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Identification of a balanced complex chromosomal rearrangement involving chromosomes 3, 18 and 21 with recurrent abortion: case report

Yaping liao, changqing liu.

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Received 2014 Mar 19; Accepted 2014 Jun 2; Collection date 2014.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/4.0 ), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/ ) applies to the data made available in this article, unless otherwise stated.

Complex chromosome rearrangements (CCRs) are constitutional structural rearrangements involve more than two breakpoints on two or more chromosomes. Balanced CCR carriers are often phenotypically normal but associated with high risk of spontaneous abortion and having abnormal offspring with unbalanced karyotype. Here, we report a new familial case of complex chromosome structural aberrations involving chromosomes 3, 18 and 21 and four breakpoints.

Cytogenetic investigations showed a complex chromosomal chromosome rearrangement involving chromosomes 3, 18 and 21 with four breakpoints. 2 of 4 breakpoints were within the long arm of chromosome 18. Three-color fluorescence in situ hybridization (FISH) confirmed the complexity of the rearrangement and showed the derivative 21 to be composed of 3 distinct segments derived from chromosomes 21, 18, and 3. The karyotype of CCR carrier was determined as 46,XX,t(3;21;18)(3pter → 3q12::18q23 → 18qter;21pter → 21q22.1::18q21.1 → 18q23::3q12 → 3qter; 18pter → 18q21.1::21q22.1 → 21qter).

A new complex balanced CCR was characterized using conventional high resolution banding and molecular cytogenetic analysis. The results provided an explanation of recurrent abortion and abnormal child for balanced CCR carriers. Genetic counselling and prenatal diagnosis for couples with a balanced CCR is necessary since they have a high risk of having a child with unbalanced karyotype. Additional studies to reveal the molecular mechanism of CCRs would help reveal the rule of inherited CCRs in offspring.

Keywords: Complex chromosomal rearrangements (CCRs), Recurrent spontaneous abortions, Genetic counseling, Fluorescence in situ hybridization

Complex chromosome rearrangements (CCRs) are structural aberrations involving at least three breakpoints on two or more chromosomes and exchange of genetic material between these chromosomes. Translocation, insertion and transposition are often involved in CCRs. CCRs are rare in humans and can be familial or de novo [ 1 , 2 ]. So far, ~255 cases of CCRs involving three or more chromosomes have been reported and most are de novo [ 3 ].

It has been observed that most balanced CCRs occur in females, and about half of them are inherited [ 4 ]. In males, balanced CCRs are often subfertile or sterile due to spermatogenesis disturbance [ 2 , 3 , 5 , 6 ]. Although balanced CCR carriers are not often associated with abnormal phenotypes, a high risk of miscarriage and live born child with an unbalanced karyotype are found. It is difficult to identify CCRs correctly by conventional cytogenetics based on banding techniques without the aid of additional diagnostic tools such as fluorescence in situ hybridization (FISH) or other advanced molecular cytogenetic techniques [ 7 - 10 ].

Here, we present a family with at least three unbalanced or balanced CCR carriers involving chromosomes 3, 18 and 21 using traditional high resolution banding and three-color FISH.

Case presentation

The proband (II-3), a 31-year old woman, and her husband were referred by cytogenetic investigation because the proband had four first-trimester miscarriages (Figure  1 A). The physical examination revealed that the proband (II-3) and her husband were phenotypically normal including their reproductive systems. The proband had a 7-year old son (III-4) with typical symptoms and physical characteristics of Down syndrome including mental retardation and physical growth delay [ 11 , 12 ]. He was born at 39 weeks of gestation with a weight of 3.1 kg and a length of 47 cm. He had a height of 103 cm and a weight of 18 kg at 7-year old.

Identification of a complex translocation involving chromosomes 3, 18 and 21. A : Pedigree of the proband’s family (arrow). B : GTG banded karyotype of the proband showing three derivative chromosomes. C : BAC-probes RP11-379C23 (green) (3q27. 2), RP11-190A24 (21q22.3) (orange) and RP11-89 N1 (red) (18q23) demonstrate a translocation among chromosomes 3, 21 and 18. D : BAC-probes TRP11-7H17 (18q23) (green), BAC-probe RP11-57 F7 (18q22.2) (red) and RP11-89H21(21q11.2) (orange) show the insertion of part of chromosome 18 in derivative chromosome 21.

