The aim of this study is to observe cell proliferation, cell viability, apoptosis, and autophagy on acute myeloid leukemia (AML) cell lines, NB4 and U937, with the drug repurposing of mefloquine (MQ). Methods such as the 3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay and trypan blue staining have shown a decrease in live cells with high concentrations of mefloquine. Using their average perspective IC50 values of MQ concentration, Western blotting was applied by means of apoptosis and autophagy markers to determine if the induction of apoptosis and inhibition of autophagy was present in MQ-treated AML cells. The experiment will be continued with more cell lines, drugs, and other means of protocol in order to contribute to cancer therapy.https://scholarscompass.vcu.edu/uresposters/1285/thumbnail.jp

Corey, Seth

Garg, Bhavuk

Mehta, Hrishikesh

Phan, Jessica L.

English

VCU Scholars Compass

Virginia Commonwealth UniversityVCU Scholars CompassUndergraduate Research Posters Undergraduate Research Opportunities Program2018The Evaluation of Mefloquine Drug Repurposingon Acute Myeloid LeukemiaJessica L. PhanVirginia Commonwealth UniversityBhavuk GargMassey Cancer Center of Virginia Commonwealth UniversityHrishikesh MehtaMassey Cancer Center of Virginia Commonwealth UniversitySeth CoreyMassey Cancer Center of Virginia Commonwealth UniversityFollow this and additional works at: https://scholarscompass.vcu.edu/urespostersPart of the Chemical Actions and Uses Commons, Hematology Commons, Hemic andLymphatic Diseases Commons, Medical Pathology Commons, Oncology Commons, and thePediatrics Commons© The Author(s)This Book is brought to you for free and open access by the Undergraduate Research Opportunities Program at VCU Scholars Compass. It has beenaccepted for inclusion in Undergraduate Research Posters by an authorized administrator of VCU Scholars Compass. For more information, pleasecontact libcompass@vcu.edu.Recommended Citation[1] Arakawa, S., S. Honda, H. Yamaguchi, and S. Shimizu. 2017 Molecular mechanisms and physiological roles of Atg5/Atg7-independent alternative autophagy. Journal List. 93(6): 378–385. [2] Eriksson, A., E. Chantzi, M. Fryknäs, J. Gullbo, P. Nygren,et al. 2017 Towards repositioning of quinacrine for treatment of acute myeloid leukemia - Promising synergies and in vivo effects.Leukemia research. 63:41-46. [3] Folkerts, H., S. Hilgendorf, A.T.J. Wierenga, J. Jaques, A. B. Mulder et al. 2017 Inhibition ofautophagy as a treatment strategy for p53 wild-type acute myeloid leukemia. Cell Death and Disease. 8(7):e2927. [4] Sharma, N., S.Thomas, E.B. Golden, F. M. Hofman, T. C. Chen et al. 2012 Inhibition of autophagy and induction of breast cancer cell death bymefloquine, an antimalarial agent. Cancer Letters. 326(2):143-54 [5] Staines, H.M., B.C. Dee, M.R. Shen, and J.C. Ellory. 2004 Theeffect of mefloquine and volume-regulated anion channel inhibitors on induced transport in Plasmodium falciparum-infected humanred blood cells. Blood Cells Mol Dis. 32(3):344-8The Evaluation of Mefloquine Drug Repurposing on Acute Myeloid LeukemiaJessica L. Phan, BS1; Bhavuk Garg, PhD2; Hrishikesh M. Metha, PhD3; Seth J. Corey, MD, PhD41Department of Biology, Virginia Commonwealth University, Richmond, Virginia, USA 2,3,4Division of Pediatric Hematology Oncology and Stem Cell Transplantation, Massey Cancer Center of Virginia Commonwealth University, Richmond, VA, USAMTT Assay1. Serial dilute 50 mM stock solution of MQ with DMSO. Seed NB4 and U937 with RPMI media (10% FBS, 1% PS) at 2 x 105 cells/mL and 1 x 105 cells/mL, respectively. Total volume per well: 100 µL.2. Incubate plates for 48 hours. Add 10 µL of MTT reagent, incubate for 4 hours, then add 100 µL of detergent. 