Molecular Cytogenetic Abnormalities Research Articles

Does Double Mean Trouble? Coexistence of Myeloproliferative and Lymphoproliferative Neoplasms.

Background: The occurrence of myeloproliferative neoplasms (MPNs) that evolve into each other is well-described, as is this occurrence of lymphoproliferative neoplasms (LPNs). However, less is known about rare MPN/LPN coexistence, and the aim of our study was to analyze charachteristics of these patients after long term follow-up. Methods: Fourteen patients with MPN/LPN coexistence were diagnosed and treated according to guidelines at a single university center across two decades. Results: The overall median age was 53 years (22-69). MPNs patients with subsequent LPNs had a shorter period of second malignancy development and a more aggressive course of LPN, which can cause fatal outcomes. Polycythemia vera and chronic lymphocytic leukemia were most commonly associated (36%). The JAK2V617F mutation had 2/3 and cytogenetic abnormalities occurred in 1/3 of patients. MPN/LPN coexistence cases had significantly higher thrombotic potential (42.8%) and a higher third malignancy accruement frequency (21.4%) versus those without such malignancies. Conclusions: Considering the younger ages at MPN diagnosis, it is recommended to check regularly for blood lymphocytosis or lymphadenopathy occurrences and organomegaly progression faster than expected for MPN, with the aim of timely LPN diagnoses. The presence of molecular-cytogenetic abnormalities in a majority of patients indicate possible genetic instability and increased risk of development of multiple neoplasms, thus elevating thrombotic risk.

Survival Outcomes Associated with Molecular and Cytogenetic Alterations in AML Patients Treated with Hypomethylating Agent and Venetoclax

Molecular profiling has improved prognostication and helped identify novel therapeutic targets in AML. Nonetheless, it has also created a complex web of molecular profiles and cytogenetic abnormalities and in many cases it remains unclear which specific alterations primarily drive leukemogenesis. We sought to review our patients (pts) treated with a hypomethylating agent (HMA) + Venetoclax (Ven), to evaluate molecular and cytogenetic profiles with their associated outcomes. We identified 142 pts with AML treated with HMA + Ven at Northwell Health between January 2019 and December 2023. Through retrospective chart review, we obtained patient and disease characteristics, treatment regimens, clinical outcomes as well as molecular and cytogenetic profiling (using commercial AML panels) prior to the initiation of HMA + Ven. Bone marrow biopsy pathology reports were reviewed to identify the subtype of AML and grouped into De novo AML, AML with MDS related changes (AML-MRC) and therapy-related AML (t-AML). Median relapse free survival (mRFS) and median overall survival (mOS) in months (m) as defined by ELN 2022 (Dohner et al, Blood 2022), were estimated with Kaplan-Meier curves and compared utilizing log-rank testing. T-test was used to evaluate significant differences between categorical data. Of the 142 pts identified, the median age was 77. The majority of patients had adverse (n = 99) or intermediate risk (n = 31) genetics as defined by the 2022 ELN genetic risk classification. The overall response rate (ORR) was 42% with mOS of 9.6m. AML-MRC patients (n=80) had mOS of 9.1m and mRFS of 11.6m with 61 pts receiving HMA + Ven as frontline treatment. 47 pts with De novo AML had mOS of 13.4m and mRFS of 16.1m with 36 pts receiving HMA + Ven as frontline therapy. Only 15 pts had t-AML with mRFS of 12.3m and mOS of 8.8m with 10 pts receiving HMA + Ven as frontline therapy. There was no significant difference in mRFS or mOS between these groups, although there was a trend favoring De novo AML. 47 different molecular alterations were identified among the 142 patients. Figure 1 illustrates outcomes associated with the 16 most prevalent molecular alterations found in our AML population. Mutational burden was similar between AML types (2.7 vs 3.0 vs 2.4 mutations per pt). De novo AML was associated with an increased mutation burden in signaling molecules (60%) and transcription factors (47%) compared to AML-MRC (29%, 31%). AML-MRC had a higher occurrence of mutations in epigenetic regulatory genes (79%) and splicing factors (31%) compared to De novo AML (55%, 19%). Mutations in splicing factors were associated with a significant increase in both mRFS and mOS(p = 0.01) compared to pts without mutations. Pts with DNMT3A, NPM1 or SRSF2 mutations had the highest ORR to HMA + Ven (63%, 64% and 63% respectively). RUNX1, TP53, complex cytogenetics and del 7q were associated with worse ORR (46%, 36%, 46% and 42% respectively) as well as decreased mOS. SRSF2, mutated in both AML-MRC and De novo AML, not only had an elevated ORR, but improved mRFS and mOS. Contrarily, mRFS and mOS were not significantly improved despite the high ORR in DMT3A mutated pts. DNMT3A was mutated most often in AML-MRC and was associated with a higher rate of other epigenetic regulatory gene co-mutations. The same is true for patients with KMT2A mutations (ORR 50%, mOS 2.8m, mRFS 6.1m) which suggests ineffectiveness of HMA+ Ven in maintaining remissions with this alteration with possible early selection for resistant clones. RUNX1 mutations were associated with an improved mRFS (16.1m) despite lower mOS (8.8m). NPM1 mutations had the highest mRFS of 34.4m. The survival advantage may be secondary to the absence of adverse cytogenetics in 88% of these pts. IDH2 mutations was associated with a significantly improved mOS compared to unmutated IDH2; however this may be confounded by the use of an IDH2 inhibitor at relapse. Our review of molecular alterations in AML pts identified several molecular alterations such as SRSF2, DNMT3A, RUNX1 which we found to be associated with favorable responses when treated with HMA + Ven. In addition, we demonstrated that NPM1 mutation remains favorable in patients treated with HMA + Ven. Our findings help contribute to the ever-growing, complex molecular and genomic profiling of AML patients.