The proband (II-3) had two sisters. The eldest sister, 43-year old, had one miscarriage (~22 weeks) and two phenotypically normal boys. The second elder sister (II-2), 40-year old, had eight first-trimester miscarriages and one 12-year old boy who had normal phenotype and normal or balanced karyotype. Recurrent abortion at first-trimester and one abnormal child occurred in this family suggested a possible chromosomal aberration.

The blood karyotype from the proband (II-3) revealed a translocation involving chromosomes 3, 21 and 18 (Figure  1 B). Additionally, it seems that a segment from 18q21 ~ q23 inserted to der 21(q22) when high resolution staining karyotype analysis was used, but it could not be karyotypically determined. The husband had a normal karyotype both by GTG banding and high resolution staining (data not shown). Analysis of the siblings revealed different cytogenetic anomalies. A sister (II-2) showed the same chromosome rearrangement as that of the proband (II-3), whereas the son (III-4) of the proband had unbalanced karyotype carrying not only the der (3), der (18) and der (21) but also two normal chromosome 21. So he was diagnosed as Down syndrome according to karyotype and typical symptoms.

To identify complex chromosomal rearrangements or subtle translocations, three-color FISH was performed with regional specific BAC-probes (Table  1 ) as described previously [ 13 ]. The BAC-probe RP11-379C23 (3q27. 2), RP11-190A24 (21q22.3) and RP11-89 N1 (18q23) located near to telomeres confirmed a translocation between chromosomes 3, 21 and 18 (Figure  1 C). The BAC-probe TRP11-7H17 (18q23) was used as a control, and it showed hybridization to the der (3) (Figure  1 D). However, the BAC-probe RP11-57 F7 (18q22.2) was used as a second control, it showed hybridization to the der (21) (Figure  1 D). Based on FISH and cytogenetic results, there are two breakpoints located in abnormal chromosome 18q. One breakpoint was located in 18q22.2 ~ 18q23, and the other one was located in 18q21.1. The middle fragment inserted in der (21q22) was derived from chromosome 18. Thus, the present case is not a simple three-way CCR and the karyotype was readjusted and assigned according to ISCN 2013 as follows: 46,XX,t(3;21;18)(3pter → 3q12::18q23 → 18qter;21pter → 21q22.1::18q21.1 → 18q23::3q12 → 3qter; 18pter → 18q21.1::21q22.1 → 21qter).

Fluorescence in situ hybridization analysis with the following probes used

Although CCRs are rarely found in screened populations, they are often associated with mental retardation, congenital abnormalities, recurrent spontaneous abortions and infertility [ 14 - 16 ]. The application of FISH and its derivative techniques facilitated the characterization of CCRs and become essential for further delineation of chromosomal breakpoints [ 3 , 17 ]. The aim of this study was to identify whether a complex abnormal karyotype was apparent or not, and whether possible breakpoints were involved in chromosomes using high-resolution chromosome analysis and FISH. This is especially important for prenatal diagnosis and appropriate genetic counseling [ 18 ].

In the present case, the proband (II-3) was found to carry balanced CCRs involving chromosomes 3, 18 and 21 with four breakpoints. The four breakpoints were located in 3q12, 18q21.1, 18q23 and 21q22.1. To the best of our knowledge, this is the first report of a balanced CCR involving chromosomes 3, 18 and 21 with 2 breakpoints on chromosome 18. By combining FISH and high resolution banding data, we changed the previous description of this CCR from a simple three-way translocation to the extra complex CCR according to the category of CCRs proposed by Kausch et al. [ 19 ] and Madan [ 4 ]. In this family, we proposed that one of the proband’s parent was carrying the balanced CCR since the complete balanced CCR was present in at least two of their offspring: II-2 and II-3 (Figure  1 A). It was previously observed that transmission of CCR was usually maternal since male carriers have an increased risk of primary infertility or subfertility [ 16 ]. We inferred the familial CCRs possibly inherited from her mother. However, this hypothesis could not be confirmed because her parent’s karyotype could not be studied due to the unavailability of blood samples.