3. Store in the dark for 24 hours. Record absorbance at 595 nm and determine IC50.Trypan Blue Staining1. Perform similar pre-incubation steps as the MTT protocol2. After 48 hours, perform a cell count with Trypan Blue to view cell viability at increasing MQ concentrations (include IC50s: 7 µM for NB4 and 8 µM for U937)Western Blot1. Prepare lysates of NB4 and U937 cells with MQ treatment for 0,  2, 4, and 24h. Run gel electrophoresis.2. Perform electrotransfer of the proteins using polyvinylidene fluoride (PDVF) membrane 3. Wash membrane in Ponceau S staining solution. Block membrane in 10% milk (with TBST) to rid of background “noise.” 4. Wash in 5% milk with the primary antibody (1:1000) overnight in 4oC shaker. Wash membrane three times in TBST for 10 min. 5. Wash in 5% milk with secondary antibody (1:3000) for 1 hour. Wash membrane three times in TBST for 10 min.6. Prepare ECL mix (1:2 dilution of solution A and B).7. Visualize results in dark room for chemiluminescent detection.MTT Dose Response Results (48h)Trypan Blue Results (48h)Statistical Comparisons:Fig. 2 - Standard deviations (SD) and standard error of means (SEM) displayed in the table served as separate comparisons between the MTT and trypan blue assay trials from the graphs. The two-tailed p-value in paired t-tests were then computed to see the statistical significance between MTT and trypan blue; p-values were less than 0.05 for both NB4 and U937.BACKGROUNDAcute leukemia is a fast-growing blood cancer with an early onset in hematopoietic stem cells. DNA mutations block normal development in lymphoid and myeloid cells, leading to an accumulation of blast cells in the bone marrow. This can result in either acute lymphocytic leukemia (ALL) or acute myeloid leukemia (AML) as shown in Fig. 1. With 2-year survival rates of less than 20% in the elderly and 60% in children [2], AML became the main focus for this project.The growth maintenance of cancer cells depend partially on autophagy, a catabolic process that degrades and recycles unwanted proteins and organelles in the lysosome for cell survival. Based on previous studies, leukemic cells undergoing autophagy inhibition experience metabolic stress, triggering apoptosis [3]. The use of an antimalarial drug, mefloquine (MQ), has sparked interest in drug repurposing for leukemic cells due to its effective reduction of parasitemia from blood. Mefloquine’s current mechanism of action for malaria includes damaging the membrane of free heme, interacting with phosphatidylinositol, and affecting volume-regulated anion channels (VRAC) to bring toxicity to parasites in blood [5]. A high-throughput screening panel of leukemic cell lines identified mefloquine as an anti-leukemic compound by lysosome disruption, releasing cathepsins into the cytosol and causing permeability in the lysosome membrane [2].HYPOTHESIS The repurposing of mefloquine (MQ) as an anti-leukemic drug will result in decreased myeloid leukemia cell proliferation due to lysosome disruption, thereby inhibiting autophagy and inducing apoptosis. METHODSWe found that AML cells, NB4 and U937, had mean IC50 values respectively at 7µM and 8µM mefloquine (MQ) treatment through verification in MTT and trypan blue assays. The MTT assay uses tetrazolium dye reduction to measure metabolism as a marker for cell proliferation. Cells with increasing MQ concentration had the least metabolism by losing their ability to convert the MTT into formazan and revealing less color as quantified by absorbance. The trypan blue assay