ABBREVIATIONS: AIHA = autoimmune hemolytic anemia; B-- CLL - B-chronic lymphocytic leukemia; B-PLL = B-- prolymphocytic leukemia; CLL = chronic lymphocytic leukemia; DAT = direct antiglobulin test; PLL = prolymphocytic leukemia; TPN = total parenteral nutrition.
INDEX TERMS : chronic lymphocytic leukemia; leukemia.
Clin Lab Sci 2001;14(4):233
A 66-year-old male whose major complaint was general fatigue and weight loss was admitted to a local hospital in November 1999 for a routine physical examination. The initial blood work demonstrated a leukocyte count of 63.7 x 10^sup 9^ with many immature lymphocytes, hemoglobin 1.15 mmol/L (7.4 gm/dL), platelets 125 x 10^sup 9^, reticulocyte count of 2.8%, and 73 smudge cells per 100 WBCs (Table 1).
A diagnosis of chronic lymphocytic leukemia (CLL) was initially made even though the hematologic picture did not actually correlate with CLL. Results of a chemistry panel included elevated results for the following constituents: BUN 12.1 mmol/L (34 mg/dL); creatinine 132.6 (mu)mol/L (1.5 mg/dL); AST 1.33 uktal/L (78 U/L); LD 10.9 uktal/L (640 U/L); and, total bilirubin 29.1 (mu)mol/L (1.7 mg/ dL). A decreased haptoglobin (
Blood bank testing revealed the patient to be B Positive with a positive antibody screen and positive direct antiglobulin test (DAT). The antibody was determined to be a warm-reacting autoantibody with undetermined specificity. Coagulation studies were all within normal range (Table 3).
A review of the peripheral smear revealed the presence of a prominent population of prolymphocytes. Immunophenotyping of peripheral blood cells demonstrated a monoclonal population of Lambda positive B-cells consistent with a lymphoproliferative disorder of B-cells. Immunophenotyping demonstrated B-lymphocytes expressing CD 19, CD20 (bright), CD 22, CD23 (heterogeneous), CD5(dim), CD25, FMC-7, and lambda light chains. The cells were negative for CD 10, CD 110, and CD34.
This phenotype is consistent with B-cell lymphoproliferative disorder. The phenotype does not support a diagnosis of CLL, hairy cell leukemia, mantle cell lymphoma, or acute lymphoblastic leukemia. The morphology and phenotype are consistent with but not diagnostic of prolymphocytic leukemia.
Flow cytometry results confirmed complex hyperdiploid anomalies consistent with B cell leukemia/lymphoma. Translocations involving the heavy chain Ig locus at 14q32 that is specifically involved with B-cell neoplasia were noted.
Bone marrow biopsy was performed and demonstrated diffuse infiltration of prolymphocytes, resulting in a confirmatory diagnosis of B-cell prolymphocytic leukemia and resulting warm autoimmune hemolytic anemia. Thoracic and abdominal computerized tomography (CT) scans were normal.
During early treatment, consisting of cyclophosphamide (600 mg/ m^sup 2^) and fludarabine (20 mg/m^sup 2^), the leukocyte count ranged from 21 to 69 x 10^sup 9^/L until the treatment regimen was changed once confirmatory diagnosis was made. Chlorambucil and prednisone were administered (dosages unknown), with a resulting decrease in the leukocyte count to a variable leukopenic state (0.5 to 25 x 10^sup 9^/L). The patient continued to be refractory to red cell transfusions and was started on a second cycle of cyclophosphamide and fludarabine.
The autoimmune hemolytic anemia was confirmed to be caused by a warm-reacting autoantibody that resulted in the lysis of the patient's RBCs. When his red blood cell count fell below normal, blood transfusions were administered. This would increase his red blood cell count initially, but the autoimmune process continued to hemolyze the red cells. Reticulocyte counts were performed on a daily basis and ranged from 0.5% to 3.5%. As the autoimmune anemia and B-PLL progressed, total bilirubin, direct bilirubin, LD, and BUN remained elevated.
