The IL6(-174G/C) polymorphism is a prognostic factor for survival after treatment initiation in Waldenström macroglobulinemia patients aged 65 years or less

Waldenström macroglobulinemia (WM) is characterized by the production of serum monoclonal immunoglobulin (Ig) M and by lymphoplasmacytic bone marrow infiltration.1 Upregulation of interleukin 6 (IL6) has been demonstrated in WM,2,3 in accordance with the increased IL6 serum concentrations previously observed.4,5 Moreover, clonal blood B cells from patients with WM differentiated spontaneously in vitro to plasma cells via an autocrine pathway involving IL6.6 Subjects with the C allele in the IL6–174 promoter region had lower serum IL6 concentration.7,8 Therefore, we aimed to investigate the prognostic value of the IL6(-174G/C) polymorphism in WM.

One hundred patients with WM (M/F ratio: 1.7, median age: 67 years, range: 39–91) entered the study with the following inclusion criteria: (i) proven WM and (ii) initiation of therapy before January 2006 only in symptomatic patients according to the second international workshop on WM recommendations; (iii) informed consent obtained according to the protocol submitted to the Institution Review Board. Seventy-five patients required first-line therapy (at diagnosis: 48 patients and 4–114 months later: 27). Fifty-seven patients received chlorambucil, 3 combination chemotherapy, 9 fludarabine alone, 3 fludarabine in combination with other chemotherapy, and 3 rituximab. Median follow-up was 42 months (range: 17–180 months) in alive patients with a stopping date set at January 1st, 2007. IL6 genotype was assessed using an allelic discrimination assay performed on an ABI PRISM 7000 (Applied Biosystems, USA) as previously described.9 The clinical and laboratory characteristics of the different genotypic groups and survival curves were compared using the Yates modified χ test or the Fisher exact test and the log-rank test with bootstrap resampling (1,000 replicates) respectively. Characteristics associated with a significant prognostic value were introduced in a Cox proportional hazard model, after assessment of the validity of the assumption of this model. These analyses and differences in the distribution of age in genotypic subgroups prompted us to assess the prognostic value of the IL6 polymorphism in subgroups defined by age. All statistical analyses were carried out using the Splus 6.2 (MathSoft, Cambridge, MA, USA) software.

Table 1.Clinical characteristics of 75 patients at the time of initiation of therapy according to IL6(-174G/C) polymorphism.

The distribution of the IL6-174 polymorphism genotypes for the 100 patients were as follows: G/G, 33%; G/C, 52%; and C/C, 15%. The frequency of the IL6-174G allele was 0.59. This distribution did not differ from that expected from Hardy Weinberg equilibrium and agreed with previous reports.7,9–11 The IL6(-174G/C) genotype of the 75 patients who received treatment and their main clinical characteristics at the time of first treatment initiation are reported in Table 1. Patients homozygous for the G allele (G/G) were younger than remaining patients (p=0.007). Mean monoclonal component concentration was 32.7 g/L in IL6(-174GG) patients and 23.3 g/L in the remaining patients (p=0.10). Thirty-two of the 75 patients (43%) who received treatment had died at the stopping date and the median survival after treatment initiation was 106 months (95CI: 74–197 months). Advanced age >65 years (p=0.005) and intermediate or high-risk according to the International Scoring System for WM (ISSWM, p=0.05) were associated with an adverse prognostic value for survival after treatment initiation. Multivariate analysis demonstrated the presence of an interaction between IL6(-174G/C) polymorphism and age. Indeed, the IL6(-174G/G) genotype was associated with an adverse prognostic value only in patients aged 65 years or less (6-year survival probability after treatment initiation: 55% for IL6(-174G/G) vs. 87% for IL6(-174C/G) or (C/C) patients, p=0.02, Figure 1). Three of the 25 remaining untreated patients had died from causes not related to the disease and the IL6(-174G/C) genotype had no prognostic value for the overall survival in the 100 patients.

Figure 1.Actuarial survival curves of patients with Waldenström’s macroglobulinemia aged 65 years or less, according to their IL6 (-174G/C) polymorphism status. The solid line indicates the IL6(-174 G/G) patients, the dashed line IL6(-174 G/C) or IL6(-174 C/C) patients (p=0.02).

For the first time, we describe an influence of the IL6(-174GG) polymorphism, previously associated with high expression level of IL6, on the outcome of WM after treatment initiation in patients aged less than 65 years. This observation was achieved in a large biological series of WM patients with prolonged follow-up and validated with bootstrap resampling, given the rarity of the disease. Previous results may support this finding: IL6 is expressed by tumoral cells,2,3 IL6 favor the differentiation of blood B cells from WM patients via an autocrine pathway.5,6 Moreover, IL6 induces rapid Akt phosphorylation5,12 that promotes survival and growth of tumor cells.5,12 Unfortunately, serum was not available and, thus, IL6 measurements could not be performed. Conflicting results were reported on the relationships between serum IL6 concentration and IL6(-174G/C) polymorphism, using various experimental protocols.7–9 The numerous circumstances influencing IL6 levels, especially advanced age, the high clearance of serum IL610,11 and probable tumoral specific deregulations of IL6 in bone marrow support the analysis of the IL6(-174G/C) polymorphism, in place of serum analysis, to assess the overall expression profile of IL6. We cannot exclude a linkage dysequilibrium between the informative IL6 locus and one or more other informative genes that contribute to the pathogenesis of the disease. However, the presence of the C allele in the IL6–174 promoter region has also been associated with improved outcome in patients with breast cancer.10 The IL6(-174GG) genotype was associated with young age. This pattern of distribution of age may counteract the adverse prognostic value associated with the IL6(-174GG) genotype in the 75 symptomatic patients. We recorded deaths not related to the disease in asymptomatic patients, whatever their IL6(-174G/C) polymorphism status. These findings probably explain the lack of prognostic value of the polymorphism in the whole series.

The present results should be viewed as exploratory and require confirmation in larger prospective studies. They indicate that IL6(-174G/G) genotype may be a new adverse prognostic factor in WM patients aged less than 65 years who require therapy. This finding supports the clinical significance of the genetic background of the IL6 pathway in WM.

Acknowledgments

the authors wish to thank Valérie Grandières, Claudine Delsaut and Catherine Vanicatte for their excellent technical assistance.

Footnotes

• Funding: this work was supported by the Comité du Pas de Calais de la Ligue contre le Cancer.