Presence of multiple abnormal immunologic markers is an independent prognostic factor of diffuse large B-cell lymphoma

Abstract

Autoimmune diseases (ADs) increase the risk of non-Hodgkin’s lymphoma and contribute to poor prognosis of patients. However, the association between immunologic markers and clinical outcome has rarely been investigated. This study aims to analyze the prognostic value of pretreatment immunologic markers in newly diagnosed patients with diffuse large B-cell lymphoma (DLBCL). We retrospectively reviewed the data on 502 patients with DLBCL treated in our institution from January 2013 to March 2018. Survival functions were estimated using Kaplan–Meier method and Cox regression model. The 3-year progression free survival (PFS) and overall survival (OS) rates were 70.2% and 80.9%, respectively, and the complete remission (CR) rate was 78.1%. Among the patients, those with multiple (≥3) abnormal immunologic markers had significantly shorter 3-year PFS (52.7% vs. 77.3%, P<0.001) and OS (68.5% vs. 85.8%, P=0.001) than those without multiple abnormal immunologic markers. Multivariate analysis revealed that the presence of multiple abnormal immunologic markers and the elevated serum levels of lactate dehydrogenase were the independent adverse prognostic factors for PFS (P=0.008, P<0.001) and OS (P=0.003, P<0.001). Meanwhile, advanced Ann Arbor stage was an independent adverse prognostic factor for PFS (P=0.001) and age>60 years for OS (P=0.014). In conclusion, the immunologic status was closely related to lymphoma progression, and this study provides new insights into the risk stratification of patients with DLBCL.

Keywords

immunologic marker / diffuse large B-cell lymphoma / prognosis

Introduction

Chronic immune stimulation and autoimmune disorders are risk factors for non-Hodgkin’s lymphoma [¹]. For example, systemic lupus erythematosus (SLE) and primary Sjögren syndrome (pSS) are associated with an increased risk of B cell lymphoma, especially marginal zone lymphoma. High activity of rheumatoid factor (RF) predisposes to diffuse large B cell lymphoma (DLBCL) [¹,²]. RF, anti-double stranded DNA IgG (anti-dsDNA IgG), and anti-nuclear antibody (ANA) levels are substantially higher in patients with DLBCL than in non-DLBCL lymphoma patients [³]. Low C3 and C4 levels contribute to lymphoma development in pSS [⁴].

The correlation between abnormal immunologic status and clinical outcome in DLBCL remains unclear. In this study, we showed that the presence of multiple abnormal immunologic markers is significantly related to disease progression in newly diagnosed patients with DLBCL.

Patients and methods

Patients

From January 2013 to March 2018, 684 consecutive patients with de novo DLBCL (not otherwise specified, NOS) diagnosed based on registry data were enrolled in this study. Histological diagnosis was established according to World Health Organization (WHO) 2008 classification [⁵], with exclusion of mediastinal large B cell lymphoma (n = 66), primary central nervous system DLBCL (n = 62), and patients with DLBCL who received only supportive care (n = 54). Finally, 502 de novo patients with DLBCL were evaluated. The clinical features of the patients were described in Table 1.

Informed consent was obtained from all patients in accordance with the regulations of the Hospital and Institutional Review Boards and the Declaration of Helsinki.

Immunologic marker detection

Immunologic markers related to autoimmune diseases (ADs) and immunologic status were included. RF, anti-dsDNA IgG, and anti-Sjögren’s-syndrome-related antigen (anti-SSA) are the representative markers in the diagnosis of rheumatoid arthritis (RA), SLE, and pSS, respectively [⁶–⁸]. ANA is commonly used in the diagnosis of ADs, including SLE and pSS [⁹], whereas antistreptolysin “O” (ASO) is utilized in the diagnosis of rheumatic diseases [¹⁰]. Circulating immune complex (CIC) is a prominent feature of several autoimmune diseases, such as RA, SLE, and pSS [¹¹], and decreased complements (C3 and C4) occur in the activate stage of ADs [¹²]. Serum immunoglobulins G, M, A, E (IgG, M, A, E) are elevated in various ADs.