P-448 Impact of molecular cytogenetic abnormalities on the risk of disease progression in solitary bone plasmacytomas

P-448 Impact of molecular cytogenetic abnormalities on the risk of disease progression in solitary bone plasmacytomas

Study of Molecular Cytogenetic Abnormalities in Lymphoma and Their Clinical Relevance

Objective: To investigate the issue of molecular cytogenetic abnormalities in lymphoma and to study their clinical relevance. Methods: First, medium-term chromosomes of healthy men and women were prepared. Normal human peripheral blood lymphocytes were stimulated with PHA, treated with MTX synchronization, blocked with colchicine, cultured for 3 days, routinely treated with hypotonicity, cells were obtained, and steam-dried method was used to drop slices for backup. Next, comparative genomic hybridization. Preparation of probes; processing of normal human mid-term split images; hybridization; washing of slices after hybridization; staining; fluorescence microscopy observation, uptake and analysis of fluorescence signals. Results: Lymphoma CGH test results; Relationship between CGH abnormalities and lymphoma histological subtypes:Hodgkin's lymphoma CGH abnormalities, diffuse large B-cell lymphoma (DLBCL) CGH abnormalities, follicular lymphoma (FL) CGH abnormalities, and follicular lymphoma (FL) CGH abnormalities; Correlation between CGH test results and lymphoma clinical features: CGH test results and lymphoma patients, the type of CGH abnormality correlated with the clinical features of lymphoma, simple CGH abnormalities and complex CGH abnormalities correlated with the clinical features of lymphoma patients, and CGH abnormalities at specific chromosomal sites correlated with the clinical features of lymphoma. Conclusion:Conventional model analysis examines both chromosomal translocations and gene deletions or spreads, fails to observe the full picture of the lymphoma genome, and fails to detect new chromosomal abnormalities.

ОПРЕДЕЛЕНИЕ РАДИОЧУВСТВИТЕЛЬНОСТИ БОЛЬНЫХ РАКОМ СЛИЗИСТОЙ ОБОЛОЧКИ ПОЛОСТИ РТА ПО ЧАСТОТЕ КЛЕТОК С ПОЛИСОМИЕЙ ХРОМОСОМ 7 И 11 ПРИ ФРАКЦИОНИРОВАННОЙ ГАММА-ЛУЧЕВОЙ ТЕРАПИИ

Purpose: To study the dynamics of frequency of tumor cells with polysomy 7 and 11 chromosomes in patients with oral cancer during fractionated gamma-radiotherapy. Material and methods: The study was carried out using fluorescence in situ hybridization (FISH) on smears of 19 patients with oral cancer before treatment, after first total focal dose of gamma-radiotherapy (up to 18 Gy) and after second total focal dose (up to 32 Gy). As for control oral smears of 12 healthy donors were taken. Molecular cytogenetic abnormalities were examined in taken samples. Results: The average group frequency of cell with polysomy 7 and 11 chromosomes in controls was 0.7 ± 0.2 % and 0.3 ± 0.1 % respectively. The average frequency of cells with chromosome 7 polysomy in group oral cancer patients before treatment was 32.3 ± 4.5 %, after first focal dose was 15.7 ± 2.9 % and after second focal dose was 8.0 ± 2.3 %. Dynamics analysis of these values revealed the significant decreasing (р < 0.05) in cells with chromosome 7 polysomy during radiotherapy. The average group frequency of cells with polysomy 11 in cancer patients before treatment was 25.8 ± 5.6 %, 12.2 ± 2.4 % and 5.4 ± 0.9 % after first and second focal dose respectively. These values was significantly (р < 0.01) higher compare with the one in the control group. After radiotherapy the 16 (84 %) patients with polysomy chromosomes 7 and/or 11 had significant decreasing (р < 0.05) in aberrant levels. Some of them (6 persons – 32 %) had decreasing frequency up to the control levels. 4 persons (21 %) observed firstly the increasing of cells with polysomy but after there was the reduction. 5 patients (26 %) over full course of gamma-radiotherapy demonstrated the gain in frequency of cells with polysomy or the same value as it was before treatment. Conclusion: Thus, the study of molecular cytogenetic abnormalities in cells of oral cancer patients before and after radiotherapy was shown the polysomy of chromosomes being the marker of genome instability could indicate the tumor response ongoing therapy.

Therapy-related acute myeloid leukemia: A case series.

Patients with primary cancer receiving chemotherapy and/or radiotherapy may develop therapy-related acute leukemia (t-AL). Therapy-related acute myeloid leukemia (t-AML) accounts for the majority of these cases and is frequently associated with a variety of cytogenetic and molecular abnormalities. The aim of the present study was to explore the clinical characteristics, treatments and prognosis of patients with t-AML. A total of 272 cases of AML treated at our institution between 2016 and 2020 were reviewed, among which nine cases of t-AML were identified for analysis. All patients had received alkylating or topoisomerase II inhibitor chemotherapy drugs for primary cancer treatment and three patients had received radiotherapy. A total of nine patients had been administered recombinant human granulocyte colony-stimulating factor (G-CSF). The median latency period for the nine patients with t-AML was 25 months (range, 10–240 months). The molecular cytogenetic abnormalities included t(15:17)(q22:q21), inv(16)(p13q22), del(5)(q22), CBFB/MYH11(+), FLT3(+), NARS(+), IDH(+), TET2(+), and TP53(+). Out of nine patients with t-AML, eight received chemotherapy, two of whom underwent HSCT. The median survival time of the nine patients with t-AML was 10 months and the 2-year-survival rate was 44.4%. Greater clarity around the diagnosis and treatment is required to improve the outcomes of patients with t-AML.

Evolution in Myeloid Neoplasms Clonal Architecture Post Allogeneic Hematopoietic Cell Transplantation