It is difficult to obtain the exact rate of meiotic segregation from the ovum of balanced CCR carriers because most of ovum cannot be obtained. In an attempt to understand the mechanisms of meiotic segregation, Loup et al. [ 20 ] analyzed meiotic segregation in the sperm of a patient with a three-way familial CCR. They found a high rate of unbalanced sperm (75.9%) including 34.1% from 3:3 segregation, 38.2% from 4:2 segregation, 3.5% from 5:1 segregation and 0.05% from 6:0 segregation. Only 14.8% of sperm were normal or balanced. We think the mode of meiotic segregation may be various in different type of CCRs. Recently, Madan [ 4 ] reviewed the mode of segregation of 68 offspring of 63 CCR carriers. They found a clear difference between the simple type of CCR (three-way or four-way CCR) where 4:2 segregation was more frequent (14/26) and the extra complex CCRs (number of breaks > number of chromosomes) where adjacent 1 segregation took place in the majority of cases (39/42).In the present case with extra complex CCR, there are several possibilities for a high incidence of abnormal pregnancy outcome. At the pachytene stage, the CCR will form a possible hexavalent configuration different from three-way translocation (Figure  2 ). Generally, unbalanced 3:3, 4:2, 5:1 and 6:0 segregations often produce greater genomic imbalance, and early pregnancy loss are expected.. In addition, recombination in the inserted segment can result in gametes with new unbalanced karyotype. However, live born abnormal child with Down syndrome is possible though 4:2 segregation because chromosome 21 is involved in this CCR. The proband (II-3) had a son (III-4) who had unbalanced CCR with abnormal phynotype. According to G-banding karyotype, he gained the der (3), der(18), der(21) and normal chromosome 21 from his mother (Figure  2 ). Therefore, we confirmed that the mode of meiotic segregation is 4:2 in our patient.

Theoretical pachytene configuration. Pachytene diagram of the proband (II-3) and segregation mode of her son (III-4). The arrows indicate the direction of separation to each pole.

Genetic counseling for CCR carriers is very important and can be offered before and after pregnancy. Madan et al. [ 16 ] reviewed 60 familial and de novo cases of balanced CCRs, and estimated that carriers have a 50% risk of spontaneous abortion and a 20% risk of having a child with an unbalanced karyotype. Although published data are a good rough guide for counseling, we think it cannot be applied to individual CCRs as the risk estimates remains highly empirical. The category of CCRs and the number of chromosomes involved can vary greatly giving a wide variety of possible gametes [ 19 , 20 ]. In our case, 4/5 pregnancies ended in a spontaneous abortion and a child with unbalanced karyotype (1/5). One of the proband’s sister (II-2) with the same karoytpe had high frequent spontaneous abortion (8/9) and one phenotype normal child (1/9). A total of 12/14 pregnancies from two carriers ended in spontaneous abortion indicated an increased risk of miscarriage in this family higher than previously reported [ 16 , 21 ]. Considering the high incidence of abnormal pregnancies, natural pregnancy of the balanced carriers is not encouraged in this family. Even if the balanced carriers conceive a child naturally, prenatal diagnosis for balanced CCR carriers is necessary owing to an estimated risk of 7.1% (1/14) having a child with Down syndrome in this family. It has been thought preimplantation genetic diagnosis (PGD) is impossible for CCRs carries as the highly complexity of meiotic segregation. Recently, several studies reported that healthy live birth after successful PGD of CCR carries. These data indicate that CCRs are amenable to PGD analysis as well as egg donation, after a proper genetic counselling [ 22 - 24 ].

We report here a familial case of CCRs possibly inherited from her mother. The balanced CCRs resulted in recurrent abortion and an abnormal child with unbalanced karyotype. It is very important to identify the chromosomes and the breakpoints involved in CCRs as accurately as possible to understand the mechanism underlying the formation of CCRs and to provide correlation between phenotype and chromosomal aberration. With accurate characterization of CCRs, correct prenatal diagnosis and efficient genetic counseling can be made. Our data indicate that the extra complex CCRs have higher incidence of abnormal pregnancy outcome, and that it is difficult to predict the exact risk of having a child with unbalanced karyotype. Additional studies to reveal the molecular mechanism of CCRs would help reveal the rule of inherited CCRs in offspring.

