Abstract 2823
Poster Board II-799
AL amyloidosis (AL) is characterized by the deposition of amyloid fibrils in diverse tissues due to an underlying monoclonal plasma cell dyscrasia. In a previous ...study (Bochtler et al, Blood 2008) we have demonstrated that in AL cytogenetic aberrations were detectable in about 90% of patients (pts). Translocation t(11;14) proved to be the most frequent aberration in AL found in 45% of the pts. In this study we evaluated whether the concept of hyperdiploidy and non-hyperdiploidy as major pathogenetic pathways in monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM) is also applicable to AL. Our study was based on the largest patient group tested for cytogenetics in AL thus far including 184 pts with AL - among them 21 pts with concomitant MM I. They were assessed for their ploidy status by interphase fluorescence in situ hybridization (FISH). 179 MGUS and MM I pts not requiring therapy served as controls. We used a well established score (Wuilleme et al, Leukemia 2005), which requires extra copies for at least two out of the three probes 5p15/5q35, 9q34 and 15q22 as criterion for hyperdiploidy. The hyperdiploidy frequency was very low in AL with 14% as compared to 32% in MGUS / MM I (p<0.001). Among AL pts those with a concomitant MM I displayed a higher hyperdiploidy frequency than those without (43% versus 10%, p<0.001) suggesting that chromosomal gains reflect progression of the monoclonal plasma cell clone.
Addressing hyperdiploidy probes in detail, we could show that both in the 184 pts. with AL and 179 pts. with MGUS / MM I gains of 11q23, 17p13 and 19q13.3 closely clustered with the three hyperdiploidy defining probes 5p15/5q35, 9q34 and 15q22 (p'0.01 for all probes after adjusting for multiple testing). However, gain of 11q23 was also frequently detected in association with t(11;14). The group with gain of 11q23 subdivides into a t(11;14) positive and a hyperdiploidy positive subgroup in both the AL (p<0.001) and the MGUS / MM I (p<0.001) entities. As revealed by additional probes for 11p15 and 11cen, gain of 11q23 in hyperdiploid pts reflected a gain of the whole chromosome 11 in all tested pts (10 AL and 31 MGUS / MM I). On the contrary, gain of 11q23 in t(11;14) positive pts was merely due to the translocation involving chromosome 11 (with 25 out of 26 AL and 5 out of 7 MGUS / MM I pts displaying a normal diploid status for 11p15 and 11cen). Therefore, gain of 11q23 is a poor indicator of hyperdiploidy in AL, where t(11;14) frequencies are particularly high and hyperdiploidy frequencies are particularly low.
Addressing the cytogenetic clustering of hyperdiploidy with other cytogenetic aberrations we observed a strong inverse association of hyperdiploidy with t(11;14) in both AL and MGUS / MM I (p<0.001 in both entities). Accordingly, both aberrations were allocated to branches separating from each other already at the root in the oncogenetic tree model (see figure 1). Del13q14/t(4;14) and IgH translocations with an unknown partner also separated as distinct branches early from the root. These similar clustering patterns of both AL and MGUS / MM I with 4 major cytogenetic groups suggests common pathogenetic mechanisms in both entities despite their differing hyperdiploidy and t(11;14) frequencies.
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No relevant conflicts of interest to declare.
Abstract 332
PURPOSE: In Multiple Myeloma (MM), the combination of serum beta-2-microglobulin level with serum albumin concentration has been proposed as an outcome predictor in the International ...Staging System (ISS). More recently, subgroups of MM defined by genetic and cytogenetic abnormalities have been associated with unique biologic, clinical, and prognostic features.
PATIENTS AND METHODS: We analyzed the prognostic value of 12 chromosomal abnormalities by fluorescent in situ hybridization (FISH) in a series of 354 MM patients treated within the HOVON-65/GMMG-HD4 trial. Patients with newly diagnosed MM were randomized to receive either three cycles of VAD (arm A; vincristine, adriamycin, dexamethasone) or PAD (arm B; bortezomib, adriamycin, dexamethasone). All patients underwent autologous stem cell transplantation (ASCT) followed by maintenance therapy with thalidomide 50 mg daily (arm A) or bortezomib 1.3 mg/m2 once every 2 weeks (arm B), respectively. In addition, a second cohort of patients was analyzed as a control group (n=462), undergoing ASCT at the University of Heidelberg between September 1994 and December 2010.
