A 30-year old Nepalese man presented to emergency department with 2 weeks' history of fever, abdominal pain, and fatigue. Physical examination revealed fever, pallor, jaundice, and hepatosplenomegaly. Peripheral blood analysis revealed moderate normochromic normocytic anemia (hemoglobin, 9 g/dL; normal, 13-17 g/dL), severe thrombocytopenia (platelets, 25 x 109/L; normal, 150-400 x 109/L), and marked leukocytosis of 164 x 109/L (normal, 4-10 x 109/L). Peripheral blood smear showed many blast cells with left shift and increased monocytic cells with a differential count of neutrophils 7%, lymphocytes 3%, eosinophils 1%, monocytes 16%, promyelocytes 8%, promonocytes 23%, and blasts 42%. The blasts were medium to large in size with fine chromatin and prominent nucleoli, some with irregular/convoluted nuclear contour (Figure 1A).
Initial bone marrow (BM) aspirate stained with Wright stain was hypercellular with 39% blasts, 13% promonocytes (blasts equivalent), and increased monocytes (17%). No Auer rods were noted. Maturing myeloid cells comprised (28%) severe dysplastic features and included 2% eosinophils (Figure 1B). Bone marrow biopsy showed markedly hypercellular BM with diffuse infiltration by blasts (Figure 1C), positive for CD34, lysozyme, myeloperoxidase, and CD68, with partial positivity for CD117. There were prominent eosinophilic cells, scattered and in small groups with marked suppression of normal trilineage hemopoiesis.
Multicolor flow cytometry (FCM) analysis was performed on BM aspirate using CD45-gating strategy to identify the immunophenotype of the blasts. Acute leukemia panel of 28 antibodies in a 4-color combination (FITC/PE/ECD/PC5 fluorescent conjugates) was used, as follows: (1) CD34/CD117/CD45/CD19, (2) CD14/CD13/CD45/CD64, (3) HLA-DR/CD7/CD45/CD5, (4) CD34/CD33/CD45/CD56, (5) CD19/CD10/CD45/CD3, (6) CD15/CD33/CD45/CD2, (7) CD9/CD19/CD45/CD4, (8) CD20/CD10/CD19/CD45, (9) cMPO/cCD79a/cCD3/sCD45, (10)TdT/sCD19/sCD3/sCD45, (11) CD36/CD11c/CD45/CD11b, and (12) CD41/glycophorin A/CD45/CD61(PC7).
Flow cytometry analysis revealed approximately 56% myeloid blasts with moderate CD45 and expressing CD34, cMPO, CD33, CD13, and CD9, and the majority was positive for CD117 and HLA-DR, with partial expression of CD15 (on approximately 35% of the cells) (Figure 2). The blasts were negative for CD56, CD14, CD64, CD11b, CD11c, CD36, CD61, CD41, glycophorin A, TdT, and B-cell and T-cell markers. Monocytic cells increased (23%) expressing CD64, and the majority was positive for CD14. Cells in the granulocytic gate comprised approximately 17% showing loss of CD10 expression (features of dysmaturation).
The morphology and FCM findings were consistent with the diagnosis of AML with increased monocytic cells. Cytogenetic analysis by fluorescent in situ hybridization (FISH) using a probe for CBFB (BAR) gene revealed an abnormal hybridization signal pattern, indicating rearrangement in 56% of the cells. This was confirmed by conventional karyotype which showed inv(16)(p13.1q22), concluding the diagnosis of AML with inv(16)(p13.1;q22) CBFB:MYH11 (Figure 3).
Patient initial laboratory workup revealed elevated total bilirubin of 82.6 micromol/L (normal, 3-20 micromol/L), mainly direct bilirubin of 55.8 micromol/L (normal, 0-8.6 micromol/L), and high liver enzymes, with alanine transaminase (ALT) of 187 IU/L (normal, 0-40 IU/L), aspartate transaminase (AST) of 87 IU/L (normal, 0-37 IU/L), and alkaline phosphatase of 378 IU/L (normal, 40-150 IU/L). Renal function tests were normal with creatinine of 94 micromol/L (normal, 70-115 micromol/L). Hepatitis serology screening showed hepatitis E virus IgG and IgM with evidence of past hepatitis B virus infection, for which lamivudine was given before starting chemotherapy. Magnetic resonance imaging of the liver and magnetic resonance cholangiopancreatography done to exclude biliary obstruction revealed hepatomegaly with no focal lesion, multiple porta hepatis, peripancreatic, mesenteric, and para-aortic lymph nodes with no evidence of biliary obstruction. All septic workup and quantiferon testing proved negative.
