HLA alleles, haplotypes frequencies, and their association with hematological disorders: a report from 1550 families whose patients underwent allogeneic bone marrow transplantation in Egypt.

HLA alleles are representative of ethnicities and may play important roles in predisposition to hematological disorders. We analyzed DNA samples for HLA-A, -B, -C, -DRB1, and -DQB1 loci, from 1550 patients and 4450 potential related donors by PCR-SSO (Polymerase chain reaction sequence-specific oligonucleotides) and estimated allele frequencies in donors and patients from 1550 families who underwent bone marrow transplantation (BMT) in Egypt. We also studied the association between HLA allele frequencies and incidence of acute myeloid leukemia, acute lymphoblastic leukemia, and severe aplastic anemia. The most frequently observed HLA class I alleles were HLA- A*01:01 (16.9%), A*02:01 (16.1%), B*41:01 (8.7%), B*49:01 (7.3%), C*06:02 (25.1%), and C*07:01 (25.1%), and the most frequently observed class II alleles were HLA-DRB1*11:01 (11.8%), DRB1*03:01 (11.6%), DQB1*03:01 (27.5%), and DQB1*05:01 (18.9%). The most frequently observed haplotypes were A*33:01~B*14:02 ~ DRB1*01:02 (2.35%) and A*01:01~B*52:01~DRB1*15:01 (2.11%). HLA-DRB1*07:01 was associated with higher AML odds (OR, 1.26; 95% CI, 1.02-1.55; p = 0.030). Only HLA-B38 antigen showed a trend towards increased odds of ALL (OR, 1.52; 95% CI, 1.00-2.30; p = 0.049) HLA-A*02:01, -B*14:02, and -DRB1*15:01 were associated with higher odds of SAA (A*02:01: OR, 1.35; 95% CI, 1.07-1.70; p = 0.010; B*14:02: OR, 1.43; 95% CI, 1.06-1.93; p = 0.020; DRB1*15:01: OR, 1.32; 95% CI, 1.07-1.64; p = 0.011). This study provides estimates of HLA allele and haplotype frequencies and their association with hematological disorders in an Egyptian population.

Downregulation of IRAIN long non-coding RNA predicts unfavourable clinical outcome in acute myeloid leukaemia patients.

BACKGROUND: Although it has been shown that the long non-coding RNA (lncRNA) insulin-like growth factor type 1 receptor (IGF1R) antisense imprinted non-protein coding RNA (IRAIN) is downregulated in leukaemia cell lines, its usefulness as a prognostic marker in acute myeloid leukaemia (AML) has not yet been thoroughly investigated. Here, we sought to determine whether the expression of IRAIN is associated with clinical outcome of AML patients. SUBJECTS & METHODS: Using quantitative real-time polymerase chain reaction (qRT-PCR), IRAIN expression levels were assessed in peripheral blood leukocyte samples from 150 patients with AML and 50 healthy controls. Analysis was done on the relationship between IRAIN expression and clinical outcomes in AML patients. RESULTS: When compared to healthy controls, IRAIN expression was markedly reduced in AML patients (P = 0.019). IRAIN expression could distinguish French-American-British (FAB) subtypes of AML (P = 0.024). Low IRAIN expression status was associated with shorter event-free survival (EFS) in the non-t(15;17) cytogenetically abnormal AML subset (P = 0.004). IRAIN downregulation was identified as an independent adverse prognostic marker for complete remission (CR) not only in the in the non-t(15;17) cytogenetically abnormal AML subset (P = 0.006) but also in the AML-M4/M5 subgroup (P = 0.033). CONCLUSION: Aberrantly low IRAIN expression is closely associated with lower CR rates in AML patients, particularly in non-t(15;17) cytogenetically abnormal AML and M4/M5 AML, suggesting that the determination of IRAIN expression level at diagnosis provides valuable prognostic information, serves as a promising biomarker for evaluating treatment response, and helps predicting clinical outcome of AML patients.

ABCB1 Single Nucleotide Polymorphism Genotypes as Predictors of Paclitaxel-Induced Peripheral Neuropathy in Breast Cancer.