was used to evaluate anti-proliferative effects based on cell viability. The mean IC50 values between the MTT and trypan blue assays were statistically insignificant because p > 0.05 for both cell lines (Fig. 2). Having little difference in procedural methods, 7µM for NB4 and 8µM for U937 were used for future experiments. Western blotting was performed to detect protein expression using apoptosis and autophagy markers. These cells were treated at 0, 2, 4, and 24 hours of MQ treatment to investigate molecular processes before 48 hours. Apoptosis MarkersThe p53 tumor suppressor gene induces apoptosis by protease enzymes in the Caspase family. Cleaved Caspase-3 is needed in order to activate the proteolytic cleavage of PARP for the induction of apoptosis [4]. Since there was no Caspase-3 cleavage in the results, there was no induction of apoptosis for NB4 and U937 with MQ. PARP-1 repairs single-stranded DNA (ssDNA) breaks, helping to maintain cell viability. Unlike Caspase-3, there was an indication of PARP-1 cleavage, which allows for the likelihood of autophagy inhibition instead if cathepsins proteases are involved in autophagosomes [4]. Elevated levels of CHOP initiate apoptosis through ER stress. Since CHOP levels were inconsistent in the results, apoptosis may not have occurred. (Fig. 3)Autophagy MarkersAtg-5 and Atg-7 are important molecules for the induction of autophagy through the formation of autophagosomes [1]. Although only Atg-5 was cleaved, it served as an indicator of apoptosis. A different pathway may be involved since there was no cleavage of Atg-7. Due to the inclusion of p62 in signal transduction pathways and degradation of ubiquitinated proteins, having no p62 cleavage in the cell lines implies that autophagy was not inhibited in this pathway [4]. LC3, a marker of autophagosomes, is the main protein in the autophagy pathway. A higher protein expression in LC3B-II means increased chances for autophagy [4], so based on the results, LC3 lipidation is not a contributor to autophagy inhibition. (Fig. 3)CONCLUSIONS Although IC50 values were obtained for NB4 and U937 cells, there was little evidence that mefloquine (MQ) inhibited autophagy or induced apoptosis. A distinct DMSO control for Western blots, longer time points, IC90 trials, other cell lines, and different assays such as flow cytometry will have to be included to evaluate the drug repurposing of mefloquine. Additional anticancer drugs will also be tested. We hope that our results will eventually provide the rationale to continue with clinical trials.RESULTSRESULTS (cont.)Western Blot ResultsFig. 3 - Apoptosis Markers: PARP-1, Caspase-3, CHOP; Autophagy Markers: Atg-7, Atg-5, p62, LC3B DISCUSSION/CONCLUSIONS[1] Arakawa, S., S. Honda, H. Yamaguchi, and S. Shimizu.  2017  Molecular mechanisms and physiological roles of Atg5/Atg7-independent alternative autophagy.  Journal List. 93(6): 378–385.[2] Eriksson, A., E. Chantzi, M. Fryknäs, J. Gullbo, P. Nygren, et al.  2017  Towards repositioning of quinacrine for treatment of acute myeloid leukemia - Promising synergies and in vivo effects.  Leukemia research. 63:41-46.[3] Folkerts, H., S. Hilgendorf, A.T.J. Wierenga, J. Jaques, A. B. Mulder et al. 2017  Inhibition of autophagy as a treatment strategy for p53 wild-type acute myeloid leukemia. Cell Death and Disease. 8(7):e2927.[4] Sharma, N., S. Thomas, E.B. Golden, F. M. Hofman, T. C. Chen et al. 2012  Inhibition of autophagy and induction of breast cancer cell death by mefloquine, an antimalarial agent. Cancer Letters. 326(2):143-54[5] Staines, H.M., B.C. Dee, M.R. Shen, and J.C. Ellory.  