Chronic Myelogenous Leukemia Research Articles

Abstract 6918: Ba/F3-BCR-ABL engineering cell line panel, a useful platform for novel drug discovery

Abstract In the past few years, with the great success of Imatinib, the first kinase inhibitor, Imatinib rapidly became the first-line treatment for CML (Chronic myelogenous leukemia) carrying the BCR-ABL mutation. It opened a new chapter in targeted cancer treatment. Despite the increase in overall survival allowed by imatinib, the onset of drug resistance led scientists to investigate imatinib's fine structural mechanism of action to develop new and more effective compounds against mutated forms of Bcr-Abl. 7 BCR-ABL inhibitors have been approved, they are Imatinib, Dasatinib, Nilotinib, Bosutinib, Ponatinib, Olverembatinib, and Asciminib. However, acquired resistance has been a major challenge in the treatment of CML, and some patients still experience disease progression or even death due to drug resistance. BCR-ABL kinase domain mutations are one of the important mechanisms of acquired resistance, with the T315I mutation being the most common type of drug resistance mutation, accounting for approximately 25% of drug-resistant CML cases. Patients with T315I mutations in CML are resistant to all current first- and second-generation BCR-ABL inhibitors. Asciminib is designed as an allosteric inhibitor combined with an ATP-competitive TKI agent, which may hinder the development of mutation-driven acquired resistance or restore efficacy against highly resistant BCR-ABL mutants. Our group has generated 38 Ba/F3-BCR-ABL engineering cell lines as useful models for novel drug discovery in vitro and in vivo, including wild-type BCR-ABL and inhibitors leading drug resistance mutations, such as T315I, E255, Y253, F317, F359. V299, G250, A337T, V468F, and double mutations. Each cell was validated by cell sequencing and cell efficacy verification using approved BCR-ABL inhibitors. The Ba/F3-BCR-ABL engineering cell line panel can be useful for developing and evaluating next-generation BCR-ABL inhibitors and exploring newly acquired resistance mutations in BCR-ABL. Citation Format: Guoqian Wang, Yao Peng, Yu Wang, Yao Tang, Jingxiao Xu, Jinying Ning, Feng Hao. Ba/F3-BCR-ABL engineering cell line panel, a useful platform for novel drug discovery [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6918.

Abstract 1320: Engineered expression of HLA-E and HLA-G protects iPSC-derived cells from killing by primary NK cells

Abstract Background: A major challenge to off-the-shelf allogeneic cell therapies is the HLA-targeted elimination of genetically dissimilar cells. To combat this, HLA Class I can be deleted through transgene insertion at the beta-2 microglobulin (B2M) locus, while HLA Class II can be deleted by disruption of the gene encoding MHC Class II Transactivator (CIITA). Lack of HLA Class I expression, however, marks cells for elimination by natural killer (NK) cells, which normally receive inhibitory signals from this protein family. HLA-E and HLA-G are non-classical HLA Class I molecules that are far less polymorphic than HLA-A/B/C. They can inhibit natural killer cell-mediated activation by engaging NKG2A, KIR2DL4, ILT2, and ILT4. The objective of this study was to assess allo-evasion from NK cells by iPSC-derived cells engineered to overexpress HLA-E and -G. Methods: Allo-evasion by HLA-E, -G or both was first assessed in the chronic myelogenous leukemia K562 cell line, which expresses very low HLA Class I and is highly susceptible to NK-mediated killing. K562 cells were engineered using CRISPR/CAS9 to express chimeric molecules encoding either HLA-E or HLA-G heavy chains tethered to a defined nominal self-peptide and B2M, or HLA-E/peptide/B2M-P2A-HLA-G/peptide/B2M. Along with parental K562, these were used as targets in co-cultures with allogeneic PBMCs (n=22 donors) at different E:T ratios. Target killing was observed using flow cytometry and normalized to parental K562. Phenotyping of effector PBMCs for NK frequency and expression of HLA-E and -G receptors (NKG2A, KIR2DL4, ILT2, and ILT4) was performed to study correlations with killing efficiency and allo-evasion. IPSCs with endogenous B2M knocked out were also engineered to overexpress HLA-E and -G with the constructs described above. These were then differentiated into NK cells and gamma delta T cells using proprietary processes. Allo-evasion of these cells from killing by the same PBMC cohort was evaluated by flow cytometry. Results: While NK cells across donors expressed heterogeneous combinations of HLA-E and -G receptors, the strongest correlation with PBMC allo-responsiveness was NK frequency. K562 and iPSC-derived cells lacking HLA were susceptible to killing by PBMCs. Overexpression of HLA-E and -G offered protection to K562 and iPSC-derived cells against all tested donors. HLA-E offered more protection than HLA-G, and the combination of both HLA-E and -G was most potent. Conclusion: When genetically dissimilar HLA Class I protein family must be deleted to prevent graft rejection, expression of the more-conserved HLA-E and -G can effectively protect allogeneic drug products from elimination. These Allo-evasionTM edits may increase the in vivo persistence and enable multi-dosing of our iPSC-derived allogeneic cell therapies. Citation Format: Andriana Lebid, Dae Hwan Kim, Gabrielle Greco, Heidi Jessup, Alyssa Suarez, Shelby Brown, Nicholas Alexander, Mitan Desai, Buddha Gurung, Barry Morse, Daniel J. Perry, Michael F. Naso, Hyam Levitsky. Engineered expression of HLA-E and HLA-G protects iPSC-derived cells from killing by primary NK cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1320.