Materials and methods

Karyotyping.

Cytogenetic analysis was performed on peripheral blood cultures after 72 h of incubation. Metaphase spreads were prepared for GTG banding and high-resolution staining according to standard procedures. Karyotypes were obtained from the proband, her husband, her son, and her second elder sister. The proband’s parent (I-1 and I-2) and three children (III-1, III-2 and III-3) of proband’s sisters could not be studied due to the unavailability of blood samples from these members. Twenty metaphases were analyzed from each subject by GTG banding. Olympus microscope (BX41) was used for karyotyping and metaphase images were captured using VideoTesT-Karyo software (Metasystems, Altlussheim, Germany).

To identify balanced CCR or subtle translocation, three-color FISH analysis was performed for the proband following conventional protocols as previously described [ 13 ]. Specific bacterial artificial chromosome (BAC) clones were selected from the human library RPCI-11 according to the UCSC Human Genome Assembly (UC Santa Cruz, USA, assembly February 2009) and provided by State Key Laboratory of Medical Genetics, Central South University. BAC DNA was directly labeled with spectrum Green, spectrum Orange and Spectrum Red-dUTP using nick translation (Table  1 ). The chromosomes were made fluorescent by 4′, 6-diamidino-2-phenylindole (DAPI). Each of the 20 metaphase spreads was analyzed using a fluorescence microscope (Leica DM5000B). Images were captured and processed by using Leica CW 4000 software.

Written informed consent was obtained from the patient for publication of this Case report and any accompanying images.

Abbreviations

CCRs: Complex chromosomal rearrangements; FISH: Fluorescence in situ hybridization; BAC: Bacterial artificial chromosome; PGD: Preimplantation genetic diagnosis.

Competing interests

The authors declare that they have no competing interests.

Authors’ contributions

LP and LQ drafted the paper and participated in the molecular cytogenetic analysis. WQ evaluated the family with examination and genetic counseling. ZD did the cytogenetic analysis. All authors read and approved the final manuscript.

Contributor Information

Yaping Liao, Email: [email protected].

Liqun Wang, Email: [email protected].

Ding Zhang, Email: [email protected].

Changqing Liu, Email: [email protected].

Acknowledgements

Research supported by the Scientific Research Foundation of Anhui Province, China (No. 1208085QC67). We thank all of the patient family members’ participation in this study. We thank Dr. Shuqi Wang for providing language help.

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Introduction to the Analysis of the Human G-Banded Karyotype

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There is an internationally agreed system for describing the banding pattern of chromosomes, such that if an abnormality is accurately described in one laboratory then it can be recognized in another. This is known as the ISCN, the International System for Human Cytogenetic Nomenclature. Since its first appearance in 1965, it has been tried, tested, and modified, and the 1995 edition remains the standard version in current use ( 1 ). This is an essential reference for the definition of cytogenetic abnormalities. Within the ISCN are the formal descriptions of how to describe chromosome bands and abnormalities. There is also a schematic representation of the human karyotype, and several illustrations of karyotypes of normal chromosomes, stained in different ways. Every cytogeneticist needs to become familiar with the correct way of describing chromosomes and their abnormalities.

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Swansbury, J. (2003). Introduction to the Analysis of the Human G-Banded Karyotype. In: Swansbury, J. (eds) Cancer Cytogenetics. Methods in Molecular Biology™, vol 220. Humana Press. https://doi.org/10.1385/1-59259-363-1:259

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Conventional Karyotyping and Fluorescence In Situ Hybridization for Detection of Chromosomal Abnormalities in Multiple Myeloma

Matthew crabtree, jennifer cai.

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Corresponding Author: Xin Qing, Hematology and Flow Cytometry Laboratories, Department of Pathology, Harbor-UCLA Medical Center, Torrance, CA 90502, USA. Email: [email protected]

Received 2022 Apr 22; Accepted 2022 Jun 18; Issue date 2022 Jun.