2004  The effect of mefloquine and volume-regulated anion channel inhibitors on induced transport in Plasmodium falciparum-infected human red blood cells.  Blood Cells Mol Dis. 32(3):344-8REFERENCESPARP-1Caspase-3CHOPINTRODUCTION116 kDa89 kDa35 kDa27 kDap62LC3B-ILC3B-II62 kDa16 kDa14 kDaActin (control) 45 kDa78 kDa37 kDa55 kDaAtg-7Atg-5Cell Line Trypan Blue Mean IC50 (48h)NB4 7 µMU937 9.5 µMCell Line MTT Mean IC50 (48h)NB4 6.9 µM (~ 7 µM)U937 8.2 µM (~ 8 µM)Two-tailed p-value t-test value Standard Deviation (SD) Standard Error of Mean (SEM)P = 0.40 t = 1.1 (df = 2)MTT: 1.4Trypan Blue: 2.0MTT: 0.82Trypan Blue: 1.2Two-tailed p-value t-test value Standard Deviation (SD) Standard Error of Mean (SEM)P = 0.33 t = 1.3 (df = 2)MTT: 0.8Trypan Blue: 1.2MTT: 0.5Trypan Blue: 0.7NB4: MTT vs. Trypan Blue IC50 Values (48h)U937: MTT vs. Trypan Blue IC50 Values (48h)The aim of this study is to observe cell proliferation, cell viability, apoptosis, and autophagy on acute myeloid leukemia (AML) cell lines, NB4 and U937, with the drug repurposing of mefloquine (MQ). Methods such as the 3-(4,5-Dimethylthiazol-2-Yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay and trypan blue staining have shown a decrease in live cells with high concentrations of mefloquine. Using their average perspective IC50 values of MQ concentration, Western blotting was applied by means of apoptosis and autophagy markers to determine if the induction of apoptosis and inhibition of autophagy was present in MQ-treated AML cells. The experiment will be continued with more cell lines, drugs, and other means of protocol in order to contribute to cancer therapy.ABSTRACTLymphoidProgenitorMyeloidProgenitorMyeloblastLymphoblastErythrocytes (Red Cells)PlateletsBasophilNeutrophilEosinophilMonocyteT LymphocyteB LymphocyteALLAMLHematopoieticStem CellFig. 1 - Hematopoiesis for Acute Leukemia

The Evaluation of Mefloquine Drug Repurposing on Acute Myeloid Leukemia

Virginia Commonwealth University
VCU Scholars Compass
Undergraduate Research Posters Undergraduate Research Opportunities Program
2018
The Evaluation of Mefloquine Drug Repurposing
on Acute Myeloid Leukemia
Jessica L. Phan
Virginia Commonwealth University
Bhavuk Garg
Massey Cancer Center of Virginia Commonwealth University
Hrishikesh Mehta
Massey Cancer Center of Virginia Commonwealth University
Seth Corey
Massey Cancer Center of Virginia Commonwealth University
Follow this and additional works at: https://scholarscompass.vcu.edu/uresposters
Part of the Chemical Actions and Uses Commons, Hematology Commons, Hemic and
Lymphatic Diseases Commons, Medical Pathology Commons, Oncology Commons, and the
Pediatrics Commons
© The Author(s)
This Book is brought to you for free and open access by the Undergraduate Research Opportunities Program at VCU Scholars Compass. It has been
accepted for inclusion in Undergraduate Research Posters by an authorized administrator of VCU Scholars Compass. For more information, please
contact libcompass@vcu.edu.
Recommended Citation
[1] Arakawa, S., S. Honda, H. Yamaguchi, and S. Shimizu. 2017 Molecular mechanisms and physiological roles of Atg5/
Atg7-independent alternative autophagy. Journal List. 93(6): 378–385. [2] Eriksson, A., E. Chantzi, M. Fryknäs, J. Gullbo, P. Nygren,
et al. 2017 Towards repositioning of quinacrine for treatment of acute myeloid leukemia - Promising synergies and in vivo effects.
Leukemia research. 63:41-46. [3] Folkerts, H., S. Hilgendorf, A.T.J. Wierenga, J. Jaques, A. B. Mulder et al. 2017 Inhibition of
autophagy as a treatment strategy for p53 wild-type acute myeloid leukemia. Cell Death and Disease. 8(7):e2927. [4] Sharma, N., S.
