ALBERT

Description: Aberrant methylation of CpG island of the promoter regions of genes is an important epigenetic mechanism, alternative to deletions or mutations, that cause the silencing of genes involved in carcinogenesis. We therefore investigated epigenetic events involved in multiple myeloma by screening 6 genes of interest for evidence of promoter hypermethylation. PATIENTS AND METHODS: The methylation patterns of the tumor suppressor genes p16INK4a (p16), E-cadherin (ECAD), FHIT and PTEN (negative regulator of AKT/PKB signalling pathway), Wnt signalling pathway antagonist genes WIF-1 and DKK-3 were determined in the bone marrow aspirates of 50 patients with Multiple Myeloma (MM) (37 at diagnosis; male 22; female 28; 68.8 median age. ISS stage: 47% stage I, 8% stage II and 45% stage III.), using the methylation-specific polymerase chain reaction after DNA bisulphite modification. Ten patients with monoclonal gammopathy of undetermined significance (MGUS) and 4 peripheral blood from controls were also evaluated. For statistical analysis Student t and Chi squared or Fisher exact tests were used. RESULTS: We found at least one hypermethylated gene in all MM patients. Gene methylation frequencies varied from 6% to 74%. ECAD, FHIT and p16 genes demonstrated a relatively high frequency of aberrant methylation with frequencies of 74%, 69% and 42%, respectively. WIF-1, PTEN, DKK-3 genes showed a low frequency of methylation with values of 28%, 14%, 6%, respectively. The methylation frequencies detected in MGUS were 91.6% for ECAD and FHIT, 41.6% for WIF-1, 16,6% for p16 and DKK3. The difference in methylation profile between MM and MGUS was statistically significant (X2= 37, df=5; p=0.0000). PTEN was found hypermethylated in 7/50 (14%) MM but none of MGUS or control samples were methylated. Of these 6 patients had IgG K isotype, 4 had abnormal cariotype with 13q deletion, one hyperdiploidia, one IgH rearrangements. Two patients were simultaneously hypermethylated in p16 gene promoter. No statistically significant correlation between methylation data and clinical parameters: gender, age, isotype of M component, type of light chain, haemoglobin, serum albumin level, calcium, β2 microglobulin, serum creatinine level, LDH, lytic bone lesions were found for any of examined genes. When correlation with Durie-Salmon Stage disease was tested hypermethylation of p16 and WIF-1 resulted more frequently associated with stage IIIA/B (p values 0.006 and 0.000, respectively). When gene methylation status and cytogenetics abnormalities were compared, we found that aberrant methylation of p16 and ECAD were significantly more frequent in MM patients with chromosome 13 abnormalities as sole entity or with 13q deletion associated to IgH translocations. By contrast, promoter hypermethylation of Wnt signalling antagonist genes was associated with 13q deletion co-occurring with a hyperdiploide cariotype. (X2=17; df=8 p value: 0.000). According to the number of methylated genes observed in each sample 48% MM had 3–4 hypermethylated genes. When differences between methylator phenotype and cytogenetics abnormalities were analyzed we found that 3–4 hypermethylated genes have the tendency to be associated with chromosome 13 abnormalities more often than with hyperdiploide cariotype (p value 0.05). CONCLUSION: Our results indicate that the accumulation of epigenetic events affecting genes regulating cell cycle control, cell adhesion and apoptosis is a common phenomenon in plasma cell disorders and may further contribute to the phenotype of MM cells. This is the first demonstration of PTEN inactivation as a result of promoter hypermethylation in MM patients. Since talidomide seems to have a role in the PTEN/PI3K/AKT pathway, the PTEN function is worthy to be further investigated in Multiple Myeloma.

