Exploring Genetic Alterations in Hodgkin Lymphoma using Comparative Genomic Hybridization on DNA Microarrays

doi:10.21203/rs.3.rs-2429456/v1

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Exploring Genetic Alterations in Hodgkin Lymphoma using Comparative Genomic Hybridization on DNA Microarrays

https://doi.org/10.21203/rs.3.rs-2429456/v1

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The genetic alterations can contribute to the development and progression of the disease. In the case of c HL, a type of cancer that affects the immune system, understanding the genetic changes that occur may inform the development of more targeted and effective treatments.

In this study, we used comparative genomic hybridization CGH on DNA microarrays to perform a whole-genome analysis of 30 individuals, 15 patients with HL and 15 healthy controls.

Aberrations were divided into three groups based on size: large aberrations, regional aberrations, and microstructural aberrations

Our results showed that there were more duplications of genetic material (gains) than deletions (losses), and that specific genetic additions and deletions were present in multiple tumors. Aberrations were divided into three groups based on size: large, regional, and microstructural. Large aberrations included trisomy of whole chromosomes and gains or losses of chromosome arms. Regional aberrations were greater than 5 Mb in size and were found on multiple chromosomes. Microstructural aberrations were smaller than 5 Mb and had high amplitudes. Amplification of specific loci, such as 5q31.1 and 8q24.3, were found in multiple tumors, and loss of a single region, such as 8q22.1, was found in multiple tumors as well. it appears that genetic alterations of chromosome 8, specifically the 8q21-8q22 and 8q24.2 regions, are common in individuals with Hodgkin lymphoma. In particular, the MYC and MALT1 genes, as well as the RAD54B protein, may play a role in the development of Hodgkin lymphoma. and could potentially be targeted for therapeutic purposes.

Overall, the results of this study provide insight into the genetic alterations present in Hodgkin lymphoma and may inform future research and treatment approaches. Further research is needed to fully understand the functional consequences of these genetic changes and to explore their potential as therapeutic targets.

Exploring Genetic Alterations in Hodgkin Lymphoma using Comparative Genomic Hybridization on DNA Microarrays

Hodgkin Lymphoma

CGH

Microarray. Genetic Alterations

Abnormality of chromosomes

Hodgkin's lymphoma (HL) is a type of cancer that affects the lymphatic system, a network of vessels and organs that helps to fight infection and disease. HL is relatively rare, accounting for about 1% of all cancers in the United States. ) is incidence of about 3 new cases per 100,000 individuals per year. It is more common in people between the ages of 15 and 44, and is more common in men than in women (Küppers, R.,.et al., 2012). The incidence of HL has been increasing over the past few decades, although the reasons for this increase are not fully understood.HL is generally very treatable, with cure rates of around 80–90% using a combination of chemotherapy and radiation therapy(Piris, M.A.,.et al., 2020 ). More recent treatment approaches, such as immune checkpoint inhibitors and drug-conjugated anti-CD30 antibodies, have shown promise in improving cure rates and reducing the side effects and long-term toxicity of conventional therapy (Ansell, S.M., 2015).

Molecular genetics plays a significant role in the understanding and treatment of Hodgkin's lymphoma (HL). The Reed-Sternberg cells that are characteristic of HL have been found to have specific genetic abnormalities, including chromosomal translocations, mutations, and gene expression changes (Küppers, R., 2009).

One of the most well-known genetic abnormalities in HL is the t(2;8)(p23;q24) translocation, which involves the fusion of the MYC and KHSRP genes. This translocation is present in about 15% of HL cases and is associated with a poorer prognosis. Other genetic abnormalities that have been identified in HL include mutations in the TP53 and CDKN2A genes and alterations in the expression of genes involved in immune system function and cell growth and division (Lourenco, C.,.et al.,2021).

