11q Aberration Research Articles

Biologic Features and Treatment Outcome of Secondary Acute Lymphoblastic Leukemia - Review of 101 Cases.

Secondary acute lymphoblastic leukemia (sALL) is a rare disease and its biologic features are not well described. Data suggested that sALL occurs more frequently at older age. Since leukemia at older age in general is associated with worse outcome, we wanted to assess the biology and outcome of patients with sALL by age at time of primary diagnosis. We describe a cohort of 7 patients and found additional 94 cases in the literature on whom biological parameters were described. Patients were stratified (at least 5 patients per strata) according to their age at initial diagnosis (<18, 18–59, and ≥60 years of age), initial diagnosis [acute myeloid leukemia (AML), Hodgkin's disease (HD), neuroblastoma, breast and prostate cancers], cytogenetic groups [diploid, t(9;22), 11q23 aberrations, complex karyotype], immunophenotypes (B vs. T), and Burkitt's defined by either morphology and/or cytogenetic analysis demonstrating c-myc rearrangement. A total of 101 patients were evaluated; 29 were <18, 54 were 18–59 and 18 were ≥60 years old. The distribution of primary diagnoses was as expected: neuroblastoma was seen only in the <18 age group (P=0.003), HD was more common in the 18–59 age group (75% of all HD cases; P=0.084) while breast (P=0.003) and prostate (P=0.005) cancers were prevalent only in the >18 year old patients. The time interval to develop sALL was similar among the three age groups (3, 2.2 and 1.8 years, P=0.561). However, the time interval to develop sALL was longer for HD (5.5 years) and neuroblastoma (3.7 years) as compared to AML (1 year), breast (1.6 years) and prostate (1.98 years) cancers (overall P=0.0003). Further, the time interval to develop sALL was significantly longer for patients with complex karyotype (5.3 years) as compared to all other aberrations [11q23 − 1.78; t(9;22) − 1.9; diploid − 1.98 years; overall P=0.0497]. Disease characteristics at diagnosis were as follows: T cell immunophenotype was more common in the <18 age group (P=0.016) and the presence of 11q23, t(9;22), complex and normal karyotypes was equally distributed among the three age groups (P=0.2, 0.073, 0.635 and 0.271 individually). Complete remission was infrequent in the ≥60 age group (22.22%) compared to the other groups (73.9% for <18 and 67.7% for 18–59; P=0.025). Even though only patients <60 years old were transplanted (33.3% for <18 and 19.4% for 18–59; P=0.102), the overall survival was poor in all age groups [probability of survival at 1 year for <18=0.222, 18–59=0.226 and ≥60=0.3 (P=0.7941)]. Primary diagnoses, cytogenetic subgroups and immunophenotype did not affect outcome. In summary, the time interval to develop sALL is significantly longer for HD, neuroblastoma and complex karyotype. However, sALL is associated with very poor outcome regardless of age and any of the biologic features. Therefore, identification of prognostic factors to prevent the occurrence of sALL is needed.

Prognostic Factors in Adult Patients up to 60 Years with AML and Translocations of Chromosome 11q23: Individual Patient Data Based Analysis of the German AML-Intergroup.

The MLL gene on chromosome 11q23 is translocated to a variety of fusion partners in about 5–10% of adults with acute myeloid leukemia (AML). The heterogeneity of these genetic aberrations hampers risk stratification of the patients. To identify prognostic parameters in AML patients up to 60 years with 11q23 aberrations, a pooled data analysis of 8 trials for the treatment of adults with AML was performed. All patients (pts) received double induction with araC and an anthracycline followed by intensive consolidation with either a high dose araC based chemotherapy or an autologous or allogeneic stem cell transplantation (alloSCT). 180 pts with 11q23 translocations were identified by cytogenetics and/or molecular techniques. 76 pts (42%) had a t(9;11), 35 (19%) a t(6;11), 17 (9%) a t(11;19), 14 (8%) a t(10;11) and 10 (6%) a t(11;17). 28 pts (16%) had other translocation partners. 77 pts (43%) had secondary chromosomal aberrations. Median age was 39 years (range 16–60). Median platelet and PB blast count was 45G/l and 7.5G/l. Median follow up is 53 months. Complete remission rate was 71%. Logistic regression identified t(9;11) (OR 2.6; 95% CI 1.2–5.9), secondary chromosomal aberrations (OR 0.4; 95% CI 0.2–0.8) and secondary leukemia (OR 0.3; 95% CI 0.1–0.6) as significant factors for induction success. Four year overall survival (OS) was 29% (95% CI 25–33%). Multivariate analysis identified presence of a t(9;11) (HR 0.6; 95% CI 0.4–0.9), platelets above the median (HR 0.6; 95% CI 0.4–0.9), secondary leukemia (HR 1.9; 95% CI 1.2–2.9) and PB blasts above the median (HR 1.9 95% CI 1.3–2.8) as significant factors for OS. Combination of these factors separated three risk groups for OS (p=0.0001): Good, pts with de novo AML, t(9;11) and PB blasts below median (4y OS 61%); intermediate, remaining pts with platelets above median (4y OS 32%); poor, remaining pts with platelets below median (4y OS 15%). Four year relapse-free survival (RFS) was 29% (95% CI 24–34%). Cox regression analysis identified the presence of a t(6;11) (HR 1.8; 95% CI 1.1–3.0) and PB blasts above the median (HR 1.7; 95% CI 1.0–2.6) as independent risk factors for RFS. By combination of these factors, three risk groups for RFS were defined: Good, pts without t(6;11) with PB blasts below median (4y RFS 63%); intermediate, pts without t(6;11) with PB blasts above median (4y RFS 23%); poor, pts with t(6;11) (4y RFS 9%). The effect of different consolidation strategies was analyzed “as treated” in CR-pts who had received at least one cycle of consolidation. In these pts, RFS was improved by an alloSCT (MRD or MUD) in first CR (p=0.04). However, this effect was restricted to the RFS-intermediate risk group (p=0.029), whereas pts in the good or poor risk group did not benefit from an alloSCT in first CR. In conclusion, the prognosis of patients with translocations of chromosome 11q23 is heterogenous. Risk stratification of these patients is feasible based on the MLL-fusion partner and clinical parameters. This prognostic model can be used as a basis for the selection of consolidation therapy.