Description: Abstract 2956 Poster Board II-932 Background. The B-cell leukemia 11A gene (BCL11A/Evi9/CTIP1) is essential for normal lymphoid development and genetic association studies have shown its potential regulator effect in blood related phenotypes. BCL11A encodes a Krüppel-like zinc-finger protein and functions as a transcriptional repressor through its interaction with several proteins including BCL6. The corresponding mouse gene is a common site of retroviral integration in myeloid leukemia, and may function as a leukemia oncogene. It is down-regulated during hematopoietic cell differentiation and abnormalities involving this gene have been detected in a variety of B-cell malignancies in humans. We genotyped SNP rs11886868 in the BCL11A gene, which has been previously associated with HbF production, in patients with hematological malignancies from Sardinia to investigate a possible contribution of this gene in determining genetic susceptibility to onco-hematological diseases. Patients and Methods. We screened a total of 325 patients with hematological malignancies for rs11886868 SNP at the BCL11A locus using the TaqMan allelic discrimination assay: 51 B-cell Non Hodgkin's lymphoma (NHL), 27 Hodgkin's disease (HD), 42 Chronic Lymphocytic Leukemia (CLL), 52 Multiple Myeloma, 35 Cutaneous T-cell Lymphomas (CTCL), 11 Acute Lymphoblastic Leukemia (ALL), 19 Myelodysplastic Syndromes (MDS), 31 Acute Non Lymphoblastic Leukemia (ANLL), 36 Philadelphia negative Myeloproliferative Disorders (MPD), 21 Chronic Myelogenous Leukemia. Fifty–four DNAs from healthy individuals were used as population controls. Both patients and controls originated from central Sardinia. The frequencies comparisons between controls and cases were performed using chi-square test and Odds Ratio (OR) analysis with Cornfield 95% confidence intervals (CI). Results. Allele frequencies for BCL11A rs11886868 were 22% for the “C” allele and 78% for the “T” allele. No statistically significant difference was observed between cases and controls. All genotypes were in Hardy-Weinberg equilibrium for both patients and controls groups. The genotype frequencies were 65% (T/T), 26% (C/T) and 9% (C/C) in controls and 53% (T/T), 40.5% (C/T), and 6.5% (C/C) in hematological malignancies. When compared with the genotype frequencies reported for Caucasian and healthy controls from Sardinia no statistically significant difference was observed (p=0.4). However, the C/T genotype was more frequent in cases than controls (41% vs 26%) conferring an increased risk for hematological malignancies with an estimated OR=1,9 (95%CI 1.08-3.6; p=0.03). In detail, statistically significant differences in genotype distribution were observed in CTCL (p〈 0.0001), MPD (p=0.0006), NHL (p=0.008), HD (p=0.002) and ALL patients (p=0.02). The C/C genotype was not observed in CTCL and HD patients, while heterozygousity conferred an increased risk of 4.2 (2.3-7.7; p value

Description: Background. Array-based technology has been showing a great impact on clinical cancer cytogenetic, especially on genetically heterogeneous disease, such as MM, where relevant lesions might be the hallmarks of different patients’ subgroups, thus becoming of clinical relevance as well. We present herein the results of a molecular sub-study of the EMN02 phase III study (EMN02_HOVON95) which was designed to compare consolidation therapy Bortezomib, Melphalan and Prednisone versus upfront autologous stem cell transplantation, both applied after induction therapy with bortezomib-cyclophosphamide-dexamethasone (VCD). The sub-study was aimed at developing a comprehensive, high throughput genomic profile to be used to stratify uniformly treated MM patients according to their genomic background at baseline and to perform correlations with response to induction therapy. Patients and methods. Data obtained from 170 patients who consecutively entered the study and received three 21-day cycles of VCD induction therapy were analyzed. Baseline patients’ characteristics, including cytogenetic abnormalities, were comparable with those of 717 patients enrolled by participating Italian centres. Highly purified CD138+ bone marrow plasma cells were profiled by SNPs array (Affymetrix 6.0 and CytoScanHD® chip). ChAS (Affymetrix) and Nexus Copy NumberTM 7.5 (Biodiscovery) software were used to perform Copy Number Alterations (CNAs) analyses and clinical correlations, respectively. Results. After induction therapy, 66 out of 170 (38.8%) patients achieved a very good partial response (VGPR) or better, including 15 (8,8%) who attained a complete response (CR). On the contrary, 104/170 (61.1%) patients achieved

Description: Introduction Chronic Lymphocytic Leukemia (CLL) is a heterogeneous disease with a variable clinical course. Therapeutic decisions are based mainly on clinical grounds. Several prognostic markers based on genetic, phenotypic and molecular lesions have emerged in the past decade as relevant, alone or in combination with each other, in order to predict the clinical course and the best treatment option for CLL patients. Among these, NOTCH1 and TP53 mutation, have been described as new prognostic markers and predictive of survival in CLL. In this study, a real-life cohort of 165 CLL patients from Sardinia (Italy), was retrospectively analyzed for clinical, laboratory data (including NOTCH-1, TP53 mutation), first line treatment and followed up for survival. Methods Patients (99 males, 66 females, age ranged 31-89 y, 24% VH3〉VH2〉VH1〉VH7, whereas IGHV 3 was the most frequently used in the unmutated group, being expressed in 13 patients. IGHV 1-69 genes were present in 3 CLL patients overall (2 unmutated). Correlation between cytogenetic categories and mutation status showed that the poor prognostic marker 17p deletion was present in 4/36 (11,0%) of unmutated CLLs and in 2/55 (3,6%) of mutated cases while 13q14 deletion was statistically associated with the 45% of mutated cases (p=0.0089). Conclusion In our cohort of unselected patients NOTCH1 mutation didn’t affect outcome of CLL patients, while TP53 deletion/mutation and IGHV mutational status maintain a strong prognostic negative impact. Treatment heterogeneity is likely the reason of absence of difference in outcome in our study. In current clinical practice prognostic markers need yet to be validated in large clinical trials, not biased by stringent selection criteria. Disclosures: No relevant conflicts of interest to declare.

