Fluorescence methods to assess multidrug resistance in individual cells

Nelson, Eric J.; Zinkin, Noah T.; Hinkle, Patricia M.

doi:10.1007/s002800050820

Cited by 19 publications

(13 citation statements)

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“…60 AML cells were incubated with Rh123 and the efflux measured by flow cytometry as previously described. 60 Saturation binding studies. To assess binding of lintuzumab to AML cells, the antibody was linked to Alexa Fluor 488 reactive dye (Molecular Probes, Eugene, OR).…”

Section: Methodsmentioning

confidence: 99%

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Anti-leukemic activity of Lintuzumab (SGN-33) in preclinical models of acute myeloid leukemia

Sutherland

Lewis

et al. 2009

mAbs

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Section: Methodsmentioning

confidence: 99%

“…Cells that are MDR + do not retain the Rh123 and efficiently pump it out, indicative of Pgp or other pump activity. 60 AML cells were incubated with Rh123 and the efflux measured by flow cytometry as previously described. 60 Saturation binding studies.…”

Section: Methodsmentioning

confidence: 99%

Anti-leukemic activity of Lintuzumab (SGN-33) in preclinical models of acute myeloid leukemia

Sutherland

Lewis

et al. 2009

mAbs

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“…In this context, it should be pointed out that lyophilic derivatives of intracellular fluorescent probes are substrates of P-glycoprotein (Pgp) and MRP1 [47]. Furthermore, MDR expression is affected by intracellular variation of glutathione (GSH) [61] and oxidative stress [62–65], as well as by various dietary factors [66–69], inflammatory cytokines [70–72], disease states [73–75], and drugs [76–81]. In particular, aspirin, indomethacin, and ibuprofen are substrates for MRP4 [76] and may interfere with fluorescent probe staining.…”

Section: Measurement Of Reactive Species In Leukocytes and Platelementioning

confidence: 99%

Measurement and Clinical Significance of Biomarkers of Oxidative Stress in Humans

Marrocco

Altieri

Peluso

2017

Oxidative Medicine and Cellular Longevity

565

500

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Oxidative stress is the result of the imbalance between reactive oxygen species (ROS) formation and enzymatic and nonenzymatic antioxidants. Biomarkers of oxidative stress are relevant in the evaluation of the disease status and of the health-enhancing effects of antioxidants. We aim to discuss the major methodological bias of methods used for the evaluation of oxidative stress in humans. There is a lack of consensus concerning the validation, standardization, and reproducibility of methods for the measurement of the following: (1) ROS in leukocytes and platelets by flow cytometry, (2) markers based on ROS-induced modifications of lipids, DNA, and proteins, (3) enzymatic players of redox status, and (4) total antioxidant capacity of human body fluids. It has been suggested that the bias of each method could be overcome by using indexes of oxidative stress that include more than one marker. However, the choice of the markers considered in the global index should be dictated by the aim of the study and its design, as well as by the clinical relevance in the selected subjects. In conclusion, the clinical significance of biomarkers of oxidative stress in humans must come from a critical analysis of the markers that should give an overall index of redox status in particular conditions.

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“…The presence of drug transporter activity was functionally determined via efflux of the substrate dye, rhodamine 123 (supplemental Figure 1, available on the Blood Web site). 23 …”

Section: Human Aml Cell Linesmentioning

confidence: 99%

SGN-CD33A: a novel CD33-targeting antibody–drug conjugate using a pyrrolobenzodiazepine dimer is active in models of drug-resistant AML

Sutherland

Walter

Jeffrey

et al. 2013

Blood

347

290

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• SGN-CD33A is a novel antibody-drug conjugate, consisting of an engineered anti-CD33 mAb conjugated to a potent DNA cross-linking cytotoxin.• SGN-CD33A is highly active in a broad panel of preclinical AML models and, in contrast to GO, is active despite MDR or poor-risk cytogenetics.Outcomes in acute myeloid leukemia (AML) remain unsatisfactory, and novel treatments are urgently needed. One strategy explores antibodies and their drug conjugates, particularly those targeting CD33. Emerging data with gemtuzumab ozogamicin (GO) demonstrate target validity and activity in some patients with AML, but efficacy is limited by heterogeneous drug conjugation, linker instability, and a high incidence of multidrug resistance. We describe here the development of SGN-CD33A, a humanized anti-CD33 antibody with engineered cysteines conjugated to a highly potent, synthetic DNA cross-linking pyrrolobenzodiazepine dimer via a proteasecleavable linker. The use of engineered cysteine residues at the sites of drug linker attachment results in a drug loading of approximately 2 pyrrolobenzodiazepine dimers per antibody. In preclinical testing, SGN-CD33A is more potent than GO against a panel of AML cell lines and primary AML cells in vitro and in xenotransplantation studies in mice. Unlike GO, antileukemic activity is observed with SGN-CD33A in AML models with the multidrug-resistant phenotype. Mechanistic studies indicate that the cytotoxic effects of SGN-CD33A involve DNA damage with ensuing cell cycle arrest and apoptotic cell death. Together, these data suggest that SGN-CD33A has CD33-directed antitumor activity and support clinical testing of this novel therapeutic in patients with

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Fluorescence methods to assess multidrug resistance in individual cells

Cited by 19 publications

References 0 publications

Anti-leukemic activity of Lintuzumab (SGN-33) in preclinical models of acute myeloid leukemia

Anti-leukemic activity of Lintuzumab (SGN-33) in preclinical models of acute myeloid leukemia

Measurement and Clinical Significance of Biomarkers of Oxidative Stress in Humans

SGN-CD33A: a novel CD33-targeting antibody–drug conjugate using a pyrrolobenzodiazepine dimer is active in models of drug-resistant AML

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