RESULTS: For the entire group of patients treated within the HOVON-65/GMMG-HD4 trial, we identified 233 patients with 2 copies (67.7%), 95 patients with 3 copies (27.6%) and 16 patients (4.7%) with more than three copies of the chromosomal region 1q21. In addition to del(17p13) and t(4;14), we added +1q21 (>3 copies) to the group of high-risk aberrations, since the outcome of these patients was almost as poor as it was observed for patients with del(17p13). Subsequently, we analyzed whether combining the ISS score with information on the presence of high-risk aberrations could improve the prognostic value with regard to patients’ outcome. A combination of the presence or absence of del(17p13), t(4;14), or +1q21 (>3 copies) with the ISS score allowed patients to be stratified into three distinct groups: low-risk absence of del(17p13)/t(4;14)/+1q21 (>3 copies) and ISS I, high-risk presence of del(17p13)/t(4;14)/+1q21 (>3 copies) and ISS II/III, and intermediate-risk (all remaining patients). Most of the patients belonged to the low- (33%) and intermediate-risk (49%) groups, whereas 18% were allocated to the high-risk group. The median PFS times for the low-, intermediate-, and high-risk groups were 41.9 months, 31.1 months (HR=1.7; p=0.0018) and 18.7 months (HR=3.6; p<0.0001), respectively. The 3yr-overall survival (OS) decreased from 94% in the low-risk group to 80% (HR=4.6; p=0.0001) and 43% (HR=12.8; p<0.0001) in the intermediate- and high-risk groups, respectively.
These results were confirmed in the independent cohort of patients: From date of first ASCT, the median PFS times for the low-, intermediate-, and high-risk groups were 43.3 months, 23.0 months (HR=1.5; p=0.015) and 13.8 months (HR=2.4; p=0.0003), respectively. The 4yr-OS decreased from 84% in the low-risk group to 71% (HR=2.1; p=0.0043) and 49% (HR=3.84; p<0.0001) in the intermediate- and high-risk groups, respectively.
CONCLUSION: In our series, the ISS/FISH-based score/algorithm predicted PFS and OS much better than the ISS alone. Our results with molecular cytogenetic techniques may already have implications for the risk-adapted clinical management of patients with MM particularly in younger patients. Display omitted
van de Velde:Ortho Biotech Oncology Research & Development: Employment. Sonneveld:Celgene: Membership on an entity’s Board of Directors or advisory committees, Research Funding; Janssen-Cilag: Membership on an entity’s Board of Directors or advisory committees, Research Funding; Onyx: Membership on an entity’s Board of Directors or advisory committees, Research Funding; Millennium: Membership on an entity’s Board of Directors or advisory committees.
Multiple Myeloma (MM) is a malignant lymphoproliferative B-cell disease characterized by the accumulation of monoclonal plasma cells in the bone marrow. Acquired genomic aberrations have been shown ...to significantly impact response to chemotherapy and survival in MM. The aim of our study was to assess the clinical relevance of genomic abnormalities in 306 MM patients treated with high-dose chemotherapy (HDCT) and peripheral stem cell transplantation (PBSCT) in our center.
We analyzed 171 males and 135 females with a median age of 60 years (range 25 – 73 years). According to the international staging system (ISS), MM patients were classified as stage I (46.6%), stage II (36.1%) and stage III (17.4%) at the onset of chemotherapy. All patients underwent frontline HDCT with 200 melphalan mg/m2 and PBSCT according or in analogy to the GMMG-HD3- or GMMG-HD4-trials. Interphase-FISH-analysis was performed on CD138-purified plasma cells using probes for chromosomes 1q21, 6q21, 8p21, 9q34, 11q23, 13q14.3, 15q22, 17p13, 19q13, and 22q11, as well as IgH-translocations t(4;14)(p16.3;q32.3) and t(11;14)(q13;q32.3). For the entire group, the median overall survival (OS) and progression-free survival (PFS) after HDCT was 6.4 and 2.2 years, respectively.
Table 1. Chromosomal abnormalities with significant results (a-level=0.05) on PFS or OS (univariate analysis, unadjusted p-values)
Aberration yesvs.noFrequency %3-year PFS %P value3-year OS %P valuedel(8p21)1926 vs. 370.0158 vs. 780.02del(13q14.3)4623 vs. 53<0.00165 vs. 830.03del(17p13)1022 vs. 400.0244 vs. 82<0.001t(4;14)1410 vs. 42<0.00154 vs. 810.04+1q213520 vs. 46<0.00167 vs. 850.002+19q135449 vs. 220.0376 vs. 730.92
In a first step, we analyzed the prognostic impact of each individual chromosomal aberration on PFS and OS (Table 1). After adjustment for the ISS-score, del(8p21), del(13q14.3), del(17p13), t(4;14) as well as gains of 1q21 and 19q13 preserved significant impact on PFS, while del(17p13), t(4;14) and gain of 1q21 were of statistical significance for OS, indicating that these chromosomal aberrations give prognostic information in addition to the ISS-score. Subsequently, we performed a multivariate analysis including all the chromosomal aberrations analyzed. While del(17p13) and gain of 1q21 showed significant results on OS, del(13q14.3), del(22q11) and gain of 15q22 were significant for PFS.