Description: Background: The role of Minimal residual disease (MRD) as surrogate for survival in Multiple Myeloma (MM) patients is well established. Therefore, new response criteria recommend including Multiparameter Flow cytometry (MFC) and next generation Sequencing (NGS) MRD negativity (minimum sensitivity of 1 in 105 nucleated cell) to deeply characterize complete remission (CR) [Kumar SK, Lancet Oncol 2016]. Here we analyzed and compared MRD data from the FORTE trial both by MFC and NGS techniques. Methods: Newly diagnosed MM patients ≤65 years were randomized to receive carfilzomib, lenalidomide, dexamethasone (KRd) induction - autologous stem cell transplantation (ASCT) intensification - KRd consolidation (KRd_ASCT); KRd12 and carfilzomib, cyclophosphamide, dexamethasone (KCd) induction - ASCT intensification - KCd consolidation (KCd_ASCT). Thereafter, patients were randomized to maintenance therapy with lenalidomide alone or lenalidomide plus carfilzomib. MRD evaluation was performed by 8-color second generation flow cytometry (sensitivity 10-5) in patients who achieved at least a very good partial response (VGPR) before maintenance. In a subgroup of these ≥ VGPR patients, next generation flow (NGF; sensitivity 10-5 - 10-6) was performed. Moreover, in patients achieving ≥ CR, MRD pre-maintenance was also assessed by NGS (Adaptive Biotechnologies, Seattle, WA; sensitivity 10-5 - 10-6). Therefore, only patients achieving ≥ CR have both MFC and NGS evaluations, and we focused on this ≥ CR population to compare MRD results with the 2 techniques. Spearman's rank correlation coefficient was used to measure the concordance between MFC and NGS. Fisher or Pearson's Chi-squared tests were adopted, where appropriate, to evaluate the statistical significance, at the level of 0.05. Results: ≥ CR pre-maintenance was achieved in 233 patients enrolled in the trial; at data cut-off, MFC and NGS data were available for 176/233 (76%) patients who were included in this analysis (62 received KRd_ASCT, 65 KRd12 and 49 KCd_ASCT). Median age of this ≥ CR population is 57 years (IQR 52-62), 14% have International Staging System (ISS) III stage and 26% high risk cytogenetics by FISH features [either del(17p) or t(4;14) or t(14;16)] reflecting baseline clinical features of the entire FORTE population. 139/176 (79%) ≥ CR patients were MFC negative at a cut-off of at least 10-5. Rate of MRD negative MFC among ≥ CR negative in the 3 arms were: 53/62 (85%) in KRd_ASCT, 51/65 (78%) in KRd12 and 35/49 (71%) in KCd_ASCT, reflecting the higher rate of MCF MRD negativity pre-maintenance reported by ITT in the overall population in the 2 KRD arms [Gay F, ASH 2018]. NGS negativity at a cut-off of at least 10-5 was detected in 87/176 (49%) of ≥ CR patients. Rate of MRD negative NGS at least 10-5 among ≥ CR negative in the 3 arms were: 35/62 (56%) in KRd_ASCT, 31/65 (48%) in KRd12 and 21/49 (43%) in KCd_ASCT. NGS negativity at a cut-off 10-6 was detected in 34/123 (28%) of ≥ CR patients (for 53/176 CR patients 10-6 sensitivity was not reached). Rate of MRD negative NGS at least 10-6 among ≥ CR negative in the 3 arms were:14/41 (34%) in KRd_ASCT, 10/44 (23%) in KRd12 and 10/38 (26%) in KCd_ASCT. Thereafter, we have analyzed the concordance between MRD results by the two techniques. Overall (samples at 10-5 and 10-6 sensitivity), Spearman's coefficient is 0.63 (p 〈 0.001); discordances between the two methods have been observed in 54/176 paired samples (30%). In particular, 53 samples (30%) are NGS positive but MFC negative, of which 34/53 (64%) have reached 10-6 sensitivity by NGS. Only 1 MFC positive sample is NGS negative. In a subgroup of patients evaluable both by NGS and NGF (26 paired samples, sensitivity 10-6), Spearman' s coefficient is 0.83 (p 〈 0.001), although a higher sample size is needed to confirm these preliminary concordant results. Conclusion: In patients who achieved ≥ CR, rate of at least 10-5 MRD MCF negativity was 79% and rate of at least 10-5 NGS negativity was 49%. Assessment both by MFC and NGS showed a good concordance, particularly if the same sensitivity is reached. Longer follow up is needed to assess the impact of MFC in comparison with NGS on patients' outcomes, particularly to evaluate if 10-6 NGS or NGF sensitivity may provide further clinical information, possibly identifying patients with very long survival or potentially cured. Disclosures Oliva: Celgene: Honoraria; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Adaptive Biotechnologies: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees. Belotti:Janssen: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees. Galli:Bristol-Myers Squibb: Honoraria; Takeda: Honoraria; Leadiant (Sigma-Tau): Honoraria; Janssen: Honoraria; Celgene: Honoraria. Offidani:Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees. Vozella:Amgen: Honoraria; Celgene: Honoraria. Zambello:Janssen: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees. Kirsch:Adaptive Biotechnologies: Employment. Jacob:Adaptive Biotechnologies: Employment, Other: shareholder. Ballanti:Amgen: Honoraria; Bristol-Myers Squibb: Honoraria; Janssen: Honoraria; Celgene: Honoraria. Corradini:Roche: Honoraria; Sanofi: Honoraria; KiowaKirin: Honoraria; Kite: Honoraria; Novartis: Honoraria, Other: Travel Costs; BMS: Other: Travel Costs; Servier: Honoraria; Takeda: Honoraria, Other: Travel Costs; Gilead: Honoraria, Other: Travel Costs; AbbVie: Consultancy, Honoraria, Other: Travel Costs; Amgen: Honoraria; Celgene: Honoraria, Other: Travel Costs; Daiichi Sankyo: Honoraria; Jazz Pharmaceutics: Honoraria; Janssen: Honoraria, Other: Travel Costs. Omedé:Janssen: Membership on an entity's Board of Directors or advisory committees. Cavo:takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: travel accommodations, Speakers Bureau; janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: travel accommodations, Speakers Bureau; bms: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; novartis: Honoraria; AbbVie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Musto:Amgen: Honoraria; Celgene: Honoraria. Boccadoro:Sanofi: Honoraria, Research Funding; Celgene: Honoraria, Research Funding; Amgen: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; AbbVie: Honoraria; Mundipharma: Research Funding. Gay:AbbVie: Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees. OffLabel Disclosure: The presentation includes discussion of off-label use of a drug or drugs for the treatment of multiple myeloma.