Genetic abnormalities in Hodgkin cells have been found to play a role in the NF-κB and JAK/STAT pathways and in immune evasion. These genetic abnormalities, in combination with the unique cellular origin of Hodgkin cells, may contribute to the development of Hodgkin lymphoma. The interaction of these genetic abnormalities with cells in the microenvironment can lead to the activation of multiple signaling pathways, often in complex ways (Weniger, M.A. and Küppers, R., 2021).

Studies of the genetic changes in HL have helped to improve our understanding of the disease and have led to the development of targeted therapies that can more specifically attack cancer cells. For example, the BTK inhibitor ibrutinib has been approved for the treatment of HL based on the finding that many HL cells have a genetic alteration that leads to overactive BTK signaling (Farrell, K. and Jarrett, R.F., 2011).

Overall, the genetic basis of HL is complex and continues to be the subject of active research. Further understanding of the genetic changes that occur in HL may help to identify new treatment targets and improve the prognosis for patients with this disease.

Objectives

The aim of this study was to perform CGH on DNA microarrays to identify and characterize genetic alterations in HL.

In the present work was implemented method of comparative genomic hybridization (CGH) on DNA microarrays, covering the entire genome was performed in which whole-genome analysis of unbalanced genetic alterations in 30 individuals. 15 patients with Hodgkin lymphoma (HL) 10 female and 5 males and 15 samples as healthy The material was taken after tumor resection and diagnosis in the Department of Oncology in AL-Najaf provinces respectively from November 2017 to May 2018. The collection of samples was approved by the Institutional Ethics Committee of Department of Oncology and all participants signed informed consent forms. All tumors were staged postoperatively according to the classification system of the Union for International Cancer Control (UICC). In 3o of the tumor samples and healthy were used Oligonucleotide microarrays with densities 105.072 sequences covering the entire genome with a resolution of 35 Kb,

DNA extraction

We extracted total DNA from tissue samples using the QIAamp DNA minikit (Qiagen, Hilden, Germany) and measured its concentration spectrophotometrically using a NanoDrop ND-2000 spectrophotometer with sample volumes of 1–2 mL. The resulting 260/280 ratio for each sample was within the range of 1.8-2.0. As a further quality control measure, we also checked the DNA in a 1% agarose gel, where the presence of high molecular weight DNA (> 50 kbp) indicated its suitability for use."

Comparative genomic hybridization on DNA microarrays:

"Microarray-based comparative genomic hybridization (CGH) is a technique that compares the DNA of a tumor and normal tissue using labeled DNA probes on slides with spotted DNA sequences (oligonucleotides or bacterial artificial chromosomes (BAC)). For our analysis, we used the genomic array CytoChip (BlueGnome, Cambridge, UK), which is an oligonucleotide microchip with a density of 105.072 sequences covering the entire genome with a resolution of 35 Kb. The chip has two separate fields for hybridization, allowing us to simultaneously analyze two samples on one chip. Each chip also has a unique code at the bottom of the glass."

We used the Blue Gnome Fluorescent Labelling System to label the control and reference DNA with random priming. The labeled products were purified using AutoSeqTM G50 columns. For the hybridization, we dissolved the labeled probes in a hybridization buffer and added the labeling mix to each tube containing DNA and primers. The test DNA was labeled with the fluorochrome Cy3 and the control DNA with Cy5. The hybridization process was performed according to the manufacturer's instructions."

After hybridization, we washed the arrays in standard saline citrate solutions with decreasing concentrations and scanned them using a GenPix 4100A. We then used the BlueFuse Multi software version 2.2 (BlueGnome, Cambridge) to process the data. This software generates base 2 logarithm (log2) ratios of the Cy3 and Cy5 intensities for all hybridized clones. We considered normal copy numbers to be in the range of -0.3 to 0.3. Values above 0.3 were evaluated as gains or amplifications, while values under − 0.3 were considered losses or deletions. The genomic profiles were plotted with logarithmic ratios on the Y-axis and the 23 chromosomes on the X-axis. The individual chromosomal profiles were depicted with clone positions on the Y-axis and logarithmic ratios on the X-axis.

