Coexistance of Multiple Myeloma and Myelodysplastic Syndrome or Lymphoid Diseases At Diagnosis: Evidence for Two Clonal and Independent Chromose Abnormalities

Abstract Abstract 5063 Introduction, We describe the simultaneous presentation of multiple myeloma (MM) and yeloproliferative disorders (MPD) or lymphoid diseases (LD) at diagnosis. Therapy-related myelodysplasia (tMDS) occurring during the course of MM is generally believed as a result from hematopoietic stem cell-toxic therapies, such as ionizing radiation and alkylating agent-based chemotherapies (melphalan, nitrosoureas).Patients and methods, We study a total of 342 patients (151 F, 191 M; median age 68.1 years; range 42 to 93 Years), diagnosed with MM based on the International Staging System. The basis for inclusion of patients in this study was with previous untreated MM ones. The study was performed in accordance with the declaration of Helsinki. To determine whether chemotherapies for MM factors play the critical role in the development of secondary disease, simultaneously two different cultures were processed, an unstimulated 96 hours culture (U96HC) on whole BM(WBM), a short-time 24 hours culture (ST24HC) after CD138+ plasma cells (PCs) depleted on negative fraction (CD138- cells) of BM and the FISH was investigated on purified CD138+.All samples were enriched in PCs by the Automated Magnetic Cell Sorter (Miltenyi technology)proceeded with anti-CD138 specific antibodies applied. The CD138+ PCs and the CD138- cells were collected in different tubes. The CD138− cells were used for a ST24HC. FISH was performed on the purified CD138+, PCs with a recommended FISH panel (MM International Working Group). Screening was performed systematically for the following unbalanced alterations and reciprocal rearrangements: del(13)(q14)(D13S25), del(17)(p13)(TP53),+3(D3Z), +9(D9Z1), +15(D15Z14), t(4;14)(p16;q32)/IGH-FGFR3, t(11;14)(q13;q32)/IGH-CCND1 (Abbott).After observing the results of U96HC on whole BM (CD138+ and CD138− cells), ST24HC (CD138− cells) and FISH for each patient, two clone cytogenetically were distinct and unrelated chromosomal abnormalities were found in 40 (11.7%) of the 342 MM patients (6 F, 34 M; median age 74 years; range 42 to 87 Years) 34 had a MPD and 6 had a LD. A second immunophenotyping analysis confirmed the presence of those LD/MM simultaneous haematological malignancy. In the cases of the patients with MM/ MPD, the frequency of cytogenetic abnormality unrelated to the myeloma clone was respectively; the 20q deletion, detected for 13 the 34 patients, the 20q- is a sole abnormality for 12 cases and associated with a complex caryotype in 1 case. The trisomy of chromosome +8 was observed in 7 cases, the del(7q) or monosomy 7 in 5 cases, loss of gonosome Y in 4 cases, del(11) for 2 cases, translocation t(9;22) in one case, 5q abnormality in one case and trisomy 9 with JAK2 V617F mutation in one case. For the patients with MM/LD, 5 patients had a trisomy +12 and or trisomy +18 like sole abnormality or associated with others cytogenetics abnormalities and one patient had 6q deletion. Discussion, Whereas in the literature the most common cytogenetic abnormalities typifying MPD after alkylator-based therapy include partial or complete deletions of chromosomes 5, 7, and 20 as well as trisomy 8. In our study we observed those abnormalities with the same frequency for the patients had simultaneous MPD associated in untreated MM at diagnosis. Six patients had simultaneous LD and MM. The marginal zone lymphoma was confirmed for 3 patients. The CC observed a trisomy +12 for those three patients associated with +18 and +19 for 2 cases and del(13) and trisomy 3 for one among them. We demonstrated in untreated MM patients the coexistence of MM and MPD or LD at diagnosis with MPD-type or LD-type chromosome abnormalities within MM signature karyotype. We hence recommend that CC studies, 96 hours WBM, 24 hours on negative fraction CD138− cells and FISH on purified CD138+ PCs, the three should be an integral part of the evaluation of patients with MM at diagnosis into clinical trials using HDT is warranted to determine whether patients who are predisposed to developing tMDS/sAML, they can be identified prospectively. Disclosures: No relevant conflicts of interest to declare.

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Genome Wide Profiling Of Chromosomal Abnormalities In 37 Patients With Monoclonal Gammopathy Of Undetermined Significance (MGUS)

Blood ◽

10.1182/blood.v122.21.1874.1874 ◽

2013 ◽

Vol 122 (21) ◽

pp. 1874-1874

Author(s):

Aneta Mikulasova ◽

Vladimira Vallova ◽

Jan Smetana ◽

Henrieta Greslikova ◽

Renata Kupska ◽

...

