A Potential Mechanism for the Antioxidant Compound Kushen Injection Targeting the ROS Signaling Pathway in Acute Myeloid Leukemia

Abstract Abstract It is a challenge for treatment of acute myeloid leukemia (AML) in clinic. The increase in the levels of reactive oxygen species (ROS) in AML patients has been previously described; thus, it is important to regulate ROS levels in AML. In this study, we found that intracellular ROS levels in AML cells were decreased, the total antioxidant capacity (T-AOC) and glutathione (GSH) contents were increased and xanthine oxidase (XOD) vitality was decreased when treated with the compound kushen injection (CKI). This shows that CKI inhibited the proliferation of AML cell lines and patient cells and enhanced the cytotoxicity of AML cells, which has few toxic effects on haematopoietic stem cells (HSCs) and T cells. At the single-cell level, individual AML cells died gradually by CKI treatment on optofluidic chips. CKI could promote apoptosis and arrest cell cycle at G1/G0 phase in U937 cells. Furthermore, higher peroxiredoxin-3 (Prdx3) expression levels were found in CKI-treated U937 cells through quantitative proteomics detection. Mechanically, the expression of Prdx3 and peroxiredoxin-2 (Prdx2) was up-regulated in CKI-treated AML cells, while thioredoxin 1 (Trx1) was reduced. Laser confocal microscopy showed that the Prdx2 and Trx1 proteins could be co-localized by CKI treatment. In vivo, in the CKI-treated group, survival was longer in an AML patient-derived xenograft model in B-NSG mice. The disease was partially alleviated by decreased CD45+ immunophenotyping in peripheral blood and bone marrow smear analysis. After CKI injection, the T-AOC vitality, GSH content and CAT activity increased and the concentration of H2O2decreased in mouse plasma. A bone marrow biopsy and immunohistochemistry analysis showed that Prdx3 and Prdx2 expression was increased, while Trx1 expression was decreased. In a conclusion, we provided a model for the anti-leukaemic effects of CKI. CKI decreased intracellular ROS levels by up-regulating the expression of Prdx2 in the cytoplasm and Prdx3 in the mitochondria and down-regulating Trx1 expression, which maintained the intracellular REDOX and further inhibited AML cell proliferation. Therefore, antioxidant CKI is a promising drug for the treatment of AML in the clinic. We aimed to explore the therapeutic efficacy of low toxicity natural antioxidants against AML by targeting ROS pathways and providing new strategies to improve survival in AML patients. Disclosures No relevant conflicts of interest to declare.

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CXCR4 Invalidation Limits the MLL-ENL Induced Leucemogenesis in Vivo

Blood ◽

10.1182/blood.v120.21.4089.4089 ◽

2012 ◽

Vol 120 (21) ◽

pp. 4089-4089

Author(s):

Yanyan Zhang ◽

Hadjer Abdelouahab ◽

Aline Betems ◽

Monika Wittner ◽

William Vainchenker ◽

...

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Survival Time ◽

Median Survival ◽

Median Survival Time ◽

Myeloid Leukemia ◽

Conflicts Of Interest ◽

Hematopoietic Stem ◽

Acute Myeloid

Abstract Abstract 4089 The receptor CXCR4 and its ligand SDF-1 play major physiological roles especially on hematopoietic stem cells homing and retention. Many studies have implicated CXCR4 in the invasion by tumor cells of organs that produce SDF-1. In acute myeloid leukemia, the physiological role of CXCR4 is not fully understood. We used retrovirus to express MLL-ENL oncogene in CXCR4+/+ and CXCR4−/− hematopoietic primitive cells (Lin- isolated from fetal liver) and showed that CXCR4 is dispensable for generation of immortalized colonies in vitro. To determine CXCR4 function in vivo, CXCR4+/+ and CXCR4−/− transformed cells were transplanted into lethally irradiated mice. Whatever their phenotype, the recipient developed a myelo-monocytique leukemia characterized by their expression of Gr-1 and Mac-1. As expected, all recipients of MLL-ENL transduced CXCR4+/+ cells were moribund within 35 to 80 days post transplant (median survival time: 50 days). Strikingly, recipients of MLL-ENL transduced CXCR4−/− cells showed significantly increased lifespan, with a median survival time of 90 days. The cellularity of the peripheral blood of recipients of MLL-ENL transduced cells displayed considerable increases over time although this increase was much lower in CXCR4−/− than in CXCR4+/+ chimera. Bone marrow of MLL-ENL transduced CXCR4−/− chimera had moderately decreased numbers of mononuclear cells. There were important numbers of leukemic CD45.2+/Gr1+/Mac1+/c-kit+ cells in spleen from MLL-ENL CXCR4+/+ chimera which suggested that CXCR4 is important for leukemic progenitors cells retention in the bone marrow and especially in the spleen. The homing capacity of transduced CXCR4+/+ cells is comparable to the CXCR4−/− cells. Finally, more DNA damages were found in the BM cells of MLL-ENL CXCR4−/− chimera. All these results were confirmed by treating of MLL-ENL CXCR4+/+ chimera with CXCR4 inhibitor (TN140). These results demonstrated that in absence of CXCR4, the cells transduced by oncogene MLL-ENL are capable of generating leukemia in the recipients. However, mice transplanted with MLL-ENL transduced CXCR4−/− FL cells developed acute myeloid leukemia with reduced aggressiveness and organ infiltration, which is associated with induced differentiation and DNA instability. These results indicated that the MLL-ENL progenitors are dependent on CXCR4 for their maintenance in the BM and spleen suggesting that CXCR4 inhibitors might have potential therapeutic applications. Disclosures: No relevant conflicts of interest to declare.