CIC, IgG, IgM, IgA, IgE, C3, C4, RF, anti-SSA, and ASO were assessed by turbidimetric inhibition immunoassay (Beckman Coulter, California, USA). Anti-dsDNA IgG and ANA were assessed by ELISA (Inova, California, USA). Serum immunologic markers were detected in patients with DLBCL at diagnosis.

Treatment and response criteria

All 502 patients received rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone-based (R-CHOP) chemotherapy. Treatment response was evaluated according to the WHO response criteria [¹³]. Complete remission (CR) is defined as no evidence of residual disease, partial response (PR) with at least a 50% reduction in tumor burden from the onset of treatment. Progressive disease (PD) or relapse is defined as the appearance of any new lesion more than 1.5 cm at the end of therapy or at least a 50% increase in the longest diameter of any single or from nadir in the SPD of previously involved nodes. Stable disease (SD) is defined as the state of neither PR nor PD.

Statistical analysis

Baseline characteristics of patients were analyzed using two-sided t2 test for categorical data. Logistic regression analysis was performed to analyze the risk factors of achieving CR. Progression-free survival (PFS) was calculated from the date when treatment began to the date when the disease progression was recognized or the date of the last follow-up. Overall survival (OS) time was measured from the date of diagnosis to the date of death or the last follow-up. Survival functions were estimated using the Kaplan–Meier method and compared by log-rank test. Univariate hazard estimates were generated with unadjusted Cox proportional hazards models. Covariates indicating significance on univariate analysis were included in the multivariate model. Statistical significance was defined as P<0.05. All statistical analyses were conducted using Statistical Package for the Social Sciences (SPSS) 22.0 software (SPSS Inc., Chicago, IL, USA).

Results

Abnormal immunologic markers frequently occur in patients with DLBCL

Among the 502 patients, 71.9% had abnormal immunologic markers. With regard to serum immunoglobulins and complements, elevated CIC, IgG, IgM, IgA and IgE levels were found in 147 (29.3%), 76 (15.1%), 21 (4.2%), 20 (4.0%), and 98 (19.5%) patients, respectively.

Decreased levels of C3 and C4 were significantly associated with SLE [¹²], whereas their increased levels were observed in tumor or inflammatory cases [¹⁴]. Therefore, both changes in C3 and C4 were regarded as abnormal and found in 110 (21.9%, high: 19, low: 91) and 95 (18.9%, high: 13, low: 82) patients, respectively. Among the 377 patients with pretreatment results of serum RF, anti-dsDNA IgG, anti-SSA, ANA and ASO, 41 (10.9%) had high serum RF, 3 (0.8%) were positive for anti-dsDNA IgG, 6 (1.6%) were positive for anti-SSA, 73 (19.4%) had elevated ANA, and 18 (4.8%) had elevated ASO level.

We further divided the patients into different groups according to the number of abnormal immunologic markers. Seventy-six patients (20.2%) had multiple abnormal immunologic markers (i.e.,≥3 abnormal immunologic markers). Among them, 11 patients had pre-diagnosed ADs, including RA (6 cases), SLE (2 cases), and pSS (3 cases). The medical history of the patients was recorded, and the diagnosis was thereafter confirmed according to the criteria [⁶–⁸].

These immunologic markers had relationship with the clinical features of the patients. IgM was increased in patients with multiple extranodal involvement (P= 0.008), and IgE was increased in patients with IPI scores 3–5 (P = 0.034). C3 was abnormal in patients with poor ECOG (P = 0.018), RF was significantly increased in patients with IPI scores 3–5 (P = 0.012) and those with elevated LDH (P = 0.029), anti-SSA was positive in patients with poor ECOG (P = 0.029), and ANA was increased in patients with poor ECOG (P = 0.044) (Table 2).