Update on the development of the international neuroblastoma risk group (INRG) classification schema

9503 Background: Modern treatment strategies for neuroblastoma (NB) are tailored according to patient risk. However, it is not currently possible to compare the results of clinical studies conducted around the globe because the criteria used to define risk are not uniform. A committee of international investigators with expertise in NB have worked during the past 2 years to develop a uniform International NB Risk Group (INRG) Classification System for pre-treatment stratification. Methods: Investigators from North America and Australia (COG); Europe (SIOPEN and Germany), and Japan collated data on 8,800 children with NB diagnosed between 1990 and 2002. Survival tree regression analyses tested 13 potential prognostic factors. Tumor differentiation, MKI, and diagnostic category were evaluated individually in lieu of the International NB Pathology Classification (INPC) system to determine if these histologic features had prognostic value independent from age. To stage patients at the time of diagnosis prior to surgery, a new staging system was developed (INRGSS) based on the presence or absence of image-defined risk factors (IDRFs) and metastases. Results: Since statistical analyses demonstrated support for an optimal age cut- off between 14–19 months, 18 months was selected. In addition to age, stage, MYCN amplification, tumor differentiation, ploidy, and genetic aberrations of 11q were found to be the most highly prognostically significant factors. These clinical and biological factors were combined to define 15 INRG pre-treatment groups. Patients with low- (3 groups), intermediate- (4 groups), high- (4 groups), or ultra-high-risk NB (4 groups) had EFS of ≥85%, &gt;70–85%, &gt;50–70%, or &lt;50%, respectively. Conclusion: International collaborative studies in NB will be greatly facilitated by the INRG classification system which will allow comparisons of different risk-based therapeutic approaches in homogeneous patient cohorts. No significant financial relationships to disclose.

Prognosis of Adult Patients ≤60 Years with AML and Aberrations of Chromosome 11q23: Pooled Data Analysis of the German AML-Intergroup.

Aberrations of the MLL gene on chromosome 11q23 occur in about 5–10% of adult patients with acute myeloid leukemia (AML). Most frequently, the MLL gene is fused to the genes AF9 on chromosome 9 or AF6 on chromosome 6 resulting in the translocation t(9;11) and t(6;11), respectively. The remaining patients with aberrations of chromosome 11q23 constitute a heterogeneous group with numerous fusion partners. This heterogeneity hampers risk stratification of the patients. Accordingly, AML patients with 11q23 aberrations have varyingly been stratified as intermediate or high risk patients by different study groups. To analyze the prognostic impact of different 11q23 aberrations in AML patients up to 60 years, a pooled data analysis of 6 consecutive studies for the treatment of adult AML patients was performed. All patients received double induction treatment with araC and an anthracycline followed by an intensive consolidation with either a high dose araC based chemotherapy regimen or an autologous or allogeneic stem cell transplantation. In total, 150 patients with 11q23 aberrations were identified by cytogenetics and/or molecular techniques. 53 patients (35%) had a t(9;11), 27 patients (18%) a t(6;11), 10 patients (7%) a t(11;19), 8 patients (5%) a t(11;17) and 6 patients (4%) a t(10;11). 46 patients (31%) had other fusion partners or deletions of 11q23. Median age of all patients was 39 years (range 16–60). Overall complete remission rate was 73% with no significant difference between the groups (79% for t(9;11), 74% for t(6;11) and 66% for others). Altogether, 9% of the patients had treatment failure and 7% died during induction. Relapse-free survival (RFS) and overall survival (OS) at 5 years for the entire group was 24% and 26%, respectively. RFS and OS of the t(6;11) group was 6% and 13% and thus was significantly inferior to patients with t(9;11) (RFS: 48%, OS: 31%) and patients with other 11q23 translocations (RFS: 20%, OS: 18%) or 11q23 deletions (RFS: 38%, OS: 33%). Within the t(9;11) group there was no difference in RFS or OS between patients who had a t(9;11) as a sole aberration (n=36) and patients with additional aberrations (n=17) and between patients with de novo or secondary AML. Moreover, in t(9;11) no difference in RFS was observed between treatment with high dose araC, autologous or allogeneic stem cell transplantation as late consolidation. Regarding other translocations of 11q23, there was no difference in RFS and OS between patients with t(11;17), t(11;19), t(10;11) and patients with various other fusion partners. In conclusion, these data demonstrate that the prognosis of patients with 11q23 is heterogeneous. Patients with t(9;11) who enter CR have a relatively good outcome independent of the consolidation therapy used. In contrast, the prognosis of patients with t(6;11) is extremely poor. Therefore, patients with t(6;11) should be regarded as high risk and alternative treatment strategies for this subgroup are required.