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Acute myeloid leukemia induces protumoral p16INK4a-driven senescence in the bone marrow microenvironment

Blood ◽

10.1182/blood-2018-04-845420 ◽

2019 ◽

Vol 133 (5) ◽

pp. 446-456 ◽

Cited By ~ 17

Author(s):

Amina M. Abdul-Aziz ◽

Yu Sun ◽

Charlotte Hellmich ◽

Christopher R. Marlein ◽

Jayna Mistry ◽

...

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Stromal Cells ◽

Myeloid Leukemia ◽

Stromal Cell ◽

Cell Senescence ◽

Senescent Phenotype ◽

Age Related ◽

Acute Myeloid

Abstract Acute myeloid leukemia (AML) is an age-related disease that is highly dependent on the bone marrow (BM) microenvironment. With increasing age, tissues accumulate senescent cells, characterized by an irreversible arrest of cell proliferation and the secretion of a set of proinflammatory cytokines, chemokines, and growth factors, collectively known as the senescence-associated secretory phenotype (SASP). Here, we report that AML blasts induce a senescent phenotype in the stromal cells within the BM microenvironment and that the BM stromal cell senescence is driven by p16INK4a expression. The p16INK4a-expressing senescent stromal cells then feed back to promote AML blast survival and proliferation via the SASP. Importantly, selective elimination of p16INK4a+ senescent BM stromal cells in vivo improved the survival of mice with leukemia. Next, we find that the leukemia-driven senescent tumor microenvironment is caused by AML-induced NOX2-derived superoxide. Finally, using the p16-3MR mouse model, we show that by targeting NOX2 we reduced BM stromal cell senescence and consequently reduced AML proliferation. Together, these data identify leukemia-generated NOX2-derived superoxide as a driver of protumoral p16INK4a-dependent senescence in BM stromal cells. Our findings reveal the importance of a senescent microenvironment for the pathophysiology of leukemia. These data now open the door to investigate drugs that specifically target the “benign” senescent cells that surround and support AML.

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Selective targeting of NAMPT by KPT-9274 in acute myeloid leukemia

Blood Advances ◽

10.1182/bloodadvances.2018024182 ◽

2019 ◽

Vol 3 (3) ◽

pp. 242-255 ◽

Cited By ~ 8

Author(s):

Shaneice R. Mitchell ◽

Karilyn Larkin ◽

Nicole R. Grieselhuber ◽

Tzung-Huei Lai ◽

Matthew Cannon ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Treatment Options ◽