Porous Zinc Oxide and Functionalized Multiwalled Carbon Nanotubes Composite as Electrode Material for Enhanced Electrochemical Sensing of an Anticancer Drug, Dasatinib

An anticancer drug, dasatinib (DAS), is used to treat prostate cancer, chronic myelogenous leukaemia, and acute lymphoblastic leukaemia. In the present work, we describe the preparation of zinc oxide (ZnO) nanoparticles (NPs) using Averrhoa bilimbi leaf extract. These ZnO NPs were combined with functionalized multi-walled carbon nanotubes (f-MWCNT) to prepare a ZnO@f-MWCNT nanocomposite for electrochemical sensing of DAS. The prepared electrode materials, ZnO NPs, f-MWCNT, and ZnO@f-MWCNT, were characterised by powder X-ray diffraction (XRD), scanning electron microscopy (SEM), the Fourier-transform infrared (FT-IR) spectroscopy, atomic force microscopy (AFM), zeta potential, and the dynamic light scattering (DLS) techniques. DAS exhibited a significant enhancement (18-fold) in the oxidation peak current at ZnO@f-MWCNT, 10-fold at f-MWCNT, and 6-fold at ZnO NPs as compared to that at bare GCE. Under optimised conditions, a linear relationship was noticed among the peak current and concentration of DAS in the range of 0.03–82.65 μM and 0.01–122.45 μM for the differential pulse voltammetric (DPV), and square wave voltammetric (SWV) methods, respectively. Further, the applicability of the proposed sensor was demonstrated by analysing pharmaceutical and biological samples containing DAS. Higher percentage recovery values, and lower relative standard deviation (RSD) values highlighted the accuracy and precision proposed electrochemical methods.

Novel Mn(II) and Cu(II) metal complexes with sulfa drug-derived ligands as potent antimicrobial and anticancer agents: In vitro studies, ADMET profile and molecular docking