Problems were ever present with obtaining accurate results for the complete blood count, especially the platelet and reticulocyte counts. Repeated communication with the nursing staff was necessary to insure that the correct blood sample was received for hematologic studies, especially since the patient also developed an antibody that clumped platelets in EDTA that necessitated collection of a sodium citrate tube for accurate platelet counting. The hemolytic process became so intense as indicated by a total bilirubin of 428 to 513 (mu)mol/L (25 to 30 mg/dL) that plasma replacement with normal saline had to be performed in order to obtain accurate results. The patient's red cells became so fragile that agitation times had to be decreased to 10 and 15 minutes to obtain an accurate reticulocyte count rather than following the normal 30-minute incubation.
Following completion of chemotherapy, the patient developed marked increases in his liver enzymes and total bilirubin. This was associated with a drop in hematocrit from 22 to 14 in a 24-hour period. The patient was begun on prednisone (1 mg/kg/day) and IV Immunoglobulin (1 gm/kg/day) due to profound immunosuppression. Additionally, the patient was started on prophylactic therapy with Levaquin, Acyclovir, Diflucan, and Bactrim. The bilirubin decreased over the next several days but he continued to require sporadic transfusions for severe anemia. His white cell count did decrease somewhat but revealed evidence of peripheral blasts consistent with refractory disease.
The patient was noted to have a 17-pound weight loss and was started on total parenteral nutrition (TPN). He subsequently developed chest pain and was evaluated for myocardial infarction that was ruled out. Iron studies were obtained and the ferritin level was noted to be extremely elevated and the patient was begun on Desferal (1,000 mg over four hours) to begin iron unloading.
Extensive discussions were held with the patient regarding persistence of peripheral blood blast cells and he was subsequently begun on Rituxan (375 mg/m^sup 2^). Following the first day of Rituxan, his hematocrit dropped from 22 to 12 without significant explanation. This was accompanied by significant hyperphosphatemia that was managed and resolved with Amphojel. The patient received additional transfusions to maintain a hematocrit of 14. He developed a fever and was pan cultured and begun on prophylactic Zosyn and Gentamycin. He continued to exhibit fever and was started on empiric Amphotericin B along with Neupogen for profound neutropenia. At this point he developed some epistaxis and ultimately developed lower gastrointestinal bleeding; however, due to the patient's neutropenia, it was not felt prudent to pursue a gastrointestinal evaluation since the patient was already receiving Prevacid. All cultures remained negative. Additionally, the patient developed a diffuse purpura but the skin biopsies were nondiagnostic. The purpura was felt to be a sign of evolving sepsis in the setting of profound immunosuppression. The patient's clinical status began to deteriorate rapidly and he expired the next day from complications resulting from B-cell prolymphocytic leukemia, warm autoimmune hemolytic anemia, and combination chemotherapy. No autopsy was performed.
Leukemia is the predominant malignant disease of the leukocytes that affects the white blood cell lines whereas Hodgkin's disease, non-Hodgkin's lymphoma, and multiple myeloma manifest as tumor masses. Leukemia is a diffuse proliferation of neoplastic hematopoietic cells that infiltrate the bone marrow, blood, and other tissues of the body. Most leukemias are either acute or chronic, lymphocytic or myeloid.1
In acute leukemia, the cell type is poorly differentiated and lineage is difficult to identify morphologically. In chronic leukemia the cell type is more mature and readily identified. Untreated acute leukemias run a fatal course in a few months. Untreated chronic myeloid leukemias have a three to four year survival rate whereas CLL patients may have a many years survival rate.1
CLL vs PLL
B-prolymphocytic leukemia (B-PLL) is a morphological variant of B-chronic lymphocytic leukemia (B-CLL). CLL is predominantly a disease of adults over age 40. Diagnosis is easily made from the peripheral blood by finding an absolute lymphocytosis between 10 to 15 x 10^sup 9^/L. The nucleus of the lymphocyte is round with block type chromatin clumping and a small amount of light blue cytoplasm. Lymphocytes are very fragile and will rupture as the slide is made, resulting in 'smudge' cells. Large lymphocytes may be seen with looser, less clumped chromatin and a single nucleolus, which are actually prolymphocytes. As the disease progresses, lymphocytes in bone marrow replace normal cells resulting in normocytic normochromic anemia, neutropenia, and thrombocytopenia.2
Examination of the bone marrow demonstrated a focal or diffuse infiltration of lymphocytes of the same morphology as the peripheral blood. Approximately 90% of the lymphocytes were of B-cell origin with a uniform distribution of surface immunoglobulin whose density was less than normal lymphocytes. The heavy chain is usually associated with a monoclonal light chain. B-CLL cells usually exhibit markers for CD 19, CD20, CD24, and T-cell marker CD5.(3)