Tumor Development ◽

Xenograft Model ◽

Nicotinamide Phosphoribosyltransferase ◽

P21 Activated Kinase ◽

Induced Apoptosis ◽

Acute Myeloid

Abstract Treatment options for acute myeloid leukemia (AML) remain extremely limited and associated with significant toxicity. Nicotinamide phosphoribosyltransferase (NAMPT) is involved in the generation of NAD+ and a potential therapeutic target in AML. We evaluated the effect of KPT-9274, a p21-activated kinase 4/NAMPT inhibitor that possesses a unique NAMPT-binding profile based on in silico modeling compared with earlier compounds pursued against this target. KPT-9274 elicited loss of mitochondrial respiration and glycolysis and induced apoptosis in AML subtypes independent of mutations and genomic abnormalities. These actions occurred mainly through the depletion of NAD+, whereas genetic knockdown of p21-activated kinase 4 did not induce cytotoxicity in AML cell lines or influence the cytotoxic effect of KPT-9274. KPT-9274 exposure reduced colony formation, increased blast differentiation, and diminished the frequency of leukemia-initiating cells from primary AML samples; KPT-9274 was minimally cytotoxic toward normal hematopoietic or immune cells. In addition, KPT-9274 improved overall survival in vivo in 2 different mouse models of AML and reduced tumor development in a patient-derived xenograft model of AML. Overall, KPT-9274 exhibited broad preclinical activity across a variety of AML subtypes and warrants further investigation as a potential therapeutic agent for AML.

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Silencing long non-coding RNA XIST suppresses drug resistance in acute myeloid leukemia through down-regulation of MYC by elevating microRNA-29a expression

Molecular Medicine ◽

10.1186/s10020-020-00229-4 ◽

2020 ◽

Vol 26 (1) ◽

Author(s):

Chong Wang ◽

Lingling Li ◽

Mengya Li ◽

Weiqiong Wang ◽

Yanfang Liu ◽

...

Keyword(s):

Bone Marrow ◽

Drug Resistance ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Bone Marrow Cells ◽

Cellular Viability ◽

Down Regulation ◽

Acute Myeloid

Abstract Background Long non-coding RNAs (lncRNAs) are biomarkers participating in multiple disease development including acute myeloid leukemia (AML). Here, we investigated molecular mechanism of X Inactive-Specific Transcript (XIST) in regulating cellular viability, apoptosis and drug resistance in AML. Methods XIST, miR-29a and myelocytomatosis oncogene (MYC) expression in AML bone marrow cells collected from 62 patients was evaluated by RT-qPCR and Western blot analysis. Besides, the relationship among XIST, miR-29a and MYC was analyzed by dual luciferase reporter assay, RIP, and RNA pull down assays. AML KG-1 cells were treated with anti-tumor drug Adriamycin. The role of XIST/miR-29a/MYC in cellular viability, apoptosis and drug resistance in AML was accessed via gain- and loss-of-function approaches. At last, we evaluated role of XIST/miR-29a/MYC on tumorigenesis in vivo. Results XIST and MYC were up-regulated, and miR-29a was down-regulated in AML bone marrow cells. Silencing XIST inhibited cellular activity and drug resistance but promoted cellular apoptosis of KG-1 cells by down-regulating MYC. XIST inhibited miR-29a expression to up-regulate MYC. Moreover, silencing XIST inhibited tumorigenesis of AML cells in vivo. Conclusions Overall, down-regulation of XIST decreased MYC expression through releasing the inhibition on miR-29a, thereby reducing drug resistance, inhibiting viability and promoting apoptosis of AML cells.

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Cellular pharmacokinetics of daunorubicin: relationships with the response to treatment in patients with acute myeloid leukemia.

Journal of Clinical Oncology ◽

10.1200/jco.1988.6.5.802 ◽

1988 ◽

Vol 6 (5) ◽

pp. 802-812 ◽

Cited By ~ 32

Author(s):

E Kokenberg ◽

P Sonneveld ◽

W Sizoo ◽

A Hagenbeek ◽

B Löwenberg

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Response To Treatment ◽

Remission Induction ◽

Pharmacokinetic Parameters ◽

Induction Treatment ◽

Blast Cells ◽

Acute Myeloid

In an attempt to identify pharmacokinetic factors that determine the response of acute myeloid leukemia (AML) patients to induction chemotherapy, we determined the concentrations of daunorubicin (DNR) and the main metabolite daunorubicinol (DOL) in vivo and particularly evaluated the concentrations in blood and bone marrow nucleated cells. Cell measurements were obtained in 37 evaluable patients during their first remission induction treatment with DNR and cytarabine (ara-C) and directly compared with the plasma distribution kinetics of DNR. We show that (1) plasma DNR concentrations do not correlate with DNR concentrations in bone marrow nucleated cells; but (2) plasma area under the curve (AUC) values of DNR correlate inversely (P less than .01) with AUC values of DNR in WBCs; (3) concentrations of DNR in WBCs correlate positively (P less than .01) with DNR concentrations in bone marrow nucleated cells; and (4) the concentrations of DNR in WBCs show a negative correlation (P less than .01) with the numbers of peripheral blast cells at diagnosis. We then tested whether the pharmacokinetic parameters had predictive value for the clinical outcome of therapy, but none of the plasma levels or WBC and bone marrow concentrations of DNR predicted treatment outcome. The inverse correlation between the concentrations of DNR in WBC and the numbers of peripheral blast cells suggests that the effective DNR concentrations achieved intracellularly are mainly a function of the tumor load so that lesser amounts of DNR accumulate intracellularly when the AML cell numbers in blood are higher.