The present work deals with the synthesis, characterization, and pharmaceutical potential evaluation of new biologically active Mn(II) and Cu(II) complexes with Schiff bases derived from sulfamethoxazole. The coordination mode of the complexes, geometry, and general formula were determined using elemental analysis, emission atomic spectroscopy, molar conductance, magnetic measurements, TGA, FTIR, electronic absorption spectroscopy, SEM-EDX, and EPR spectroscopy. The results indicate that the Schiff base act as bidentate ligands binding to the metal ions via the nitrogen atoms of the sulfonamide and oxazole groups which leads to a distorted octahedral geometry around the central metals.The in vitro antimicrobial properties of the Schiff base ligands and their respective Mn(II) and Cu(II) complexes were evaluated against Gram-negative and Gram-positive (Escherichia coli klebsiella pneumoniae, Staphylococcus aureus, and Bacillus subtilis)) bacteria, and two fungal strains (Candida albicans, Aspergillus niger). The results assert that the complexes display excellent antimicrobial properties with low MIC values. Additionally, DPPH-scavenging assay results suggest that the coordination compounds could act as efficient antioxidant species.The in vitro cytotoxicity assay conducted on chronic myelogenous leukaemia cell lines (K562) showed that the coordination compounds exhibit potent anticancer activities. Furthermore, the pharmaceutical properties, including pharmacokinetics and toxicity, were examined by ADMET simulations and molecular docking studies were employed to assess interactions between the complexes and drug targets. The in vitro experimental data and in silico computational analysis revealed that the ligands and their respective coordination compounds as noteworthy candidates for drug development.

Synthesis of Novel Nilotinib Analogues and Biological Evaluation of Their Antiplatelet Activity and Functionality towards Cancer Cell Proliferation In Vitro.

Nilotinib, a second-generation tyrosine kinase inhibitor for the treatment of chronic myelogenous leukemia (CML), inhibits Bcr-Abl tyrosine kinase activity and proliferation of Bcr-Abl-expressing cells, as well as other malignancies. In the present study, new nilotinib analogues were synthesized and fully characterized. A platelet aggregation assay was performed, and the expression of P-selectin and PAC-1, as well as the effect on the proliferation of healthy endothelial cells, were evaluated. The expression and antimetastatic effects of E-cadherin and N-cadherin were assessed. The analogues inhibited platelet aggregation in a statistically significant manner compared to nilotinib, while they exhibited a strong inhibitory effect on P-selectin and PAC-1 expression when activated by AA. All three analogues caused arrest in the mitosis phase of the HepG2 cell cycle, while analogue-1 exhibited the most potent apoptotic effect compared to nilotinib. Interestingly, none of them promoted apoptosis in HUVECs. All the analogues reduced the expression of E- and N-cadherin in different amounts, while the analogues-1 and -3 exhibited similar antimigratory effects on HepG2 cells. The results of this study reveal considerable potential to develop new tyrosine kinase inhibitors with improved antiplatelet and antitumor properties.

Hydroxychavicol sensitizes imatinib-resistant chronic myelogenous leukemia cells to TRAIL-induced apoptosis by ROS-mediated IAP downregulation: Erratum.

Hydroxychavicol sensitizes imatinib-resistant chronic myelogenous leukemia cells to TRAIL-induced apoptosis by ROS-mediated IAP downregulation: Erratum.

Identification, Isolation, and Characterization of a Novel Degradation Impurity of Busulfan Using Preparative Chromatography, NMR, and LC-MS.

Busulfan is the most effective medication for treating chronic myelogenous or granulocytic leukemia because it has cytotoxic properties that harm or kill hematopoietic cells. It cannot absorb light in the Ultraviolet range due to its structure. Because of this, it is very challenging to quantify using traditional HPLC coupled with UV/Photodiode Array detectors. So, using sodium diethyldithiocarbamate, a derivatization method was developed to quantify related impurities. A significant unknown impurity was identified in derivatized samples of busulfan and a noticeably high percentage level was discovered during routine drug testing. We aimed to isolate, and characterize the unknown impurity observed in the samples and to identify its root cause. Preparative HPLC was used to isolate the unidentified, derivatized impurity, and 1H NMR, 13C NMR, and MS were used to decipher its structural components. The spectral characterization data analysis showed that the unknown impurity was related to busulfan. Additionally, it was noted that the impurity developed as a result of the residual buffer used to prepare the derivatizing reagent. The isolated impurity was found to be same as comparable to that found in busulfan drug substances, according to the results of the characterization tools. An alternative method of reagent preparation was optimized and deemed satisfactory because the buffer used in reagent preparation is the only factor contributing to the formation of impurities. Using cutting-edge analytical characterization tools, it was possible to explain the structural characteristics of an unknown impurity and discover that it was a novel impurity, which undoubtedly contributes to the comprehension of drug substance reaction properties.