In conclusion, our results show that the heterogeneity seen in the clinical course of MM patients after HDCT can be correlated with distinct chromosomal aberrations. This analysis may have implications for the risk-adopted management of patients with MM.
The aim of the present study was to investigate whether dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) allows visualization of a change in microcirculation between healthy controls on ...the one side and early/advanced stages of plasma cell disease on the other. We examined a group of 222 individuals consisting of 60 patients with monoclonal gammopathy of undetermined significance (MGUS), 65 patients with asymptomatic multiple myeloma (aMM), 75 patients with newly diagnosed symptomatic MM (sMM) and 22 healthy controls with DCE-MRI of the lumbar spine. A continuous increase in the microcirculation parameter amplitude A reflecting vascular volume was detected from normal controls over MGUS and asymptomatic to symptomatic MM. Significant differences were found between controls and aMM (p = 0.03), controls and sMM (p = 0.001) and between asymptomatic and symptomatic MM (p = 0.02) respectively. While diffuse microcirculation patterns were found in healthy controls as well as MGUS and MM, a pattern with focal hot spots was exclusively detected in 42.6 % of sMM and in 3 patients with MGUS and 3 patients with aMM. Patients with MGUS and aMM with increased microcirculation patterns showed a significantly higher bone marrow plasmocytosis compared to patients with a low microcirculation pattern. Our investigations substantiate by means of a non invasive assessment of bone marrow microcirculation the concept of an angiogenic switch from early plasma cell disorders to symptomatic MM. Pathological DCE-MRI findings can be identified and correlate with an adverse prognostic parameter.
The introduction of rituximab into the treatment of malignant lymphomas of the B-cell lineage has had a major impact on the management of these diseases. In diffuse large B-cell lymphomas (DLBCLs) ...and follicular lymphomas (FLs) several multicenter prospective randomized trials consistently demonstrated an improved outcome when rituximab was added to chemotherapy. In addition, prolonged exposure to rituximab as maintenance therapy has been benefical in patients with FL and mantle cell lymphoma (MCL). For patients, the effect of any prolonged antitumor therapy on the quality of life (QoL) is a very important question. However, so far the question whether rituximab maintenance therapy may impair QoL in patients with Non-Hodgkins-lymphoma remains unanswered. To investigate this subject, we have performed a prospective randomized trial of rituximab maintenance therapy (8 cycles rituximab 375 mg/m2 every 3 months) versus observation in patients with CD20+ B-cell Non-Hodgkins-Lymphoma in our institution. Between July 2002 and December 2005, 106 patients (pts) were included into the trial. QoL was assessed with the standardized questionnaires EORTC-QLQ-C30 and EuroQol-5D. After statistical analysis with the Wilcoxon signed-rank test, we found no significant differences of the QoL between the rituximab treatment group and the observation group. We conclude that rituximab maintenance therapy is safe and does not impair quality of life in this patient population.
Introduction: Cytogenetic aberrations (CA) have emerged as important pathogenetic and prognostic factors in plasma cell disorders. However, in AL amyloidosis (AL) only a few reports with small ...numbers of patients have been published.
Methods: Using interphase FISH analysis in CD138+ cells, we evaluated the role of CA in a series of 85 AL patients as compared to 146 patients with a monoclonal gammopathy without treatment requirement in a prospective manner. Our panel included IgH translocations t(11;14), t(4;14), t(14;16) and translocation of 14q32 with an unidentified partner, gains of 1q21, 11q23 and 19q13 as well as deletions of 8p21, 13q14 and 17p13. Using these probes we could detect at least one of these aberrations in 95% of AL and in 88% of the control group. Age, gender and plasma cell content were statistically equally distributed among both groups.