The Effects of Anti-High Molecular Weight-Melanoma Associated Antigen (HMW-MAA) Monoclonal Antibodies (mAb) Against 11q23 Positive Acute Leukemia Cells.

Acute leukemia with 11q23 aberrations is associated with especially poor outcome to standard treatment. HMW-MAA is a membrane bound proteoglycan that has been targeted in the treatment of malignant melanoma. It is widely distributed on 11q23-positive leukemic cells and is not expressed on normal hematopoietic cells. Finally, HMW-MAA is believed to function by signaling through the focal adhesion kinase (Fak)/proline-rich tyrosine kinase 2 (Pyk2) pathways and by supporting angiogenesis via its expression on pericytes. We hypothesized that inhibition of HMW-MAA with mAb directed against specific HMW-MAA determinants would effectively limit the growth of 11q23-positive acute leukemia cells in vitro and in vivo, by affecting Fak/Pyk2 pathways. We examined an 11q23-positive acute myeloid leukemia (AML) cell line (ML-2) and leukemia cells from two patients (one AML, one acute lymphoblastic leukemia) with 11q23 aberrations. Our results demonstrate that ML-2 cells expressed only the VT68.2 and 225.28 determinants, while primary acute leukemia cells expressed VT68.2, 225.28, and 763.74 determinants. Treatment of ML-2 cells with mAb against VT68.2 and 225.28 in vitro did not affect overall proliferation but downregulated phosphorylated (p) Pyk2 expression as determined by Western blotting. Further, anti-HMW-MAA mAb treatment enhanced the anti-proliferative effect of cytarabine (y=0.7875) in vitro. Based on these results, we then evaluated the dose-response relationships and toxicities of anti-HMW-MAA mAb alone and in combination with cytarabine in subcutaneous ML-2 xenografts grown in SCID mice. A trend towards tumor inhibition was noted following high dose (1 mg/kg 2x/week) anti-225 mAb therapy but was not seen at lower (0.25 and 0.5 mg/kg) anti-225 mAb doses. Mice demonstrated no side effects or weight loss as a result of anti-225 therapy. No downregulation of pPyk2 in tumor cells was observed following anti-225 therapy at any dose. Furthermore, combination anti-225.28 HMW-MAA mAb and cytarabine treatment (at increasing cytarabine doses) did not result in improved tumor inhibition as compared to cytarabine alone (p>0.05). We then evaluated the effects of HMW-MAA anti-225.28 and anti-763.74 mAb alone or together for treatment of NOD/SCID mice engrafted with primary 11q23-positive leukemia patient cells. Combination anti-HMW-MAA mAb therapy resulted in no prolongation in survival of engrafted mice as compared to control treated animals. We were able to confirm the presence of high levels of circulating HMW-MAA mAb in the serum of animals 4 days following mAb injection. In addition, explanted ML-2 cells harvested following xenograft growth still maintained expression of the HMW-MAA determinants. We conclude that naïve anti-HMW-MAA mAb neither caused a statistically significant inhibitory effect nor enhanced the effect of cytarabine on 11q23-positive acute leukemia cells in vivo. Future studies using radiolabelled or toxin-conjugated antibodies may enhance the anti-tumor effects of targeted HMW-MAA immunotherapy for 11q23-positive acute leukemia.

Array-CGH Based Genomic Profiling in B-Cell Chronic Lymphocytic Leukemia Reveals Specific Correlations of Genomic Imbalances and Prognostic Subgroups and Underlines the Consistency of Chromosomal Aberration Patterns within This Disease.