Statistical analysis

To investigate the relationship between gene copy number changes and tumor phenotype (i.e. tumor stage), we used contingency table analysis and the x2 test. Results were considered statistically significant if the p-value was less than 0.05.

Comparative genomic hybridization on DNA microarrays

In the recent work there was implemented method of comparative genomic hybridization (CGH) on DNA microarrays covering the entire genome was performed in which whole-genome analysis of unbalanced genetic alterations in 30 individuals, 15 patients with Hodgkin lymphoma (HL) 10 female and 8 males and 15 samples as healthy, the duplication of genetic material (gain) is more than deletion (lose), a tumor No. 16 has 11 copy number variations CNV (Fig. 1) and tumor No. 17 has 9 CNV. For greater precision, aberrations were divided into three groups depending on their size: Large aberrations - affecting whole chromosomes or chromosome arms (when covering more than half a chromosome arm). Regional aberrations - ones that are larger than 5 Mb, but less than half of chromosome arm. Microstructural aberrations - less than 5 MB.

3.1 Large aberrations

Analysis of the most common large aberrations involving whole chromosomes or chromosome arms in HL. The additional copies of whole chromosomes (trisomy) are found as follows: Trisomy 2 (in tumor № 20) and Trisomy 13 (in tumor № 17), trisomy 18 and (in tumor № 26) – (Fig. 2). There are genetic additions to the following chromosome arms the short arm of chromosome 5 (5p +) in tumor № 19 and the short arm of chromosome 8 (8p +) in tumor № 22 (Fig. 3A.) and the short arm of chromosome 10 (10p +) in tumor № 16 and the short arm of chromosome 1 (1p +) in tumor № 21.There are also gain of the long arm of chromosome 8 (8q +) in tumor № 23 and of long arm of chromosome 11 (11q +) in tumor № 18 and in tumor № 24, and of chromosome 10 (10q +) in tumor № 26 (Fig. 3B.) and in ch18 (18q +) in tumor № 30.

Also there are genetic losses following chromosome arms: Deletion of the short arm of chromosome 7 (7p-) in tumor № 22, deletion of the short arm of chromosome 1 (1 p-) in tumor №23, deletion of the short arm of chromosome 12 (12 p-) in tumor №3(Fig. 4. B) deletion of the short arm of chromosome 11 (11 p-) in tumor №25 deletions of the long arm of chromosome 8 (8q-) in tumor № 28, 7(7q-) in tumor № 19,and 12(12q) in tumor № 17 (Fig. 4.A). see more detail in (table:7).

3.2. Regional aberrations

Analysis of regional aberrations (greater than 5 Mb) in Hodgkin lymphoma (HL)

Regional genetic additions were found on chromosomes 2, 4,6,8,9,11,12,14 and 16 - Table (1.). The highest frequency excelled regional supplements to 8q21.13- in 15% of tumors Regional genetic Deletion (loses) are found on chromosomes 1, 5, 7, 8, 9, 11, 13,14,15,17 and X chromosome Table 2

3.3. Microstructural Aberrations

Analysis of aberrations, which are smaller than 5 Mb and high amplitude

In order to fine mapping of the most important microstructural aberrations and to identify possible candidate genes, we applied the following approach: Identified aberrations that are of a size less than 5 Mb. Selected those aberrations that have a high amplitude (log2 ratio T / H > 0.5 for genetic additions and <-0.5 for genetic losses). Following this approach were selected 61 aberrations. Among them are 34 genetic additions (Table 3.), and losses − 27 (Table 4.). Amplification of the same locus 5q31.1 was reported in three of the analyzed tumors ( HL 20,21,23 ) locus 8q24.3 was reported in three of the analyzed tumors( HL16 ,17,25 ) – ( Table 3). Also three of the analyzed tumors were found loss of a single region – 8q22.1 in HL 17, 21 and 30 (Table 4.).