Multiple abnormal immunologic markers do not affect treatment response in DLBCL

Upon treatment, 392 patients (78.1%) achieved CR. For 76 patients with multiple abnormal immunologic markers, their CR rate was 71.1%. No significant difference in CR rate was observed in patients with and without multiple abnormal immunologic markers (P = 0.117). According to logistic regression, the factors statistically associated with lower CR rate were IPI scores 3–5 (P<0.001), elevated LDH (P<0.001), and advanced Ann Arbor stage (P = 0.032) (Table 3).

Multiple abnormal immunologic markers reflect poor clinical outcome in DLBCL

The 3-year PFS and OS rates of the 502 patients with DLBCL were 70.2% and 80.9%, respectively (Fig. 1A and 1B). After a median follow-up of 30 months, patients with multiple abnormal immunologic markers showed significantly inferior prognosis compared with those without multiple abnormal immunologic markers (control group). The 3-year PFS and OS rates of the two groups were 77.3% and 52.7% (P<0.001) and 85.8% and 68.5% (P = 0.001), respectively (Fig. 2A and 2B).

Presence of multiple abnormal immunologic markers is an independent adverse prognostic factor in DLBCL

Univariate analysis showed that the factors significantly associated with shorter PFS and OS were elevated LDH (both P<0.001), ECOG≥2 (both P<0.001), multiple extranodal involvement (P = 0.001 and P = 0.015), advanced Ann Arbor stage (both P<0.001) and multiple abnormal immunologic markers (both P = 0.001). The presence of age>60 was of predictive value only for OS (P<0.001) (Table 4).

The significant parameters in the univariate analysis were inputted into the Cox regression model for multivariate analysis. Elevated LDH (P<0.001), advanced Ann Arbor stage (P = 0.001), and multiple abnormal immunologic markers (P = 0.008) predicted shorter PFS. However, age>60 (P = 0.014), elevated LDH (P<0.001), and multiple abnormal immunologic markers (P = 0.003) retained their independent prognostic effect on inferior OS (Table 5).

Discussion

DLBCL is a heterogeneous subtype of aggressive lymphoma. Approximately 30%–40% of the patients eventually relapse after receiving first-line chemotherapy [¹⁵,¹⁶]. Investigating the factors leading to poor clinical outcome is necessary to develop reasonable individualized treatment plans. In this study, we showed that the presence of multiple abnormal immunologic markers is related to resistance to conventional immunochemotherapy and is an independent unfavorable factor for PFS or OS in DLBCL.

Accumulating data revealed the association between specific ADs and the risk of non-Hodgkin lymphoma development [¹⁷]. However, the influence of immune disorders on the prognosis of the patients with DLBCL has seldom been studied. Here, we confirmed poor prognosis in patients with DLBCL who had multiple abnormal immunologic markers. Dysregulation of T regulatory, T helper, and B cells occurred in patients with lymphoma and ADs [¹⁸,¹⁹]. B cell proliferation and auto-antibody production are the features of RA, SLE, and pSS [²⁰–²²]. Lymphomas may arise when the B cells evolve in an uncontrolled manner due to somatic hypermutation, which plays an important role in lymphomagenesis. BCL6 mutations are associated with a high frequency in DLBCL [²³]. Another possible reason for the similar pathogenesis of immune disorder and lymphoma is their similar genetic backgrounds. Low expression of HLA Class I and II is related to decreased survival time of B cell lymphoma and determines the susceptibility to a range of ADs, such as RA, ulcerative colitis, and type I diabetes [²⁴–²⁷]. Hence, antigen-driven immune response is also relevant to the development of DLBCL. These results may explain the presence of multiple abnormal immunologic markers and their contribution to lymphoma progression in DLBCL.

Further investigation in multicenter studies should be conducted to confirm these findings. In conclusion, immunologic status is closely related to lymphoma progression, and this work provides new insights into the risk stratification of patients with DLBCL.

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