Background: Paclitaxel is a key antineoplastic agent in the treatment of breast cancer and many other malignancies. However, paclitaxel-induced peripheral neuropathy (PIPN) is a common adverse event that occurs with paclitaxel therapy and frequently causes considerable pain and a decline in patients' quality of life. Single nucleotide polymorphisms (SNPs) in the ABCB1 gene have been frequently associated with increased severity of PIPN. However, the validity of ABCB1 SNP markers to predict the incidence of PIPN has not been confirmed. Methods: We extracted genomic DNA from samples collected from 92 Egyptian female breast cancer patients receiving weekly paclitaxel and used them to genotype ABCB1 G1236A (rs1128503) and ABCB1 G3435A (rs1045642). Markers that correlated with PIPN, together with baseline clinical factors, were used to fit additive, dominant, overdominant, and recessive genetic models. We applied a repeated k-fold cross-validation algorithm to select the model with the highest predictive accuracy. We finally performed model diagnostics and receiver operating characteristics (ROCs) analysis for the model with the highest classification accuracy. Results: The additive model achieved the highest classification accuracy. The G1236A homozygous AA variant correlated with grade ≥2 PIPN (p = 0.018). PIPN also correlated with body surface area (BSA) (p = 0.003) and history of diabetes before treatment (p = 0.015). ROCs analysis showed a sensitivity of 76.9%, a specificity of 86.8%, a positive predictive value of 83.64%, and a negative predictive value of 81.08% for the additive model. Conclusion: The ABCB1 G1236A, BSA, and history of diabetes are valid predictors of PIPN, which can enable the personalization of paclitaxel dosing to prevent PIPN.

Expression and prognostic relevance of long noncoding RNAs CRNDE and AOX2P in adult acute myeloid leukemia.

INTRODUCTION: Several long noncoding RNAs (lncRNAs) have been demonstrated to play a critical role in the tumorigenesis of acute myeloid leukemia (AML), and altered expression of certain lncRNAs has been recognized as a potential prognostic marker in AML patients. Here, we sought to determine whether the expression of the lncRNA colorectal neoplasia differentially expressed (CRNDE) and aldehyde oxidase 2 pseudogene (AOX2P) is associated with clinicopathological features and clinical outcome of patients with AML. METHODS: CRNDE and AOX2P expression levels were measured in diagnostic blood samples from 200 adult patients with de novo AML, along with 50 healthy control blood samples, using quantitative real-time polymerase chain reaction (qRT-PCR). The association of CRNDE and AOX2P expression with the clinicopathological characteristics and outcome of AML patients was analyzed. RESULTS: Upregulated CRNDE expression was independently associated with lower complete remission (CR) rates in the whole cohort of AML (P < .001). AOX2P overexpression was identified as an independent adverse prognostic marker for CR in the CN-AML (P = .009) and non-t (15;17) AML (P < .001) subgroups. Patients with high CRNDE expression had a significantly shorter event-free survival (EFS, whole cohort of AML: P = .017; CN-AML: P = .001; non-t (15;17) AML: P = .006) and inferior overall survival (OS, whole cohort of AML: P = .002; t(15;17) AML: P = .001) than those with low CRNDE expression. EFS and OS did not differ significantly between patients with high AOX2P expression and those with low expression. CONCLUSION: Aberrantly upregulated CRNDE expression and, to a lesser extent, AOX2P overexpression, are associated with poor prognosis in AML patients, suggesting that the determination of CRNDE and, perhaps, AOX2P, expression level at diagnosis provides valuable prognostic information, allows refinement of risk stratification, and helps clinical decision-making in AML.

IDH Mutations in AML Patients; A higher Association with Intermediate Risk Cytogenetics.