Analysis of the smallest regions of overlap

Another approach that is applied for fine mapping of aberrations and to identify candidate genes responsible for the onset and progression of tumors, including determination of the smallest common regions of overlap of aberrations (2q14, 3p21.1, 8q21-8q22, 8q24.2, 11q23, 14q32, 16p12) .These position for overlapping addition (gain) and for overlapping deletions (lose) throughout their length genes are shown in Table 5., for Hodgkin lymphoma (HL).

The copy number of variation (CNV) in chromosomes for the most small areas of overlapping addition (gain) and of overlapping deletions (lose) of Hodgkin lymphoma (HL) were obtained in chromosomes number Eight there where 17 aberrations in HL, 10 aberrations in short arm and 7 aberrations in long arm, following its chromosome number Tow which has 15 aberrations 13 in short arm and 2 in long arm.

Author Contributions: B.A.: Conceptualization, writing, review, editing, manuscript drafting including figures.visualization, manuscript drafting.

M.AL. Supervision, project administration, funding acquisition, review,editing. Writing, review, editing, visualization.

All authors have read and agreed to the published version of the manuscript.

Funding: This research received no external funding.

Acknowledgments: The authors are grateful to the Molecular Genetics Lab., Faculty of Science , University of Kufa, Najaf, Iraq and Middle Euphrates Oncology Center. Najaf, Iraq, to provide the research platform.

Conflicts of Interest: The authors have no conflicts of interest to declare that are relevant to the content of this article.

Availability of data and materials: The data were strictly obtained from medical records according to the privacy policy and ethics code of our Faculty.

Ethical approval and consent to participate: Ethics approval for this study was obtained from the Medical Ethics Committee Faculty of Science . University of Kufa All participants provided written informed consent prior to participating in the studya according the code 15012018/HE.Sc

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Table 1 to 2 are available in the Supplementary Files section.

Table 3.: Genetic Addition of HL in size <5 Mbp and high amplitude (log2 ratio T / H <-0.5) and some of candidate genes which observed in cytoband region.

