Trisomy 8 in Acute Myeloid Leukaemia

Two cases of acute myeloid leukaemia with trisomy 8 in all examined bone marrow cells are reported. The occurrence and the prognostic significance of trisomy C in myeloproliferative disorders are discussed. The published reports of myeloproliferative disorders with chromosomal abnormalities identified by the banding technique are reviewed. It is to be noted that among group C anomalies in the acute myeloid leukaemias, only involvement of chromosome no. 7, 8 and 9 have been reported so far.     

Trisomy 8 in Acute Myeloid Leukaemia: A Non-Random Event

6 cases of AML with a supernumerary chromosome 8 as the only aberration in practically all bone marrow mitoses and 10 cases with a normal chromosome composition of the marrow cells were investigated in order to evaluate the possible influence of trisomy 8 on some clinical and cytokinetic parameters. No significant differences between the groups were found. In our laboratory a supernumerary chromosome 8 is present in 36% of AML cases with chromosomal aberrations.     

8/21 Translocation in Acute Myeloid Leukaemia

8;21 translocation was found in 10 AML patients. These patients exhibited a distinct clinical and haematological picture, characterized by M2 bone marrow, with rather good maturation, a high count of mature granulocytes, splenomegaly, and the absence of DIC. Complete remission was easily obtained. It was reported that the median survival is better than for other AML patients with abnormal karyotypes, but this could not be substantiated in our small series. The loss of a sex chromosome was found to be frequent and of poor prognostic significance.     

Serum and Plasma Myeloperoxidase,Elastase and Lactoferrin Content in Acute Myeloid Leukaemia

Myeloperoxidase (MPO) and elastase, restricted to azurophil granules of neutrophils, as well as lactoferrin, restricted to specific granules of neutrophils, were determined in plasma and serum from patients with acute myeloid leukaemia (AML). Highly sensitive radio immuno assays were developed for detection of these proteins. Serum MPO was increased in 12/35 and decreased in 2/35 patients without correlation to WBC or neutrophil counts; these levels may reflect an abnormal production by leukaemic blasts or ineffective granulopoiesis in the bone marrow. Serum elastase was increased in 6/22 patients. Serum lactoferrin was decreased in 12/25 patients without correlation to neutrophil counts probably reflecting abnormal production. Serum elastase and MPO showed a covariation in chronic myeloid leukaemia but not in AML; the latter finding may indicate that the synthesis of these two proteins is not synchronized in AML-cells. Sequential studies of patients with AML demonstrated fluctuations of serum MPO and lactoferrin during remission most likely because of chemotherapeutic pertubation. Although a limited number of patients has been studied it is suggested that serum lactoferrin may be of help for prediction of relapse in AML.     

Mitotic Activity of the Granulopoietic Precursor Cells in the Peripheral Blood in Chronic Myeloid Leukaemia

The mitotic indices (MI) of granulopoietic precursor cells in peripheral blood and bone marrow were studied in 38 patients with typical chronic myeloid leukaemia (CML) in the chronic phase. The MI in the peripheral blood were very low, in the median 0.07 %, compared to those in the bone marrow with a median of 1.07 %. The blood MI were significantly increasing with raising WCC and the values of the MI above the median were combined with short survival times. The bone marrow MI were negatively correlated to the blood MI and it is suggested that this is a sign of an increased exchange of cells between bone marrow and blood.     

The Incidence and Characteristics of Acute Myeloid Leukaemia Arising in Hodgkin's Disease

In 201 consecutive patients with Hodgkin's disease treated from 1964 through 1975 with intensive irradiation and/or combination chemotherapy 3 cases of acute myeloid leukaemia were observed. The observed number of cases was 75 times over the expected (P < 0.01). An analysis of 47 reported cases of acute myeloid leukaemia arising in Hodgkin's disease shows that these cases differ considerably from ‘spontaneous’ cases of acute myeloid leukaemia by appearing in a much younger age group, by a very poor response to anti-leukaemic chemotherapy, and by a relatively low male/female ratio (0.84). The intensification of radiotherapy and cytostatic therapy of Hodgkin's disease during the last decade is considered the explanation of the increased incidence of acute myeloid leukaemia in these patients.     