Keyword(s):

Structural Changes ◽

Plasma Cells ◽

Array Cgh ◽

Chromosomal Abnormalities ◽

Conflicts Of Interest ◽

Multiple Displacement Amplification ◽

Chromosome 8 ◽

Genome Wide ◽

Technique Optimization ◽

Chromosomal Changes

Abstract Introduction The incidence of clonal DNA copy number alterations (CNAs) in plasma cells (PCs) is considered as one of the most important and independent prognostic factors in patients with multiple myeloma (MM). Also in the premalignancy MGUS, there are specific chromosomal changes in PCs such as del(13)(q14), IGHtranslocation, gain(1)(q21) and hyperdiploidy. However, not much is known about their significance in relation to malignant transformation. Also, MGUS research is further complicated by small number of malignant cells that could be obtained from patients. Aim Array-CGH technique optimization and DNA CNAs analysis in relation to prognosis at genome-wide level in MGUS patients. Material and methods We have analysed 37 MGUS patients (22M/15F; median age 62) using array-CGH, Agilent platforms “Human Genome CGH, 4×44K” (n=10) and “SurePrint G3 CGH+SNP, 4×180K” (n=27). DNA was isolated from separated PCs (using CD138, CD19 and CD56 markers) and amplified by multiple displacement amplification (MDA). Results CNAs were observed in 57% (21/37) MGUS patients. Numerical and structural CNAs were found in 46% (17/37) and 41% (15/37) MGUS patients, respectively. We distinguished two genetic subgroups similar to MM patients: hyperdiploid and non-hyperdiploid. Hyperdiploidy was present in 32% (12/37) MGUS patients. The most frequent whole-chromosome gains were: 9 (83%, 10/12), 19 (83%, 10/12), 3 (75%, 9/12), 11 (67%, 8/12) and 15 (58%, 7/12). Non-hyperdiploidy was detected in 68% (25/37) MGUS patients. In both, hyperdiploid and non-hyperdiploid subgroups we have found loss of chromosomes 13 (25% vs. 20%) and Y (17% vs. 4%). Loss of chromosome 8 was detected only in hyperdiploid subgroup (8%) and losses of chromosomes 21, 22 and X only in non-hyperdiploid subgroup (8%, 4% and 8%, respectively). Structural changes were significantly more often present in hyperdiploid than in non-hyperdiploid patients (67% vs. 28%, p<0.05). We have identified whole-arm chromosome changes 1q gain and 16q loss in both hyperdiploid and non-hyperdiploid patients (17% vs. 12%, 8% vs. 8%, respectively), but 16p gain was seen only in hyperdiploid patients (8%). Segmental chromosome changes were also present in both hyperdiploid and non-hyperdiploid patients (58% vs. 24%). Interestingly, we have detected more segmental losses (15 vs. 8) and gains (6 vs. 2) in hyperdiploid than in non-hyperdiploid patients, but size median of these losses (3.33 Mb vs. 19.6 Mb) and gains (36.1 Mb vs. 63.5 Mb) was smaller in hyperdiploid than in non-hyperdiploid patients. Moreover, we found one MGUS patient who has progressed to MM and required therapy after 6 months from MGUS diagnosis. Genome-wide profile of MGUS patient was especially unique by high number of structural changes (n=11) compared to other 15 MGUS patients with structural CNAs (median 1; 1 – 6). MGUS patient’s profile showed hyperdiploidy (gains of chromosomes 3, 5, 9, 11, 15 and 19), losses of chromosomes 8, 13 and Y, 7 segmental losses in areas 1p34.2-p13.1, 6p23, 6q12-q27, 7q36.3, 12p12.1-p11.23, 12q12, 12q21.2-q23.3 and 4 segmental gains in areas 6p25.3-p23, 6p23-p11.1, 6q11.1-q12, Xq21.33-q28. In addition, gains and losses, which included whole or large parts of chromosomes, showed unusual profile associated with other alterations. These findings have suggested a complex karyotype. Summary In our study we have optimized protocol of array-CGH analysis from amplified DNA and we have used it in first 37 MGUS patients. We found there are various chromosomal changes (numerical, whole-arm and segmental) in more than half of MGUS patients. Similar to MM, hyperdiploid and non-hyperdiploid genetic subgroups were identified. We also described one MGUS case with unique genome-wide profile indicating unfavourable prognosis. Support NT13492, NT13190, NT11154, OPVK CZ.1.07/2.3.00/20.0183, MSM0021622434, GAP304/10/1395 Disclosures: No relevant conflicts of interest to declare.