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In vivo administration of vascular endothelial growth factor (VEGF) and its antagonist, soluble neuropilin-1, predicts a role of VEGF in the progression of acute myeloid leukemia in vivo

Blood ◽

10.1182/blood.v100.13.4622 ◽

2002 ◽

Vol 100 (13) ◽

pp. 4622-4628 ◽

Cited By ~ 100

Author(s):

Gunter Schuch ◽

Marcelle Machluf ◽

Georg Bartsch ◽

Masashi Nomi ◽

Henri Richard ◽

...

Keyword(s):

Bone Marrow ◽

Vascular Endothelial Growth Factor ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Leukemic Cells ◽

Vascular Endothelial ◽

Neuropilin 1 ◽

Endothelial Growth Factor ◽

Acute Myeloid

Recent findings implied that the progression of hematologic malignancies, like that of solid tumors, is dependent on neovascularization. Recent studies on patients with acute myeloid leukemia (AML) showed increased levels of leukocyte-associated vascular endothelial growth factor (VEGF) and neovascularization of the bone marrow. Murine (32D, M1) and human (HEL, U937, and UKE-1) leukemic cell lines and freshly isolated leukemic cells were analyzed for the expression of VEGF and VEGF receptor mRNA. The expression of VEGF and VEGF receptors KDR and neuropilin-1 (NRP-1) was detected in these cells. In a murine chloroma model, delivery of VEGF165using microencapsulation technology resulted in enhanced tumor growth and vascularization, whereas treatment with a VEGF antagonist soluble NRP-1 (sNRP-1) inhibited tumor angiogenesis and growth. In a systemic leukemia model, survival of mice injected with adenovirus (Ad) encoding for Fc-sNRP-1 (sNRP-1 dimer) was significantly prolonged as compared with mice injected with Ad-LacZ. Further analyses showed a reduction in circulating leukemic cells and infiltration of liver and spleen as well as bone marrow neovascularization and cellularity. Taken together, these results demonstrate that angiogenic factors such as VEGF promote AML progression in vivo. The use of VEGF antagonists as an antiangiogenesis approach offers a potential treatment for AML. Finally, our novel in vivo drug delivery model may be useful for testing the activities of other peptide antiangiogenic factors.

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Mesenchimal Stroma Cells From Patients with Myelodysplastic Syndrome and Acute Myeloid Leukemia Show Distinct Cytogenetic and DNA-Mutation Data as Compared with Bone Marrow Leukemic Blasts.

Blood ◽

10.1182/blood.v114.22.4697.4697 ◽

2009 ◽

Vol 114 (22) ◽

pp. 4697-4697

Author(s):

Olga Blau ◽

Wolf-Karsten Hofmann ◽

Claudia D Baldus ◽

Gundula Thiel ◽

Florian Nolte ◽

...

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Conflicts Of Interest ◽

Control Analysis ◽

Npm1 Mutation ◽

Dna Mutation ◽

Healthy Donors ◽

Acute Myeloid

Abstract Abstract 4697 Bone marrow mesenchymal stroma cells (BMSC) are key components of the hematopoietic microenvironment. BMSC from patients with acute myeloid leukemia (AML) and myelodisplasic syndrome (MDS) display functional and quantitative alterations. To gain insight into these questions, we carried out cytogenetic analyses, FISH, FLT3 and NPM1 mutation examinations of both hematopoietic (HC) and BMSC derived from 53 AML and 54 MDS patients and 35 healthy donors after in vitro culture expansion. Clonal chromosomal aberrations were detectable in BMSC of 12% of patients. Using FISH we have assume that cytogenetic markers in BMSC were always distinct as the aberrations in HC from the same individual. 17% and 12% of AML patients showed FLT3 and NPM1 mutations in HC, respectively. In BMSC, we could not detect mutations of NPM1 and FLT3, independent from the mutation status of HC. For control analysis, BMSC cultures from 35 healthy donors were prepared under the same conditions. BMSC from healthy donors did show normal diploid karyotypes and absence of specific DNA-mutations of NPM1 and FLT3. Our data indicate that BMSC from MDS and AML patients are not a part of malignant clone and characterized by genetic aberrations. Lack of aberrations as detected in HC and appearance of novel clonal rearrangements in BMSC may suggest enhanced genetic susceptibility and potential involvement of BMSC in the pathogenesis of MDS and AML. Disclosures: No relevant conflicts of interest to declare.

