Alteration of Biochemical Immunological and Molecular Markers of Chronic Lymphocytic leukemia in Sample of Iraqi Patients

   Chronic lymphocytic leukaemia (CLL) is a monoclonal malignancy characterized by an accumulation of small and mature looking B lymphocytes in the blood, bone marrow and other tissues. The Blymphocytes developed in a wrong way lead to an immune disorder, and are  typically  characterized by expression of some of CD markers as (CD5,CD38 and ZAP-70) determined by immunophenotyping. The aim of this study was to investigate the extent to which immunological, biochemical, and molecular parameters are altered in CLL patients, and the potential of applying these alterations as biomarkers for CLL in Iraqi patients. This study was conducted on randomly selected 55 CLL patients referred to the National Center for Blood Diseases, AL-Mustansiyria  University for evaluation and treatment during the period extending from Oct. 2013 to Sept. 2015. CLL  patients included 18 newly diagnosed untreated patients (12 males and 6 females), with a mean age of 55±12 yr and age range of(40-80)yr. According to the Binet staging system for CLL, 10 of these patients were in stage B and 8 in stage C. Besides, this study includes 37 already diagnosed CLL patients who are currently receiving treatment. They included 28 males and 9 females, with a mean age of 57±9 yr and on age range of 45-71 yr. Twenty three 23 patients of this group were in stage B and 14 patients in stage C. Nineteen  apparently healthy subjects were also involved in this study. They included 9 males and 10 females, with a mean age of 61 ± 17 yr. and an age range of 30-90 yr.  A Complete peripheral blood picture examination in the ntreated newly diagnosed CLL patients revealed absolute lymphocytosis, mild anemia, decreased platelet count and elevated ESR level as compared to controls. A Follow-up analysis of the data obtained from those patients in response to combination therapy (rituximab, fludaribine, cyclophosphamide(RFC)) for 1-8 months (median 4.5 months) normalized WBC, and Lymphocytes counts and anemia, but failed to correct the lowered platelet counts or the elevated ESR values. The significant biochemical changes observed in the untreated CLL patients were those of slight elevations of albumin and uric acid levels, which were not corrected by RFC therapy. Nevertheless, these values remained within the reference values. Serum levels of the major antioxidant, glutathione (GSH) showed a significant decline and serum levels of malondialdehyde (MDA) were significantly elevated in untreated CLL patients. RFC therapy failed to normalize these values, suggesting the presence of sustained increased endogenous oxidative stress in CLL patients. Furthermore, five out of the (18) (27.8%) newly diagnosed CLL patients were diabetic, and showed hyperglycemia and elevated HbA1c levels. The immunological profile of the newly diagnosed CLL patients revealed  the presence of increased expression of  CD5, CD 38, and ZAP-70. Similarly, the levels of IL-6 and IL-10 were elevated in the serum of untreated CLL patients. RFC  therapy  managed  to normalize only the expression of ZAP-70. Real Time-PCR results showed the presence of mutated in immunoglobulin heavy-chain variable region (IgVH) in 3 out of 8 untreated and 4 out of 8 treated CLL patients, while healthy controls did not show any mutation in IgVH. Another 37 old CLL patients who were previously diagnosed and received therapy for a range of 4 months to 8.5 years (median 21.5
months) showed comparable results to those obtained after a median 4.5 months of RFC therapy. Furthermore, 11 out of 37 (29.7%) old treated CLL patients were diabetic and showed hyperglycemia and elevated    The immunological profile of the newly diagnosed CLL patients revealed  the presence of increased expression of  CD5, CD 38, and ZAP- 70. Similarly, the levels of IL-6 and IL-10 were elevated in the serum of untreated CLL patients. RFC  therapy  managed  to normalize only the expression of ZAP-70.Real Time-PCR results showed the presence of mutated in immunoglobulin heavy-chain variable region (IgVH) in 3 out of 8 untreated and 4 out of 8 treated CLL patients, while healthy controls did not show any mutation in IgVH.HbA1c levels compared to euglycemic old treated patients and control
subjects. In conclusion, CLL patients of the present study showed various immunological, biochemical, and molecular changes.  The interesting finding that 16out of 55 CLL patients (i.e. 29%) had hyperglycemia and elevated HbA1c levels deserves further studies to assess its clinical significance and whether or not it may be utilized as a potential risk factor for the incidence of CLL in Iraqi patients.