Relapse-related mortality remains one of the main causes of death in patients (pts) with myeloid neoplasms (MN) who undergo allogeneic hematopoietic cell transplantation (allo-HCT). Molecular mutations and cytogenetic abnormalities present in hematopoietic cells are important determinants of relapse risk, with MRD predictive of relapse after allo-HCT. We studied the clonal hierarchy of MN at diagnosis compared to the relapsed state, to determine the effects of the ancestral hit and subclones on the impact on post-transplant recurrence.DNA from marrow/PB samples of MN pts was subjected to analysis at the time of diagnosis, post, and at the time of relapse to evaluate clonal expansion or evolution using a targeted multi-amplicon deep NGS panel of all ORFs of the top 60 most commonly mutated genes in MDS.We identified 43 pts with MDS and MDS/MPN who underwent allo-HCT from 2000 to 2016, whom were assessed serially in non-relapsed MN (NR-MN) and relapsed (R-MN). Median age at diagnosis was 54 years (y, range, 22-69) and 54y (range, 22-70) at time of HCT; 56% were male. NR-MN vs. R-MN pts median IPSS and IPSS-R scores at the time of diagnosis, 1.0 vs. 1.25 (P=0.5) and 4.0 vs. 4.0 (P=0.6), respectively. 53% underwent MAC and 58% had a PBSC. Pre-HCT treatments included HMA (42%), ESA (29%), 7+3 (10%), IMiD (10%), and HU (10%). Serial cases had a median follow up of 4 y (range 0.7 - 14.8) from diagnosis (Dx) and 2.3 (0.1-10.7) y from HCT. Rates before HCT of CR (30% vs. 30%), progressive disease (13% vs. 5%), stable disease (25% vs 25%), and disease progression (35% vs. 40%) (P=0.89). Median follow up post allo-HCT was 1.6y vs. 2.5y (P= 0.26). The incidence of acute GVHD in NR-MN vs. R-MN was 43% vs. 65% (P= 0.269) and 39% vs. 15% for chronic GVHD (P= 0.156), respectively. No difference in median blast percentage pre allo-HCT 4% vs 3.5% (P=0.5722).In NR-MN pts (N=23) after transplant, their pre-HCT characteristics showed 45% (10/23) had abnormal cytogenetics, with the most common abnormalities being del17p, del7q, and/or monosomy 7. SNP analysis showed pathogenic lesions in 43% of assessable cases (6/14), with most frequent abnormalities being loss of 14q11.2 (N=3) and aberrations (loss/UPD) of chr 7 (N=3). Furthermore, 17/21 pts had ≥1 mutation in decreasing frequency TET2 (N=4) , BCOR/BCORL1 (3) , SRSF2 (3), STAG2 (3), ASXL1 (3) , TP53 (2). Post-HCT NGS analyses are ongoing.In R-MN pts (n=20) after transplant, their pre-HCT characteristics showed 65% had abnormal cytogenetics at diagnosis, with the most common chromosomal aberrations involving monosomy 7/7q, trisomy 8, del5q, and del20q. Pre-HCT, 14/20 pts had ≥1 mutation in decreasing frequency TP53 (N=4) , SF3B1 (3), U2AF1 (3) , DNMT3A (2) , NRAS (2), and ASXL1 (2) . NGS at disease diagnosis, post-HCT, and at relapse was performed for 9 pts. In 5 pts relapse of the original ancestral hit (PHF6, SF3B1, NRAS, TP53, and DNMT3A) evolution of additional mutations (ASXL1, KRAS, EZH2) (4 featured in Figure 1). Clinical management of relapsed pts included a 2nd allo-HCT in 7 pts (35%), of which 2 are alive, and DLI in 6 (30%), of which 3 are alive.Logistic regression analysis demonstrated that mutations in DNMT3A and TP53 were non-significantly associated with higher relapse rates (OR= 3.27; P=0.1403). Univariable analysis for time to relapse demonstrated a significant association with earlier relapse with chromosome 11 aberrations (HR=6.4; P=0.005), but DNMT3A and TP53 together (HR=2.285; P=0.1071), TP53 alone (HR=2.257; P=0.1576), and complex cytogenetics (HR=2.15; P=0.1733) with relapse did not reach statistically significance, due to limited sample size.In conclusion, serial analysis of R-MN pre and post-HCT indicated recurrences of the original ancestral mutation, but often included acquisition of new mutations. Retraction but not elimination of the initial ancestral clone in R-MN pts demonstrated the relevance of an earlier ontogenetic stage. The initial hit could potentially drive the phenotype and resultant characteristics of the disease of subsequent lesions or vice versa. We plan to apply ultra-deep sequencing followed by an in-house bio-analytical pipeline to exclude false positive low frequency hits thereby determining the true MRD for the mutation identified in the original MN pre allo-HCT. Deeper NGS of MN pts in the pre & post allo-HCT setting will expand understanding of primary clonal aberrations and their subsequent re-emergence or regression with disease evolution. [Display omitted] DisclosuresGerds:CTI BioPharma: Consultancy; Incyte: Consultancy. Advani:Pfizer: Consultancy; Takeda/ Millenium: Research Funding. Sekeres:Celgene: Membership on an entity's Board of Directors or advisory committees. Majhail:Sanofi: Honoraria; Anthem, Inc.: Consultancy.

Acquisition Patterns and Interaction of Molecular, Obvious and Cryptic Cytogenetic Abnormalities in Myelodysplastic Syndromes

Abstract Introduction: The acquisition of cytogenetic aberrations (CA) and/or of molecular mutations (clonal evolution, CE) is known to be associated with poor outcome in pts with myelodysplastic syndromes (MDS) (Jabbour et al, Am J Hematol, 2013; Cevik et al, abstract, Onkologie, 2013). The aim of our study was to identify additional cases with CE by taking cryptic CA, which cannot be detected by conventional chromosomal banding analysis (CCB), and their interaction with molecular mutations, into account. Methods: We retrospectively reviewed 103 pts with proven MDS (10x RCUD, 40x RCMD, 10x RAEB-1, 16x RAEB-2, 11x sAML after MDS, 3x MDS-U, 7x CMML, 6x MDS with unknown subtype). Karyotype from CCB and molecular karyotype from SNP-array analysis (SNP-A) were available for all pts. SNP-A and fluorescence in situ hybridization (FISH) analyses were used to detect cryptic CA. For 57 pts results from mutational analysis from a 17-genes Sanger and/or a 54-genes next generation sequencing panel were included. Genetic follow-up and survival data were available for 57 pts, 22 pts were genetically analyzed under treatment with disease modifying therapies. Genetic follow-up comprised analysis of bone marrow and/or frequent sequential genetic monitoring of CD34+ peripheral blood cells. Results: CA were detected at diagnosis in 65/103 pts (63%). By CCB (aberrant in 55/103 pts; 53%) the median number of CA was 1 (range: 0-4). One to 6 (median 1) cryptic CA were detected in 31/103 (30%) pts, 10/31 pts (32%) showed a normal karyotype in CCB. The cryptic CA included cryptic copy number variations (CNV) in 12 pts and CN neutral losses of heterozygosity (CN-LOH) in 19 pts as the sole cryptic CA (14x) or in addition to cryptic CNVs (5x). In 12 pts with CN-LOHs we could identify a homozygous molecular mutation or a cryptic deletion in the region of a CN-LOH. Genes mutated in the region of a CN-LOH were DNMT3A (2p23.3, 2 pts), EZH2 (7q36.1, 2 pts), CBL (11q23.3, 2 pts), RUNX1 (21q22.12, 2 pts), TET2 (4q24), IKZF1 (7p12.2), TP53 (17p13.1), and SRSF2 (17q25.1). The formation of this phenomenon was observed in consecutive samples of 3 pts. From the 57 pts with follow-up data available, 8 showed manifestations of antecedent CE at first diagnosis (2x sub-clones; 6x homozygous mutations in CN-LOHs), from which 7 showed further genetic progression during the course of the disease. CE after first diagnosis was also observed in 20 of the 49 other pts with available follow-up data. The new aberrations of the biggest clone at first CE after diagnosis were partial or complete LOH of chromosome (chr) 7 (6 pts), trisomy 8 (5 pts), chr 21 abnormalities (2 pts), chr 12 abnormalities (2 pts), CN-LOH in 4q, 11p-, 20q-, -X, an additional marker chromosome, tetraploid cells, and mutations in ASXL1 (2 pts), CDKN2A, ETV6, IDH1, and JAK2 . Further steps of CE in these pts are currently being evaluated. Median survival time from detection of the first CE after diagnosis (25 months, 12/20 deaths) did not differ significantly from median survival time from first diagnosis for pts with signs of antecedent CE at first diagnosis (25 months, 5/8 deaths, P=0.92, n.s.). Median survival time of pts without detection of CE was significantly longer (173 months, 8/29 deaths, P=0.005 and Conclusions: In our cohort of MDS pts the frequency of cryptic CA was 30%. A homozygous mutation in a region of CN-LOH, which indicates antecedent CE, was detected in 12% of pts. The presence of such evolutionary patterns at first diagnosis might be indicative of a genetically instable disease as these pts had a shorter survival time and a shorter time to further genetic progression. There is a clear tendency that cytogenetic LOHs preferentially affect regions with molecularly mutated genes, resulting in homozygous mutations. The development of these patterns seems to be an important step towards progression. Our data imply that the relevance of cryptic CA for CE and disease progression in MDS has been underestimated as yet. Cryptic CA should be taken into account for developing prediction models for clinical progress and for improving individualized patients care. Download : Download high-res image (124KB) Download : Download full-size image Disclosures Al-Ali: Gilead: Consultancy, Honoraria; Alexion: Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding. Metz: Celgene: Other: study projects, advanced training support; Novartis: Consultancy, Other: study projects, advanced training support. Germing: Janssen: Honoraria; Novartis: Honoraria, Research Funding; Celgene: Honoraria, Research Funding. Platzbecker: Janssen: Consultancy, Honoraria, Research Funding; Acceleron: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Research Funding; Celgene: Consultancy, Honoraria, Research Funding. Haase: Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy, Honoraria, Research Funding.