Immunological and Molecular Investigation of BCR-ABL1 in Samples of Iraqi Chronic Myeloid Leukemia Patients

Background: Chronic myelogenous leukemia (CML) is a blood malignancy that affects hematopoietic stem cells. Also, it is commonly identified by the Philadelphia chromosome. This chromosome produces a protein with strong tyrosine kinase activity and functions as a tumorigenic factor. Objective: This study aimed to investigate the Immunological and Molecular of BCR-ABL1 gene expression and ABL1 protein in patients admitted to the Oncology and Hematology Center of the Kirkuk Governorate, Iraq. Materials and Methods: A study was conducted in Kirkuk Governorate, Iraq, from the beginning of November 2022 to May 2023 on Chronic Myeloid Leukemia patients in the Oncology and Hematology Center, which was hospital-based and cross-sectional. The study comprised fifty-three patients with Chronic Myeloid Leukemia. They were diagnosed using a complete blood count (CBC), a molecular test BCR-ABL1, and an immunological test ABL1 protein. The research did not include patients with missing data and those without consent. Fifty-three peripheral blood samples from 53 CML patients at chronic stages of CML were taken, and using automated Gene Xpert BCR-ABL1 Ultra assay, BCR-ABL1 transcript quantification was conducted on them. Results: The research included fifty-three patients in total (27 males, 28 females), with a mean age of 47.3 12.8 years. The average of BCR\_ABL1 by RT\_ qPCR 3.22 0.09 and the mean of ABL1 by Enzyme-linked Immune sorbent assay (ELISA) reigned is 981 891. Conclusion: Values of the quantitative real-time reverse transcription polymerase chain reaction (RT\_ qPCR) by Gene Xpert Ultra level range between 38.62 and 0.00 with a mean of 3.22 0.09. The RT\_ qPCR level showed that 22% of the patients were in good condition, 11% of them had a strong response to treatment, and other 11% showed a complete cytogenetic response (CCyR), which is also a good outcome; however, 78% of the patients either not taking their medication, or there is true resistance. These two methods used different techniques; the former is a nuclear-derived method that detects the presence of specific genetic materials of any pathogen; in the current research, BCR-ABL1 is the target. The latter is a test that detects and measures antibodies, antigens, proteins, and glycoproteins in biological samples, so ABL1 is the target.

The Mozart effect in chronic myeloid leukaemia.

Clinical research has not been able to establish whether the differences between first- and second-generation BCR-ABL 1 kinase inhibitors are clinically relevant with regard to outcome. In the study by Alcazer etal., a relevant difference seems to emerge-paradoxically in the absence of the drugs-as demonstrated by differences in the relapse kinetics after cessation of therapy. Commentary on: Alcazer etal. Kinetics of molecular recurrence after tyrosine kinase inhibitor cessation in chronic phase chronic myelogenous leukaemia patients. Br J Haematol 2024;204:1536-1539.

Biochips to measure ultraviolet light absorption by leukemia cells

During the treatment of leukemia, side effects create a great burden for patients. Therefore, new therapies to replace conventional chemotherapy and hematopoietic stem cell transplantation are highly desirable. In a previous study, I found that UV irradiation at 365 nm induced apoptosis in 98% of leukemia (Jurkat T) cells. To determine the most suitable wavelength for such treatments, it is necessary to measure the UV light absorption of cancer cells. In this study, a biochip was fabricated and evaluated for this purpose. Three types of leukemic cells (human acute myelogenous leukemia cells, human acute lymphoblastic leukemia cells, and human chronic myelogenous leukemia cells) were injected into the biochip, and light absorption was measured, revealing that individual leukemic cells have different light absorption characteristics. This variation is important in order to develop an implant care device to treat leukemia more effectively.