Tumor / №	lymphoma	Cytoband	start	end	Gene
16	HL	8q24.3	131,439,284	131,979,761	TRAPPC9, CHRAC1, EIF2C2
16	HL	3p21.31	49,108,045	49,176,486	USP19, KIAA0891 , LAMB2, LAMS, NPHS5,KLHDC8B, CHL,
16	HL	5p13.2	35856848	35879602	DNAJC21, DNAJA5, BMFS3, PRLR, MFAB, HPRL, IL7R, IL7RA, CD127, NADK2, C5orf33, DECRD
16	HL	13q22.1	72,490,874	73,265,427	___
17	HL	2q14.1	113,099,364	113,134,015	IL1B ,IL1RN, MVCD4, DIRA, PAX8
17	HL	8q21.13	78675869	78806829	IL7, ZFHX4, ZFH4, PKIA, STMN2, SCGN10, SCG10, PAG, FABP5, PAFABP, EFABP
18	HL	Xpter-p22.32	X:0-	6,100,000	ANIB5 , ARSC2 , EMWX , FGS3, HDPA SHOX , LECD
18	HL	9p13	34,551,431	34,689,569	CNTFR, DCTN3, DCTN22, GALT CCL27, SCYA27, ILC, CTACK, DNAJB5, KIAA1045, HSC40
19	HL	3q28	190,368,588	191,292,238	GMNC, OSTN, UTS2D, CCDC50, PYDC2
19	HL	3q25.33	159,988,835	159996018	GFM1, EFG1, GFM, COXPD1, IL12A, IFT80, KIAA1374, WDR56, SRTD2, ATD2, TRIM59, RNF104, MRF1, IFT80L
19	HL	2q14.1	113,005,460	113,011,070	SF3B3, SF3B130, SAP130, IL34, C16orf77, CMTR2, MTR2, FTSJD1, MTSS1L, ABBA1
20	HL	8p12	36,900,922	37,452,354	ZNF703, ERLIN2, PROSC, GPR124, RAB11FIP1, BRF2, ADRB3, GOT1L1, , ASH2L, LSM1, BAG4, DDHD2, PPAPDC1B, WHSC1L1, LETM2
20	HL	5q31.1	132539193	132539193	MAR ,SLC22A5, OCTN2, CDSP, SCD RIL , IRF1,IL5
20	HL	17q21.31	40,086,578.1	41,174,222.9	ACLY, TTC25, CNP, DNAJC7, NKIRAS2, ZNF385C, , HCRT, GHDC, STAT5B, STAT5A, STAT3, PTRF, ATP6V0A1, NAGLU, HSD17B1, COASY, MLX, TUBG1 CNTNAP1, , CNTD1, G6PC, BECN1, PSME3, RAMP2, WNK4,
21	HL	5q31.1	132539193	132539193	MAR ,SLC22A5, OCTN2, CDSP, SCD RIL , IRF1,IL5
21	HL	12q15	68201348	68225790	IL26, CAND1, TIP120A, TIP120, KIAA0829, DYRK2, IFNG, IFG, IFI, IL22, IL21, ILTIF, MDM2, ACTFS
22	HL	8p11.23 -8p11.1	39,647,643	41,599,834	C8orf4, ZMAT4, SFRP1, GOLGA7, GINS4, AGPAT6, ANK1, KAT6A, AP3M2, PLAT, IKBKB, VDAC3, SLC20A2, POLB, DKK4, CHRNB3, CHRNA6, THAP1, RNF170, HOOK3, FNTA, SGK196, HGSNAT, POTEA, MIR486, MIR4469
22	HL	1q22.1	169,819,902	172,950,759	RASSF5, NORE1 IL10, CSIF, GVHDS
22	HL	11p13	30,582,677.3	35,334,211	HIPK3, CD59, FBXO3, LMO2, CAPRIN1, NAT10, ABTB2, CAT, ELF5, EHF, APIP, PDHX, CD44, SLC1A2, PAMR1, MIR1343 ,ELP4, PAX6, RCN1, WT1, EIF3M, CCDC73, PRRG4,
23	HL	5q31.1	132656523	132661108	SLC22A5, OCTN2, CDSP, SCD, IRF1, MAR, IL5, IL13, ALRH, BHR1, GDF9, POF1,
23	HL	9q34.2	122,724,330	123,060,355	CAMSAP1, UBAC1, NACC2, LHX3, QSOX2
23	HL	16p13.2	6,957,553	7,062,587	METTL9,RBFOX1
24	HL	1p13.3	109,727,865	109,778,742	NBPF4,SLC25A24
24	HL	2q37.3	242,514,623	242,656,003	THAP4, ATG4B, DTYMK, ING5
25	HL	8q21.13	78675869	78806829	IL7, ZFHX4, ZFH4, PKIA, STMN2, SCGN10, SCG10, PAG,
26	HL	16p12.1	27,313,667	27364777	AQP8, NSMCE1, NSE1, IL4R, IL4RA
26	HL	17p13.3	1,197,564	2,535,589	RILP, PRPF8, SERPINF2, TLCD2, WDR81, SERPINF1, SMYD4, RPA1, RTN4RL1, DPH1, OVCA2, HIC1, SMG6, SRR, TSR1, SGSM2, MNT, METTL16, PAFAH1B1, MIR22, MIR132, MIR212
27	HL	11q23	117,427,771	117,836,975	DSCAML1, KIAA1132FXYD6 , AMICA1,
28	HL	1q31.2	194,985,271	195,163,895	EIF2D, HCA56 MAPK,APK2, MK2
28	HL	16p12.1-p11.2	28499361	28526729	IL27, IL30, GSG1L, CLN3, BTS, NUPR1, COM1, p8
28	HL	8q21.13	78675869	78806829	IL7, ZFHX4, ZFH4, PKIA, STMN2, SCGN10, SCG10, PAG, FABP5, PAFABP, EFABP
29	HL	2q33.1	201,182,880	201,233,442	CASP10, MCH4, ALPS2,CASP8, MCH5, ALPS2B
30	HL	13q32.1	95,503,342	96,093,412	ZNF90,GPC6
30	HL	1q21.1	169,011,374	172,065,896	CSIF