Description: Background The role of single vs double autologous stem cell transplantation (ASCT) in patients with newly diagnosed (ND) multiple myeloma (MM) needs to be prospectively investigated in the novel agent era. Methods The phase III EMN02/HO95 study was designed to compare (first randomization, R1) (stratification according to ISS stage) standard-dose intensification therapy with bortezomib-melphalan-prednisone (VMP) vs high-dose intensification therapy with melphalan at 200 mg/m2 (HDM) followed by ASCT after 3-4 cycles of bortezomib-cyclophosphamide-dexamethasone as induction therapy. A second randomization to consolidation therapy vs no consolidation was performed after intensification therapy, to be followed by lenalidomide maintenance until progression or toxicity in both arms. A primary study endpoint was progression-free survival (PFS) from R1. In centers with a policy of double ASCT, patients were randomized in a 1:1:1 ratio to either VMP or single ASCT (ASCT-1) or two sequential courses (administered 2 to 3 months apart) of HDM and double ASCT (ASCT-2) in order to prospectively compare ASCT-1 with ASCT-2, which was an additional study objective. Results From February 2011 to April 2014, 1510 pts aged ≤65 years with symptomatic NDMM were registered and 1192 of these were eligible for R1. According to the design of the study, 614 eligible patients who received the diagnosis of MM in centers with a double intensification policy were randomly assigned to either VMP (n=199) or ASCT-1 (n=208) or ASCT-2 (n=207). Patients randomized to ASCT-1 or ASCT-2 were included in the current pre-specified analysis. Median age was 59 years for patients in the ASCT-1 group and 57 years for those in the ASCT-2 group. The frequency of ISS stage III was 18% and 19%, while revised ISS stage III was 9% and 11%, respectively. Cytogenetic abnormalities were detected by FISH analysis of CD138+ plasma cells. A high-risk (HR) cytogenetic profile, defined by t(4;14) and/or del(17p) and/or t(14;16) (HR cyto-3), was observed in 26% of evaluable patients who were randomized to ASCT-1 and in 21% of those randomly assigned to ASCT-2. If amp(1q) and/or del(1p) were added for the definition of high-risk disease, a HR cytogenetic profile that included at least 1 of the 5 above mentioned chromosomal abnormalities (HR cyto-5) was reported in 55% of evaluable patients in the ASCT-1 group and in 54% of those in the ASCT-2 group. Median follow-up from R1 was 27 (IQR: 20-35) months. On an intention-to-treat basis, the median PFS was 45 months in the ASCT-1 arm and was not yet reached for patients in the ASCT-2 arm; 3-year estimates of PFS were 60% and 73%, respectively (HR=0.66; 95% CI=0.45-0.96; P=0.030). PFS benefit with ASCT-2 was retained across predefined subgroups, including patients with β2-microglobulin 〉3.5 mg/L (HR=0.59; CI=0.34-0.99; P=0.005), bone marrow plasma cells 〉60% (HR=0.41; CI=0.22-0.77; P=0.006), LDH values above the upper limits (HR=0.52; CI=0.28-095; P=0.034), revised ISS stage II (HR=0.50; CI=0.31-0.80; p=0.004), HR cyto-3 (HR=0.49; CI=0.24-1.02; P=0.057) and HR cyto-5 (HR=0.57; CI=0.35-0.93; P=0.024). In a multivariate Cox regression analysis stratified by ISS stage, randomization to ASCT-2 (HR=0.62; CI=0.40-0.95; P=0.027) and HR cyto-5 (HR=2.63; CI=1.63-4.16; P