Results of a randomized trial in children with Acute Myeloid Leukaemia: medical research council AML12 trial

The Medical Research Council Acute Myeloid Leukaemia 12 (MRC AML12) trial (children) addressed the optimal anthracenedione/anthracycline in induction and the optimal number of courses of consolidation chemotherapy. 504 children (<16 years) with AML were randomized between mitoxantrone/cytarabine/etoposide or daunorubicin/cytarabine/etoposide as induction chemotherapy and 270 entered a second randomization between a total of four or five courses of treatment. Ten‐year event‐free (EFS) and overall survival (OS) was 54% and 63% respectively; the relapse rate was 35%. There was no difference in complete remission rate between the induction regimens, but there was a benefit for mitoxantrone with regard to relapse rate [32% vs. 39%; Hazard ratio (HR) 0·73; 95% confidence interval (CI) 0·54, 1·00] and disease‐free survival (DFS; 63% vs. 55%; HR 0·72; 95% CI 0·54, 0·96). However, this did not translate into a better EFS or OS (HR 0·84; 95% CI 0·63, 1·12). Results of the second randomization did not show a survival benefit for a fifth course of treatment (HR 1·01; 95% CI 0·63, 1·62), suggesting a ceiling of benefit for conventional chemotherapy and demonstrating the need for new agents. EFS was superior compared to the preceding trial AML10, partly due to fewer deaths in remission, highlighting the importance of supportive care.     

P-Glycoprotein Expression in Acute Myeloid Leukaemia Cells at Diagnosis: Its relationship to Daunorubicin or Idarubicin Induction Therapy and Survival

We investigated the expression of P-glycoprotein (P-gp) in 50 adults with de novo diagnosed acute myeloid leukaemia (AML) and the relationship between presence of P-gp in leukaemic cells and efficacy, as remission induction and survival rate, of two different anthracyclines, daunorubicin (DNR) and idarubicin (IDR).

We found that 30 out of 50 patients (60%) were negative (Group 1) and 20 (40%) were positive (Group 2) for P-gp expression evaluated by mean of MRK16 MoAb using a cut-off of 10% positive cells. Thirty-five out of 50 patients (70%) obtained complete remission (CR); depending on P-gp expression, the CR rate was 80% for group 1 and 45% for group 2 (p lt; 0.005).

The median duration of overall survival was 20 months for patients in Group 1 as compared with 10 months for patients of Group 2 (p < 0.005).

Regarding the anthracycline used, no significant difference in CR was observed in patients of Group 1 (75% of CR with DNR vs. 90% with IDR); Group 2 obtained 40% of CR with DNR vs. 70% with IDR (p < 0.005). The median duration of overall survival (OS) with the two regimens was comparable in Group 1, while it was significantly longer in patients of Group 2 treated with IDR compared with DNR regimen (p < 0.005).

These results confirm the prognostic value of P-gp expression in AML at first appearance and we suggest that idarubicin could be a valid anthracycline drug in the treatment of AML to be evaluated as potential drug of choice in patients with primary or drug-induced multidrug resistance.     

Correlation between Chromosomal Pattern,Cytological Subtypes,Response to Therapy,and Survival in Acute Myeloid Leukaemia

Chromosomal karyotypes were determined with standard G-banding in 103 patients with acute myeloid leukaemia (AML). Abnormal clones were present in 52 (50.5%). Higher frequencies of abnormalities were observed in male than in female patients and in erythro-leukaemia (EL) than in other subtypes of AML. Abnormalities were more frequent in myeloblastic (AMyL) than in myelomonocytic leukaemias (AMML) and mixtures of both normal and abnormal karyotypes were more common among elderly patients; these differences, though of marginal statistical significance, are consistent with previous reports. 9 of 10 cases with 5 or more aberrant chromosomes and 6 of 8 cases with unidentified marker chromosomes were either AMML or EL. Remission rates, median survivals and relative death rates were collated in 82 patients, in relation to the karyotype patterns NN (all normal), AN (mixed normal and abnormal) and AA (all abnormal). The differences between the groups did not reach statistical significance. Serial cytogenetic studies were performed in 10 patients. New karyotype changes emerged in only 1 of 6 relapses. 4 examples of t(8;21) and 2 of t(15;17) were found. 1 case of AMML showed trisomy 8 and double minute chromosomes. 1 case of EL showed 2 marker chromosomes with homogeneously staining regions.     

Chromosome Abnormalities Identified by Banding Technique in a Patient with Acute Myeloid Leukaemia Complicating Hodgkin's Disease

Chromosome analyses using the Giemsa banding technique were performed on bone marrow cells in a patient with the association of Hodgkin's disease and acute myeloid leukaemia. All cells had an abnormal karyotype showing an extra chromosome No. 14, loss of one chromosome No. 17 and gain of one chromosome No. 18. These abnormalities are in many respects similar to the karyotype changes of lymphoid cells in malignant lymphomas, suggesting a pathogenetic relationship between the two disorders.     