This article is distributed under the terms of the Creative Commons Attribution Non-Commercial 4.0 International License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Multiple myeloma (MM) is a genetically heterogeneous disease, with cytogenetic findings that determine disease behavior. Genetic abnormalities can be assessed by fluorescence in situ hybridization (FISH) analysis and/or G-banded karyotyping. The two methods produce unique and overlapping information, and the clinical utility of using both is investigated here.

Seventy patients diagnosed with MM at a hospital in Southern California were retrospectively reviewed for the FISH and G-banded karyotyping results obtained from bone marrow specimens.

Karyotype was normal in 71% (50/70), abnormal in 27% (19/70), and inadequate in 1% (1/70). Among patients with abnormal karyotype, FISH provided additional information about genetic aberrations in 95% of cases (18/19). Among cases with abnormal FISH, karyotype provided additional information about genetic aberrations in 27% of cases (18/66). When numerical abnormalities were present (detected by FISH and/or karyotype), FISH detected them in 95% (54/57), of which karyotype missed 70% (38/54) of the time. Karyotyping detected numerical abnormalities in 33% (19/57), which FISH missed 16% (3/19) of the time.

Conclusions

Karyotyping and FISH analysis in MM each provide unique information. For most patients, performing both tests together will provide more information than either test alone.

Keywords: Multiple myeloma, Cytogenetics, FISH, Karyotype

Introduction

Multiple myeloma (MM) is a clonal bone marrow disease characterized by the neoplastic transformation of differentiated B cells. MM represents about 10-15% of all hematopoietic neoplasms and 1% of all cancer cases. The median age at diagnosis is 60 years old. Prognosis has been steadily improving over the last two decades alongside the introduction of new therapeutic strategies, which has prolonged the median survival from 3 to 8 - 10 years [ 1 , 2 , 3 ].

MM is diagnosed by bone marrow biopsy showing clonal bone marrow plasma cells ≥ 10% or biopsy showing plasmacytoma, and any one or more of the following myeloma-defining events: hypercalcemia, renal insufficiency, anemia, and bone lesions [ 1 , 2 ].

Risk stratification in MM is a function of cytogenetic information and stratifies patients into three prognostic groups: high risk, intermediate risk, and standard risk [ 4 - 7 ]. This information can be acquired by conventional metaphase karyotyping or interphase fluorescence in situ hybridization (FISH) technology, and the two methodologies are partially overlapping in the information they provide of prognostic significance in MM [ 8 ]. We aimed to review a series of MM cases with both tests performed and summarize the extent to which these two tests provide unique information.

Materials and Methods

This research study was conducted retrospectively from data obtained for clinical purposes. Cytogenetic analysis was performed on 95 patients from 2013 to 2021 at Harbor-UCLA Medical Center in Southern California who had a bone marrow biopsy showing MM, monoclonal gammopathy of undetermined significance (MGUS), or residual disease after treatment. Of these 95, 70 satisfied the inclusion criteria of this study: clonal plasma cell percentage ≥ 10%, a diagnosis of MM for the first time, and both karyotyping and FISH performed on the corresponding specimen. Selected patients were aged 42 - 85 years (median 62) with 59% being male (41/70) and 41% being female (29/70) ( Table 1 ). The 25 reviewed patients who were not included had a plasma cell percentage of < 10% (n = 17) or karyotyping was not performed (n = 20).

Table 1. Demographics.

ISS: International Staging System; R-ISS: revised International Staging System.

Karyotyping was performed at Quest Diagnostics Nichols Institute (San Juan Capistrano, CA). Fresh bone marrow aspirate samples were cultured as 48- and 72-h unstimulated cultures following standard cytogenetic methods. To increase the mitotic index of the cultures, stimulation with 2 µg/mL phorbol 12-myristate 13-acetate and 200 µL/mL phytohemagglutinin was also used. Chromosomal analysis was performed on cultured bone marrow samples using the standard G-banding technique. At least 20 metaphase cells were used for karyotyping.