CLL has the capacity to transform into other disease processes. It can transform into prolymphocytic leukemia (PLL), with a predominance of prolymphocytes, and into Richter's syndrome (a large cell lymphoma).3 In PLL, abnormal lymphoid cells proliferate and accumulate in spleen, bone marrow, and liver. Hypogammaglobulinemia is frequently present in these patients. PLL is characteristically seen in men over age 60 who present with symptoms of fatigue, weakness, night sweats, and fever. Presenting symptoms in PLL are usually acute, whereas in CLL onset may be insidious. The most common physical sign is greatly enlarged spleen and liver. However, lymphadenopathy is uncommon. The prognosis for PLL is much poorer than CLL.1
PLL has B- and T-cell variants, and is characterized by larger, less mature appearing cells with condensed chromatin and prominent nucleoli. PLL is an aggressive disease with high leukocyte counts, splenomegaly, and no substantial adenopathy. Leukocyte counts in excess of 150 x 10^sup 9^/L, consisting almost entirely of prolymphocytes, is common. These cells are characterized by high-intensity surface membrane immunoglobulin and decreased mouse rosette formation, and usually express FMC-7 and CD22. The cells do not express CD5.(4)
PLL occurs less frequently than CLL with less being known about chromosomal abnormalities of the disease. A number of cases of PLL have been reported in the literature with about 50% of these having the same abnormality, a 14q marker with the breakpoint in band 14q32. The translocated material in most PLL patients has not been identified. Two rearrangements have been seen in PLL patients thus far: t(11I; 14)(q13:q32) and t(14; 17)(q32;q11). Other chromosomal aberrations include trisomy 12; deletions of chromosomes I and 6, usually at bands q32 and q21, respectively; terminal deletions of the short arm of chromosome 3, del (3) (p13); and, chromosome 12, del(12), (p12-13). Differential diagnosis of T-PLL and B-PLL can be established by immunological markers and clinical features (Table 4).4,5
LABORATORY FINDINGS AND CORRELATION WITH DISEASE
Peripheral blood leukocyte counts are usually elevated from 25 to 1000 x 10^sup 9^/L. The prolymphocytes present in the blood differ from patient to patient. The prolymphocyte is a large mononuclear lymphoid cell with an oval or round nucleus, with coarse chromatin strands and one or two large vesicular nucleoli with perinuclear concentration of chromatin. The cytoplasm is granular and basophilic with Romanowsky stains. On the basis of membrane phenotype, it is conjectured that the normal counterpart of the prolymphocyte in PLL can be found in the B mantle zones (lymphocyte corona) of the peripheral lymph nodes.2
Bone marrow examination reveals almost total replacement of the marrow with prolymphocytic infiltration, with only a few residual hematopoietic cells remaining. It is common for patients with PLL to be both anemic and thrombocytopenic.2
Treatment regimens for patients with B-PLL are defined by systemic symptoms, rapidly enlarging or painful lymphoid masses, autoimmune hemolytic anemia, thrombocytopenia, and progressive bone marrow failure. Lymphocytosis with or without symptoms is not an indication for therapy.6
Therapy is guided to reduce the lymphocyte mass in the blood, marrow, and tissues; to reduce symptoms; and, to improve hematopoiesis in patients who are anemic or thrombocytopenic. PLL responds poorly to the therapies that are successful with CLL, such as corticosteroids and alkylating agents (worsen the myelosuppression). Prednisone may cause fluid and sodium retention with increased calcium and potassium, as well as carbohydrate intolerance, glycosuria, and hyperglycemia. Some patients with PLL respond to combination of chemotherapeutic agents such as cyclophosphamide, doxorubicin (POACH), vincristine, and prednisone.6
Prognosis and response to treatment are poor, with a median survival of three years. PLL is usually less responsive to refractory treatment and has a poorer prognosis than does CLL.7-9
AUTOIMMUNE HEMOLYTIC ANEMIA (AIHA)
Approximately 10% of patients with PLL acquire an autoimmune hemolytic anemia (AIHA). AIHAs are caused by an altered immune response that results in the production of antibodies against the patient's own red blood cells, with subsequent hemolysis. The cause of antibody production is unknown. Development of AIHA in patients with lymphoproliferative or autoimmune disorders is thought to be due to an abnormality with the B cells, T cells, macrophages, or interaction between these cells.7-10
AIHAs are classified according to the thermal properties of the red cell antibody in question. Warm antibodies bind to red cells at 37 deg C; whereas cold antibodies display greater affinity for red cells as the temperature approaches 0 deg C. Warm antibodies are usually IgG; cold antibodies are predominantly IgM. Warm antibodies facilitate the sequestration of sensitized red cells by the spleen while cold antibodies cause intravascular destruction of sensitized red cells by a complement-mediated mechanism. A third group, 'mixed' warm/cold antibodies, tends to be implicated in severe hemolytic episodes characterized by a chronic intermittent course that is relatively resistant to available treatments.1,10