Identification of an Immune-Related Gene Signature Based on Immunogenomic Landscape Analysis to Predict the Prognosis of Adult Acute Myeloid Leukemia Patients.

Acute myeloid leukemia (AML) is a hematopoietic malignancy characterized by highly heterogeneous molecular lesions and cytogenetic abnormalities. Immune disorders in AML and impaired immune cell function have been found to be associated with abnormal karyotypes in AML patients. Immunotherapy has become an alternative therapeutic method that can improve the outcomes of AML patients. For solid tumors, the expression patterns of genes associated with the immune microenvironment provide valuable prognostic information. However, the prognostic roles of immune genes in AML have not been studied as yet. In this study, we identified 136 immune-related genes associated with overall survival in AML patients through a univariate Cox regression analysis using data from TCGA-AML and GTEx datasets. Next, we selected 24 hub genes from among the 136 genes based on the PPI network analysis. The 24 immune-related hub genes further underwent multivariate Cox regression analysis and LASSO regression analysis. Finally, a 6 immune-related gene signature was constructed to predict the prognosis of AML patients. The function of the hub IRGs and the relationships between hub IRGs and transcriptional factors were investigated. We found that higher levels of expression of CSK, MMP7, PSMA7, PDCD1, IKBKG, and ISG15 were associated with an unfavorable prognosis of AML patients. Meanwhile, patients in the TCGA-AML datasets were divided into a high risk score group and a low risk score group, based on the median risk score value. Patients in the high risk group tended to show poorer prognosis [P = 0.00019, HR = 1.89 (1.26–2.83)]. The area under the curve (AUC) was 0.6643. Multivariate Cox Regression assay confirmed that the 6 IRG signature was an independent prognostic factor for AML. The prognostic role of the immune related-gene signature was further validated using an independent AML dataset, GSE37642. In addition, patients in the high risk score group in the TCGA dataset were found to be of an advanced age, IDH mutation, and M5 FAB category. These results suggested that the proposed immune related-gene signature may serve as a potential prognostic tool for AML patients.

Prevalence of Genetic Abnormalities in Patients with Multiple Myeloma and Its Clinical Relevance in a Developing Country like India

Introduction Multiple myeloma (MM) is a malignancy involving terminally differentiated plasma cells. Its incidence in India is about 0.7/1,00,000 population amounting to about 6,800 new cases a year A number of genomic aberrations are associated with MM, most of which confer prognostic significance. Cytogenetic abnormalities are a part of R-ISS score for prognostication which stratifies presence of del(17p), t(4;14) or t(14;16) as stage 3, mSMART is another risk stratification tool which divides MM into high risk and standard risk groups based on genetic aberrations. Hence it is evident that determining the genetic abnormality in MM is important. however, due to limited resources genetic testing is not routinely done and the data in the Indian population is limited. Objective: To estimate the prevalence of molecular cytogenetic abnormalities by Fluorescent in situ hybridization (FISH) analysis in patients with MM and to assess the co-relation with response to induction chemotherapy, relapse and overall survival. Material and Methods: 64 patients were included from January 2016 to December 2019 and followed up till June 2020. Interphase FISH study was performed either at diagnosis or at relapse, on bone marrow aspirate with panel of probes consisting of CKS1B (1q21-22), CDKN2C (1p32.3), D13S319 (13q14.2/13q34), IGH (14q32.33), p53 (17p13.1) and trisomy (5p15/9q22/15q22) (trisomies are considered as hyperdiploidy in this study). Plasma cell purification techniques were not applied prior to FISH analysis. Patients were divided into 2 risk groups; 1) high risk group with presence of del17p, del13q, amplification 1q, del1p and two or more aberrations with either of these and 2) standard risk group with presence of hyperdiploidy or no genetic abnormality. There was no difference in chemotherapy regimen between the 2 groups; 46 (71.8%) received bortezomib-thalidomide-dexamethasone, 10 (15.6%) received bortezomib-cyclophosphamide-dexamethasone, 2(3.1%) received bortezomib-lenalidomide-dexamethasone, 1(1.5%) received daratumumab-bortezomib-dexamethasone and 5(7.8%) received 2 drug chemotherapy. Patients who did not complete minimum follow up of 6 months either due to death or lost to follow up were excluded from the study. Institutional Ethics Committee's approval was taken. Results: Mean age of the population was 60.33 years and male to female ratio was 1.65. 46.87%, 28.13% and 25% of the study population were in the age group of ≤ 60, 61 - 65 and ≥66 years respectively. 12.3%, 43.8% and 43.8% were in R ISS stage 1, 2 and 3 respectively. FISH analysis was done on 61 out of 64 patients (remaining 3 were excluded due to hemodilute bone marrow sample). 22 (36.1%) patients had abnormal genetic aberration on FISH analysis with 10 (16.39%) having two or more abnormalities. The frequency of genetic aberrations was as follows; amplification 1q (13/61, 21.31%), del13q (9/61, 14.75%), hyperdiploidy (7/61, 11.47%), del17p (4/61, 6.55%), IgH rearrangement (3/61, 4.91%), and del1p (1/61, 1.6%). All 3 patients with IgH rearrangement had associated one or more high-risk genetic aberration and hence were included in high risk group. 31.1% of the patients were high risk and 68.9% were standard risk. The response to induction chemotherapy, incidence of relapse, time to 1st relapse and total number of relapses are shown in (table;1) and there was no significant difference between high risk and standard risk group. Overall survival in standard risk group at 2 and 5 years was 89.6% ± 5.8% and 78.6% ± 13.9% and in high risk group was 60.7% ± 13.2% and 29.5% ± 23.1% respectively (p value: 0.762). Overall survival was significantly lower in age group ≥66 years as compared to age group ≤ 60 years and 61 - 65 years (p value 0.001) and it was also significantly lower in R ISS stage 3 as compared to R ISS stage 1 and 2 (p value 0.006). Conclusion: More than one third patients of MM (36.1%) showed genetic abnormality, amplification 1q being the most frequent. Overall survival was significantly lower in older age group and R ISS stage 3 patients. Response to induction chemotherapy and relapse rate were similar in high and standard risk groups. Although overall survival was lower in high risk group, it was statistically not significant. This study highlights the importance of FISH analysis for disease stratification and prognostication which should be routinely practiced. Disclosures No relevant conflicts of interest to declare.