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CIC-Mutation As a Potential Molecular Mechanism of Acquired Resistance to Combined BRAF/MEK Inhibition in CNS Multiple Myeloma

Blood ◽

10.1182/blood-2018-99-117203 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 3181-3181 ◽

Cited By ~ 2

Author(s):

Matteo Claudio Da Via' ◽

Antonio Giovanni Solimando ◽

Andoni Garitano-Trojaola ◽

Santiago Barrio ◽

Nadine Rodhes ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Exome Sequencing ◽

Plasma Cells ◽

Conflicts Of Interest ◽

Critical Role ◽

Point Mutations ◽

Sequencing Depth ◽

Resistance Development ◽

Average Sequencing Depth

Abstract Central nervous system (CNS) involvement is an extremely rare extramedullary multiple myeloma (MM) manifestation, diagnosed in less than 1% of patients. It is considered an ultimate high-risk feature, associated with unfavorable cytogenetics, and, even with intense treatment applied, survival is short, reaching less than 12 months in most cases. In June 2017 an 81 years old male with a κ light chain MM was referred to our institution for an isolated CNS MM relapse. His cerebrospinal fluid (CSF) demonstrated a high load of clonal plasma cells, however, the patient's bone marrow infiltration was very little with a percentage of plasma cells less than 5%. Imaging, including gold standard MRI and experimental 11C-methionine PET scan, was performed, and high metabolic activity was detected supra- and infratentorially as well as in the right femur and the clivus. Following CD138+ cell purification we analyzed the specimen with M3P (v3.0) a disease specific in-house customized, next generation targeted sequencing panel for MM (Ion torrent platform). This includes most commonly mutated MM genes, actionable drug targets and drug resistance associated genes. The average sequencing depth increased 700X and spatial MM heterogeneity was detected, as the CFS cells harbored a clonal BRAFV600E mutation, absent in the bone marrow. Initial intrathecal and systemic chemotherapy with Cytarabine and Thiotepa was intolerable, thus the patient underwent a combined target inhibition with Dabrafenib/Trametinib, well known specific BRAF and a MEK 1/2 inhibitors. The patient displayed a rapid complete response (Figure. 1A), however, disease relapse occurred after three months of therapy. We obtained a sequential CFS sample and Whole Exome Sequencing (Illumina platform) was applied to pre and post therapy CFS sampling. Exome sequencing of the two time points performed an average sequencing depth of 115X; a total number of 97 non-silent coding variants (missense, nonsense, indels, splice) with an allele frequency higher than 5% were detected. In detail, 19 point mutations were acquired at relapse, including a subclonal missense mutation in CIC (p.A984P, VRF 17%), recently identified as a candidate gene contributing to MEK/BRAF resistance development. Next, we established a CIC knock-down model electroporating a specific anti-CIC siRNA into U266 MM cell line. We cultured the silenced and not-silenced cells with Trametinib and Dabrafenib, either as single agents, or in combination. As expected, we observed resistance induction to the combination of the two drugs (Row Factor 85.94%; P<0.0001, Two-way ANOVA) suggesting a critical role for this patient derived mutation for his MEK/BRAF resistance development (Figure 1C, D). In order to better clarify the landscape pathway related to CIC we analyzed expression data from 647 patients enrolled in the MMRF CoMMpass trial. Remarkably, we found a significant down-regulation of ERF and ETV6 (t-test -9.95, -9.93, P <0.001, respectively), two well characterized tumor suppressor genes correlated with the re-activation of the RAS downstream pathway (Figure 1B). This is the first report giving evidence for a potential role of point mutations in CIC as a resistance mechanism to targeted MEK/BRAF inhibition in BRAF mutated MM. The performed pathway analysis significantly extends the insights of the resistance mechanisms highlighted. Our results foster a statistically powered study to corroborate the clinical relevance. Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.

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Multiple Myeloma: Clinical Implications of Cytogenetic Aberrations on Selected Plasma Cells on Conventional Cytogenetics and Interphase Flourescence In Situ Hibridization (FISH)

Blood ◽

10.1182/blood.v116.21.4987.4987 ◽

2010 ◽

Vol 116 (21) ◽

pp. 4987-4987 ◽

Cited By ~ 1

Author(s):

Rakesh Verma ◽

Lalit Kumar ◽

Ashutosh Halder ◽

Atul Sharma ◽

Ritu Gupta ◽

...

