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© Borgis - Postępy Nauk Medycznych 6/2015, s. 367-373
*Iwona Grygoruk-Wiśniowska, Sławomira Kyrcz-Krzemień
Analiza niespójności wyników badań wybranych parametrów białkowych surowicy krwi i klonalnych plazmocytów szpiku w ocenie odpowiedzi na leczenie u chorych ze szpiczakiem plazmocytowym (MM) poddanych autologicznej transplantacji krwiotwórczych komórek macierzystych (AHSCT)
An analysis of discrepancies between test results of the selected protein parameters and clonal bone marrow plasma cells in an assessment of treatment response in a group of patients with multiple myeloma (MM) prior to an autologous hematopoietic stem cell transplantation (AHSCT)
Department of Haematology and Bone Marrow Transplantation, Medical University of Silesia, Katowice
Head of Department: prof. Sławomira Kyrcz-Krzemień, MD, PhD
Streszczenie
Wstęp. Szpiczak plazmocytowy (ang. multiple myeloma – MM) jest nieuleczalną chorobą rozrostową układu krwiotwórczego wywodzącą się z limfoidalnych komórek B. Współczesna diagnostyka i monitorowanie MM obejmuje testy białkowe surowicy i moczu oraz badanie cytologiczne aspiratu szpiku kostnego. Niespójności uzyskiwanych wyników mogą być przyczyną trudności interpretacyjnych.
Cel pracy. Analiza niespójności wyników badań wybranych parametrów białkowych surowicy krwi i klonalnych plazmocytów szpiku u chorych ze szpiczakiem plazmocytowym przed zabiegiem AHSCT.
Materiał i metody. Do badania włączono 72 chorych z MM ocenianych przed rozpoczęciem procedury AHSCT. Analizą objęto 37 mężczyzn i 35 kobiet w medianie wieku 58 lat (zakres 38-81 lat). Wykorzystano następujące testy diagnostyczne: elektroforezę białek surowicy (SPE), immunofiksację (IFE), test Freelite do oceny stężenia wolnych lekkich łańcuchów kappa, lambda i ich wzajemnego stosunku oraz badanie klonalności plazmocytów szpiku przy użyciu cytometrii przepływowej.
Wyniki. Spośród 72 pacjentów poddanych analizie u 32 (44%) stwierdzono występowanie niespójności analitycznych w testach diagnostycznych stosowanych w ocenie stanu remisji przed AHSCT. U 40 (56%) pacjentów wyniki testów diagnostycznych były spójne. Zaobserwowano cztery typy niespójności.
Wnioski. Właściwa ocena stanu remisji szpiczaka plazmocytowego przed zabiegiem AHSCT wymaga wykorzystania co najmniej kilku testów diagnostycznych ze względu na obserwowane niespójności, a badanie klonalności plazmocytów przy zastosowaniu cytometrii przepływowej stanowi jedynie metodę uzupełniającą.
Summary
Introduction. Multiple myeloma (MM) is an incurable, B-cell malignancy. Modern diagnostics and disease monitoring include the assessment of serum and urine protein as well as bone marrow examination. The discrepancies of the results provided by these studies may cause the difficulties in the interpretation of disease status.
Aim. The analysis the incompatibility of test results a selected protein parameters and clonal plasma cells in bone marrow in response to treatment in MM patients prior to the AHSCT.
Material and methods. Seventy two MM patients (37 male and 35 female) at a median age of 58 years (range 38-81 years) were included in this study. The following diagnostic tests have been used: electrophoresis (SPE), immunofixation (IFE), Freelite for serum free light chains kappa and lambda assessments and clonal plasma cell analysis in bone marrow by flow cytometry (FC).
Results. 32 patients (44%) were found to have discrepancies in the diagnostic tests evaluating disease status before transplant; in the remaining 40 patients (56%) the results were consistent. The 4 types of discrepancies were detected.
Conclusions. At least several diagnostic tests are required in order to reliable assess the MM status before AHSCT and clonal plasma cell assessment by FC may remain an additional diagnostic tool.
Introduction
Multiple myeloma (MM) is an incurable, B-cell malignancy. MM represents about 10% of all hematologic malignancies and is characterized by the proliferation of single clone plasma cells (1).
The first description of the disease comes from the 80’s of the Nineteenth Century and was written by Samuela Sollye, John Dalrymple and Henry Bence-Jones. In 1889 Otto Kahler and Osip Rusticki used the term myeloma multiplex for the first time (2).
The annual incidence rate is 4-7 cases per 100 000 inhabitants. It affects slightly more men and people of certain races, such as African or Afro-Caribbean. Furthermore, the average age of the diagnosis is 65 (3).
The etiology of multiple myeloma is not clear. The causes of the disease are complex, probably following various stages and then leading to gene mutations. The development of MM is closely connected to clonal plasma cells which secrete many autocrine substances and stimulating factors such as: MIPIα, MIPIβ, TNF, M-CSF, HGF, VEGF, MMP9, MMP2, IL-1, IL-3, IL-6, IL-11 (4, 5).