Fluorescence in Situ Hybridization for Detecting Molecular Cytogenetic Abnormalities of Chronic Lymphocytic Leukemia

To investigate the value of fluorescence in situ hybridization (FISH) in the diagnosis and prognosis evaluation of patients with chronic lymphocytic leukemia (CLL). Ninty-three patients with newly diagnosed CLL were tested by five probes including RB1 (13q14.1), D13S25 (13q14.3), p53(17p13.1), ATM( 11q22.3) and CSP12, while conventional cytogenetics (CC) was used for karyotype analysis. Then the correlation of the molecular cytogenetic abnormalities with the clinical Binet stages, Rai stages and the other related laboratory examinations was analyzed. The detection rate of chromosome abnormality in 93 patients was 79.6%, out of which detection rate of 13q (13q- was the highest and accounted for 45.2%), followed by trisomy 12 (+12) 26.9%, p53 deletion (17p-) 19.4% and ATM deletion (11p-) 17.2%. There were 27 cases (29.0%) with 2 or more abnormalities, including 13 cases with 13q-/17q-, 5 with 13q-/11q-, and 4 with 13q-/+12. Compared with CC test results, the positive rate of FISH detection was significantly higher (χ2=32.127, P＜0.01). There was no significant correlation between FISH results and Rai stages (P＞0.05), meanwhile 17p- highly correlated with later stage of the Binet stages (P=0.012). The molecular cytogenetic abnormalities significantly correlated with age, absolute value of peripheral lymphocyte count and CD38 expression level (P＞0.05). The incidence of 13q- in female (65.4%) was statistically significantly higher than that in male (37.3%) (P=0.015). The unmutated IGHV rate of CLL patients with a 17p- was significantly higher than that in patients without this genetic abnormality (P=0.013). The expression of CD38 was detected among 29.0% of the patients, which significantly correlated with Binet stages (P=0.027) and unmutated IGHV (P=0.006). FISH can greatly increase the detection rate of molecular cytogenetic abnormalities in CLL patients, which, as a powerful supplement to the conventional cytogenetics, can be applied for the clinical staging and prognosis evaluation of CLL patients.

Abstract IA10: Liquid biopsies as biomarkers of disease progression in multiple myeloma

Abstract Multiple myeloma (MM) is a plasma cell dyscrasia that is almost always preceded by the precursor states of monoclonal gammopathy of undetermined significance (MGUS) and smoldering multiple myeloma (SMM). Some patients rapidly progress from MGUS/SMM to overt MM while others remain indolent for many years. Disease evolution also depends on the surrounding microenvironment, which can be either permissive or inhibitory of clonal expansion. The use of liquid biopsies for cancer detection and monitoring is rapidly gaining prominence. Current methods for the detection of circulating tumor cell-free DNA (cfDNA) include low-pass genome sequencing for copy number profiling and deep targeted sequencing for detection of recurrent somatic mutations. However, the sensitivity of these methods is limited among patients with minimal disease, such as MGUS or the minimal residual disease (MRD) state, on account of low tumor burden and a highly variable genomic landscape, whereby most patients do not carry any of the recurrent alterations. I will discuss our current data on detecting cfDNA and circulating tumor cells in the peripheral blood of patients with MGUS, SMM, and overt MM as well as novel personalized, sensitive methods to detect cfDNA with deep UMI-based sequencing for potential use to detect MRD. There is an urgent need for a liquid biopsy assay that can be integrated in the clinical diagnosis of patients for more sensitive tumor detection along with better characterization of their genomic and epigenetic alterations leading to much-needed solutions for integrative staging of MM precursor and MRD patients, with potential implications for patient monitoring and treatment.

Results of Cytogenetic and Molecular Cytogenetic Studies in Relapce/Refractory of Multiple Myeloma

Background. Multiple myeloma (MM) – a tumor of the lymphoid tissue, which is characterized by low proliferative activity of abnormal cells and a wide range of acquired chromosomal aberrations that play a leading role in predicting the course of the disease. Accumulation of new data in the relapsed/refractory of MM will make it possible to understand the nature of the formation of cloning insensitive to chemotherapy (CT), to predict the effectiveness of therapy, which will further improve the individualization of treatment for each patient. Objective. The aim of the study is to determine the peculiarities of chromosomal aberrations of abnormal clones of bone marrow (BM) cell in relapsed/refractory of MM by standard cytogenetic and molecular cytogenetic investigations. Methods. Analysis of BM cells chromosomal abnormalities in relapsed/refractory of MM for 62 patients with MM was performed. Slides of metaphase chromosomes were prepared according to standard procedure after cultivating during 24 or 96 h in the medium without stimulation. Results. Clonal chromosomal abnormalities were formed, complexity structure of karyotypes with clone structures was shown. In structural rearrangements, derivative chromosomes (34.9%) and balanced translocations (23.2%) were dominated at cytogenetic procedure. By molecular cytogenetic (i-FISH) investigation chromosomal abnormalities were detected in 26.7% cases. Del(17)(p13.1) was registered in 22.0%. Aberrations involving IGH were fixed in 5.7%. The atypical signal distribution determined by i-FISH may indicate another possible mechanism for the evolution of an abnormal clone. Conclusions. The cytogenetic peculiarities of the formation of abnormal BM cell clones in relapse/refractory of MM formations, discovered by us, supplement the understanding of the biological features of the disease and contribute to the outline of further tasks for the study of genetic aberrations in MM.