Chronic lymphocytic leukemia of B-cell type (B-CLL) is characterized by a number of typical genomic aberrations. In comparison to patients with normal karyotypes or 13q deletions patients with high risk imbalances such as deletions of 11q, 17p or unmutated IgVH status have a higher risk for advanced disease and a significantly shorter survival.For a precise mapping of chromosomal imbalances as well as to verify the number of aberrations per case in different genetic subgroups of B-CLL (e.g. del11q, del13q, del17p, +12 or unmutated IgVH status) we performed high resolution genomic profiling using a genomic DNA-chip containing 2.800 probes. Target clones compriseda large genome-wide cluster of clones covering the genome at a distance of approx. 1.5Mb andclones mapping to genomic regions or genes of possible pathogenetic relevance in lymphoma.This chip covers approximately 10% of the human genome.In 93 (70%) of 133 analyzed B-CLL cases 171 genomic imbalances were identified (between 1–7 aberrations/case). Besides the confirmation of known recurrent chromosomal aberrations, previously unknown recurrent imbalances were detectable on 2p (8%), 4p (4%), 7p-q (5%), 10q (5%) and 20p (3%). Most of these imbalances were of larger extension (> 10 Mb) and therefore impeded a further delineation of minimal aberrant regions and the identification of possible candidate genes. The mean number of chromosomal aberrations per case (= genomic complexity) in IgVH unmutated CLLs was approx. 2 times higher than in mutated cases (0,77 vs. 1,58 per case). 84% of samples with > 2 aberrations showed an unmutated IgVH status. Moreover, most of the previously unknown imbalances were identified within this group. A higher genomic complexity was also shown for samples with gain on 2p vs. balanced 2p status (2.2 times higher; 2.5 vs. 1.2) and in samples with del17p vs. balanced 17p status (3.7 times higher; 3.52 vs. 0.95). 11q aberrations had no impact on the number of genomic aberrations per case (1.6 vs. 1.2). Moreover, we found a strong association of 2p gains and an unmutated IgVH status (100%).Array based genomic profiling confirmes the chromosomal aberration structure and underlines the consistency of chromosomal aberration patterns of B-CLL. The biological and prognostic relevance of 2p gains and unmutated IgVH mutational status have to be further investigated.

Prognosis of Adult Patients ≤60 Years with AML and Aberrations of Chromosome 11q23: Pooled Data Analysis of the AML-SG and SHG-Dresden Study Groups.

Aberrations of the MLL gene on chromosome 11q23 occur in about 5–10% of adult patients with acute myeloid leukemia (AML). Most commonly, fusion of the MLL gene to the genes AF9 on chromosome 9 or AF6 on chromosome 6 are found resulting in the translocation t(9;11) and t(6;11), respectively. The remaining patients with aberrations of chromosome 11q23 constitute a heterogeneous group with numerous fusion partners. This genetic heterogeneity hampers risk stratification of these patients. Accordingly, AML patients with 11q23 aberrations have varyingly been stratified as intermediate or high risk patients by different study groups. To analyze the impact of different 11q23 aberrations on the prognosis of AML patients up to 60 years, a pooled data analysis of 5 consecutive studies for the treatment of adult AML patients was performed. All patients received double induction treatment with araC and an anthracycline followed by an intensive consolidation with either a high dose araC based chemotherapy regimen or an autologous or allogeneic stem cell transplantation. In total, 137 patients with 11q23 aberrations were identified by cytogenetics and/or molecular techniques. 51 patients (37%) had a t(9;11), 19 patients (14%) a t(6;11), 8 patients (6%) a t(11;19), 6 patients (4%) a t(11;17) and 5 patients (4%) a t(10;11). 48 patients (35%) hold other fusion partners or deletions of 11q23. For further evaluation, patients harbouring other 11q23 aberrations than t(9;11) and t(6;11) were grouped together. Median age of all patients was 39 years (range 16–60). Overall complete remission rate was 75% with no significant difference between the groups (82% for t(9;11), 74% for t(6;11) and 70% for others). Altogether, 9% of the patients had treatment failure and 8% died during induction. Relapse-free survival (RFS) and overall survival (OS) at 5 years for the entire group was 34% and 28%, respectively. RFS and OS of the t(6;11) group was 0% and 11% respectively and thus was significantly inferior to patients with t(9;11) (RFS: 48%, OS: 33%) and patients with other 11q23 aberrations (RFS: 32%, OS: 28%). Within the t(9;11) group there was no difference in RFS or OS between patients who had a t(9;11) as a sole aberration (n=35) and patients with additional aberrations (n=16). Moreover, in t(9;11) no difference in RFS was observed between treatment with high dose araC, autologous or allogeneic stem cell transplantation as late consolidation. In conclusion, these data demonstrate that prognosis of patients with 11q23 is heterogeneous. Patients with t(9;11) have a relatively good outcome independent of the consolidation therapy used. In contrast, the prognosis of patients with t(6;11) is extremely poor. Therefore, patients with t(6;11) should be regarded as high risk and alternative treatment strategies for this subgroup are required.

Risk estimation in localized unresectable single copy MYCN neuroblastoma by the status of chromosomes 1p and 11q

In localized neuroblastoma, the identification of patients requiring intensive treatment is still difficult. We retrospectively analyzed data of 280 single copy MYCN stage 2 and 3 neuroblastoma patients with gross residual tumor after initial surgery. The 3-year-event free survival of the total group was 83±2%, and 3-year-overall survival was 92±2%. Patients ≤1.5 years had a better outcome than older children. Deletions/imbalances of chromosome 1p were found in 9/90 patients and were associated with a higher event rate but not with a higher death rate. Aberrations of chromosome 11q in 14/91 patients were correlated with a higher event and death rate. Multivariate analysis identified 1p aberrations as important for event free survival and 11q aberrations for overall survival.