Treatment-related deaths during induction and first remission of acute myeloid leukaemia in children treated on the Tenth Medical Research Council Acute Myeloid Leukaemia Trial (MRC AML10)

Between 1988 and 1995, 341 children with acute myeloid leukaemia (AML) were treated on the Medical Research Council Acute Myeloid Leukaemia Trial (MRC AML10). The 5-year overall survival was 57%, much improved on previous trials. However, there were 47 deaths (13. 8%), 11 of which were associated with bone marrow transplantation (BMT). The treatment-related mortality was significant at 13.8%, but decreased in the latter half of the trial from 17.8% in 1998-91 to 9. 6% in 1992-95 (P = 0.03%). The main causes of death were infection (65.9%), haemorrhage (19.1%) and cardiac failure (19.1%). Fungal infection was a significant problem, causing 23% of all infective deaths. Haemorrhage occurred early in treatment, in children with initial white cell counts >100 x 109/l (P = 0.001), and was more common in those with M4 and M5 morphology. Cardiac failure only occurred from the third course of chemotherapy onwards, with 78% (7/9) in conjunction with sepsis as a terminal event. Some deaths could be prevented by identifying those most at risk, and with prompt recognition and aggressive management of complications of treatment. Future options include the prophylactic use of antifungal agents, and the use of cardioprotectants or alternatives to conventional anthracyclines to decrease cardiac toxicity.     

Generation of Procoagulant Activity (PCA) by Macrophage-Like Cells Derived from Acute and Chronic Myeloid Leukaemia Cells in Response to Phorbol Esters

Normal human monocytes and macrophages, as well as in vitro human leukaemic promyelocytic cell line (HL-60) transformed into macrophage-like cells by 12-0-tetradecanoyl-phorbol-13-acetate (TPA) generate potent procoagulant activity (PCA) similar to tissue thromboplastin. In the present study, only mild PCA was detected in primary cultures of cells from the peripheral blood of patients with acute lymphatic leukaemia (ALL), acute myeloid leukaemia (AML) and chronic myeloid leukaemia (CML). After exposure to TPA, AML and CML cells assumed characteristics specific to monocytes and macrophages. Differentiation was associated with the generation of PCA. PCA was not found in ALL cells exposed to TPA. The PCA of TPA-induced macrophages derived from AML and CML cells resembled tissue thromboplastin and normal monocyte and macrophage PCA in several aspects: (a) accelerated clotting via the extrinsic coagulation pathway, (b) inhibition by concanavalin A and protection against lectin inhibition by methyl-α-D-mannopyranoside, (c) localization in the cell membrane. The capacity for PCA generation is additional evidence for the similarity between TPA-induced macrophages derived from AML and CML cells and normal human monocytes and macrophages.     

Bone marrow stroma‐mediated resistance to FLT3 inhibitors in FLT3‐ITD AML is mediated by persistent activation of extracellular regulated kinase

A consistent pattern of response has been observed when FMS‐like tyrosine kinase 3 (FLT3) tyrosine kinase inhibitors (TKIs) have been used as monotherapy to treat patients with relapsed or refractory FLT3‐ internal tandem duplication (ITD) acute myeloid leukaemia (AML). Circulating blasts are cleared from the peripheral blood, while bone marrow blasts are either unaffected or are cleared from the marrow at a much slower rate. We used an in vitro model of FLT3‐ITD AML blasts co‐cultured with normal human bone marrow stromal cells to investigate the basis for this dichotomous response pattern to FLT3 inhibitors. We have found that in blasts on stroma, potent FLT3 inhibition predominantly results in cell cycle arrest rather than apoptosis. The anti‐apoptotic effect is mediated through a combination of direct cell‐cell contact and soluble factors. The addition of exogenous FLT3 ligand (FL) augments the protection, primarily by shifting the 50% inhibitory concentration for FLT3 inhibition upwards. Cytokine‐activated extracellular regulated kinase (ERK), rather than STAT5, appears to be the most important downstream signalling protein mediating the protective effect, and inhibition of MEK significantly abrogates stromal‐mediated resistance. These findings explain the phenomenon of peripheral blood versus bone marrow blast responses and suggest that the combination of potent FLT3 inhibition and MEK inhibition is a promising strategy for the treatment of FLT3‐ITD AML.     