Major clinical manifestations are: renal failure, hypercalcemia, anemia, osteolitic bone lesion or pathological fractures. Non-specific symptoms are: weakness, weight loss, fever, susceptibility to infection, neuropathy (6).
Each case of MM is preceded by an asymptomatic malignant stage, termed MGUS – monoclonal gammopathy of undetermined significance. MGUS is characterized by the presence of the M protein in serum (< 30 g/L) and the appearance of clonal plasma cells in the bone marrow which is greater than or equal to 10% (7). The risk of progression to myeloma is 1% per year (8). The next stage in the development of MM is smoldering or asymptomatic multiple myeloma (SMM). In order to identify SMM, monoclonal protein concentration must be over 30 g/L and/or clonal plasma cells in the bone marrow must exceed 10% in the absence of end-organ damage (CRAB symptoms). The estimated risk of progression of SMM to multiple myeloma is on the level of 10% per year for the first 5 years since recognition (9).
A number of tests are used to help confirm multiple myeloma. The diagnosis require the presence of the clonal plasma cell in the bone marrow (> 10%), the presence of monoclonal proteins in the serum and/or urine and so-called CRAB symptoms.
International Myeloma Working Group has established criteria for the diagnosis and monitoring response to treatment in patients with monoclonal gammopathy. This organization recommends the following diagnostic tests performed in the serum and urine: electrophoresis (SPE), immunofixation (IFE), a serum kappa and lambda free light chains assessment along with a flow cytometry analysis (FC) of clonal bone marrow plasma cells (10-12).
Aim
An analysis of discrepancies in the test results of the selected protein parameters and clonal bone marrow plasma cells in MM patients prior to the AHSCT.
Material and methods
Seventy two MM patients (37 male and 35 female) with a median age of 58 years (range 38-81 years) were included in this study. All of them were treated with chemotherapy in the Department of Haematology and Bone Marrow Transplantation Silesian University in Katowice, between 2011 and 2013. The following diagnostic tests were used: electrophoresis (SPE), immunofixation (IFE), a serum kappa and lambda free light chains assessment (Freelite) and a flow cytometry analysis (FC) of clonal plasma cells in the bone marrow.
Results
Out of 72 patients, 32 (44%) were found to have discrepancies in the diagnostic tests evaluating disease status before the transplant; in the remaining 40 patients (56%) the results were consistent.
4 types of discrepancies were detected: 1 – Freelite and FC tests were negative, but SPE/IFE was positive (n = 10) (tab. 1, item 1-10), 2 – Freelite test was negative whereas FC and SPE/IFE were positive (n = 10) (tab. 1, item 11-20), 3 – FC was negative with positive Freelite test and SPE/IFE (n = 8) (tab. 1, item 21-28) and 4 – FC and SPE/IFE were negative with positive Freelite test (n = 4) (tab. 1, item 29-32).
Table 1. The discrepancies in the results of the diagnostic tests.
Patient Clonal plasma cells FLCr (FLCsκ/FLCsλ)IFESPE
10011
20010
30011
40011
50011
60010
70011
80010
90010
100011
111010
121011
131010
141011
151011
161011
171010
181011
191010
201010
210111
220111
230110
240110
250111
260111
270110
280110
290100
300100
310100
320100
FLCr – FLC ratio κ/λ; IFE – serum protein immunofixation; SPE – serum protein electrophoresis
0 – negative result; 1 – positive result
The analysis of patients with MM identified two subgroups: Group A – without the presence of discrepancies in the study and Group B – with the presence of discrepancies in the study.
Considering parameters such as: age, sex, multiple myeloma subtype, response to treatment, type of treatment, the number of therapy lines, blood morphology parameters, recovery times in each line after AHSCT and duration of the disease – no significant differences were proved between the parameters of the patients from group A (without the presence of discrepancies) and those from group B (with the presence of discrepancies). The comparison of the MM patients whose tests results did not have discrepancies in protein tests (group A) with the patients whose tests had discrepancies (group B) was illustrated in table 2.
Table 2. The comparison of the MM patients whose tests results did not have discrepancies in protein tests (group A) with the patients whose tests had discrepancies (group B).
ParameterGroup A (n = 40)Group B (n = 32)p
Age (median, range); years58.2 (44-74)58.7 (38-80.5)0.27
Sex (W/M) (n (%))19 (47%)/21(53%)16 (50%)/16 (50%)0.99
Subtype MM (n (%)):
IgG kappa
IgG lambda
IgA kappa
IgA lambda
 