Warm antibodies do not bind complement. Destruction of the red cells occurs as a result of the red cell-bound autoantibodies to the Fc receptors on monocytes and macrophages; phagocytosis and cytotoxic lysis of the red cells subsequently occur. AIHA occurs in both sexes, at any age, but the onset increases after age 40. The course of the anemia ranges from mild with gradually developing symptoms to acute with fulminating symptoms. Physical findings include weakness, fever, pain, hemoglobinuria, jaundice, hepatosplenomegaly, and lymphadenopathy.1,6
Hemoglobin and hematocrit values vary with the severity of the disease process. The red cells are often macrocytic, reflecting a young population of cells, and exhibit anisocytosis. Spherocytes indicate the presence of the hemolytic process. Reticulocytosis indicates that the bone marrow is attempting to compensate for the hemolysis; however, infiltration of the marrow by malignant cells can result in a marked decrease in reticulocyte count. Thrombocytopenia may also be present and the WBC may vary from moderate leukopenia to moderate leukocytosis.1,3,5
The serum bilirubin level is moderately increased (usually 2.5 to 5 mg/dL) with the indirect bilirubin being predominant. Haptoglobin may be at very low levels when hemolysis is severe in spite of extravascular red cell destruction. Urine and fecal urobilinogen levels are elevated. The osmotic fragility is usually increased in direct proportion to the number of spherocytes in the peripheral smear.1,3,5
The DAT is usually positive, confirming the presence of IgG antibodies with or without complement on the red cells. Warm antibodies may have specificity for Rh antigens.1,9
TREATMENT OF AHIA
Treatment of the underlying malignancy may result in a reversal of the hemolytic process. Often it is necessary to treat the hemolytic anemia as a separate entity. One or more of the following measures may be employed:
In patients with severe disease, as an emergency measure, blood transfusion may be necessary. When transfusion is utilized for severe anemia, problems are evident. There may be difficulty in accurate typing of blood to find a compatible unit. The destruction of the transfused blood is usually as rapid as that of the patient's own blood. It is prudent to use only the amount of blood needed to stabilize patient. It is recommended that the patient be phenotyped and that red blood cells negative for the same antigens as the patient be transfused to avoid formation of alloantibodies which would further complicate the picture.1,3
Adrencortical Hormones or Steroids
The adrenocortical hormones comprise the initial treatment of choice in patients with warm-antibody type of hemolysis. Response to therapy is quickly evident, but hematologic improvement may not be seen for up to a week. Corticosteroids inhibit clearance of IgG-sensitized red cells and suppress synthesis of antibodies.1,6
For those patients who will not respond to corticosteroid therapy, splenectomy is recommended. The spleen is the major site where IgG-sensitized cells are sequestered. The results of splenectomy improve greatly by selecting patients who have demonstrated excessive splenic sequestration of sensitized red cells.1,6
Splenic irradiation has been recommended for patients with warm-- AIHA who are resistant to treatment and in whom splenectomy is contraindicated.11
Many cytotoxic drugs are potently immunosuppressive, which provides theoretic basis for treatment of patients with AIHA; however, many of these chemotherapeutic agents are contraindicated in treatment of B-PLL. Several drugs, including thiopurines (azathioprine, 6-mercaptopurine, thioguanine) as well as alkylating agent cyclophosphamide, have achieved intermediate success. Other drugs include 2-chloro-2'-deoxyadenosine, and cycloporine. Cyclophosphamide (Cytoxan) has been the most effective immunosuppressive agent available for treatment. But there have been no controlled studies conducted in clinical trials to determine clear guidelines for treatment with these cytotoxic agents. These agents are reserved for patients with AIHA who do not respond to steroid therapy or who require extremely high maintenance doses of steroids and are poor candidates for splenectomy.6
Prognosis varies depending on severity of the hemolytic episodes and complexity of underlying diseases.2,6
The case study reported is of a patient initially misdiagnosed as chronic lymphocytic leukemia. Immunophenotyping studies ultimately identified the nature of the disease as B-cell prolymphocytic leukemia with concomitant warm autoimmune hemolytic anemia. The leukemia and hemolytic anemia were refractory to all conventional treatments administered. The patient survived a significantly shorter period of time than the median time of three years reported in the literature. The patient expired from complications resulting from B-cell PLL, warm autoimmune hemolytic anemia, and combination chemotherapy.