Keyword(s):

Multiple Myeloma ◽

Plasma Cells ◽

Chromosomal Abnormalities ◽

Conflicts Of Interest ◽

Response To Therapy ◽

Novel Agents ◽

Cytogenetic Aberrations ◽

Bone Marrow Plasma ◽

Conventional Cytogenetics

Abstract Abstract 4987 Purpose: We prospectively studied patients of multiple myeloma (MM) for presence of t(14q32), and del13q14 on FISH and numerical chromosomal abnormalities on conventional cytogenetics(CC). Methods: Between February 2007 and June 2009, 90 previously untreated patients of MM were enrolled, Patient's median age was 55 years (range, 34 to 75 years) and 67 were males (M: F: 2.9: 1). 24 patients had ISS stage I, stage II-46 & 20 patients had stage III. Purified bone marrow plasma cells, using magnetic activated cell sorter with CD138 micro beads were used. Conventional cytogenetics for numerical chromosomal abnormalities and FISH for deletion 13q14 (RB-1) and 14q32 (IgH) translocations was carried out on these purified plasma cells. 65/90 (72%) patients received therapy; using novel agents, n=54 (thalidomide-dexamethasone (n=50), Lenalidomide-dexamethasone (n=4)) and melphalan, prednisolone and thalidomide (MPT, n=11). Response was evaluated post 4 cycles using EBMT response criteria. Results: Metaphases could be obtained on CC in 53/90 patients (58%); with numerical abnormalities in 21: hyperdiploid -16 & hypodiploid in 5. Del 13q14 and t (14q32) by FISH was present in 51 (56%) and 75 (83%) patients, respectively. t (14q32) was present more in patients with ISS-III (p<.008). Presence of del13q14 correlated with BM plasma cells >40% (P<.007) and Hb ≤9.2G/dl (p=.06). Following therapy, 49/65 patients (75.2%) achieved significant response (CR+VGPR+PR). Patients aged ≤55 years (P = 0.07), those with ISS- I (P <0.05), IgG-kappa (P<0.04) and with absence of del13q14 (P = 0.04) responded better. The median overall survival (OS) has not reached yet, median event-free survival (EFS) is 33 months (95% CI 18–48). Estimated OS and EFS at 2 years is 77% and 50 %, respectively. Durie Salmon stage IIIB (HR 3.6) and response to therapy (HR 7.8) were significant predictors of OS. For EFS - del13q14 (HR 2.7, P <0.036), response to therapy (HR 2.7,P<0.01) and use of novel agents (HR 3.1, (P<0.008) were important predictors. Conclusion: Chromosomal abnormalities including IgH translocations and del13q14 with numerical chromosomal abnormalities are common in MM patients at diagnosis and can predict the treatment outcome and survival. Disclosures: No relevant conflicts of interest to declare.

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Extramedullary Relapse of Multiple Myeloma - Plasma Cells Characteristics.

Blood ◽

10.1182/blood.v120.21.2935.2935 ◽

2012 ◽

Vol 120 (21) ◽

pp. 2935-2935

Author(s):

Ludek Pour ◽

Sabina Sevcikova ◽

Lucie Rihova ◽

Lenka Kubiczkova ◽

Henrietta Greslikova ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Soft Tissues ◽