Table 4. Genetic Deletion of HL in size <5 Mbp and high amplitude (log2 ratio T / H <-0.5) and some of candidate genes which observed in cytoband region.

Tumor / №	lymphoma	Cytoband	start	end	Gene
16	HL	2q14.1	113,099,364-	113,134,015	IL1RN, MVCD4, DIRA
16	HL	12q13.3	56,230,035	56,316,222	SLC39A5, MYP24, CS, CNPY4, TMEM4, MSAP, PAN2, USP52, KIAA0710
16	HL	9p21.3	21,077,104	21,186,617	IFNB1, IFNW1, IFNA21, IFNA4
17	HL	16p12.1	25,313,667	27,464,777	AQP8 , KDM8, JMJD5,IL4R, IL4RA, L21R, IMD56
17	HL	8p23.1	7,506,931	8,290,301	DEFB4A, , TM2D2, ADAM9, ADAM32, ADAM2, IDO1, , , DEFB104A, , SPAG11A, , FAM90A7, MIR548I3
17	HL	21q22.11	30,200,001	30,356,981	IFNAR1
18	HL	2p25.1	11,467,751	12,585,722	IL1RN, MVCD4, DIRA, PAX8
18	HL	16q22.1	70,579,894	70,661,681	IL34, C16orf77, MTSS1L, ABBA1
19		Xpter-p22.32	X:0	X:0	FGS3, HDPA, LECD(Hodgkin disease)
20	HL	11q23.3	117,486,390-	118,001,482	KIAA1132, FXYD2, ATP1G1, FXYD6, TMPRSS13, IL10RA, IL10R, IBD28
20	HL	8q21.13	80,034,867	80,967,809	TPD52, D52, PAG
21	HL	2q11.1	97,394,778	98,518,238	TMEM131
21	HL	8p23.1	7,140,623	8,724,821	FAM90A5, , DEFB4A, DEFB4B, DEFB103A, DEFB103B, DEFB106B, DEFB107A, DEFB107B, SPAG11A, SPAG11B, ZNF705G
22	HL	5q11.1	48,795,760	50,268,123	EMB, PARP8
22	HL	12q13.3	56,334,158	56,341,539	IL23A, SGRF, P19, IL23P19, STAT2, IMD44
23	HL	11q23.3	118,077,011	118,226,689	TMPRSS4, SCN4B, LQT10, ATFB17, AMICA1, JAML
24	HL	2q37.3	243,514,621	243,656,145	TMEM131, DTYMK, ING5,THAP4, ATG4B,
24	HL	14q11.2	23,352,808-	23,376,402	BCL2L2, BCLW, PABPN1, IL25, IL17E
25	HL	19p13.11	18,058,993	18,099,026	JAK3, JAKL, IL12RB1, IMD30, IFI30, GILT
25	HL	8p12-8p11.23	39,276,182	39,625,839	FGFR1, TACC1
26	HL	4q13.2	70,057,689	70,643,356	YTHDC1,TMPRSS11B, TMPRSS11E, UGT2B17, UGT2B15
26	HL	11q23.3	117,886,390-	118,001,482	KIAA1132, FXYD2, ATP1G1, FXYD6, TMPRSS13, IL10RA, IL10R, IBD28
27	HL	8q11.23	49,909,725	52,939,907	ST18, ZNF387, KIAA0535,RB1CC1, CC1, TCEA1, TF2S, GTF2S
28	HL	3p21.1	53,161,549	53,867,387	PRKCD, CVID9, ALPS3, TKT, IL17RB, IL17BR, IL17RH1
29	HL	6p25.2	2,948,158	3,063,840	NQO2, NMOR2,RIPK1, RIP1, RIP
30	HL	8p23.2	4,199,879	4,787,749	CSMD1, IDO1, IDO2,PLEKHA2, HTRA4, TM2D2, ADAM9, ADAM32, ADAM18, ADAM2,
30	HL	10q11.21	47,011,798	47,271,563	PPYR1, ANXA8, ANXA8L1