Thomas, E.B. Golden, F. M. Hofman, T. C. Chen et al. 2012 Inhibition of autophagy and induction of breast cancer cell death by
mefloquine, an antimalarial agent. Cancer Letters. 326(2):143-54 [5] Staines, H.M., B.C. Dee, M.R. Shen, and J.C. Ellory. 2004 The
effect of mefloquine and volume-regulated anion channel inhibitors on induced transport in Plasmodium falciparum-infected human
red blood cells. Blood Cells Mol Dis. 32(3):344-8
The Evaluation of Mefloquine Drug Repurposing on Acute Myeloid Leukemia
Jessica L. Phan, BS1; Bhavuk Garg, PhD2; Hrishikesh M. Metha, PhD3; Seth J. Corey, MD, PhD4
1Department of Biology, Virginia Commonwealth University, Richmond, Virginia, USA 
2,3,4Division of Pediatric Hematology Oncology and Stem Cell Transplantation, Massey Cancer Center of Virginia Commonwealth University, Richmond, VA, USA
MTT Assay
1. Serial dilute 50 mM stock solution of MQ with DMSO. Seed NB4 and U937 with 
RPMI media (10% FBS, 1% PS) at 2 x 105 cells/mL and 1 x 105 cells/mL, 
respectively. Total volume per well: 100 µL.
2. Incubate plates for 48 hours. Add 10 µL of MTT reagent, incubate for 4 hours, then 
add 100 µL of detergent. 
3. Store in the dark for 24 hours. Record absorbance at 595 nm and determine IC50.
Trypan Blue Staining
1. Perform similar pre-incubation steps as the MTT protocol
2. After 48 hours, perform a cell count with Trypan Blue to view cell viability at 
increasing MQ concentrations (include IC50s: 7 µM for NB4 and 8 µM for U937)
Western Blot
1. Prepare lysates of NB4 and U937 cells with MQ treatment for 0,  2, 4, and 24h. 
Run gel electrophoresis.
2. Perform electrotransfer of the proteins using polyvinylidene fluoride (PDVF) 
membrane 
3. Wash membrane in Ponceau S staining solution. Block membrane in 10% milk 
(with TBST) to rid of background “noise.” 
4. Wash in 5% milk with the primary antibody (1:1000) overnight in 4oC shaker. 
Wash membrane three times in TBST for 10 min. 
5. Wash in 5% milk with secondary antibody (1:3000) for 1 hour. Wash membrane 
three times in TBST for 10 min.
6. Prepare ECL mix (1:2 dilution of solution A and B).
7. Visualize results in dark room for chemiluminescent detection.
MTT Dose Response Results (48h)
Trypan Blue Results (48h)
Statistical Comparisons:
Fig. 2 - Standard deviations (SD) and standard error of means (SEM) displayed in the table served as separate 
comparisons between the MTT and trypan blue assay trials from the graphs. The two-tailed p-value in paired t-tests 
were then computed to see the statistical significance between MTT and trypan blue; p-values were less than 0.05 
for both NB4 and U937.
BACKGROUND
Acute leukemia is a fast-growing blood cancer with an early onset in 
hematopoietic stem cells. DNA mutations block normal development in lymphoid and 
myeloid cells, leading to an accumulation of blast cells in the bone marrow. This can 
result in either acute lymphocytic leukemia (ALL) or acute myeloid leukemia (AML) 
as shown in Fig. 1. With 2-year survival rates of less than 20% in the elderly and 60% 
in children [2], AML became the main focus for this project.
The growth maintenance of cancer cells depend partially on autophagy, a catabolic 
process that degrades and recycles unwanted proteins and organelles in the lysosome 
for cell survival. Based on previous studies, leukemic cells undergoing autophagy 
inhibition experience metabolic stress, triggering apoptosis [3]. 
The use of an antimalarial drug, mefloquine (MQ), has sparked interest in drug 
repurposing for leukemic cells due to its effective reduction of parasitemia from blood. 