The value of fluorescence in situ hybridization in the diagnosis and prognosis of chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) is the most common adult leukemia. Prognosis is related to clinical staging and cytogenetic findings. Conventional cytogenetic analysis of CLL reveals abnormalities in approximately one third of patients. Fluorescence in situ hybridization (FISH) is analytically more sensitive than conventional cytogenetics for specific chromosomal abnormalities. To evaluate the usefulness of FISH in CLL, a study of 100 CLL patients comparing conventional cytogenetics and a commercially available multiprobe FISH kit was undertaken. One hundred consecutive CLL patients (67 males, 33 females) were studied. The male–female patient ratio was ∼2.0 to 1. Twenty-eight percent (28/98) of patients had abnormal karyotypes by conventional cytogenetics (one patient had no specimen for conventional cytogenetics and one had an unanalyzable karyotype), and of those 19/100 (19%) had more than one chromosomal abnormality. Sixty-four percent (64/100) of the patients were positive for at least one abnormality by the FISH probes used. The following abnormalities were noted with FISH: 11q22 ATM, 23/100 (23%); trisomy 12, 11/100 (11%); 13q14.3, 40/100 (40%); 13q34.3, 4/100 (4%); 17p13.1, 12/100 (12%). Conventional karyotypes revealed 2 patients with abnormalities of chromosome 6 (which FISH did not address); 11 with abnormalities of 11 or 11q; 6 with trisomy 12; and 4 with abnormalities of 17. Aberrations of 11q and 17p are reported to have a poor prognosis in CLL. FISH can identify abnormalities missed with conventional cytogenetics and is helpful in diagnosis, prognosis, and evaluation of therapy for CLL. Additional chromosomal changes are identified with conventional cytogenetics that are not addressed by the multiprobe FISH kit.

Fluorescence In Situ Hybridization (FISH) Using a Subtelomeric 11q Probe as a New Diagnostic Tool for Congenital Thrombocytopenia Caused by Deletions in 11q.

Neonatal thrombocytopenia occurs in about 1% of all newborns. Inherited forms like 11q- or Jacobsen syndrome are rare. However, they may remain undetected with karyotyping because the deleted regions in 11q often involve small subtelomeric regions. Here we report on the detection of deletions in 11q in two newborns with normal routine karyotypes who were shown to carry subtelomeric deletions in 11q by means of fluorescence in situ hybridization (FISH) using a subtelomeric 11q probe (Abbott, Diagnostics, Wiesbaden, Germany). Both children showed thrombocytopenia (18.000/μl and 26.000/μl, respectively) and dysmegakaryopoiesis (absence of normal megakaryocytes and presence of micromegakaryocytes) associated with facial dysmorphism, cardiac defects and psychomotoric retardation. In the second case, the mother and the grandmother also showed mild thrombocytopenia. In both patients, FISH analyses on peripheral blood and bone marrow showed the loss of the telomere-associated region of 11q distal of the MLL gene. In the first patient, the deletion of 11q resulted from an unbalanced complex rearrangement with duplication of 11p. As the source of this chromosomal aberration, a paternal pericentric inversion of chromosome 11 was identified. The partial monosomy 11q and the partial trisomy 11p in the first patient were confirmed by comparative genomic hybridization (CGH) analysis. Array/matrix CGH assisted in determining the breakpoints at 11p15.1 and 11q24.1. No structural aberrations of 11q were found in the mother of the second patient, but further investigations are under way. These findings give further evidence that small subtelomeric deletions of 11q and probably mutations of genes located therein cause thrombocytopenia. Since it can be very difficult to detect these deletions by karyotyping, FISH using a subtelomeric 11q probe seems to be an extremely useful new diagnostic tool. This new method should be applied in children with congenital thrombocytopenia, in particular if they have additional complex dysmorphic features.

FISH analyses for alterations in chromosomes 1, 2, 3, and 11 define high-risk groups in neuroblastoma.

The prognostic chromosomal markers 1p loss and MYCN amplification (MNA) are only present in a subgroup of approximately 30% of neuroblastomas. To further characterize high and low risk subsets we investigated alterations in chromosome arms 3p and 11q, additional changes in 1p and MYCN as well as the somy-status of chromosome 1 in the same sample. Fluorescence in situ hybridization (FISH) was used as an alternative technique to PCR/LOH- or comparative genomic hybridization (CGH) analyses. Alterations in chromosomes 3p and 11q were investigated in 182 unselected tumors, 1p loss and MNA in 174 and 179 of these, respectively. The somy-status of chromosome 1 was determined in 165 tumors as it highly correlates with the tumor ploidy. Alterations in the four chromosomal regions were found in the following frequencies: 3p26: 19%, 11q23: 29%, 1p36: 29%, MNA: 19%. Fifty-two percent of all cases displayed structural aberrations in at least one chromosomal region, 83% in stage 4 and 30% in stages 1-3, 4s. All aberrations were thus correlated with stage 4 disease but were also present in a substantial subset of localized and 4s tumors. Trisomy of chromosome 1 was found in 38% of the tumors, disomy or tetrasomy in 62%. Patients with alterations in any of the four chromosomes and di/tetrasomy 1 showed a significantly increased age at diagnosis. Loss in 1p and MNA were closely associated with each other, as well as 3p and 11q aberrations but not the groups 1p/MNA versus 3p/11q. Only a small portion of trisomic tumors showed aberrations in at least one of the four chromosomal regions (14%) in contrast to the majority of the di/tetrasomic cases (74%). As already known the MYCN status discriminated between good and poor outcome in localized and metastatic stage 4 tumors. In addition alterations in 1p or 11q, deletion in 3p and di/tetrasomy 1 were associated with an unfavorable prognosis in MYCN single copy tumors of stages 1-3, 4s. Multivariate analysis revealed 11q alterations and MNA as the most important chromosomal prognostic factors in all stages. FISH analyses for chromosomal alterations in 3p and 11q as well as in 1p and MYCN allows to define different groups with an increased risk for disease progression.