Plasma Cells ◽

Chromosomal Abnormalities ◽

Conflicts Of Interest ◽

Fish Analysis ◽

Extramedullary Relapse ◽

Igh Rearrangement ◽

Lower Expression

Abstract Abstract 2935 Background: Multiple myeloma (MM) is the second most common hematological malignancy in the world. The introduction of new drugs (thalidomide, bortezomib, revlimid) has dramatically improved survival of MM patients, but MM still remains an incurable disease. Unfortunately, an increase in the incidence of extramedullary relapse of MM (EM), an aggressive mostly resistant entity with abysmal prognosis for patients has been reported. EM can affect any area of tissue - soft tissue involvement can be with or without relationship to bone. A recent study of 936 MM patients by Usmani et al (2012) reported presence of EM in the skin and soft tissues at the time of diagnosis while liver involvement was common at relapse or progression. Aims: The objective of this study was to evaluate cytogenetic and flowcytometric data of available set of EM patients, and also to compare characteristics of plasma cells isolated from bone marrow and the extramedullary tumor. Material and methods: In total, we evaluated 29 EM patients. Patients' characteristics were as follows: males/females 18/11, median age was 61.2 years, ISS stage I/II/III 1/5/23, IgG/IgA/ LC only 20/6/3. I-FISH analysis was performed on bone marrow (BM) samples obtained at the time of diagnosis of EM. Flowcytometric analysis was performed on plasma cells (PC) isolated from BM as well as the EM tumor. Results: Using flowcytometry, PC were identified as CD138+CD38+ leukocytes and surface expression of CD20, CD27, CD28, CD33, CD40, CD54, CD117, CD19 and CD56 were analysed on PC in whole BM and the tumor. We found statistically significant decrease of CD27 (60.0 vs. 9.1% positivity in BM vs. tumor, resp.; p<0.02) and CD19 (35.0 vs. 8.3%; p=0.001). Other markers were non-significantly decreased: CD33 (27.3 vs. 12.5%), CD40 (84.6 vs. 75.0%), CD54 (84.6 vs. 50.0%), CD117 (26.7 vs. 16.7%), CD56 (70.0 vs. 58.3%) while expression of CD28 was increased (13.3 vs. 33.3%) on tumor PC compared to BM PC. In the BM PC, we found del(13)(q14) in 67% (18/27), del(17)(p13) in 22% (6/27), IGH rearrangement in 58% (11/19), t(4;14) in 33% (6/18), 1q21 gain in 58% (15/26), hyperdiploidy in 43% (10/23) of EM patients. The total number of aberrations per patient was: 0–1 aberration in 31%, 2–3 aberrations in 62%, 4 aberrations in 7% of MM patients BM. For 4 patients, we were able to analyze both BM and the EM tumor. We found that in 2/4 patients, there was no agreement in chromosomal abnormalities found in the BM and EM tumor. The differences were in del(13)(q14) and IGH rearrangement. del(13)(q14) was present in all 100% (4/4) samples of BM but only 75% (3/4) of EM tumors. del(17)(p13) was present in 25% (1/4) of patients in the BM as well as EM. IGH rearrangement was present in 75% (3/4) of BM but only 25% (1/4) of EM. 1q21 gain was present in 50% (1/2) of patients in the BM and EM and hyperdiploidy was not present in the BM or EM tumor (0/2). Conclusion: Chromosomal abnormalities connected to worse prognosis are more common in EM patients. PC phenotype seems to be different in cells obtained from BM and EM tumor. PC from EM tumor had significantly lower expression of CD27 and CD19. CD27 is a tumor necrosis factor receptor and plays a key role in regulating B-cell activation and immunoglobulin synthesis. Its low expression could be one of the main reasons for resistance in MM while loss of CD19 can create a proliferative advantage for the malignant plasma cell clone. Other interesting markers are CD54 and CD56 which were non-significantly decreased. CD54 also known as ICAM-1 plays a key role in stabilizing cell-cell interactions and migration, and CD56 (NCAM) is important for adhesion of PC to the bone marrow microenvironment. CD54 and CD56 lower expression may be the reason for EM development in MM but their role needs to be further elucidated. Acknowledgment: This study was supported by grants NT12130, MSM0021622434, NS10207, NT11154. Disclosures: No relevant conflicts of interest to declare.

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The Chromosomal Abnormalities Del(17p), t(4;14), and +1q21 Predict Progression From Smoldering to Symptomatic Multiple Myeloma

Blood ◽

10.1182/blood.v120.21.1806.1806 ◽

2012 ◽

Vol 120 (21) ◽

pp. 1806-1806 ◽

Cited By ~ 1

Author(s):

Kai Neben ◽

Anna Jauch ◽

Thomas Hielscher ◽

Jens Hillengass ◽

Nicola Lehners ◽

...

Keyword(s):