Table 5.: Most small areas of overlapping addition (gain) and of overlapping deletions( lose ) throughout their length genes for Hodgkin lymphoma (HL) and candidate genes responsible for the onset and progression of tumors.

The smallest area of overlap	HL Tumor Addition	HL Tumor Deletion	Genes
2q14	17,19	16	IL1B ,IL1RN, MVCD4, DIRA, PAX8 IL36B, IL1F8, IL1H2, FIL1H, SF3B3, SF3B130, SAP130,IL1RN, MVCD4, DIRA IL36B, IL1F8, IL1H2, FIL1H, IL36RN, IL1F5, FIL1D, IL1HY1, IL1RP3, PSORP, PSORS14, IL1F10, IL1HY2, IL1RN, MVCD4, DIRA ,IL37, IL1F7, FIL1Z, IL1H4, IL1RP1, IL36G, IL1F9, IL1H1, IL1RP2, IL36A, IL1F6, FIL1E
11q23	27	20,23,26	DSCAML1, KIAA1132FXYD6 , AMICA1, KIAA1132, FXYD2, ATP1G1, FXYD6, MPRSS13, IL10RA, IL10R, IBD28,TMPRSS4, SCN4B, LQT10, ATFB17, AMICA1, JAML,KIAA1132, FXYD2, ATP1G1, FXYD6, TMPRSS13, IL10RA, IL10R, IBD28,SIDT2, TAGLN, PCSK7, PC8, PC7, LPC,KIAA1132, TMPRSS13, IL10RA, IL10R, IBD28 ,FXYD2, ATP1G1, FXYD6,
8q21.1- 8q22.	17,25,28	17,20,21 27, 28,30	DUP8q22.1, C8DUPq22.1, HTC1, TMEM67, MKS3, JBTS6, NPHP11, RAD54B SNORD13, UNC5D, UNC5H4, SCLL, ST18, ZNF387, KIAA0535,RB1CC1, CC1, TCEA1, TF2S, GTF2S,CALB1, TMEM55A, RUNX1T1, CBFA2T1, AML1T1, ETO TPD52, D52, PAG,IL7, ZFHX4, ZFH4, PKIA, STMN2, SCGN10, SCG10, FABP5, PAFABP, EFABP,PDP1, PPM2C, PDPC , CDH17, , HPT1, GEM ,RAD54Bs
8q24.2	16,22		IMD12 TRAPPC9, CHRAC1, EIF2C2 PVT1, FAM149B, ASAP1, DDEF1,CASC11, LINC00990, CARLO7, MYC,CASC21, LINC01244, CARLO2, CASC8, LINC00860, CARLO1, POU5F1B, POU5F1P1, OCT4PG1, CCD26, RAM.
14q32			GLRX5, C14orf87, PRO1238, FLB4739, PRSA, SIDBA3,TCL6, TCL1B, TML1, TCL1A, TCL1.
3p21.1	16,28		USP19, KIAA0891 , LAMB2, LAMS, NPHS5,KLHDC8B, CHL, PRKCD, CVID9, ALPS3, IL17RB, IL17BR, IL17RH1, MIR4271, USP4, UNP, GPX1, GPXD, RHOA, ARHA, ARH12, RHOH12 , RHO TCT
16p12	26,28	16	AQP8, NSMCE1, NSE1, IL4R, IL4RA, IL27, IL30, GSG1L, CLN3, BTS, NUPR1, COM1, p8

Table 6 : show the copy number variation (CNV) in chromosomes for most small areas of overlapping addition (gain) and of overlapping deletions ( lose ) of Hodgkin lymphoma (HL).