OBJECTIVE: IDH mutations diversely affect the prognosis of cyogenetically normal acute myeloid leukemia (CN-AML) adult patients. The aim of this study is to assess the frequency of IDH mutations and to evaluate its role in AML prognosis. METHODS: We have analyzed IDH1 and 2 mutations using High Resolution Melting curve analysis (HRM) in 70 denovo AML patients. RESULTS: The median age of AML patients is 40 years (16-75). Incidence of IDH mutations is 10/70 (14.3%); 2 (2.9%) IDH1 mutant and 8 (11.4%) IDH2 mutant. Median PB blasts of mutant IDH patients was 67.5% (25-96) vs. 44% (0-98) for wild type (p=0.065). Eight/10 (80%) mutant IDH patients had B.M blasts ≥50% vs. 2/10 (20%) <50% (p<0.001) and were classified as intermediate risk cytogenetics (p=0.020) with wild FLT3-ITD (p=0.001). Ten/10 (100%) mutant IDH patients showed wild NPM1 (p=0.049). Median OS of mutant IDH in the intermediate risk cytogenetics was 1.8 years (0.7-3.1) vs. 3.1 years (1.1-5.5) for wild IDH (p=0.05). CONCLUSION: IDH mutation is mainly associated with intermediate risk AML and when integrated in this specific subgroup displays a lower survival and can be considered an additional integrated molecular risk marker for AML prognosis.
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Cytogenetic features in primary myelodysplastic syndrome Egyptian patients.

Karyotype is the most important diagnostic and prognostic parameter in myelodysplastic syndromes (MDS) and is abnormal in approximately 50% of patients. We emphasized the importance of chromosomal analysis and reported the most frequent cytogenetic abnormalities in 50 MDS (29 males (58%) and 21 females (42%), median age: 57.5 years) Egyptian patients using conventional banding analysis (CBA). Karyotype description was conducted according to the International System for Human Cytogenetic Nomenclature (ISCN, 2013). Patients were diagnosed based on complete history, bone marrow (BM) aspirate, peripheral blood (PBL) examination, and Iron stain. MDS with multilineage dysplasia (MDS-MLD) was the most frequently encountered subtype; 19/50 (38%) followed by MDS with single lineage dysplasia (MDS-SLD); 11/50 (22%). 27/50 patients (54%) showed a normal karyotype while 23 patients (46%) showed clonal nonrandom chromosomal abnormalities. Most patients with MDS with excess blasts-II (MDS-EB-II) showed abnormal karyotype (3/4; 75%) followed by MDS-EB-I (3/5, 60%) and MDS-MLD (10/19, 53%). Among 50 primary MDS patients; 14/50 (28%) had a single chromosomal abnormality, 3/50 (6%) had double chromosomal abnormality, and 6/50 (12%) had complex karyotype. Male sex was more frequently associated with higher IPSS prognostic risk categories than female gender. The most common single chromosomal abnormalities were -5/del5q; 7/50 (14%) patients followed by -7; 4/50 (8%) patients. +8, del20q and delY were each detected in 1/50 patient (2%). Abnormalities of chromosome 5 (-5/del5q) as a single chromosomal abnormality was the most frequent chromosomal abnormality among Egyptian primary MDS patients followed by complex karyotype. Cytogenetic characteristics of MDS Egyptian patients were similar to North African and European patients. Karyotype offers useful information in establishing accurate diagnosis and male gender is an important predisposing factor that can predict worse prognosis in MDS patients.

MPN10 score and survival of molecularly annotated myeloproliferative neoplasm patients.

JAK2, CALR, MPL and triple-negative mutational status has a direct impact on symptom severity and disease burden assessed by MPN10 score in myeloproliferative neoplasms (MPNs). Among 93 patients; median MPN10 score was 48 (5-76) in JAK2 mutants versus 25 (4-80) in JAK2 negative (p < .001); 22.5 (4-65) in CALR mutants versus 35 (5-80) in CALR negative (p < .050) and 21 (10-48) in triple negative versus 40 (4-80) in JAK2/CALR/MPL mutants (p < .001). At three years, progression free and overall survival of JAK2-positive versus JAK2-negative patients were 62% versus 100% (p < .001); 85% versus 100% (p = .011) and were 100% versus 78% (p = .067); 100% versus 92% (p = .197) in CALR-positive versus CALR-negative patients and 100% versus 75% (p = .004); 100% versus 90% (p = .015) in triple negative versus mutant patients, respectively. MPN10 score in association with driver gene mutations can be used as a predictor of survival in MPN patients.

The prognostic significance of minimal residual disease in adult Egyptian patients with precursor acute lymphoblastic leukemia.