Mutation in the ATP-binding site of BCR-ABL in a patient with chronic myeloid leukaemia with increasing resistance to STI571

The Abl kinase inhibitor STI571 (imatinib mesylate) induces haematological remissions in many patients with chronic myeloid leukaemia (CML) but advanced stage CML usually becomes resistant to STI571. We describe a patient in whom progressive resistance to STI571 correlated with the appearance of a mutation in the Bcr-Abl kinase domain. This was a G to A transition that resulted in a glutamic acid to lysine substitution at position 255 (E255K) in the Abl type 1a protein. We suggest that the acquisition of point-mutations in the tyrosine kinase domain of Bcr-Abl may cause progressive clinical resistance to STI571.     

Allogeneic haematopoietic cell transplantation for adult acute myeloid leukaemia in second remission: a retrospective study of the Adult Acute Myeloid Leukaemia Working Group of the Japan Society for Haematopoietic Cell Transplantation (JSHCT)

To evaluate the outcomes and prognostic factors following allogeneic haematopoietic cell transplantation (HCT) for adult acute myeloid leukaemia (AML) in second complete remission (CR2), we retrospectively analysed the Japanese registration data of 1080 adult AML patients in CR2 who had received allogeneic HCT. The probability of overall survival and the cumulative incidence of relapse at 3 years was 66% and 19%, respectively. In multivariate analysis, older age, poor cytogenetics and shorter duration of first complete remission were significantly associated with a higher overall mortality. Our data demonstrated the significant efficacy of allogeneic HCT for adult AML in CR2.     

The expression of P-glycoprotein in AML cells with FLT3 internal tandem duplications is associated with reduced apoptosis in response to FLT3 inhibitors

P-glycoprotein (pgp), a membrane efflux pump, is recognized to have an anti-apoptotic function. Internal tandem duplications (ITDs) of the Fms-like tyrosine kinase 3 (FLT3) receptor are the most common mutations in acute myeloid leukaemia (AML). Both ITDs and pgp positivity confer an adverse clinical prognosis. FLT3 inhibitors induce variable apoptosis in cell lines transfected with FLT3 ITDs. We studied the effect of herbimycin A, AG1296 and PKC412 on primary AML blasts. All compounds showed significantly higher cell kill after 48-h incubation in samples with an ITD compared with wild type (Herbimicin P < 0.001; AG1296 P = 0.001, PKC412, P = 0.002). Pgp-positive samples were significantly less sensitive to herbimycin and AG1296 than pgp-negative samples, although neither molecule inhibited the efflux function of pgp. The concurrent incubation with the pgp inhibitor PSC833 resulted in an enhanced cell kill in 4/5 ITD pgp-positive samples versus two of nine ITD pgp-negative samples. PKC412 inhibited pgp function and induced cell death in FLT3 ITD/pgp-positive samples. We conclude that AML samples with a FLT3 ITD are more susceptible to these inhibitors than wild-type samples. However, the expression of pgp in cells with FLT3 ITDs can reduce their sensitivity to FLT3 inhibitors and therefore pgp expression should be assessed in clinical trials of FLT3 inhibitors.     

CXCL12 and CXCR7 are relevant targets to reverse cell adhesion‐mediated drug resistance in multiple myeloma

Cell adhesion‐mediated drug resistance (CAM‐DR) by the bone marrow (BM) is fundamental to multiple myeloma (MM) propagation and survival. Targeting BM protection to increase the efficacy of current anti‐myeloma treatment has not been extensively pursued. To extend the understanding of CAM‐DR, we hypothesized that the cytotoxic effects of novel anti‐myeloma agents may be abrogated by the presence of BM stroma cells (BMSCs) and restored by addition of the CXCL12 antagonist NOX‐A12 or the CXCR4 inhibitor plerixafor. Following this hypothesis, we evaluated different anti‐myeloma agents alone, with BMSCs and when combined with plerixafor or NOX‐A12. We verified CXCR4, CD49d (also termed ITGA4) and CD44 as essential mediators of BM adhesion on MM cells. Additionally, we show that CXCR7, the second receptor of stromal‐derived‐factor‐1 (CXCL12), is highly expressed in active MM. Co‐culture proved that co‐treatment with plerixafor or NOX‐A12, the latter inhibiting CXCR4 and CXCR7, functionally interfered with MM chemotaxis to the BM. This led to the resensitization of MM cells to the anti‐myeloma agents vorinostat and pomalidomide and both proteasome inhibitors bortezomib and carfilzomib. Within a multicentre phase I/II study, NOX‐A12 was tested in combination with bortezomib‐dexamethasone, underlining the feasibility of NOX‐A12 as an active add‐on agent to antagonize myeloma CAM‐DR.