and treatment was therefore discontinued.11
Correlation of FcγRIIA-R/H131 and FcγR IIIA-F/V158 polymorphisms with treatment response
Six patients (22%) had an FcγRIIA-R/R131 genotype, 12 patients (44%) had an FcγRIIA-H/R131 genotype and nine patients (33%) had an FcγRIIA -H/H131 genotype. FcγRIIA-R/H131 genotypes were not associated with any of the outcome measures.
Treatment outcome stratified for FcγRIIIA genotypes are summarised in table 1. Eight patients (30%) had an FcγRIIIA-F/F158 genotype, 11 patients (41%) had an FcγRIIIA-F/V158 genotype and eight patients (30%) had an FcγRIIIA-V/V158 genotype. Three of five (60%) patients with anti-MAG neuropathy and the FcγRIIIA-V/V158 genotype improved one or more points on the ODSS functional scale after treatment. In contrast, the ODSS did not change in any of the 13 (0%) patients with the FcγRIIIA-F/V158 or F/F158 genotypes (p=0.01). Two patients (40%) with anti-MAG antibodies and the FcγRIIIA-V/V158 genotype improved more than 5% on the distal motor sum score, whereas none of the patients with the FcγRIIIA-F/V158 or F/F158 genotypes had similar improvement (p=0.07). Three out of five anti-MAG neuropathy patients (60%) with the FcγRIIIA-V/V158 genotype also had improvement of the sensory sum score of more than 5%, while two out of eight (25%) anti-MAG neuropathy patients with the FcγRIIIA-F/V158 genotype and one out of five (20%) with the FcγRIIIA-F/F158 genotype improved more than 5% on the sensory sum score (p=0.27).
The two patients who experienced paradoxical worsening had FcγRIIIA-V/V158 or FcγRIIIA-F/V158 genotypes.
The presence or titres of anti-MAG antibodies were not associated with specific FcγR genotypes and results were not different after inclusion of the nine additional patients with polyneuropathy and IgM monoclonal gammopathy but without anti-MAG antibodies.
Open label and randomised studies suggest that rituximab may be effective in a subset of patients with IgM-PNP.3 ,4 ,9 Our data suggest that interindividual differences in treatment response may be determined by functional polymorphisms of FcγRIIIA that is expressed on important leukocyte subsets that mediate antibody-dependent cell-mediated cytotoxicity including natural killer cells. The FcγRIIIA-F/V158 polymorphism determines the affinity of this receptor to interact with IgG, including rituximab bound to CD20-expressing B cells.5 In vitro, antibody-dependent cell-mediated cytotoxicity and other antibody-mediated leukocyte functions are more efficiently triggered by leukocytes from donors with at least one FcγRIIIA-V158 allele.5 The association of FcγRIIIA genotypes with treatment outcome may reflect differences in efficiency of recruitment of natural killer cells by rituximab when it is bound to circulating B cells. FcγRIIIA genotypes may therefore represent novel biomarkers to predict treatment response to rituximab in patients with IgM-PNP.
Improvement of the ODSS functional score, which was used as a primary outcome measure in our patients, was only seen in patients with FcγRIIIA-V/V158 genotype. Improvement of secondary outcome measures including the sensory and distal motor sum scores were also primarily seen in patients with this genotype.9 Although the sample size of our current study is relatively small, the validity of our findings is supported by the association of FcγRIIIA-V/V158 with improvement or trends towards improvement of several outcome measures. Moreover, other studies with relatively large sample sizes found similar associations of rituximab efficacy in patients with lymphoma and generalised autoimmune disease such as rheumatoid arthritis.7 ,8 Two patients who experienced paradoxical temporary worsening of weakness during rituximab treatment had FcγRIIIA-V/V158 and FcγRIIIA-F/V158 genotypes. We cannot exclude the possibility that paradoxical worsening is associated with specific FcγR genotypes, since associations of FcγR polymorphisms with side effects of monoclonal antibody therapy have been reported previously.12–14
In one of the randomised trials, response to rituximab in patients with IgM-PNP was associated with higher anti-MAG antibody titres and the depletion of clonally expanded memory B cells.3 ,15 Anti-MAG titres were not associated with FcγRIIIA genotypes or response to treatment in our patients, suggesting that there may be several independent determinants of treatment efficacy. Moreover, FcγR polymorphisms and FcγR expression levels on monocytes may determine rituximab response, as FcγR expression levels were recently described to be associated with disease severity in other immune-mediated neuropathies.16 The lack of an alternative treatment option for patients with IgM-PNP, the cost of rituximab treatment and potentially serious side effects indicate that the predictive value of determinants of rituximab treatment efficacy should be studied in more detail.
Contributors ACJS: conception and design of study, acquisition, analysis and interpretation of data, drafting and revision of manuscript, final approval of manuscript. NCN, LHvdB, J-MFN and JHWL: conception and design of study, interpretation of data, revision of manuscript, final approval of manuscript. RRDS and MN: conception and design of study, acquisition and analysis of data, revision of manuscript, final approval of manuscript. W-LvdP: conception and design of study, acquisition, analysis and interpretation of data, drafting and revision of manuscript, final approval of manuscript. He is the principal investigator and had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis.
Competing interests LHvdB received travel grants and consulting fees from Baxter and served on advisory boards of Biogen Idec and Cytokinetics. W-LvdP received travel grants from Baxter International Inc and research support from the Prinses Beatrix Spierfonds and the Stichting Spieren voor Spieren.
Ethics approval Study approval was granted by the UMC Utrecht Medical Ethics Committee.
Provenance and peer review Not commissioned; externally peer reviewed.
Data sharing statement All relevant data are included in this study.
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