Detection of the Cytogenetic Aberrations in Multiple Myeloma by Using Microrray Comparative Genomic Hybridization

To detect the molecular cytogenetic abnormalities of multiple myeloma (MM) by using microrray-based comparative genomic hybridization (array-CGH) technology and to investigate its value of application in MM. The whole-genoine copy number variants (CNV) of bone marrow samples acquired from 20 cases of newly diagnosed MM patients were detected by genome-wide hybridization and scanning by CytoScan 750K Array (Affymetrix). At the same time, the chromosome abnormalities of bone marrow cells were detected by karyotype analysis and FISH using 9 specific probes: D13S319, RB1, p53, 1q21, IgH, IgH/CCND1, IgH/FGFR3, IgH/MAF, IgH/MAFB. Among the 20 MM patients, the incidence of chromosome abnormalities detected by karyotype analysis, FISH and array-CGH were 15%, 65% and 90%, respectively. The types of CNV detected by array-CGH included the gain (106), loss (156) or UPD (23). There were many different CNVs in every chromosomes except chromosome 5, 9, 18, 21 and Y. Comparison of chromosome abnormalities detected by FISH and array-CGH showed that, the positive ratio of del (13q) was 35% and 40% respectively; the positive ratio of amp (1q) was 40% and 50% respectively; the positive ratio of del (17p) was both 15%. FISH detection showed 8 cases with IgH rearrangement, meansahile the array-CGH detection showed that 4 cases had amp (11q13) (CCND1 gene), 3 cases had amp (16q23) (MAF gene), 1 case had amp (4p16) (FGFR3 gene) and 2 cases had amp (20q12) (MAFB gene). Besides, many other new chromosome abnormalities were found. More than half of MM patients have cytogenetic changes, and most of them are complex chromosomal abnormalities. By using array-CGH, more chromosome abnormalities can be detected and more cytogenetic information can be provided for clinician.

Inhibition of the Mitochondrial Protein Import Machinery Is Selectively Cytotoxic to Acute Myeloid Leukemia (AML) Cells and Stem Cells

A subset of AML and stem cells have increased mitochondrial stress and increased expression of mitochondrial proteases that degrade misfolded mitochondrial proteins. Given the recent findings of the interplay between mitochondrial homeostasis and mitochondrial protein import, we hypothesized that AML cells have an increased reliance on mitochondrial protein import as a compensatory mechanism for increased mitochondrial stress. To test this hypothesis, we measured expression levels of key mitochondrial protein import genes in publicly available datasets (GSE30377, GSE42414 and GSE 24759) and demonstrated their up regulation in a subset of AML cells over normal hematopoietic cells. Increased expression occurred across the spectrum of molecular mutations and cytogenetic abnormalities. Moreover, expression levels of mitochondrial protein import genes were enriched in functionally defined AML stem cells over bulk cells.To assess the impact of inhibiting mitochondrial protein import in AML, we knocked down the outer mitochondrial membrane import channel TOM40, the inner membrane import channel Tim23 and the oxidase ALR, that folds proteins through a disulfide relay system in the mitochondrial inner membrane space and is a rate limiting step for the import of a subset of mitochondrial proteins. Knockdown of these targets in OCI-AML2, TEX and U937 leukemia cells with shRNA reduced growth and viability of AML cells. Knockdown of ALR targeted the leukemia initiating cells as it abrogated engraftment of TEX leukemia cells into immune deficient mice (shRNA ALR = 5.481 +/- 0.9 % engraftment vs shRNA control= 29.44 +/- 5.4 % engraftment; p =0.0004) . Mechanistically, knockdown of mitochondrial import genes reduced levels of nuclear (ATP5A, SDHB and NDUFB8), but not mitochondrial (CoxII) encoded proteins of the OXPHOS chain. This in turn led to decreased basal oxygen consumption in leukemic cells. As a chemical approach to investigate the impact of inhibiting mitochondrial protein import in AML and normal cells, we tested the effects of MitoBloCK-6 and related analogues that selectively inhibit ALR in zebrafish, hESCs and yeast models. MitoBloCK-6 and related analogues killed leukemia cell lines (OCI-AML2, TEX, Jurkat and NB4) with an IC50of 5-10 µM. At these concentrations, MitoBloCK-6 decreased levels of nuclear (ATP5A, SDHB and NDUFB8), but not mitochondrial encoded (CoxII) proteins of the OXPHOS chain. Demonstrating the functional importance of changes in mitochondrial metabolism by these compounds, rho zero 143B rhabdomyosarcoma cells that lack mitochondrial DNA and rely solely on glycolysis were resistant to MitoBloCK-6. Finally, we tested the effects of MitoBloCK-6 on primary AML and normal cells. Treatment with MitoBloCK-6 (2 µM) inhibited clonogenic growth of (4 of 5) primary AML > 64% but produced <3% loss of clonogenic growth in normal hematopoietic cells. Finally, in an OCI-AML2 xenograft model, systemic administration of MitoBloCK-6 reduced tumor growth > 50% of control without toxicity.Thus, we have demonstrated that AML cells have a unique reliance on mitochondrial protein import and inhibition of this pathway may be a new therapeutic strategy to selectively target a subset of AML and AML stem cells. DisclosuresSchimmer:Novartis: Honoraria.

Cytogenetic Abnormalities and Outcomes of 117 Patients with Multiple Myeloma Detected by FISH

To analyze the cytogenetic abnormalities and prognostic outcomes of patients with multiple myeloma (MM) detected by fluorescence in situ hybridization (FISH). The clinical record of 117 newly-diagnosed patients with MM treated in department of hematology and geriatric hematology of our hospital for 7 years were collected, and their molecular cytogenetic abnormalities detected by FISH and the clinical outcome were analyzed retrospectively. The detected rate of cytogenetic abnormality was 76.9%(90/117), the most common abnormality deteted by FISH was 1q21+ (71.1%), followed by 13q- (56.6%). The cross comparison method showed that 13q- and 17p13-, t(11;14) and t(4;14) were related respectively. All the patients with cytogenetic abnormalities showed no significant difference in the overall survival from cytogenetic normal patients. The positive rate of molecular cytogenetic abnormalities detected by FISH in MM patients is high, but data from larger and longer studies are needed to evaluate the prognostic outcomes.