FISH studies were performed at Quest Diagnostics Nichols Institute (San Juan Capistrano, CA). Two hundred to three hundred cells were counted and CD138 enrichment was performed whenever possible. Probes that were used included IGH (14q32), TP53 (17p13.1), D17Z1 (17p10), FGFR3 (4p16.3), CCND1 (11q13), MAF (16q23) (Abbott Molecular), MAFB (20q12) (Cytocell), DLEU (13q14.3), LAMP1 (13q34), CKS1B (1q21), CHD5 (1p36) (Kreatech), 9 (D9Z1), 11 (D11Z1), 15 (D15Z4) (SureFISH, Agilent DAKO, MetaSystems). Cutoff values were selected by Quest Diagnostics: +1q (4%), +9 (5%), +11 (6%), +15 (7%), 13q- (6%), -13 (4%), IGH rearrangement (8%), deletion TP53 (5%), t(4;14) (2%), t(11;14) (2%), t(14;16) (2%), and t(14;20) (2%).

This study was conducted in compliance with the ethical standards of the responsible institution on human subjects as well as with the Helsinki Declaration.

Among patients with MM, karyotype was normal in 71% (50/70), abnormal in 27% (19/70), and indeterminate due to inadequate specimen in 1% (1/70). FISH analysis detected abnormalities in 94% of cases (66/70). For patients with a normal karyotype (71%; 50/70), there was an abnormal FISH in 94% (47/50). For patients with normal FISH (6%; 4/70), there was an abnormal karyotype in 25% (1/4). Among patients with abnormal karyotype, FISH provided additional information about genetic aberrations in 95% of cases (18/19). Among cases with abnormal FISH, karyotype provided additional information about genetic aberrations in 27% of cases (18/66) ( Table 2 ).

Table 2. Summary of Karyotype and FISH Findings.

FISH: fluorescence in situ hybridization.

Numerical abnormalities could be detected by either karyotyping or FISH and were seen in 81% (57/70). Among these, FISH detected the presence of one or more numerical abnormalities in 95% (54/57), of which karyotype missed 70% (38/54) of the time. Karyotyping detected numerical abnormalities in 33% (19/57) of overall cases with numerical abnormalities, for which FISH failed to identify any numerical abnormalities 16% (3/19) of the time ( Table 3 ). Among these three cases, the numerical abnormalities missed by FISH were: +18 in one case, -Y in another, and +3, +11, -14, +15, -16, and +19 in the third.

Table 3. Frequency of Detection of Numerical and Structural Abnormalities.

In addition to numerical abnormalities, structural abnormalities were also detected by karyotyping and/or FISH, which included single arm gains or losses, translocations, or gene deletions. Structural abnormalities detected by either karyotyping or FISH were seen in 89% (62/70). Among these, FISH detected the presence of one or more structural abnormalities in 98% (61/62), of which karyotype missed 74% (45/61) of the time. The one case with a structural abnormality by karyotype (55,XY,+Y,t(2;8)(p12;q24.1),+3,+5,+7,+9,+11,+15,+19,+21[ 2 ]/46,XY[18]) and not by FISH (monosomy 13, gain 5, gain 9, gain 11, gain 15) was due to a t(2;8)(p12;q24.1) that only conventional karyotyping identified. Karyotyping detected structural abnormalities in 27% (17/62) of overall cases with structural abnormalities, and for which FISH missed 6% (1/17) of the time ( Table 3 ).

The most common abnormalities of any kind affected chromosome 14 (76%; 53/70), with IGH rearrangements accounting for 47% (33/70) of all cases. Most common among them were t(11;14) (11%; 8/70), t(4;14) (9%; 6/70), and t(14;16) (1%; 1/70) ( Table 4 ). Among the 20 cases with chromosome 14 abnormalities unrelated to IGH translocations, 90% (18/20) were monosomies or partial deletions of IGH, and 10% (2/20) were gains.

Table 4. Types of Abnormalities Detected by Karyotype or FISH and Their Frequencies.

The next most commonly aberrant chromosomes were chromosome 13 (56%; 39/70) and chromosome 11 (50%; 35/70). The most common abnormality of chromosome 13 was monosomy 13 (40%; 28/70), in chromosome 11 it was gain 11 (34%; 24/70), in chromosome 9, it was gain 9 (44%; 31/70), in chromosome 15, it was gain 15 (40%; 28/70), in chromosome 5, it was gain 5 (36%; 25/70), in chromosome 17, it was gain 17 (16%; 11/70), and in chromosome 4, it was t(4;14) (9%; 6/70) ( Table 4 ).