High EVI1 expression predicts poor survival in acute myeloid leukemia: a study of 319 de novo AML patients

AbstractThe proto-oncogene EVI1 encodes a DNA binding protein and is located on chromosome 3q26. The gene is aberrantly expressed in acute myeloid leukemia (AML) patients carrying 3q26 abnormalities. Two mRNAs are transcribed from this locus: EVI1 and a fusion ofEVI1 with MDS1(MDS1-EVI1), a gene located 5′ ofEVI1. The purpose of this study was to investigate which of the 2 gene products is involved in transformation in human AML. To discriminate between EVI1 andMDS1-EVI1 transcripts, distinct real-time quantitative polymerase chain reaction (PCR) assays were developed. Patients with 3q26 abnormalities often showed highEVI1 and MDS1-EVI1 expression. In a cohort of 319 AML patients, 4 subgroups could be distinguished:EVI1+ andMDS1-EVI1− (6 patients; group I), EVI1+ andMDS1-EVI1+ (26 patients; group II),EVI1− andMDS1-EVI1+ (12 patients; group III), and EVI1− andMDS1-EVI1− (275 patients; group IV). The only 4 patients with a 3q26 aberration belonged to groups I and II. Interestingly, high EVI1 and notMDS1-EVI1 expression was associated with unfavorable karyotypes (eg, −7/7q−) or complex karyotypes. Moreover, a significant correlation was observed betweenEVI1 expression and 11q23 aberrations (mixed lineage leukemia [MLL] gene involvement). Patients from groups I and II had significantly shorter overall and event-free survival than patients in groups III and IV. Our data demonstrate that highEVI1 expression is an independent poor prognostic marker within the intermediate- risk karyotypic group.

Multiple myeloma with deletion of chromosome 13q is characterized by increased bone marrow neovascularization.

Anti-angiogenesis therapy with thalidomide has been reported to have marked activity in multiple myeloma (MM). As cytogenetics is an independent prognostic factor in MM, we analysed bone marrow (BM) angiogenesis and cytogenetic abnormalities in 34 patients with active MM. BM microvessel density (MVD), as determined by staining with anti-CD34, was significantly higher in MM (MVD: 221 +/- 94 per mm2) than in controls (80 +/- 36; P < 0.0001). In patients with the presence of at least one unfavourable cytogenetic abnormality (deletion of 13q14, deletion of 17p13, aberrations of 11q), a significantly increased BM MVD was observed (254 +/- 93 vs. 160 +/- 60 in patients with absence of these abnormalities; P = 0.0035). Further analyses indicated that increased BM MVD was significantly correlated with deletion of 13q14 (259 +/- 96 vs. 188 +/- 80; P = 0. 026), but not with other cytogenetic, clinical and laboratory MM parameters. We conclude that BM neovascularization is particularly high in MM with deletion of 13q14, which provides a rationale for use of anti-angiogenic strategies in the treatment of MM with high-risk cytogenetics.

Predictive role of interphase cytogenetics for survival of patients with multiple myeloma.

Recent metaphase cytogenetic studies suggested that specific chromosomal abnormalities are of prognostic significance in patients with multiple myeloma (MM). Because the true incidence of chromosomal abnormalities in MM is much higher than that detected by metaphase analysis, we used interphase fluorescence in situ hybridization (FISH) to determine the prognostic value of specific chromosomal aberrations. Bone marrow plasma cells from 89 previously untreated patients with MM were studied consecutively by FISH to detect the deletions of 13q14, 17p13, and 11q and the presence of t(11;14)(q13;q32). FISH results were analyzed in the context of clinical parameters (response to treatment and survival after conventional-dose chemotherapy), and a multivariate analysis of prognostic factors was performed. By FISH, the deletion of 13q14 occurred in 40 patients (44.9%), deletion of 17p13 in 22 (24.7%), and 11q abnormalities in 14 (15.7%; seven with t(11;14)). Deletions of 13q14 and 17p13 were associated with poor response to induction treatment (46.9% v 77.3% in those without deletions, P =.006 and 40.0% v 73.2%, P =.008, respectively) and short median overall survival (OS) time (24.2 v 88.1 months, P =. 008 and 16.2 v 51.3 months, P =.008, respectively). Short median OS time was also observed for patients with 11q abnormalities (13.1 v 41.6 months, P =.02). According to the number of unfavorable cytogenetic features (deletion of 13q14, deletion of 17p13, and aberrations of 11q) that were present in each patient (0 v 1 v 2 or 3), patients with significantly different OS times could be discriminated from one another (102.4 v 29.6 v 13.9 months, P <.001, respectively). For patients with MM who were treated with conventional-dose chemotherapy, interphase FISH for 13q14, 17p13, and 11q provides prognostically relevant information in addition to that provided by standard prognostic factors. This observation may be considered for risk-adapted stratifications of MM patients in future clinical trials.