Mefloquine’s current mechanism of action for malaria includes damaging the 
membrane of free heme, interacting with phosphatidylinositol, and affecting volume-
regulated anion channels (VRAC) to bring toxicity to parasites in blood [5]. A high-
throughput screening panel of leukemic cell lines identified mefloquine as an anti-
leukemic compound by lysosome disruption, releasing cathepsins into the cytosol and 
causing permeability in the lysosome membrane [2].
HYPOTHESIS
 The repurposing of mefloquine (MQ) as an anti-leukemic drug will result in 
decreased myeloid leukemia cell proliferation due to lysosome disruption, thereby 
inhibiting autophagy and inducing apoptosis. 
METHODS
We found that AML cells, NB4 and U937, had mean IC50 values respectively at 7µM and 8µM mefloquine 
(MQ) treatment through verification in MTT and trypan blue assays. The MTT assay uses tetrazolium dye 
reduction to measure metabolism as a marker for cell proliferation. Cells with increasing MQ concentration had 
the least metabolism by losing their ability to convert the MTT into formazan and revealing less color as 
quantified by absorbance. The trypan blue assay was used to evaluate anti-proliferative effects based on cell 
viability. The mean IC50 values between the MTT and trypan blue assays were statistically insignificant because 
p > 0.05 for both cell lines (Fig. 2). Having little difference in procedural methods, 7µM for NB4 and 8µM for 
U937 were used for future experiments. Western blotting was performed to detect protein expression using 
apoptosis and autophagy markers. These cells were treated at 0, 2, 4, and 24 hours of MQ treatment to investigate 
molecular processes before 48 hours. 
Apoptosis Markers
The p53 tumor suppressor gene induces apoptosis by protease enzymes in the Caspase family. Cleaved 
Caspase-3 is needed in order to activate the proteolytic cleavage of PARP for the induction of apoptosis [4]. Since 
there was no Caspase-3 cleavage in the results, there was no induction of apoptosis for NB4 and U937 with MQ. 
PARP-1 repairs single-stranded DNA (ssDNA) breaks, helping to maintain cell viability. Unlike Caspase-3, there 
was an indication of PARP-1 cleavage, which allows for the likelihood of autophagy inhibition instead if 
cathepsins proteases are involved in autophagosomes [4]. Elevated levels of CHOP initiate apoptosis through ER 
stress. Since CHOP levels were inconsistent in the results, apoptosis may not have occurred. (Fig. 3)
Autophagy Markers
Atg-5 and Atg-7 are important molecules for the induction of autophagy through the formation of 
autophagosomes [1]. Although only Atg-5 was cleaved, it served as an indicator of apoptosis. A different pathway 
may be involved since there was no cleavage of Atg-7. Due to the inclusion of p62 in signal transduction 
pathways and degradation of ubiquitinated proteins, having no p62 cleavage in the cell lines implies that 
autophagy was not inhibited in this pathway [4]. LC3, a marker of autophagosomes, is the main protein in the 
autophagy pathway. A higher protein expression in LC3B-II means increased chances for autophagy [4], so based 
on the results, LC3 lipidation is not a contributor to autophagy inhibition. (Fig. 3)
CONCLUSIONS
 Although IC50 values were obtained for NB4 and U937 cells, there was little evidence that mefloquine (MQ) 
inhibited autophagy or induced apoptosis. A distinct DMSO control for Western blots, longer time points, IC90 
trials, other cell lines, and different assays such as flow cytometry will have to be included to evaluate the drug 
repurposing of mefloquine. Additional anticancer drugs will also be tested. We hope that our results will 
eventually provide the rationale to continue with clinical trials.
RESULTS
RESULTS (cont.)