Racial Differences in Molecular Cytogenetic Abnormalities in Black and White Patients with Multiple Myeloma (MM): A Single-Center Experience

Background: The incidence of MM is 2 to 3 fold higher in blacks than in whites; they present at a younger age and have better overall survival. The biological bases for these disparities remain unclear. Outcome of MM is linked to cytogenetic and molecular changes, both primary (hyperdiploidy and heavy chain (IgH) translocations) and secondary (rearrangements of MYC, activating mutations of NRAS, KRAS or BRAF, and deletions of 17p).Methods: Cytogenomic alterations in consecutive MM patients were assessed using integration of metaphase chromosome analysis by GTG-banding and interphase fluorescence in situ hybridization (iFISH) in CD138-positive cells isolated from fresh BM samples using a protocol of magnetic-activated cell sorting. Changes evaluated included monosomy 13/del(13q), monosomy 17/del(17p), gain of 1q21, and rearrangements of the IGH gene including t(4;14), t(11;14) and t(14;16).Results: Samples from 218 consecutive MM patients were analyzed (Table 1). 108 were from black and 110 were from white patients. Median age for blacks was 59 years (range: 36 - 82) and for whites, 63 years (range: 39 - 83) (p=0.008). Fewer black men than whites were observed (46.3% versus 64.6%, p=0.007). Overall, blacks had fewer abnormal karyotypes compared to whites (18.1% versus 31.8%; p=0.02). Black patients had a lower frequency of non-hyperdiploid karyotypes (8.5% versus 20.6%; p=0.01) and had a trend toward lower frequencies of rearrangements of IGH (30.8% versus 43.5%; p=0.055) than white patients. Most notably, they had significantly lower frequencies of monosomy 17/del(17p) (5.6% versus 18.5%; p=0.003) and monosomy 13/del(13q) (28.9% versus 46.3%; p=0.008).After stratification by age (Figure 1), younger patients showed significantly higher frequencies of the monosomy 17/del(17p) abnormality (p=0.001) and the t(4;14) (p=0.04) than older patients, with the difference more significant in white patients. The associations among molecular cytogenetic abnormalities (Figure 2) showed a different association pattern for black and white patients. White patients with t(11;14) were more likely to have monosomy 13/del(13q) (p=0.003) and gain of 1q21 (p=0.02), while this association was not observed in black patients.Conclusion: Black MM patients had significantly different cytogenetic profiles detected by iFISH on CD-138 selected malignant cells, compared to whites. Black MM patients had a more favorable profile, including lower frequencies of non-hyperdiploid karyotype and of IGH rearrangements. This study supports a biological basis for previously described outcome disparities between black and white patients with MM. Further studies will focus on identifying specific molecular targets and their impact on therapy and on overall outcome.Table 1Demographics and cytogenetic abnormalities of the MM patientsDemographicsBlackWhiteP-value#Total, n108110Gender, n (%)=0.007*Male50 (46.30%)71 (64.55%)Female58 (53.70%)39 (35.45%)Age (median)5963=0.008*Chromosome (karyotype)=0.022*Normal86 (81.90%)73 (68.22%)Abnormal19 (18.10%)34 (31.78%)Hyperdiploidy8 (7.6%)8 (7.4%)Non-hyperdiploidy9 (8.5%)22 (20.6%)=0.013*11;14 translocation2 (1.9%)4 (3.7%)FISH abnormality-13/del(13q)31 (28.97%)50 (46.30%)=0.008*Gain of 1q2135 (32.71%)47 (43.52%)=0.103-17/del(17p)6 (5.61%)20 (18.52%)=0.003*IGH rearrangements33 (30.84%)47 (43.52%)=0.055^t(4;14)7 (6.54%)13 (12.38%)=0.146t(11;14)15 (20.55%)15 (19.48%)=0.870t(14;16)2 (3.85%)6 (10.71%)=0.175others16 (14.95%)15 (13.89%)=0.824*means statistical significant (p-value < 0.05), where ^ means marginal significant (0.05 < p-value < 0.10). #p-values come from the Cochran-Mantel-Haenszel tests for categorical variables, and t tests for continuous variables.Associations among eight molecular cytogenetic abnormalities. Each solid black line indicates one abnormality is statistically significantly associated with another abnormality. [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Detection of Molecular Cytogenetic Aberrations by Fluorescence in Situ Hybridization in Different Bone Marrow Samples of Multiple Myeloma

To detect the molecular cytogenetic abnormalities in different bone marrow samples of multiple myeloma by using fluorescence in situ hybridization (FISH) technology. The bone marrow cells from 48 cases of MM were taken for sorting the plasma cells using CD138 magnetic beads, and the biopsy tissue from 10 cases of MM was taken and embedded in parafin, then the 2 kinds of samples were detected by using FISH. D13s319/RB1, 1q21/P53, IgH, IgH/FGFR3, IgH/MAF probes were detected in 58 patients with new diagnosed multiple myeloma (MM) by FISH technology. Fluorescent signals were both seen in 2 different types of bone marrow samples and cytogenetic aberrations were detected in 30/58 (51.7%) patients, 29.3% (17 out of 58) cases had both D13S319 and RB1 deletion. The positive rates of P53 deletion, 1q21 amplification and IgH rearrangement were 12%, 27.6% and 20.7%, respectively. Only 7 cases (23.3%) had one cytogenetic abnormality, other 23 (76.7%) cases all had 2 to 5 kinds of different abnormalities. More than half of MM patients have cytogenetic change, and most of them are complex chromosomal abnormality. The different kinds of samples can expand the useful extension of FISH technology and acquire more cytogenetic information for clinician.

Autologous Followed By Allogeneic Versus Tandem-Autologous Stem Cell Transplant in Newly Diagnosed FISH-del13q Myeloma