Granulocytic sarcomas in body cavities in childhood acute myeloid leukemias with 11q23/MLL rearrangements

Three childhood acute monoblastic leukemias (AML M5) with granulocytic sarcomas (GSs) are described. All displayed 11q23/MLL abnormalities, t(9;11)(p22;q23) in two cases and t(11;17)(q23;q21) in one case, constituting around 20% of all 11q23-positive AML cytogenetically investigated in our department. Two of the patients had GS in multiple locations, and all three had abdominal GS. In two of them, t(9;11)-positive GS was diagnosed prior to the diagnosis of AML. Fourteen (1.9%) of 752 published AML cases with 11q23 aberrations have had GS, either as a presenting feature or during disease progression. The incidence of GS has varied significantly (P < 0.05) between children (3.8%) and adults (0.8%). The most common AML subtype has been AML M5 (∼ 75%) and the most frequent GS sites have been the skin, abdomen, orbit, and thorax. Considering the possibility of underreporting of GS in published cases and the relatively high frequency in our own series, we believe that 11q23/MLL rearrangements may predispose to GS development. Although extramedullary infiltrates in the skin are known to be frequent in cases of AML M5, which is often associated with 11q23 aberrations, the present findings indicate that GS in the abdomen, orbit, and thorax may also be common, especially in pediatric AML. Thus, the possibility of 11q23/MLL-positive GS should be suspected when tumors of uncertain derivation occur in these sites. Finally, the identification of 11q23/MLL abnormalities in GSs in two patients without overt AML underscores the importance of using cytogenetic and molecular genetic investigations as a diagnostic approach in the evaluation of tumorous lesions of unknown origin. Genes Chromosomes Cancer 27:136–142, 2000. © 2000 Wiley-Liss, Inc.

Granulocytic sarcomas in body cavities in childhood acute myeloid leukemias with 11q23/MLL rearrangements

Three childhood acute monoblastic leukemias (AML M5) with granulocytic sarcomas (GSs) are described. All displayed 11q23/MLL abnormalities, t(9;11)(p22;q23) in two cases and t(11;17)(q23;q21) in one case, constituting around 20% of all 11q23-positive AML cytogenetically investigated in our department. Two of the patients had GS in multiple locations, and all three had abdominal GS. In two of them, t(9;11)-positive GS was diagnosed prior to the diagnosis of AML. Fourteen (1.9%) of 752 published AML cases with 11q23 aberrations have had GS, either as a presenting feature or during disease progression. The incidence of GS has varied significantly (P < 0.05) between children (3.8%) and adults (0.8%). The most common AML subtype has been AML M5 ( approximately 75%) and the most frequent GS sites have been the skin, abdomen, orbit, and thorax. Considering the possibility of underreporting of GS in published cases and the relatively high frequency in our own series, we believe that 11q23/MLL rearrangements may predispose to GS development. Although extramedullary infiltrates in the skin are known to be frequent in cases of AML M5, which is often associated with 11q23 aberrations, the present findings indicate that GS in the abdomen, orbit, and thorax may also be common, especially in pediatric AML. Thus, the possibility of 11q23/MLL-positive GS should be suspected when tumors of uncertain derivation occur in these sites. Finally, the identification of 11q23/MLL abnormalities in GSs in two patients without overt AML underscores the importance of using cytogenetic and molecular genetic investigations as a diagnostic approach in the evaluation of tumorous lesions of unknown origin. Genes Chromosomes Cancer 27:136-142, 2000.

Rapid and sensitive detection of all types of MLL gene translocations with a single FISH probe set.