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Fludarabine, Cytarabine, and Attenuated-Dose Idarubicin (m-FLAI) Induction for Elderly Patients with Acute Myeloid Leukemia: Results of a Multicenter Phase II Study,

Blood ◽

10.1182/blood.v118.21.3626.3626 ◽

2011 ◽

Vol 118 (21) ◽

pp. 3626-3626

Author(s):

Yoojoo Lim ◽

Youngil Koh ◽

Sung-Soo Yoon ◽

Seonyang Park ◽

Byoung Kook Kim ◽

...

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Elderly Patients ◽

Phase Ii ◽

Myeloid Leukemia ◽

Conflicts Of Interest ◽

Performance Status ◽

Free Survival ◽

Induction Regimen ◽

Acute Myeloid

Abstract Abstract 3626 Introduction: Although there have been remarkable improvements in treatment of acute myeloid leukemia (AML), the prognosis of AML in elderly patients remains poor, and the best induction chemotherapy for these patients remains yet unknown. To devise an effective induction regimen for elderly patients with AML, we conducted a phase II trial to evaluate the efficacy and safety of the modified fludarabine, cytarabine, and attenuated-dose idarubicin (m-FLAI) regimen in these patients. Patients and Methods: Elderly (60 years or older) AML patients who did not receive previous chemotherapy were enrolled. Patients received two consecutive cycles of m-FLAI chemotherapy as an induction. The m-FLAI regimen was comprised of fludarabine (25mg/m2, days 1–4), cytarabine (1000mg/m2, days 1–4), and attenuated-dose idarubicin (5mg/m2, days 1–3). The primary end point was complete remission (CR) rate. Results: A total of 108 patients (median age 68.4 years, M:F=64:44) were enrolled. CR was achieved in 62.9% of patients, and treatment-related mortality rate (TRM) was 25.8%. Median overall survival (OS) was 9.3 months, and median event-free survival (EFS) was 6.6 months. The mortality at 30 and 60 days was 18% and 24%, respectively. Performance status and comorbidity did not have prognostic value in these patients. Bone marrow expression of CD117 was related to long EFS and OS. Conclusion: In conclusion, m-FLAI is a safe and effective induction regimen for previously untreated AML in elderly patients. Bone marrow CD117 expression is an independent good prognostic factor in these patients. (ClinicalTrials.gov number, NCT01247493) Disclosures: No relevant conflicts of interest to declare.

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EZH2 Mutations Are Associated with Low Blast Percentage in Bone Marrow and −7/Del(7q) Karyotype in De Novo Acute Myeloid Leukemia.

Blood ◽

10.1182/blood.v120.21.2544.2544 ◽

2012 ◽

Vol 120 (21) ◽

pp. 2544-2544

Author(s):

Xiuli Wang ◽

Haiping Dai ◽

Qian WANG ◽

Qinrong Wang ◽

Yang Xu ◽

...

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

De Novo ◽

Conflicts Of Interest ◽

Univariate Analysis ◽

Myelogenous Leukemia ◽

Prognostic Impact ◽

Coding Region ◽

Acute Myeloid

Abstract Abstract 2544 Somatic mutation of the EZH2 gene is seen in myelodisplastic syndrome, myelofibrosis, and chronic myelomonocytic leukemia patients. The prevalence and prognostic impact of somatic mutations of EZH2 in patients with acute myelogenous leukemia (AML) remains unknown. In this study, we sought to determine the incidence and clinical implications of somatic EZH2 mutations in 714 patients with de novo AML by PCR amplification of the entire coding region followed by direct bidirectional DNA sequencing. EZH2 mutations were identified in 13/714 (1.8%) of AML patients and occurred almost exclusively in males (11/13, P=0.033). In univariate analysis, the presence of EZH2 mutations was significantly associated with lower blast percentage (21–30%) in bone marrow (P=0.0001) and −7/del(7q) (P=0.025). There was no difference in the incidence of mutations in 13 genes, including ASXL1, CBL, c-KIT, DNMT3A, FLT3, IDH1, IDH2, MLL, NPM1, NRAS, RUNX1, TET2, and WT1, between patients with and without EZH2 mutations. Complete remission, event-free survival or overall survival was similar between AML patients with and without EZH2 mutation (p>0.05). These results demonstrated EZH2 mutation as a recurrent genetic abnormality associated with lower blast percentage in BM and −7/del(7q) in de novo acute myeloid leukemia. Disclosures: No relevant conflicts of interest to declare.