Western Blot Results
Fig. 3 - Apoptosis Markers: PARP-1, Caspase-3, CHOP; Autophagy Markers: Atg-7, Atg-5, p62, LC3B 
DISCUSSION/CONCLUSIONS
[1] Arakawa, S., S. Honda, H. Yamaguchi, and S. Shimizu.  2017  Molecular mechanisms and physiological roles of Atg5/Atg7-independent alternative autophagy.  Journal List. 93(6): 
378–385.
[2] Eriksson, A., E. Chantzi, M. Fryknäs, J. Gullbo, P. Nygren, et al.  2017  Towards repositioning of quinacrine for treatment of acute myeloid leukemia - Promising synergies and in 
vivo effects.  Leukemia research. 63:41-46.
[3] Folkerts, H., S. Hilgendorf, A.T.J. Wierenga, J. Jaques, A. B. Mulder et al. 2017  Inhibition of autophagy as a treatment strategy for p53 wild-type acute myeloid leukemia. Cell 
Death and Disease. 8(7):e2927.
[4] Sharma, N., S. Thomas, E.B. Golden, F. M. Hofman, T. C. Chen et al. 2012  Inhibition of autophagy and induction of breast cancer cell death by mefloquine, an antimalarial agent. 
Cancer Letters. 326(2):143-54
[5] Staines, H.M., B.C. Dee, M.R. Shen, and J.C. Ellory.  2004  The effect of mefloquine and volume-regulated anion channel inhibitors on induced transport in Plasmodium 
falciparum-infected human red blood cells.  Blood Cells Mol Dis. 32(3):344-8
REFERENCES
PARP-1
Caspase-3
CHOP
INTRODUCTION
116 kDa
89 kDa
35 kDa
27 kDa
p62
LC3B-I
LC3B-II
62 kDa
16 kDa
14 kDaActin (control) 45 kDa
78 kDa
37 kDa
55 kDa
Atg-7
Atg-5
Cell Line Trypan Blue Mean IC50 (48h)
NB4 7 µM
U937 9.5 µM
Cell Line MTT Mean IC50 (48h)
NB4 6.9 µM (~ 7 µM)
U937 8.2 µM (~ 8 µM)
Two-tailed p-value t-test value Standard Deviation (SD) Standard Error of Mean (SEM)
P = 0.40 t = 1.1 (df = 2)
MTT: 1.4
Trypan Blue: 2.0
MTT: 0.82
Trypan Blue: 1.2
Two-tailed p-value t-test value Standard Deviation (SD) Standard Error of Mean (SEM)
P = 0.33 t = 1.3 (df = 2)
MTT: 0.8
Trypan Blue: 1.2
MTT: 0.5
Trypan Blue: 0.7
NB4: MTT vs. Trypan Blue IC50 Values (48h)
U937: MTT vs. Trypan Blue IC50 Values (48h)
The aim of this study is to observe cell proliferation, cell viability, apoptosis, and 
autophagy on acute myeloid leukemia (AML) cell lines, NB4 and U937, with the drug 
repurposing of mefloquine (MQ). Methods such as the 3-(4,5-Dimethylthiazol-2-Yl)-
2,5-Diphenyltetrazolium Bromide (MTT) assay and trypan blue staining have shown a 
decrease in live cells with high concentrations of mefloquine. Using their average 
perspective IC50 values of MQ concentration, Western blotting was applied by means 
of apoptosis and autophagy markers to determine if the induction of apoptosis and 
inhibition of autophagy was present in MQ-treated AML cells. The experiment will be 
continued with more cell lines, drugs, and other means of protocol in order to 
contribute to cancer therapy.
ABSTRACT
Lymphoid
Progenitor
Myeloid
Progenitor
Myeloblast
Lymphoblast
Erythrocytes (Red Cells)
Platelets
Basophil
Neutrophil
Eosinophil
Monocyte
T Lymphocyte
B Lymphocyte
ALL
AML
Hematopoietic
Stem Cell
Fig. 1 - Hematopoiesis for Acute Leukemia


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The Evaluation of Mefloquine Drug Repurposing on Acute Myeloid Leukemia

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