Description: Background: Multiple Myeloma (MM) is characterized by frequent and complex genetic abnormalities that contribute to the pathogenesis and its prognostic eterogeneity. There is evidence for two oncogenic pathways in the early development of clonal plasma cell disorder: i) non-hyperdiploid carring translocation of the immunoglobulin heavy-chain locus and various oncogenes ii) hyperdiploid tumors with infrequent IgH translocation. The MM clonogenic cell is positively selected during the development and reaction of the germinal center. The immunoglobulin gene (IG) repertoire in MM follows a pattern similar to that of the normal repertoire. However, available data from analysis of IGH and IGK/L genes according to cytogenetic aberrations are limited. In the present study we investigated the frequency and characteristics of IGK and incomplete DJH as well as complete VDJH rearrangements in parallel with chromosomal abnormalities in a series of untreated MM patients. Materials and Methods. Bone marrow aspirates were collected from 53 MM patients with a mean age of 69.6 (range 48–84) between 2003–2007. The serum monoclonal component was IgG and IgA in the 77% and 22% patients respectively; 1 patient presented with IgD k MM. Cytogenetics and FISH analysis were performed simultaneously in 37 MM. In 18 (50.5%) samples kariotype analysis was successful. Interphase FISH analysis was perfomed using a set of probes specific for RB-1 (13q14), D13S319 (13q14.3), IgH (14q32), and p53 (17p13.1) loci, t(4;14), t(14;16), t(11;14) and a multicolor probe set for detection of aneuploidy (Vysis, Downers Grove, IL, USA). Genomic DNA was isolated for clonality analysis. IGHV-J, IGHD-J, IGKV-J, IGKV-KDE, IGKJ-C-INTRON-KDE rearrangements were amplified by PCR and analyzed following the BIOMED-2 protocol. Results: Conventional cytogenetics allowed to detect 16 patients with a normal kariotype, 1 hyperdiploid kariotype with monosomy 13, 1 hyperdiploid kariotype with 3q21 deletion. FISH panel analysis resulted in 4 patients with hyperdiploid kariotype and 7 with abnormalities for RB-1 and/or D13S319. IGH rearrangements were detected in 3 patients and the t (4;14) was found in 1 case. The p53 deletion, t(11;14) and t(14;16) were not detected. The overall detection rate of clonality by amplifying VDJH and DJH rearrangements using family-specific primers was 90%. We found a high frequency (71.7%) of DJH rearrangements with DH3 segment under represented (4%). The DH7 segment was rearranged in the 15% of MM. Incomplete DJH and complete VDJH rearrangements were present at frequencies of 20% and 29.5%, respectively. IGK locus rearrangements were detected in 38 out of 53 MM and the 60% presented the non-productive IGKV-KDE and IGKJ-C-INTRON-KDE rearrangements. Parallel analysis of clonality pattern and chromosomal abnormalities showed that complete VDJH rearrangements were present in all hyperdiploid MM and in a small proportion (4/16) of the MM with normal karyotype. Conclusions: Our results confirm previous estimations about IgH repertoire usage. Despite the small numbers, our findings indicate that complete Ig rearrangements might be correlated with hyperdiploid MM. Combining cytogenetics and IgH clonality studies might help to identify distinct subgroups of MM and provide a framework for dissection of disease prognosis and clinical management. Research funded by Regione Autonoma Sardegna.