BACKGROUND: Minimal residual disease (MRD) studies in adult acute lymphoblastic leukemia (ALL) give highly significant prognostic information superior to other standard criteria as age, gender and total leucocytic count (TLC) in distinguishing patients at high and low risk of relapse. OBJECTIVES: We aimed to determine the value of MRD monitoring by flowcytometry (FCM) in predicting outcome in adult Precursor ALL patients. PATIENTS AND METHODS: Bone marrow (BM) samples were analyzed by 4-color FCM collected at diagnosis and after induction therapy (MRD1) to correlate MRD positivity with disease free survival (DFS) and overall survival (OS). RESULTS: Study included 57 adult ALL patients (44 males and 13 females) with a median age of 22 years (18-49). DFS showed no significant difference with age, gender and initial TLC (p=0.838, 0.888 and 0.743, respectively). Cumulative DFS at 2 years was 34% for B-lineage ALL (n: 35) and 57% for T-lineage ALL (n: 18) (p = 0.057). Cumulative DFS at 2 years was 7% for MRD1 positive (high risk, HR) versus 57% for MRD1 negative patients (Low risk, LR) (p < 0.001). Cumulative DFS at 2 years was 29% for HR patients (n: 26) versus 55% for LR (n: 27) according to GMALL classification (p = 0.064). Cumulative OS did not differ according to age, gender and TLC (p = 0.526, 0.594 and 0.513, respectively). Cumulative OS at 2 years was 36% for B ALL (n: 39) versus 77% for TALL (n: 18) (p = 0.016) and was 49% for Philadelphia chromosome (Ph) negative patients versus 0% for Ph-positive patients (p < 0.001). Regarding MRD1, OS at 2 years was 18% for MRD1 HR (n: 17) versus 65% for MRD1 LR (n: 38) (p < 0.001). OS was 35% for high-risk patients (n: 30) and 62% for low-risk patients (n: 27) classified according to GMALL risk stratification (p = 0.017). CONCLUSION: MRD by FCM is a strong independent predictor of outcome in terms of DFS and OS and is a powerful informative parameter in guiding individual treatment in ALL patients.

The prognostic impact of some cell cycle regulatory proteins in Egyptian breast cancer patients.

PURPOSE: The particular goal of this work is to study some cell cycle regulatory proteins and their potential impact on prognosis of breast cancer; p53, cyclin D1 and p27 are potential effectors being the major contributors to the control of the restriction (R) check point of the cell cycle. We also aimed to evaluate different techniques used to detect these cell cycle proteins. MATERIAL AND METHODS: Forty five breast cancer patients as well as 10 controls with non malignant pathology were assessed for cell cycle regulators each by 2 different techniques; p53 was assessed by enzyme immunoassay (EIA) and immunohistochemistry (IHC), cyclin D1 by Western Blotting (WB) and IHC and p27 by WB and IHC. The cut-off was calculated as the mean of the normal controls +2 SD. Patients were followed up for 4 years and their laboratory data were correlated with different clinical parameters and with other studied regulators. RESULTS: Eighty seven percent of cases (39/45) were positive for p53 by EIA with a range from 20 to 4300, and a mean of 464 +/- 971 pg/mg protein. By IHC, 80% (24/30) of the cases showed varying degrees of positivity. Using WB, cyclin D1 showed high expression levels above cut off values in 69% of patients (31/45) and in 67% (20/30) by IHC. The corresponding positive figures for p27 were 82% (37/45) and 73% (22/30) using the two techniques, respectively. No significant association was found between p53, cyclin D1 and p27 on one side and different clinical parameters as lymph node status, tumor size or presence of distant metastases on the other side. Survival was poor in patients with high p53 expression. Cyclin D1 positive cases showed comparable survival with negative cases, whereas high p27 levels favored a longer disease free survival. CONCLUSIONS: Techniques more suitable for assessment of each of these markers in our consideration were EIA for p53, WB for cyclin D1 and IHC for p27. Moreover, this study demonstrated that these markers were relevant to the biological behavior of the tumor cell per se with a possible impact on prognosis and survival, independent of other clinical prognostic factors.