Conventional metaphase karyotyping is a well-established test in the clinical laboratory and is available all over the world. However, due to low proliferation rate of plasma cells and the resulted limited number of metaphases, abnormal karyotype is observed in only a subset of MM patients. In addition, karyotyping typically uses 400 band G-banding; each band represents approximately 10 Mbp and contains on the order of hundreds of genes, so karyotyping cannot detect small size genetic abnormalities. In contrast, interphase FISH studies are more sensitive and can reveal genetic aberrancies in most MM patients. In a study of 27 MM patients with G-banded karyotypes, 67% revealed additional genetic aberrations by the addition of FISH [ 9 ]. Our series had more success with FISH, with 94% (66/70) of MM patients revealing additional genetic abnormalities, including 95% (18/19) of those with an abnormal karyotype.

There are likely to be cytogenetic abnormalities in this specimen set that were not measured, given that selected FISH probes and their reporting are limited to targets of known clinical significance. Karyotyping can detect cytogenetic abnormalities at any location given that they are of sufficient size to be visible, and may identify potentially prognostically relevant chromosome abnormalities that are currently unknown in MM. Our data showed that karyotype detected additional genetic aberrations in 27% (19/70) of cases, including 27% (18/66) of those with abnormal FISH results.

In this data set, structural abnormalities (which are single arm gains or losses, translocations, or gene deletions) and numerical abnormalities (which are gains or losses of whole chromosomes) were each common. FISH was more sensitive (numerical: 95%; structural 98%) than karyotyping (numerical: 33%; structural 27%) in identifying cytogenetic abnormalities among cases that demonstrated their presence using either method. Karyotyping only identified one structural abnormality not identified by FISH, but did identify numerical abnormalities among 16% (3/19) of cases that did not show numerical abnormalities by FISH. Among these three cases, the numerical abnormalities missed were: +18 (case 1), -Y (case 2), and +3, +11, -14, +15, -16, +19 (case 3). Each of these changes are documented as common genetic lesions in MM [ 10 ], but less is known about the independent prognostic significance of each. Monosomy 16 and loss of Y are associated with reduced overall survival [ 9 ]. Trisomies of 3, 7, 9, 11, 15, 19 and/or 21 (four of which were missed here) have better response rates to treatment and longer survival than patients with other aneuploidies [ 11 ]. Of the 53 patients for whom revised International Staging System (R-ISS) could be calculated, 13 had high-risk cytogenetics and FISH alone would have been sufficient to account for their high-risk status in every case. While R-ISS is a function of del(17p), t(4;14), and t(14;16), the individual role of each observed chromosomal abnormality should be evaluated in future studies to determine their relationship to disease pathogenesis and clinical course, and this in turn should inform laboratories on the suitable applications of conventional karyotyping and the development of more prognostically informative FISH panels.

This study compares the results of karyotyping and FISH analysis for patients with MM. The results show that most MM patients with normal karyotype have demonstrable cytogenetic abnormalities with FISH, and routine FISH analysis appears to be an efficient method for detection of prognostically relevant chromosomal abnormalities in MM. Conversely, FISH alone without karyotyping may occasionally miss cytogenetic abnormalities of prognostic significance (such as monosomy 16, loss of Y, or various trisomies, as examples observed in this dataset). Therefore, performing both tests together will add valuable information in the cytogenetic workup of MM, and may detect new cytogenetic aberrancies that have potential prognostic significance.

Acknowledgments

None to declare.

Financial Disclosure

Conflict of interest, informed consent.

Not applicable.

Author Contributions

Matthew Crabtree aggregated data, analyzed data, reported findings, and wrote the manuscript. Jennifer Cai aggregated data and reviewed the manuscript. Xin Qing oversaw all histopathologic work, analyzed data, and reviewed the manuscript.

Data Availability

The data supporting the findings of this study are available from the corresponding author upon reasonable request.

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