▪Background In multiple myeloma (MM), the introduction of novel compounds into first-line intensive treatment pathways has clearly improved patients’ prognosis. Very recently however, specific molecular cytogenetic abnormalities, lactate dehydrogenase elevation and International Staging System 3 disease were identified to be associated with dismal prognosis despite upfront autologous (auto) stem cell transplant (SCT). Consolidative allogeneic (allo) following initial auto SCT was shown to extend progression-free survival (PFS) as well as overall survival (OS) in some prospective studies on newly diagnosed MM patients (pts). Relatively little is known on the impact of cytogenetic features other than chromosome 13q deletion (del13q) on the outcomes of pts undergoing upfront auto followed by allo (auto/allo) SCT.Patients and methods When the DSMM V treatment program was designed del13q detected by fluorescence in situ hybridisation (FISH) was accepted as one of the distinct risk factors in MM. We therefore used FISH del13q to define the study’s “high-risk” group and aimed to compare tandem high-dose melphalan 200 mg/m² (Mel) with one cycle of Mel followed by reduced-intensity conditioning (RIC) allo SCT. Allocation to either transplant regimen was by availability of an HLA-matched (at least 9/10 matches) related (MRD) or unrelated donor (MUD). Initially, all pts underwent non-novel compound cytoreduction and chemomobilization of peripheral blood stem cells (PBSC). RIC allo SCT was prepared by fludarabine and melphalan (plus ATG in MUD cases). PFS was the primary endpoint. The study was powered to detect an improvement of 2-year PFS from 20% (tandem Mel) to 40.3% (HR, 1.769).Results 199 out of 225 del13q pts with a median age of 53 (range, 30 – 60) yrs who had been enrolled between 10/2001 and 03/2007, were included in the intent-to treat population. Allo SCT was performed in 126/199 pts (63%), 74 of whom (59%) received MUD allografts. At a median follow-up of 49.2 months (mo), 2-year PFS (calculated from day 1 of second SCT) was 59% with auto/allo SCT versus 47% with tandem Mel. Median PFS with auto/allo SCT was 34.5 mo versus 21.8 mo, respectively (p=.005). Two-year non-relapse mortality (NRM) associated with auto/allo SCT was 11.9%. As of yet, there is no difference in OS between the groups, with the median not yet reached for either transplant modality. PFS/OS in auto/allo SCT were independent of donor source (MRD vs MUD). As definitions of cytogenetic risk have evolved over time, we analyzed further FISH abnormalities in pts’ baseline samples: in addition to uniform del13q, 13.6% of pts displayed del17p. Median PFS for del13q/del17p pts after HD Mel was 6 mo versus not reached with auto/allo SCT, respectively (p=.0002). Median OS in del13q/del17p after HD Mel was 23.4 mo versus not reached, respectively (p=.011). In translocation (4;14)/del13q pts (20.7%), median PFS with tandem Mel was 19.3 mo versus 19.1 with auto/allo SCT, respectively (p=.251).Conclusions This prospective trial shows auto/allo SCT to significantly extend PFS when compared to tandem HD Mel in a large cohort of del13q MM pts. It is the first study to demonstrate allo SCT in MM can be safely performed from matched unrelated donors at a reasonable rate of NRM. Utilizing a comprehensive set of FISH cytogenetics, our data for the first time demonstrate allo SCT to specifically benefit patients with high-risk features (del13q/del17p). Incremental gain of PFS when compared to tandem Mel was more than 20 months. Extended OS data on the whole study will be presented. DisclosuresNo relevant conflicts of interest to declare.

Response to Lenalidomide, Doxorubicin and Dexamethasone (RAD) in Newly Diagnosed Multiple Myeloma Is Independent of Cytogenetic Risk and Retained after Double Stem Cell Transplant

▪Background Induction triplets with at least one of the “novel drugs” and steroids with or without chemotherapy are deemed standard of care in newly diagnosed multiple myeloma (MM). Medically fit patients (pts) remain candidates for subsequent autologous (auto) stem cell transplant (SCT) while the use of allogeneic (allo) SCT is an ongoing matter of debate. We had previously shown the RAD regimen to be well tolerated and highly effective in relapsed and relapsed/refractory disease. Based on the overall results we decided to further evaluate the combination in first-line treatment. Methods The current phase II trial was designed to include pts up to 65 years of age with newly diagnosed, symptomatic MM. Four 4-week RAD induction cycles (lenalidomide 25 mg/day, d 1-21; infusional adriamycin 9 mg/m², d1-4; oral dexamethasone 40 mg, d1-4 and 17-20; pegfilgrastim 6 mg, d 6) were followed by stem cell chemomobilization. Pts received either tandem auto SCT (melphalan 200 mg/m²; Mel) or auto followed by allo SCT. Allo SCT (conditioning regimen: treosulfan/fludarabine) was reserved for pts featuring at least one cytogenetic or serologic risk factor who had a matched sibling or unrelated donor available. Target dose for subsequent lenalidomide maintenance (R-maint; for one year) was 10 mg/d for tandem Mel and 5mg/d for auto/allo pts. Primary endpoint was response (at least VGPR) following second SCT. Results 190 pts with a median age of 55 (range, 30-66) years were recruited by 17 German centers between 8/2009 and 4/2012. 103 pts (56%) had ISS stage II/III disease and 165 pts are evaluable for molecular cytogenetic abnormalities assessed by fluorescence in situ hybridisation (FISH). Incidences were as follows: 29.6% had deletion of (del) chromosome 13q, 11.5% showed translocation (t) (4;14), 9% presented with del 17p, and 0.6% had a t(14;16). 163 pts completed all 4 RAD cycles and 47 underwent allogeneic SCT. 60 pts following tandem Mel and 15 pts after auto-allo SCT proceeded to R-maint. Median number of maintenance cycles was significantly higher in tandem Mel compared with auto-allo pts (12 versus 3; p=.01). Rate of at least (≥) VGPR increased from 47.9% following RAD induction to 60.6% following double SCT. Accordingly, post-induction complete response (CR)/stringent CR rate of 7.9% increased to 31%. In pts with FISH results, ≥ VGPR rate was 44% with t(4;14)/t(14;16)/del 17p versus 53% without those abnormalities, respectively (p=.34). Following double SCT, ≥ VGPR was increased to 63% [t(4;14)/t(14;16)/del 17 pts.] versus 65%, respectively (p=.84). Response was not different with either transplant strategy. No treatment-related mortality occurred during RAD induction, while non-relapse mortality at one year from allo SCT was 10.6%. Incidences of pneumonia, venous thromboembolism, and febrile neutropenia were 11, 7.2, and 5.3%, respectively. Preliminary overall survival results will be presented. Conclusions Our data show RAD induction to be very effective in newly diagnosed MM and to be well tolerated. By subsequent double SCT, a high number of CR/sCR was added. Correlative studies suggests response to be independent of known unfavourable cytogenetic prognosticators, such as t(4;14) and del 17p while time-to-event data will be needed to ultimately confirm a lenalidomide-based triple regimen being able to overcome adverse cytogenetic risk. Administration of R-maint to auto-allo pts was challenging despite a lenalidomide target dose of 5 mg/day. DisclosuresKnop:Celgene GmbH: Consultancy, Honoraria. Off Label Use: Use of lenalidomide, doxorubicin, and dexamethasone in newly diagnosed multiple myeloma. Einsele:Celgene GmbH: Consultancy, Research Funding. Bargou:Amgen Inc.: Consultancy, Honoraria, Other.