-Author name in bold denotes the presenting author
-Asterisk * with author name denotes a Non-ASH member
Clinically Relevant Abstract denotes an abstract that is clinically relevant.

PhD Trainee denotes that this is a recommended PHD Trainee Session.

Ticketed Session denotes that this is a ticketed session.

3075 Next-Generation Sequencing Identifies a Previously Uncharacterized Gene ANKRD36 As a Common Biomarker for Blast Crisis Chronic Myeloid Leukemia: Molecular and Protein Bio-Modeling Studies

Program: Oral and Poster Abstracts
Session: 632. Chronic Myeloid Leukemia: Therapy: Poster III
Hematology Disease Topics & Pathways:
CML, Diseases, cellular interactions, Biological Processes, Technology and Procedures, genomics, Clinically relevant, Myeloid Malignancies, hematopoiesis, molecular testing, NGS, pathogenesis
Monday, December 7, 2020, 7:00 AM-3:30 PM

Zafar Iqbal, MPhil, PhD1,2,3,4,5, Muhammad Absar, BSc, MSc, PhD6,7*, Abid Jamil, MD PhD8,9*, Tanveer Akhtar, Ph.D10*, Salman Basit, PhD11*, Sibtain Afzal12*, Khushnooda Ramzan13*, Kulsoom Qureshi14*, Muhammad Farooq Sabar, Ph.D15*, Muhammad Arshad16*, Aamer Aleem, MBBS, FRCP, FRCPath17*, Ullah Anhar12,18*, Zeenat Mirza, PhD19*, Sajjad Karim19*, Naila Batool Khalid20*, Mahmood Rasool21*, Muhammad Asif, PhD22*, Mujib Ullah, PhD23*, Ahmad M Khalid, PhD24*, Amer Mahmood, PhD25* and Nawaf Alanazi, MBBS FRCPC26,27*

1Cancer & Medical Genetics, CoAMS-A, King Saud Bin Abdulaziz University for Health Sciences & King Abdullah International Medical Research Centre (KAIMRC) / SSBMT, King Abdulaziz Medical City, National Guard Health Affairs, Al-Ahsa, Saudi Arabia
2Centre for Advanced Molecular Biology, University of the Punjab, Lahore, Pakistan
3Qarshi University, HOPES, Department of Biotechnology & Genomic Medicine, Lahore, Pakistan
4Univ. of The Punjab, Pakistan, Lahore, Pakistan
5Pakistan Society for Molecular and Clinical Hematology, Lahore, Pakistan
6Department of Pathology, King Abdulaziz Hospital, King Abdulaziz Medical City, National Guard Health Affairs, Al Ahsa, Saudi Arabia
7Hematology, Oncology and Pharmacogenetics Engineering Sciences (HOPES) Group, Department of Zoology, University of the Punjab, Lahore, Pakistan
8Hayatabad Medical Complex, Peshawar, Pakistan
9HMC Peshawar, Peshawar, Pakistan
10HOPES Group, Hematology, Oncology and Pharmacogenetic Engineering Sciences (HOPES) Group Pakistan & HOPES Centre, Department of Zoology, University of the Punjab, Lahore, Pakistan
11College of Medicine, Taiba University, Madinah, Saudi Arabia
12College of Medicine, King Saud University, Riyadh, Saudi Arabia
13King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia
14Medical Oncology Unit, HMC, Peshawar, Khyber Pakhtunkhwa, PAK
15Core Genomics Laboratories, Centre for Applied Molecular Biology (CAMB), University of the Punjab, Lahore, Pakistan., Lahore, Pakistan
16Department of Biological Sciences, International Islamic University, Islamabad, Pakistan
17Hematology/Oncology Division, Department of Medicine, College of Medicine & King Khalid University Hospital, King Saud University, Riyadh, Saudi Arabia
18Hematology, Oncology and Pharmacogenetic Engineering Sciences (HOPES) Group Pakistan & HOPES Centre, Department of Zoology, University of the Punjab, Lahore, Pakistan
19Faculty of Applied Medical Sciences & Centre of Excellence in Genomic Medicine Research, King Abdulaziz University, Jeddah, Saudi Arabia
20IMBB, University of Lahore, Lahore, Pakistan
21Faculty of Applied Medical Sciences & CEGMR, King Abdulaziz University, Jeddah, Saudi Arabia
22Department of Biotechnology, Balochistan University of Information Technology, Engineering and Management Sciences, (BUITEMS), Quetta, Pakistan
23Interventional Regenerative Medicine and Imaging Laboratory, Department of Radiology, Stanford University School of Medicine, Stanford, CA
24Department of Biotechnology & Genomic Medicine, Qarshi University, Lahore, Pakistan
25College of Medicine & King Khalid University Hospital, King Saud University (CoM KSU), Riyadh, Saudi Arabia
26Division of Pediatric Hematology /Oncology, Department of Medicine, King Abdulaziz Hospital, National Guard Health Affairs, Al Ahsa, Saudi Arabia
27CAMS-A (College of Applied Medical Sciences Al AHSA) King Saud Bin Abdulaziz University of Health Sciences & King Abdulaziz Hospital, National Guard Health Affairs, Al Ahsa, Saudi Arabia