Evaluation the Anti-leukemia effect of Methanol extracts of Camellia cuongiana on the K562 cell line

This study aims to examine the anti-proliferation effects of Camellia cuongiana methanolic extracts on the K562 chronic myelogenous leukaemia cell line. The crude extract was obtained by soaking the sample with methanol, following concentration using a rotary evaporator. Trypan blue exclusion test was performed in five concentrations of extract to detect the anti-cancer characteristic of Camellia cuongiana. Morphological changes in the treatment cell were observed using a phase-contrast microscope. After 72 hours of treatment, the extracts inhibited cell proliferation, with IC50 values of 72.26±5.75µg/mL. Increasing the extract concentration decreased the number of living cells. The most frequently observed abnormal morphology was cell shrinkage. Evidently, Camellia cuongiana could inhibit cancer cell proliferation and induce programmed cell death.

Inhibition of LATS kinases reduces tumorigenicity and increases the sensitivity of human chronic myelogenous leukemia cells to imatinib

Chronic myelogenous leukemia (CML) is a clonal hematologic malignancy of the myeloid lineage caused by the oncogenic BCR/ABL fusion protein that promotes CML cell proliferation and protects them against drug-induced apoptosis. In this study, we determine LATS1 and LATS2 expression in CML cells derived from patients who are resistant to imatinib (IM) treatment. Significant upregulation of LATS1 and LATS2 was found in these CML patients compared to healthy donors. To further explore whether the expression of LATS1/2 contributes to the IM-resistant phenotype, IM-resistant CML cell lines generated by culturing CML-derived erythroblastic K562 cells in increasing concentrations of IM were used as in vitro models. Up-regulation of LATS1 and LATS2 was observed in IM-resistant K562 cells. Reduction of LATS using either Lats-IN-1 (TRULI), a specific LATS inhibitor, or shRNA targeting LATS1/2 significantly reduced clonogenicity, increased apoptosis and induced differentiation of K562 cells to late-stage erythroid cells. Furthermore, depletion of LATS1 and LATS2 also increased the sensitivity of K562 cells to IM. Taken together, our results suggest that LATS could be one of the key factors contributing to the rapid proliferation, reduced apoptosis, and IM resistance of CML cells. Targeting LATS could be a promising treatment to enhance the therapeutic effect of a conventional BCR/ABL tyrosine kinase inhibitor such as IM.

Effect of Extracellular Vesicles Derived From Peripheral Blood Mononuclear Cells on K562 Leukemia Cell Line

Recently, extracellular vesicles (EVs) are generating considerable interest in terms of their ability. EVs defined themselves as a route for intercellular communication between the origin cells and the recipient ones by transferring information. This paper investigates the influence of peripheral blood mononuclear cells derived EVs on proliferation and apoptosis of the chronic myelogenous leukemia cell line k562. Trypan blue staining was used to detect cell viability subsequently, metabolic activity was assessed by the MTT assay. Cell apoptosis and cell cycle progression were evaluated using flowcytometric assay after treatment of k562 cell line with MNC derived EVs. Our results showed that MNC-EVs have no inhibitory effect on k562 cell growth and proliferation. Our data did not reveal any significant variations in the case of enhancing k562 cell line growth following treatment with MNC derived EVs. It seems tumour-derived extracellular vesicles support tumour cells growth by communicating with each other through their extracellular vesicles.

Stem-cells Research and Therapies

Studies with leucogenenol, a compound isolated from the metabolic products of Penicillium gilmanii and human liver showed that leucogenenol accelerated the rate at which the “functional” cells, neutrophils, red blood cells and lymphocytes developed in the bone marrow from their corresponding committed precursor cells. It was found that the neutrophils that appear in the peripheral blood circulation following injection of leucogenenol originated from the neutrophils in the bone marrow. A scheme was presented to show that leucogenenol and adroxazine play key roles in regulating the circulation of blood cells. Stem cells act as the building blocks for the tissues and organs of physiologic systems. Research studies have shown stem-cells potential to treat a wide range of diseases. The Stem-cells play important roles in the treatment of various medical conditions and monitoring of bodily functions. Stem-cell therapy has shown results in treating conditions via peripheral blood circulation in the growth of new blood vessels such as heart disease, diabetes, cancer, Parkinson’s disease. Clinical trials are being conducted to assess the safety and efficacy of stem cell based-therapies in treating various diseases such as Chronic Myelogenous Leukemia (CML) which is a type of blood cancer that affects the bone marrow and blood cells characterized by the abnormal growth of white blood cells that affects the bone marrow and blood cells known as myeloid cells. Stem cell therapies have great potential to become one of the most important tools of regenerative medicine and untreatable diseases. In this review, different types of stem-cell-based therapies are described, which are used in treating various diseases.
 Keywords: Blood Stem cells, therapies, clinical trials, CRISPR Therapeutics

New Derivatives of 1-(3-Methyl-1-Benzofuran-2-yl)Ethan-1-one: Synthesis and Preliminary Studies of Biological Activity.