Multiple Myeloma ◽

Risk Score ◽

Chromosomal Aberrations ◽

Plasma Cells ◽

Chromosomal Abnormalities ◽

Conflicts Of Interest ◽

Statistical Significance ◽

Entire Group ◽

Fish Analysis ◽

Prognostic Impact

Abstract Abstract 1806 Background: Smoldering Multiple Myeloma (SMM) is a plasma cell disorder defined by the presence of ≥10% plasma cells in bone marrow and/or a monoclonal protein level of ≥3 g/dl in serum without organ damage. The aim of the study was to analyze the prognostic impact of chromosomal aberrations on time to progression (TTP) from SMM to symptomatic MM. Design and Methods: For selection of the patients, we used the same criteria as previously described by Kyle (Kyle et al., NEJM, 2007). We analyzed the prognostic value of 5 chromosomal abnormalities and hyper-/non-hyperdiploidy (HD and NHD, respectively) in a series of 231 patients with SMM by fluorescent in situ hybridization (FISH). Gains of at least 2 of the 3 chromosomes 5, 9, and 15 defined HD status. Results: Interphase-FISH analysis on CD138-enriched plasma cells detected gains of chromosomes 1q21 (29.4%) as well as deletions of chromosomes 13q14 (19.3%) and 17p13 (6.1%). Furthermore, the IgH-translocations t(4;14) and t(11;14) were observed in a frequency of 9.2% and 22.3%, respectively. The presence of t(4;14) was correlated with the serum heavy chain IgA (p<0.001). For the entire group, the median TTP was 4.9 years (95% CI, 3.9 – NA). Of all analyzed chromosomal abnormalities, del(17p13), t(4;14), and +1q21 showed a significant impact on TTP, whereas the presence of t(11;14) and del(13q14) was of no statistical significance. The median TTP for patients with del(17p13) was 2.7 years (vs. 4.9 years without, p=0.019), with t(4;14) 2.9 years (vs. 5.2 years without, p=0.021), and with +1q21 3.7 years (vs. 5.3 years without, p=0.013), respectively. In addition, HD was associated with a statistically shorter median TTP of 3.9 vs. 5.7 years in patients with NHD, respectively (p=0.036). A multivariate analysis identified t(4;14), +1q21, HD, reduction of uninvolved immunoglobulins (no.), and the risk score defined by Kyle et al. as independent factors for adverse outcome. Conclusions: The study shows that the overall risk of progression in SMM is significantly influenced by markers for tumor burden (i.e. Kyle risk score) as well as the presence of the chromosomal aberrations del(17p13), t(4;14), and +1q21. Our findings provide evidence that specific chromosomal aberrations are not only associated with early tumor progression and drug resistance in patients with overt MM but also to drive transition from asymptomatic into symptomatic stage of disease. Disclosures: No relevant conflicts of interest to declare.

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Interphase cytogenetic analysis of in vivo differentiation in the myelodysplasia of Down syndrome

Blood ◽

10.1182/blood.v84.7.2278.2278 ◽

1994 ◽

Vol 84 (7) ◽

pp. 2278-2282 ◽

Cited By ~ 15

Author(s):

A Zipursky ◽

H Wang ◽

EJ Brown ◽

J Squire

Keyword(s):

Down Syndrome ◽

Chromosomal Abnormalities ◽

Chromosome 8 ◽

Chromosome 7 ◽

Leukemic Cells ◽

Trisomy 8 ◽

Monosomy 7 ◽

Megakaryoblastic Leukemia

Abstract In Down syndrome, acute megakaryoblastic leukemia (AMKL) occurs frequently during the first 4 years of life and is usually preceded by a period of myelodysplasia (MDS), often associated with chromosomal abnormalities. Archival peripheral blood and/or bone marrow films of six patients with Down syndrome and MDS whose leukemic cells contained monosomy 7 or trisomy 8 were studied to determine whether the abnormal precursors produce mature cells in vivo. Using fluorescence in situ hybridization (FISH) of interphase nuclei with chromosome-specific centromere probes for either chromosome 7 or 8, we were able to determine which cells had one, two, or three signals indicative of one, two, or three no. 7 or 8 chromosomes. In five patients with trisomy 8, 80% to 100% (94.5% +/- 6.2%) of the megakaryoblasts had three signals using a chromosome 8 probe; in one patient with monosomy 7, 96.5% of the megakaryoblasts had one signal using a chromosome 7 probe. In all six patients, the myeloid and lymphoid series did not have evidence of the chromosomal abnormality present in the blasts. In three of five patients with trisomy 8, three signals were observed in 27%, 33%, and 41% of normoblasts, respectively. These data are evidence that the abnormal cell in MDS is a progenitor cell with the potential of forming cells of megakaryocyte and erythroid lineages.

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cIg-FISH On Patients with Multiple Myeloma – A Modified and Simple Technique Easily Incorporated Into Routine Clinical Service

Blood ◽

10.1182/blood.v120.21.4792.4792 ◽

2012 ◽

Vol 120 (21) ◽

pp. 4792-4792

Author(s):

Leena Gole ◽

Adeline Lin ◽

Constance Chua ◽

Sok Peng Chua ◽

Hui Bao Ma ◽

...