Chromosome	No.Short arm aberrations	No. Long arm aberrations	HL Sample	Total aberrations
1	4	6	22,28.30	10
2	13	2	16,17,18,19,21 , ,29	15
3	5	4	16,19,28	9
4	1	0	24,26	1
5	4	1	16,20,21,22,23	5
6	3	2	29	5
7	0	3		3
8	15	2	20,25	17
9	1	4	16,18,23	5
10	2	0	30	2
11	5	1	20,22,23,26,27	6
12	5	0	16,21,22	5
13	3	0	16,30	3
14	6	0	24	6
15	0	0	0	0
16	2	5	17,18,23,26,28	7
17	3	2	20,26	5
18	1	0		1
19	0	3	25	3
20	0	0	0	0
21	1	0	17	1
22	0	0	0	0
X	0	2	17,18	2
y	0	0		0

Table 7.: Descript the all cytogenetic aberration and copy number variations (CNV) of sample of patients HL , AT(Addition trisomy), Ap (Addition p arm), Aq(Addition q arm), DM (Deletion monsomy ) Dp (Deletion p arm ) Dq ( Deletion q arm) and copy number variations (CNV).

No. Sample	AT	AP	Aq	DM	Dp	Dq	Long Addition	Long Deletion	Bands		CNV
No. Sample	AT	AP	Aq	DM	Dp	Dq	Long Addition	Long Deletion	Add.	Del.	CNV
HL16	13	2p			8p		14q11.2-q24.1	Xp22.33-p21.2	3	3	11
HL17	13					12q	2q24.1-q24.3	1q23.3-q25.3	2	3	9
HL18			9q					17q11.1-q12	2	2	6
HL19		5p				7q	4q11-4q21.1	8p12-p11.23	3	1	8
HL20	2							14q24.2-q32.33	3	2	7
HL21		1p					11q23.3-q25		2	2	6
HL22		8p			7p		6q14.1-q14.3	5q14.1-q21.1	2	2	8
HL23			8q		1p		8p23.1-p21.3	9p23-p22.1	3	1	8
HL24		10p				8q	12q13.13-q21.3		2	2	7
HL25					11p		16q11.2-q22.1		1	2	5
HL26	18		10q				8q24.13-q24.3	15p12-p11.2	2	2	8
HL27					3p		9q33.1-q34.3		1	1	4
HL28						8q	18q12.3-q22.1		3	1	6
HL29		13p			2p		11q24.2-qTer	7q31.32-q34	1	1	6
HL30			18q					11p13-q12.2	2	2	6
Total CNV	4	6	4		6	4	12	10	32	27	105

Table 8: Show The number and Average of Aberrations patients of HL in Whole chromosome , Regional aberrations and microstructure and the total of average of all aberrations .

Type of Aberrations	Addition	Average gain (Addition) in genetic copies	Deletion	Average lose (deletion ) in genetic copies in HL
Whole chromosome Aberrations	14	0.93/ tumor	10	0.6/ tumor
dominate large Aberrations	12	0.8 / tumor	10	0.6/ tumor
microstructures Aberration	32	2.1 / tumor	27	1.8/ tumor
Total	58	3.86/ tumor	47	3.1/ tumor

No competing interests reported.

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Exploring Genetic Alterations in Hodgkin Lymphoma using Comparative Genomic Hybridization on DNA Microarrays

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Abstract

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1. Introduction

2. Material And Methods

3. The Results

3.1 Large aberrations

3.2. Regional aberrations

3.3. Microstructural Aberrations

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