Results: The most frequent aberration in AL was t(11;14), which was detected in 45% of AL patients as compared to 26% of the control group (p=0,056). It was strongly associated with the lack of an intact immunoglobulin (p<0,001), thus accounting for the frequent light chain only subtype in AL. Markedly, t(11;14) was more frequently found in combination with gain 11q23 in AL than in the control group (20% versus 6%, p = 0.005). Other frequent aberrations in AL included deletion 13q14 (32%) and gain 1q21 (21%), which were observed in the control group at comparable frequencies (34% and 20%). The overlapping character of the underlying plasma cell disorder in both disease entities was also emphasized by the similarities of branching patterns of the five major CA in cluster analysis applied in 169 patients (figure 1). The relation of clinical parameters and chromosomal aberrations was also evaluated. The analyzed CA had no impact on the organ involvement pattern in AL patients.
Conclusions: We observed a high frequency for t(11;14) in AL. Apart from this finding, the cytogenetic patterns known in monoclonal gammopathy of unknown significance and multiple myeloma were widely shared by AL amyloidosis.
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Introduction: Dynamic contrast enhanced MRI (dceMRI) is an imaging technique detecting changes in local microcirculation reflecting increased angiogenesis. The dceMRI parameters Amplitude A and ...exchange rate constant kep have been shown to be significantly increased in patients with active multiple myeloma (MM) compared to healthy controls and to correlate with osteolytic bone involvement and prognosis. We now compared the parameters and infiltration patterns of dceMRI in patients with monoclonal gammopathy of undetermined significance (MGUS) as well as in patients with asymptomatic MM not requiring chemotherapy (NRC-group) with those in patients with symptomatic MM requiring chemotherapy according to international standards.
Methods: The NRC group contained 71 patients: 29 patients with MGUS, 39 patients with MM stage I and 3 patients with MM stage IIA according to Durie and Salmon. 24 patients had a diagnosis of a MM in stage II in progression (n = 3) or stage III (n = 21). All patients underwent standardized dceMRI with high temporal resolution (T1w-turboFLASH) of the lumbar spine before start of therapy. Color coded pharmacokinetic maps of imaged area were classified according to three distinct patterns of microcirculation: “normal” (as in healthy controls), “diffuse” or “focal”. The contrast uptake was quantified using a two compartment model with the output parameters Amplitude A and distribution constant rate kep reflecting bone marrow microcirculation.
Results: 63 % of patients in the NRC group were found to have changes in the microcirculation pattern with 26 patients (37%) displaying a normal, 43 (60%) a diffuse and 2 (3%) a focal pattern. Within the NRC group 11 MGUS patients (38%) were found to have a normal pattern, 17 patients (59%) had a diffuse and 1 patient (3%) presented with a focal pattern. 79 % of patients with symptomatic MM had an abnormal microcirculation pattern with 5 (21%) MM patients showing a normal, 12 (50%) a diffuse and 7 (29%) a focal pattern. Statistical comparison did not reveal a significant difference in the total incidence between NRC and symptomatic MM group.
Comparison of quantitative microcirculation parameters did not show a significant difference of Amplitude A (p=0.87) and exchange rate constant kep (p=0.3) in MGUS patients compared to patients with asymptomatic MM. Comparing the NRC group with the symptomatic myeloma group revealed a significant higher Amplitude A in the symptomatic MM group (p=0.01). There was no significant difference in exchange rate constant kep.
Conclusion: Our investigations revealed a group of patients with asymptomatic myeloma and MGUS that display significant increase in bone marrow microcirculation. Our findings could be the basis for stratified treatment of patients with novel therapeutics targeting the vascular system. Prognostic implications for systemic and local development of the malignant disease are topic of current investigations.
BACKGROUND. Angiogenesis is a hallmark of active multiple myeloma and a therapeutic target, e.g. by thalidomide. Others and we have shown this to be due to a differential and novo expression of ...pro/antiangiogenic genes in MM as well as an effect of increased plasma cell number. AIM of this study was to investigate based on the expression of (anti)angiogenic genes alone (consensus list determined by medline search),i.whether for respective samples being PPC, BMPC or MMC can be predicted,ii.what expression differences between these entities exist,iii.if differences between the expression pattern of MMC from early stage vs. therapy requiring MM can be used for prediction, andiv.whether the survival of MM–patients undergoing HDT is different in two groups distinguished by their angiogenic signature.