Keyword(s):

Multiple Myeloma ◽

Bone Marrow ◽

Bone Marrow Aspirate ◽

Plasma Cells ◽

Chromosomal Abnormalities ◽

Conflicts Of Interest ◽

Clinical Cytogenetics ◽

Set Up ◽

Selection Of

Abstract Abstract 4792 Chromosomal abnormalities associated with unfavorable prognosis in multiple myeloma patients include involvement of the TP53, FGRFR3 and MAF genes. It is important to accurately detect these chromosomal abnormalities to aid clinicians in the decision of therapeutic plans. Chromosomal abnormalities are not detected by traditional karyotyping due to low proliferative rate of myeloma cells. Conventional Fluorescence In-situ Hybridization (FISH) enhances the sensitivity but lacks the specificity as it does not distinguish plasma cells (PC) from the other hematopoetic cells. This can be overcome by identification of PCs by cytoplasmic immunoglobulin staining followed by FISH (cIg-FISH). Currently, cIg-FISH is done by two methods – lysing red cells in the bone marrow aspirates and spinning the pellet onto charged slides using the Cytospin machine (Ahmann et al Cancer Genet Cytogenet 1998). The second technique uses cultured and fixed cells stored in Carnoy's fixative (Filkova et al 2006 http://www.myeloma.cz/res/file/archiv/2007-cytogen-sbornik-workshop.pdf). This second technique is more easily incorporated into the routine cytogenetic protocols used for chromosomal analysis. However, clumping and the small size of plasma cells seemed to be a major setback with the protocol. We have made minor modifications to this technique, with a different approach to fixing and dropping the cells on to the slides to give nicely separated plasma cells. These when subjected to immunostaining with kappa or/and lambda antibodies, followed by FISH, result in easily identifiable plasma cells with good bright signals under the fluorescence microscope. Twenty samples from patients with multiple myeloma were subjected to routine FISH, cIg-FISH, chromosomal karyotyping along with flow cytometry and the results were compared. Three FISH probes from Vysis for t(4;14), t(14;16) and deletion of TP53 were used. To test the utility of the technique for stored cell pellets, 4 fixed pellets stored from 2005–2008 were also tested with good results. Table 1. Comparative analysis of 20 patients with multiple myeloma by three different methods No % Plasma cells Conventional Karyotyping Conventional FISH Conventional cIg FISH 1 47 Normal Normal Normal 2 55 Monosomy 14 Monosomy 14 Monosomy 14 3 71 der(1;16) Monosomy 16 Monosomy 16 4 6 Normal Normal Normal 5 2 Normal Normal Normal 6 72 der(12;16) Monosomy 16 Monosomy 16 7 13 Normal Normal Trisomy 14 8 52 Normal Normal Normal 9 8 Monosomy 17 t(4;14),del17p t(4;14),monosomy 16, del17p 10 25 Normal Normal Normal 11 1 Normal Normal Normal 12 31 Normal Trisomy 4 Trisomy 4 13 Not done Not done Normal Normal 14 5 Normal Trisomy 17 Monosomy 14, Trisomy 17 15 19.5 Monosomy 14 t(4;14), monosomy 16 t(4;14), monosomy 16 16 34 Normal Normal Normal 17 50 Normal Normal Normal 18 1.8 Normal Normal Normal 19 28 Normal Normal Normal 20 47 Normal Normal Normal The percent of plasma cells seen in bone marrow aspirates ranged from 1.8–71%. Of 20 samples, 3 samples showed a positive cIg-FISH but a normal conventional FISH result. All samples showed a uniformly higher percent of abnormality with the cIg-FISH protocol due to selection of plasma cells as expected. Figure 1. Kappa/lambda positively stained plasma cells showing FGFR3/IGH translocation Figure 1. Kappa/lambda positively stained plasma cells showing FGFR3/IGH translocation In conclusion, this technique using fixed cells will be a major asset to all cytogenetic laboratories as It saves sample as the entire quantity of bone marrow aspirate can be used to set up cultures for chromosomal karyotyping and part of the fixed pellet can be used for cIg-FISH.There is no necessity of making Cytospin slides - saving time and effort.Stored pellets, especially if cells are well stored and preserved in fixative can be used for retrospective analysis. The technique can be easily incorporated as a routine targeted FISH test for MM samples, which is not practiced in most clinical cytogenetics laboratories at present. Disclosures: No relevant conflicts of interest to declare.