The Reports and Reviews Section seeks to publish information on important clinical laboratory-related topics such as technological clinical and experimental advances and innovations. Case studies and literature reviews are also included. In addition, brief reviews of books, computer programs, audiovisual materials or other materials of interest to readers are appropriate for this section. Manuscripts and literature reviews published as a Report are peer reviewed. Direct all inquiries to Virginia R. Kotlarz PhD, Peer Review Editor Medical Technology Department, Daemen College, 4380 Main Street, Amherst NY 14226. (716) 839-8425, (716) 839-8516 (fax). firstname.lastname@example.org
1. Stiene-Martin EA, Lotspeich-Steininger CA, Koepke JA. Clinical hematology. Philadelphia: Lippincott; 1998. p 428-9, p 482-6.
2. Matures, E. T-cell prolymphocytic leukemia. Cancer Control: JMCC 1998;5(1):19-24.
3. McKenzie SB. Textbook of hematology. Baltimore: Williams and Wilkins; 1996. p 263,413-4.
4. Lens D, Coignet LJ, Brito-Babapulle VI and others. B cell prolymphocytic leukemia (B- PLL) with complex karyotype and concurrent abnormalities of the p53 and c-MYC gene. Leukemia 1999;13(6):873-6.
5. Turgeon ML. Clinical hematology theories and procedures. Philadelphia: Lippincott, Williams and Wilkins; 1999. p 232.
6. Catovsky D. Current approach to the biology and treatment of chronic lymphoid malignancies other than CLL. Hematology and Cell Therapy 1996;38(2):S63-6.
7. Shvidel L, Shtalrid M, Bassous M, and others. B-cell prolymphocytic leukemia: a survey of 35 patients emphasizing heterogeneity, prognostic factors, and evidence for a group with an indolent course. Leukemia and Lymphoma 1999;33(1-2):169-79.
8. Hoffman MA, Valderrama E, Fuchs A, and others. Leukemic meningitis in B-cell prolymphocytic leukemia. A clinical, pathologic and ultrastructural case study and a review of the literature. Cancer 1995;75:1100-3.
9. Stevens ML. Fundamentals of clinical hematology. Philadelphia: WB Saunders Company; 1997. p 101-2.
10. Lehmann CA. Saunders manual of clinical laboratory science. Philadelphia: W. Saunders Company; 1998. p 516-7,860-1,888-890,935-7.
11. Yamamoto K, Hamaguchi H, Nagata K, and others. Splenic irradiation for prolymphocytic leukemia: is it preferable as an initial treatment or not? Jpn J Clin Oncol 1998;28(4):267-9.
Josie C Roberts is a Clinical Laboratory Scientist at St Francis Medical Center, Monroe LA.
George HR oberts is Professor and Department Head, Clinical Laboratory Science, The University of Louisiana at Monroe, Monroe LA.
Address for correspondence: George H Roberts EdD, The University of Louisiana at Monroe, 700 University Avenue, Monroe LA 71210,
(318) 342-1632, (318) 342-3256 (fax). email@example.com
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