19 (48%)
9 (22%)
8 (20%)
4 (10%)
 
16 (50%)
11 (35%)
3 (9%)
2 (6%)
 
0.13
Disease state (n (%)):
sCR
CR
VGPR
PR
 
6 (15%)
7 (18%)
10 (25%)
17 (42%)
 
0 (0%)
4 (13%)
20 (63%)
8 (25%)
 
0.90
Number lines of the treatment (median, range)1 (1-4)1 (1-4)0.41
Induction therapy CTD (n (%))31 (78%)27 (84%)0.42
RBC (x 106/μL) (median, range)4.1 (3-5.1)4.1 (3.1-4.7)0.37
HCT (%) (median, range)38 (31-45)38 (29-42)0.62
Hgb (g/dL) (median, range)12.8 (10.4-16)12.4 (9.8-14.9)0.74
PLT (x 109L) (median, range)207 (76-296)208 (93-358)0.16
WBC (x 109/L) (median, range)4.4 (2.5-9.7)4.5 (2-9.7)0.91
Regeneration WBC (days) (median, range)12 (8-20)13 (10-17)0.31
Regeneration ANC (days) (median, range)13 (9-20)13 (10-18)0.69
Regeneration PLT (days) (median, range)10 (8-17)11 (8-17)0.48
Time to AHSCT (months) (median, range)19 (8-149)14 (8-71)0.89
Time from diagnosis to the last follow-up (months) (median, range)45 (23-168)26 (17-83)0.79
ANC – absolute neutrophil count; CTD – cyclophosphamide , thalidomide, dexamethasone; CR – complete response; HCT – haematocrit; Hgb – hemoglobin concentration; K– woman; M – man; PLT – platelet count; PR – partial response; RBC – red blood cell count; sCR – stringent complete response; WBC – white blood cells count; VGPR – very good partial response
Discussion
Multiple myeloma is a disease that is characterized by the proliferation and accumulation of clonal plasma cells producing monoclonal protein (1).
For over one and a half centuries, the presence of Bence Jones protein has been the most important diagnostic marker of monoclonal gammopathy (13).
In 2001, a special test called the serum free light chain (FLC) assay or Freelite test was introduced into clinical practice. Since then, it has been the standard test performed in patients with MM, used in diagnosis, prognosis, monitoring of response to treatment and, finally, for a quick evaluation of recurrence/progression (14).
The next quantum leap in the history of research on multiple myeloma was the discovery of the free light chains escape phenomenon which involves an increase of FLCs in patients with relapsing multiple myeloma, while still maintaining normal serum immunoglobulin levels. The free light chains escape is associated with the proliferation of the plasma cells clone producing only light chains and is not related to the clone producing complete immuglobin molecule. This leads to a significant increase in the concentration of FLCs in the serum, while that of the complete immunoglobulin level is not altered. Early detection of the free light chains escape is of clinical relevance for several reasons:
– it is related to a significant increase in tumor weight hence is used to measure the severity of the disease,
– it helps to identify patients with a high risk of progression and poor prognosis,

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otrzymano: 2015-04-07
zaakceptowano do druku: 2015-04-30

Adres do korespondencji:
*Iwona Grygoruk-Wiśniowska
Department of Haematology and Bone Marrow Transplantation Medical University of Silesia
ul. Dąbrowskiego 25, 40-032 Katowice
tel./fax +48 (32) 259-12-81
klinhem@sum.edu.pl

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