PATIENTS AND METHODS. 187 newly diagnosed MM/MGUS–patients (65 training (TG) / 122 independent validation group (VG)), 14 normal donors (ND) and 12 in vitro generates PPC samples were included. Bone marrow aspirates were CD138-purified by activated magnetic cell sorting. RNA was in-vitro transcribed and hybridised to Affymetrix HG U133 A+B GeneChip (TG) and HG U133 2.0 plus arrays (VG). Expression data were gcrma-normalised and the empirical Bayes algorithm used. P-Values were adjusted using the Benjamini-Hochberg method (Bioconductor). The PANP-algorithm was used to identify expressed genes. iFISH was performed on purified MM-cells using probesets for chromosomes 1q21, 9q34, 11q23, 11q13, 13q14, 15q22, 17p13, 19q13, 22q11 and the translocations t(4;14) and t(11;14). Selected expression data were verified by real time RT-PCR and western blotting.
RESULTS.
i.On TG and VG, being BMPC or PPC is predicted by a 17 gene predictor (31 probe sets) without error. Being MMC is predicted with an error rate of 6.9/4.7% with an overall-error rate of 5.8/4.3% in TG/VG, respectively.ii.5 proangiogenic genes/7 probe sets (HGF hepatocyte growth factor, ADM adrenomedullin, CXCL2 chemokine (C-X-C-motif)-ligand 2, IGF1 insulin-like growth factor 2, Met HGF-receptor) and 1 antiangiogenic/1 probe set (SerpinF1, Serpin peptidase inhibitor F1) are differentially expressed between these entities concordantly in TG and VG.iii.Attribution of MMC to early or therapy requiring can not be predicted. Most patients with MGUS and MM display 1 or several over expressed angiogenic gene. Survival analysis (EFS/OAS) will now be performed based on these findings.
CONCLUSION. Our analysis reveals novel targets for antiangiogenic therapy of multiple myeloma as anti-HGF. There is evidence that angiogenesis related genes are activated early during disease progression as rationale for early intervention using antiangiogenic compounds.
BACKGROUND. Angiogenesis is a hallmark of active multiple myeloma. However, two etiologic hypotheses have been proposed:
an angiogenic switch (i.e. differential or de novo expression of ...pro/antiangiogenic genes in MM), and, alternativelyan effect of increased plasma cell number.
AIM of this study was to investigate the angiogenic signature of multiple myeloma cells (MMC), normal bone marrow plasma cells (BMPC), the bone marrow microenvironment (BMME) and cellular subfractions therein.
PATIENTS AND METHODS. 128 newly diagnosed MM-patients (65 training (TG) / 63 independent validation group (VG)) and 14 normal donors (ND) were included. Bone marrow aspirates were CD138-purified by activated magnetic cell sorting. Whole bone marrow (n=49) and FACSAria sorted subfractions thereof (n=5) were investigated. RNA was in-vitro transcribed and hybridised to Affymetrix HG U133 A+B GeneChip (TG) and HG U133 2.0 plus arrays (VG). Expression data were gcrma-normalised and the empirical Bayes algorithm used. p-Values were adjusted using the Benjamini-Hochberg method (Bioconductor). iFISH was performed on purified MM-cells using probesets for chromosomes 1q21, 9q34, 11q23, 11q13, 13q14, 15q22, 17p13, 19q13, 22q11 and the translocations t(4;14) and t(11;14). HGF expression was verified by real time RT-PCR and western blotting. Based on Medline review, we established a list of 89 pro- and 56 antiangiogenic genes and investigated their expression according to the stage of disease: BMPC vs. MGUS, SD stage I (asymptomatic myeloma) vs. SD stage II/III (symptomatic myeloma requiring therapy).
RESULTS.
BMPC express pro- (e.g. VEGFA) and antiangiogenic genes (e.g. TIMP2).Only one pro-angiogenic gene (hepatocyte growth factor, HGF) is significantly overexpressed in MMC compared to BMPC. HGF has previously been linked with myeloma progression and induction of angiogenesis.Six antiangiogenic genes (TIMP2, SERPINF1, COL18A1, PF4, THBS1, CXCL14) are downregulated in MMC compared with BMPC.Compared to healthy donors, the BMME of MM shows a significant downregulation of PLAU (urokinase, antiangiogenic) and upregulation of TNF(proangiogenic).
CONCLUSION. Upregulation of HGF-expression, downregulation of TIMP2, SERPINF1, COLA18A1, PF4, THBS1 and CXCL14 expression in MMC as well as downregulation of PLAU and upregulation of TNFα in the BMME seem to indicate an “angiogenic switch”. However, given the relatively low number of differentially expressed genes (7/145) and the expression of angiogenic genes by BMPC, an effect caused by an increasing number of plasma cells might be evenly important.