The MLL gene on chromosome 11 band q23 is frequently involved in chromosome translocations in acute lymphoblastic leukemia and acute myeloid leukemia. The translocation results in the formation of a fusion gene on the derivative 11 chromosome consisting of the 5' part of the MLL gene and the 3' part of another gene; already more than 30 different partner chromosome regions have been described. MLL gene rearrangements are generally correlated with a poor prognosis. Therefore the presence of an 11q23 aberration has direct implications for treatment stratification, making early and rapid detection of utmost importance. In this study, we developed a FISH probe set for detection of MLL gene rearrangements according to strict design criteria. The cosmid probes are derived from the flanking regions of the MLL breakpoint region on chromosome 11 and when used in dual colored FISH experiments give rise to a split of the normally colocalizing (fused) signals in case of a translocation. This split signal was observed in seven out of 10 cases with an 11q23 translocation with various partner chromosomes. In the three other cases, a deletion of the 3' part of the MLL gene, downstream of the breakpoint region was also found. A low false positive value of only 1.7% was obtained for interphase cells in contrast to conventional dual colored FISH where the creation of a fusion signal has cut off values of at least 5-10%. A major advantage of our type of probe set is the application of a single FISH experiment to detect all types of MLL translocations. Moreover, since this cosmid probe set can be used for either interphase or metaphase studies, metaphases are no longer a prerequisite for detecting the presence of an 11q23 translocation. Nevertheless, metaphase FISH with the new probe set is helpful in determining the partner chromosome and therefore may lead to the identification of new partner genes.

Molecular rearrangement of the MLL gene in adult acute myeloid leukemia without cytogenetic evidence of 11q23 aberration

Translocations involving chromosome band 11q23 are frequently found in infant acute leukemia and involve rearrangement of the MLL gene. In this study, 29 cases of adult acute myeloid leukemia (AML) were analyzed to determine the frequency of MLL gene rearrangement. Of these, 19 cases were karyotyped and none showed cytogenetic evidence of 11q23 aberration. MLL rearrangements were demonstrable in four cases, giving a frequency of 14% ( 4 29 ). Only one of the four cases with MLL rearrangement showed features typical of leukemia with 11q23 aberration; other cases were indistinguishable from those without MLL rearrangement. There was no apparent difference in presentation blast count, remission, and survival duration when cases with or without MLL rearrangement were compared. Clinicopathologic features of adult AML with MLL rearrangements may be heterogenous.

Acute leukemia with t(10;11)(p11-p15;q13-q23)

We report six patients with acute leukemia characterized by the presence of a t(10;11) (p11-p15;q13-q23), either as sole cytogenetic abnormality (three patients) or as part of a complex abnormal karyotype. The morphologic and cytochemical features of four patients were consistent with FAB-M5A, while two patients presented with FAB-L1 characteristics. By immunophenotyping, myeloid leukemia was diagnosed in five patients, including one patient with FAB-L1 leukemia who typed as terminal transferase (TdT) +, CD7 T-cell antigen + acute myelomonocytic leukemia. Differentiated acute myeloid leukemia (AML) with expression of terminal transferase was found in two of the other cases and monocytic leukemia in two, with co-expression of T-cell antigens in one of them. The second FAB-L1 patient typed as undifferentiated acute lymphocytic leukemia (ALL) expressing myeloid antigens. Serial phenotypic studies in patient 3 during the course of the disease demonstrated a switch from monocytic to lymphoid morphology at the time of first and second relapse, which was paralleled by the appearance of a pre-T ALL immunophenotype with co-expression of the myeloid antigen CD33. These phenotypic changes occurred without apparent alteration in the genotype since t(10;11)(p11.2;q23) remained the only cytogenetic aberration at all stages of the disease. Our observations suggest that the (10;11) variant of 11q aberrations occurs in a bipotential myelomonocytic/T-lymphoid stem cell.

Rearrangement of the MLL gene in acute lymphoblastic and acute myeloid leukemias with 11q23 chromosomal translocations.

Translocations involving chromosome band 11q23 are very frequent in both acute lymphoblastic and acute myeloid leukemias and are the most common genetic alteration in infants with leukemia. In all age groups and all phenotypes of leukemia, an 11q23 translocation carries a poor prognosis. A major question has been whether one or several genes on band 11q23 are implicated in these leukemias. Previously, we identified the chromosomal breakpoint region in leukemias with the common 11q23 translocations and subsequently cloned a gene named MLL that spans the 11q23 breakpoint. We isolated a 0.74-kb BamHI fragment from a complementary DAN (cDNA) clone of the MLL gene. To determine the incidence of MLL rearrangements in patients with 11q23 abnormalities, we analyzed DNA from 61 patients with acute leukemia, 3 cell lines derived from such patients, and 20 patients with non-Hodgkin's lymphoma and 11q23 aberrations. The 0.74-kb cDNA probe detected DNA rearrangements in the MLL gene in 58 of the patients with leukemia, in the 3 cell lines, and in 3 of the patients with lymphoma. All the breaks occurred in an 8.3-kb breakpoint cluster region within the MLL gene. The probe identified DNA rearrangements in all 48 patients with the five common 11q23 translocations involving chromosomes 4, 6, 9, and 19, as well as in 16 patients with uncommon 11q23 aberrations. Twenty-one different chromosomal breakpoints involving the MLL gene were detected. MLL gene rearrangements were detected with a single probe and a single restriction-enzyme digest in all DNA samples from patients with the common 11q23 translocations as well as in 16 patients or cell lines with other 11q23 anomalies. The ability to detect an MLL gene rearrangement rapidly and reliably, especially in patients with limited material for cytogenetic analysis, should make it possible to identify patients who have a poor prognosis and therefore require aggressive chemotherapy or marrow transplantation.