Chronic Myeloid Leukemia (CML) is initiated due to t (22;9) giving rise to Philadelphia chromosome and fusion oncogene BCR-ABL1. Discovery of BCR-ABL led to development of molecularly-targeted drugs called tyrosine kinase inhibitors (TKI), that have revolutionized CML treatment in first quarter of 21st century, by transforming a once fatal disease into a almost-cured cancer. Due to TKIs, survival of CML has become equal to general population, with possibility of a number of CML patients to undergo treatment-free remission. Nevertheless, TKIs are minimally effective in blast crisis CML patients (BC-CML), making this group of CML patients one of the biggest therapeutic challenge in modern cancer medicine. Unfortunately, a common biomarker for BC-CML is not available and mechanism of CML progression to advanced phases poorly understood3. Therefore, objective of our study was to find a common molecular biomarker of disease progression and specifically BC in CML.

Materials and Methods:

Patient selection: CML patients in accelerated and blast crisis phase CML (Experimental group) were subjected to whole exome sequencing (WES) along with appropriate controls (Chronic phase treatment-naïve CML patients as Control 1, Chronic phase CML long-term TKI responders (at least 2 continuous years of MMR)2 as Control group 2, CML patients with resistant to TKIs as Control group 3 and healthy controls).

Sample collection: DNA extraction and Clinical follow-up: 10 ml peripheral blood was collected from all study subjects. DNA was extracted and patient follow-up was carried out during course of this study. All criteria per ENL guidelines were adopted.

Whole Exome Sequencing (WES): WES was carried out using Illumina NGS instrument (HiSeq). bcl files were converted to fastq files by using bcl2fastqtool4. Raw reads were aligned to genome using BWA tools while whole exome variants were annotated using Illumina Variant Studio4. R package was employed to align specific gene mutants to disease phenotypes5. Variants were confirmed using Sanger sequencing. Genes mutated in all AP/BC-CML patients but not mutated in any of control groups were selected.

Results and Discussion:

We found some novel as well as known genes associated with diverse biological functions mutated in all AP/BC-CML6. We found some previously uncharacterized genes like ANKRD36; genes associated with vital life processes, for example, POTE-G (member of cancer-testis antigen family), SARM1 (apoptosis and immunity), OR9G1 (member of G-protein-coupled receptors), RNF212 (Meiotic crossing-over) etc.; genes reported in other cancers (PRSS3, MUC6, ESRR-A, RASA4, PDE5-A, DACH-1, TRAK1 etc.); DNA repair genes (FANCD2 and ATXN3) and genes involved in transcriptional regulation (unique ZNF family genes). As ANKRD36 (ENSG00000135976) has previously uncharacterized in human and its protein structure was unknown, its protein sequence was retrieved (https://www.uniprot.org/uniprot/A6QL64), computational prediction of the protein structure was performed using I-Tasser7, the mutations manually evaluated, and the wild and mutated structures superimposed using PyMOL8. ANKRD36 has maximum expression in bone marrow, specifically myeloid cells (figure 1a-c)9. Thus, it is may serve as a potential biomarker and drug target in CML. We recommend carrying out further studies to explore the role of ANKRD36 in biology and progression of CML.


1: Valent P, Herndlhofer S, Schneeweiß M, Boidol B, Ringler B, Kubicek S, et al. Oncotarget. 2017 Apr 4; 8(14): 23061-23072.

2: Annunziata M, Bonifacio M, Breccia M, Castagnetti F, Gozzini A, Iurlo A, et al. Front Oncol. 2020 ;10:883.

3: Feng XQ, Nie SM, Huang JX, Li TL, Zhou JJ, Wang W, et al. Neoplasma, 2020 ;67(1):171-177.

4: Hashmi JA, Albarry MA, Almatrafi AM, Albalawi AM, Mahmood A, Basit S. Congenit Anom (Kyoto). 2017 Apr 16. doi: 10.1111/cga.12225. [Epub ahead of print].

5: R Core Team (2012). R Foundation for Statistical Computing, Vienna, Austria. ISBN 3-900051-07-0, URL http://www.R-project.org/

6: GeneCards: The Human Gene Database, https://www.genecards.org, accessed 11th Aug 2020.

7: Zhang Y. I-TASSER server for protein 3D structure prediction. BMC Bioinformatics. 2008 Jan 23;9:40.

8: DeLano, W. L. CCP4 Newsletter On Protein Crystallography. 2002; 40:82-92.

9: Fagerberg L, Hallström BM, Oksvold P, Kampf C, Djureinovic D, Odeberg J, et al. Mol Cell Proteomics. 2014 Feb;13(2):397-406.

Disclosures: Jamil: Novartis: Honoraria, Other: Travel Support; Roche: Honoraria, Other: Travel Support.

*signifies non-member of ASH