The patient was started on cytarabine (100 mg/m2) for 7 days. Anthracycline was omitted from the first induction because of elevated direct bilirubin. The patient had persistent fever during the neutropenia phase despite treatment with broad-spectrum antimicrobial and antifungal drugs. Miliary tuberculosis (TB) was suspected, for which liver biopsy was performed and was consistent with cholestatic hepatitis with the absence of granulomas or leukemic infiltration. The patient had no clinical findings, suggesting central nervous system (CNS) involvement; however, intrathecal chemotherapy was planned upfront as the patient was considered to be at high risk due to CNS involvement. Examination of cerebrospinal fluid was delayed as the patient was critically ill; it was done just before the first consolidation and revealed few blasts confirmed by FCM analysis. The patient was treated with triple intrathecal chemotherapy twice weekly for total of 8 doses.
Bone marrow examination post first chemotherapy cycle showed cellular aspirate with approximately 19% blasts, and BM biopsy showed interstitially increased primitive cells. Immunohistochemical stains showed increased CD34 positivity roughly estimated at 20% to 30% and overall increased positivity for CD68 with focal collections of CD117-positive cells interpreted as persistence of the leukemic process. After recovery of neutropenia, the patient was started on second induction 3+7 protocol. The hospital course was complicated with massive pleural effusion. Therapeutic pleural tapping was done and showed white blood count of 1000/microL with 65% lymphocytes, no blast cells, glucose of 6.2 mmol/L, protein of 50.6 g/L, lactate dehydrogenase of 104 IU/L, pH 7.5, and negative for acid-fast bacilli and TB by polymerase chain reaction (PCR). The patient was treated empirically with anti-TB drugs and steroids. Few days later, fever subsided, jaundice resolved, and the patient's condition improved.
Evaluation of BM after second induction showed cellular BM with 3% blasts, indicating remission. However, the BM aspirate showed the presence of morphologically atypical MC comprising approximately 2% of the total cells, mostly with irregular nuclear contour (kidney-shaped, bilobed, or multilobed nuclei), pale cytoplasm with cytoplasmic hypogranulation or irregular metachromatic granule distribution, some with fine nuclear chromatin (promastocyte morphology), together with some cells showing eccentric nucleus or spindle shape (Figure 4A and B). Examination of the BM biopsy revealed multiple perivascular and randomly distributed focal collections of MC (in clusters of >15 cells) with some interstitially increased MC with spindle forms and some with lobed nucleus. Eosinophils were prominent around few of the MC aggregates. The MC were positive for tryptase, CD117, CD68, and CD25. CD2 immunostain appeared negative (Figure 5A to C). These findings established the diagnosis of SM with AML. When reexamining the BM at diagnosis, MC were not impressive in the aspirate; however, re-evaluation of the marrow biopsy with the aid of tryptase and CD25 immunostain revealed many scattered and multiple dense clusters of cells positive for CD117, tryptase, and CD25, indicating that mastocytosis was present from the start but masked by the extensive infiltration by blasts.
Serum tryptase was elevated at 38.5 microg/L (negative <11 microg/L), and allele-specific oligonucleotide polymerase chain reaction (ASO-PCR) for KIT Asp816Val gene mutation was performed on peripheral blood and proved to be negative.
First consolidation with high-dose cytarabine 3 g/m2 on days 1, 3, and 5 along with idarubicin 12 mg/m2 was given. The course was complicated with pleural effusion recurrence which required therapeutic thoracentesis. Cytology showed no blast cells. Subsequently, the patient received 2 further consolidation courses with high-dose cytarabine that was complicated with febrile neutropenia with Enterobacter cloacae bacteremia on third cycle that required intensive care admission and antimicrobial treatment with meropenem. Bone marrow evaluation after completion of therapy revealed no increase in blast cells, but there was persistence of mastocytosis. As there was a high risk of relapse, allogeneic stem cell transplant was planned. However, it was not done as the patient traveled back to his home country.