A set of nine derivatives, including five brominated compounds, was synthesized and the structures of these novel compounds were confirmed using 1H and 13C NMR as well as ESI MS spectra. These compounds were tested on four different cancer cell lines, chronic myelogenous leukemia (K562), prostate cancer (PC3), colon cancer (SW620), human kidney cancer (Caki 1), and on healthy human keratocytes (HaCaT). MTT results reveal that two newly developed derivatives (6 and 8) exhibit selective action towards K562 cells and no toxic effect in HaCat cells. The biological activity of these two most promising compounds was evaluated by trypan blue assay, reactive oxygen species generation, and IL-6 secretion. To investigate the proapoptotic activity of selected compounds, the two following types of tests were performed: Annexin V Apoptosis Detection Kit I and Caspase-Glo 3/7 assay. The studies of the mechanism showed that both compounds have pro-oxidative effects and increase reactive oxygen species in cancer cells, especially at 12 h incubation. Through the Caspase-Glo 3/7 assay, the proapoptotic properties of both compounds were confirmed. The Annexin V-FITC test revealed that compounds 6 and 8 induce apoptosis in K562 cells. Both compounds inhibit the release of proinflammatory interleukin 6 (IL-6) in K562 cells. Additionally, all compounds were screened for their antibacterial activities using standard and clinical strains. Within the studied group, compound 7 showed moderate activity towards Gram-positive strains in antimicrobial studies, with MIC values ranging from 16 to 64 µg/mL.

Correlation analysis of bone marrow microvessel density and miRNA expression on drug resistance in patients with chronic myelogenous leukemia after tyrosine kinase inhibitor treatment

ABSTRACT Objective: This study analyzed the relationship between bone marrow microvessel density (MVD) and the expression of four miRNAs with chronic myelogenous leukemia (CML) resistance after tyrosine kinase inhibitor (TKI) treatment. Methods: 234 CML patients were divided into resistance and non-resistance groups in terms of the results of the 5-year follow-up. Patients were divided into the Optimum response group and the Warning/Failure group based on TKI response. MVD was determined by immunohistochemistry, and the expression levels of four miRNAs (miR-106a, miR-155, miR-146a, and miR-340) in bone marrow biopsy specimens were examined by qPCR. We evaluated the association of MVD with four miRNAs and them predictive value for CML resistance after TKI treatment. Results: The MVD and the levels of miR-106a, miR-155, and miR-146a were significantly higher while the miR-340 level was lower in the resistance group than the non-resistance group. Besides, MVD had a significant correlation with the levels of miR-340 and miR-155. According to the results of survival analysis, MVD as well as miR-340 and miR-155 levels were observably correlated with 5-year survival of patients without TKI resistance. The results of the ROC curve indicated that the MVD, miR-106a, miR-340, and miR-155 had good predictive accuracy for CML resistance after TKI treatment. As for the results of multivariate analysis, disease stage, risk level (high risk), high MVD, low miR-340 expression, and high miR-155 expression were all independent risk factors for CML resistance. Conclusion: MVD and the expression of miR-340 and miR-155 are closely associated with CML resistance after TKI treatment.

Asciminib a new player in treatment of TKI-resistant/intolerant chronic phase chronic myelogenous leukemia

Asciminib a new player in treatment of TKI-resistant/intolerant chronic phase chronic myelogenous leukemia

"Combination treatments of imatinib with astaxanthin and crocin efficiently ameliorate antioxidant status, inflammation and cell death progression in imatinib-resistant chronic myeloid leukemia cells".