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Interphase cytogenetic analysis of in vivo differentiation in the myelodysplasia of Down syndrome

Blood ◽

10.1182/blood.v84.7.2278.bloodjournal8472278 ◽

1994 ◽

Vol 84 (7) ◽

pp. 2278-2282

Author(s):

A Zipursky ◽

H Wang ◽

EJ Brown ◽

J Squire

Keyword(s):

Down Syndrome ◽

Chromosomal Abnormalities ◽

Chromosome 8 ◽

Chromosome 7 ◽

Leukemic Cells ◽

Trisomy 8 ◽

Monosomy 7 ◽

Megakaryoblastic Leukemia

In Down syndrome, acute megakaryoblastic leukemia (AMKL) occurs frequently during the first 4 years of life and is usually preceded by a period of myelodysplasia (MDS), often associated with chromosomal abnormalities. Archival peripheral blood and/or bone marrow films of six patients with Down syndrome and MDS whose leukemic cells contained monosomy 7 or trisomy 8 were studied to determine whether the abnormal precursors produce mature cells in vivo. Using fluorescence in situ hybridization (FISH) of interphase nuclei with chromosome-specific centromere probes for either chromosome 7 or 8, we were able to determine which cells had one, two, or three signals indicative of one, two, or three no. 7 or 8 chromosomes. In five patients with trisomy 8, 80% to 100% (94.5% +/- 6.2%) of the megakaryoblasts had three signals using a chromosome 8 probe; in one patient with monosomy 7, 96.5% of the megakaryoblasts had one signal using a chromosome 7 probe. In all six patients, the myeloid and lymphoid series did not have evidence of the chromosomal abnormality present in the blasts. In three of five patients with trisomy 8, three signals were observed in 27%, 33%, and 41% of normoblasts, respectively. These data are evidence that the abnormal cell in MDS is a progenitor cell with the potential of forming cells of megakaryocyte and erythroid lineages.

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Genome Instability in Multiple Myeloma: Facts and Factors

Cancers ◽

10.3390/cancers13235949 ◽

2021 ◽

Vol 13 (23) ◽

pp. 5949

Author(s):

Anna Y. Aksenova ◽

Anna S. Zhuk ◽

Artem G. Lada ◽

Irina V. Zotova ◽

Elena I. Stepchenkova ◽

...

Keyword(s):

Multiple Myeloma ◽

Complex Disease ◽

Plasma Cells ◽

Genome Instability ◽

Hematologic Malignancy ◽

Point Mutations ◽

Malignant Neoplasm ◽

Chemotherapeutic Agents ◽

Hematopoietic Stem ◽

Social Burden

Multiple myeloma (MM) is a malignant neoplasm of terminally differentiated immunoglobulin-producing B lymphocytes called plasma cells. MM is the second most common hematologic malignancy, and it poses a heavy economic and social burden because it remains incurable and confers a profound disability to patients. Despite current progress in MM treatment, the disease invariably recurs, even after the transplantation of autologous hematopoietic stem cells (ASCT). Biological processes leading to a pathological myeloma clone and the mechanisms of further evolution of the disease are far from complete understanding. Genetically, MM is a complex disease that demonstrates a high level of heterogeneity. Myeloma genomes carry numerous genetic changes, including structural genome variations and chromosomal gains and losses, and these changes occur in combinations with point mutations affecting various cellular pathways, including genome maintenance. MM genome instability in its extreme is manifested in mutation kataegis and complex genomic rearrangements: chromothripsis, templated insertions, and chromoplexy. Chemotherapeutic agents used to treat MM add another level of complexity because many of them exacerbate genome instability. Genome abnormalities are driver events and deciphering their mechanisms will help understand the causes of MM and play a pivotal role in developing new therapies.

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Successful treatment of gastric relapse in multiple myeloma with bortezomib after autologous hematopoietic stem cell transplantation (autoHSCT)

Mediterranean Journal of Hematology and Infectious Diseases ◽

10.4084/mjhid.2013.006 ◽

2013 ◽

Vol 5 (1) ◽

pp. e2013006

Author(s):

Serdar Sivgin

Keyword(s):

Multiple Myeloma ◽

Stem Cell ◽

Hematopoietic Stem Cell Transplantation ◽

Stem Cell Transplantation ◽

Cell Transplantation ◽

Hematopoietic Stem Cell ◽

Plasma Cells ◽

University Hospital ◽

Hematopoietic Stem ◽

Relapsed Disease

We report a case of 59-year-old Turkish man with history of MVR and COPD whom was diagnosed with stage IIIA IgG lambda multiple myeloma (MM) in 1997. He underwent autologous hematopoietic stem cell transplantation following melphalan 200mg per body area(m2)in February 2006. On 18th of February 2011; he was admitted to the emergency service of university hospital with complaints of hematemesis and melena. In gastric biopsy obtained from the lesion; pathological evaluation showed monoclonal lambda light chain infiltration originated from neoplastic plasma cells in gastric mucosa. The patient was considered as local gastric relapsed disease and was treated with 2 cycles of bortezomib. An excellent response was achieved after 2 cycles of BEP regimen, his paraprotein levels dropped below 10 g/L and there was no recurrence of the hematemesis or melena.

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