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Activity of the Hypoxia-Activated Prodrug, TH-302, in Human Acute Myeloid Leukemia Models

Blood ◽

10.1182/blood.v120.21.3611.3611 ◽

2012 ◽

Vol 120 (21) ◽

pp. 3611-3611

Author(s):

Scott Portwood ◽

Deepika Lal ◽

Yung-Chun Hsu ◽

Rodrigo Vargas ◽

Meir Wetzler ◽

...

Keyword(s):

Overall Survival ◽

Acute Myeloid Leukemia ◽

Myeloid Leukemia ◽

Conflicts Of Interest ◽

Flow Cytometric Analysis ◽

Chemotherapeutic Agents ◽

Scid Mice ◽

Hypoxic Conditions ◽

Acute Myeloid

Abstract Abstract 3611 Recent evidence has demonstrated the bone marrow (BM) microenvironment, the principal site of acute myeloid leukemia (AML) initiation and expansion, is characterized by intrinsically low oxygen tension. Theoretically, such microenvironmental changes may lead to the selective outgrowth of AML clones which are “better adapted” to survive within a severely hypoxic microenvironment and/or may confer resistance to chemotherapeutic agents, similar to solid tumor cells. We report here that human AML cells (HL60, ML-2) cultured under chronic hypoxic conditions mimicking the marrow microenvironment (1% O2, 72 hours) exhibited reduced sensitivity to cytarabine-induced apoptosis as compared with normoxic cells, as determined by flow cytometric analysis, western blot analysis, and cell viability assays. Similar results were noted in primary AML samples treated with cytarabine under normoxic and hypoxic conditions in colony formation assays (n=3 samples, p=0.01). In order to improve upon chemotherapy outcomes, we investigated the effects of TH-302, a hypoxia-activated bromo-isophosphoramidate mustard prodrug, which is currently undergoing clinical trial evaluation in multiple tumor types. Treatment of AML cell lines (HL60, HEL) and primary AML samples with TH-302 (at doses ranging from 0.1– 5 mM, p values ranging from <0.05–0.0001) resulted in dose- and hypoxic-dependent inhibition of AML proliferation and apoptosis. In vivo TH-302 treatment significantly decreased disease burden, as measured by total animal bioluminescence, and prolonged overall survival in two systemic human AML xenograft models (HEL-luciferase, HL60-luciferase) (Figure 1). Immunohistochemical studies demonstrated that TH-302 treatment reduced numbers of hypoxic (pimonidazole-positive) cells within the leukemic marrow microenvironment. Because prior data in animal models has shown that AML progression within the marrow is associated with expansion of hypoxic BM areas, we examined the effects of TH-302 treatment on systemic AML growth when initiated early (prior to AML inoculation) or late (several days following AML engraftment) in the disease process. TH-302 was equally effective at both time points. Although anti-vascular therapy has been shown to enhance tumor hypoxia in other cancer types, we noted no synergistic or additive in vivo effects when TH-302 therapy was combined with sorafenib, an inhibitor of vascular endothelial growth factor receptors (VEGFR), in our models. TH-302 therapy administered for two weeks in non-leukemic and leukemia-engrafted mice was not associated with hematologic toxicities. In summary, our results demonstrate the anti-leukemic activity of TH-302 in preclinical AML models and suggest that the efficacy of this and other drugs for AML therapy may be uniquely affected by the BM microenvironment. Further clinical development of TH-302 and other hypoxia-targeted drugs for AML therapy are warranted. Based on our data, higher TH-302 doses and/or chronic drug administration may be needed for optimal in vivo anti-leukemic activity. Figure 1. Effects of TH-302 treatment on systemic AML growth and overall survival in HL60-luciferase engrafted SCID mice. Figure 1. Effects of TH-302 treatment on systemic AML growth and overall survival in HL60-luciferase engrafted SCID mice. Disclosures: No relevant conflicts of interest to declare.

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