Imatinib resistance remains a major obstacle in the treatment of chronic myelogenous leukemia (CML). Crocin (CRC) and astaxanthin (ATX) are phytochemicals with anti-cancer properties. This study aimed to explore the effects of combination treatment of Imatinib with CRC and ATX on Imatinib-resistant K562 (IR-K562) cells. After the establishment of IR-K562 cells, growth inhibitory activity was determined by the MTT assay. To test the regeneration potential, a colony formation assay was performed. Cell cycle analyses were examined by flow cytometry. Cell injury was evaluated by lactate dehydrogenase (LDH) leakage. Real-time PCR was applied to assess the expression of IL6, TNF-α, STAT3, BAD, CASP3, TP53, and Bcl-2 genes. Caspase-3 activity was determined by a colorimetric assay. Antioxidant activity was measured using a diphenylpicrylhydrazyl (DPPH) assay. After 48h of treatment, ATX (IC50 = 30µM) and CRC (IC50 = 190µM) significantly inhibited cell proliferation and colony formation ability, induced G1 cell cycle arrest and cell injury, upregulated the expression of apoptosis-associated genes, and downregulated the expression of anti-apoptotic and inflammatory genes. The combination of IM with ATX and/or CRC synergistically reduced cell viability (combination index [CI] < 1). Our data suggest that IM shows better therapeutic efficacy at lower doses when combined with ATX and/or CRC.

SiRNA-mediated downregulation of BATF3 diminished proliferation and induced apoptosis through downregulating c-Myc expression in chronic myelogenous leukemia cells.

Despite considerable improvement in therapeutic approaches to chronic myeloid leukemia (CML) treatment, this malignancy is considered incurable due to resistance. However, investigating the molecular mechanism of CML may give rise to the development of extremely efficient targeted therapies that improve the prognosis of patients. Basic leucine zipper transcription factor ATF-like3 (BATF3), as transcription factor, is considered a key regulator of cellular activities and its function has been evaluated in tumor development and growth in several cancer types. This study aimed to evaluate the potential of the cellular impact of siRNA-mediated downregulation of BATF3 on CML cancer cells through cell proliferation, induction of apoptosis, and cell cycle distribution. The transfection of BATF3 siRNA to K562 CML cells was performed by electroporation device. To measure cellular viability and apoptosis, MTT assay and Annexin V/PI staining were carried out, respectively. Also, cell cycle assay and flow cytometry instrument were applied to assess cell cycle distribution of K562 cells. For more validation, mRNA expression of correlated genes was relatively evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). The data indicated that siRNA-mediated BATF3 inactivating severely promoted the cell apoptosis. Also, the targeted therapy led to high expression of Caspase-3 gene and Bax/Bcl-2 ratio. Silenced BATF3 also induced cell cycle arrest in phase sub-G1 compared to control. Finally, a noticeable decrement was obtained in c-Myc gene expression through suppression of BATF3 in CML cells. The findings of this research illustrated the suppression of BATF3 as an effective targeted therapy strategy for CML.

Approach to the Diagnosis and Management of Essential Thrombocytosis in a Resource-Limited Setting

Essential thrombocytosis (ET) is a myeloproliferative neoplasm (together with polycythemia vera, chronic myelogenous leukemia, and primary myelofibrosis) characterized by clonal proliferation of megakaryocytes, usually due to the presence of JAK2, CALR, or MPL mutations. Whereas a bone marrow aspirate and genetic testing for these mutations are necessary for an accurate diagnosis of ET, in resource-limited settings, these are often either inaccessible or limited by prohibitive costs. A nuanced clinical approach is necessary to diagnose and manage ET in these settings. In this study, we present a case of ET diagnosed and managed in a resource-limited rural setting in Kenya with hydroxyurea and aspirin. We highlight the importance of individualized therapeutic targets, the challenges with dose adjustments in the setting of hydroxyurea-induced neutropenia, and how extrapolated data from the use of hydroxyurea elsewhere may help guide such dose adjustments. Finally, we propose a rationalized approach to treating ET in a resource-limited clinical setting.