Description: BackgroundSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been responsible for the coronavirus disease 2019 (COVID-19) pandemic, which started as a severe pneumonia outbreak in Wuhan, China, in December 2019. Italy has been the first European country affected by the pandemic, registering a total of 300,363 cases and 35,741 deaths until September 24, 2020. The geographical distribution of SARS-CoV-2 in Italy during early 2020 has not been homogeneous, including regions severely affected as well as administrative areas being only slightly interested by the infection. Among the latter, Sardinia represents one of the lowest incidence areas likely due to its insular nature.MethodsNext-generation sequencing of a small number of complete viral genomes from clinical samples and their virologic and phylogenetic characterization was performed.ResultsWe provide a first overview of the SARS-CoV-2 genomic diversity in Sardinia in the early phase of the March–May 2020 pandemic based on viral genomes isolated in the most inner regional hospital of the island. Our analysis revealed a remarkable genetic diversity in local SARS-CoV-2 viral genomes, showing the presence of at least four different clusters that can be distinguished by specific amino acid substitutions. Based on epidemiological information, these sequences can be linked to at least eight different clusters of infection, four of which likely originates from imported cases. In addition, the presence of amino acid substitutions that were not previously reported in Italian patients has been observed, asking for further investigations in a wider population to assess their prevalence and dynamics of emergence during the pandemic.ConclusionThe present study provides a snapshot of the initial phases of the SARS-CoV-2 infection in inner area of the Sardinia Island, showing an unexpected genomic diversity.

Description: Background. In multiple myeloma (MM), the role of minimal residual disease (MRD) by multiparameter flow cytometry (MFC) and next-generation sequencing (NGS) is well established (H. Avet-Loiseau et al. IMW 2019, S. Oliva et al. EHA 2020). Aims. The aims of this analysis were the evaluation of (1) the rate of conversion from MRD-positivity (MRD-pos) to MRD-negativity (MRD-neg) with MFC and NGS during maintenance and (2) the impact on progression-free survival (PFS) and overall survival (OS) of MRD-neg with both techniques in different subgroups including different treatment arms. Methods. Newly diagnosed (ND)MM patients (pts) aged ≤65 years were randomized (R1) to receive carfilzomib (K)-lenalidomide (R)-dexamethasone (d) induction followed by autologous stem-cell transplantation (ASCT) and KRd consolidation (KRd_ASCT), 12 KRd cycles (KRd12), or K-cyclophosphamide(C)-d induction followed by ASCT and KCd consolidation (KCd_ASCT). After consolidation, pts were further randomized (R2) to KR vs R maintenance. MRD was assessed every 6 months (m) by 8-color second-generation flow cytometry (sensitivity 10-5) in pts with ≥very good partial response (VGPR). In pts achieving at least a complete response (≥CR), MRD was also assessed by NGS at the same time points (Adaptive Biotechnologies, Seattle, US-WA; sensitivity 10-5-10-6). Logistic regression analysis adjusted for International Staging System (ISS) stage (I vs II/III) and R1 was performed to evaluate the conversion rate from MRD-pos to MRD-neg during maintenance (KR vs R). PFS and OS of MRD-neg vs MRD-pos in the intention-to-treat (ITT) population were evaluated. For these analyses, MFC-pos pts included those who were positive by MRD plus those