Biochemical assay reagent/probe

Response correlation with drug efflux [2]
We evaluated the efflux of 3, 3'-diethyloxacarbocyanine iodide/DiOC2 from pretreatment leukemic blast cells as a measure of functional drug resistance. Figure 3 shows the relationship between measured leukemic blast cell drug efflux levels and saturation of CD33 sites with response. Elimination of leukemia appeared to be correlated with a low capacity by leukemic blast cells to extrude DiOC2. For example, of the 30 patients evaluated with the assay in whom doses of CMA-676 saturated >75% of available CD33 sites on peripheral blood blast cells, 8 of 17 patients with leukemic blast cells that showed ≤40 channel numbers of DiOC2 efflux had <5% blasts in the bone marrow after treatment. In contrast, none of the 13 patients with leukemic blast cells expressing >40 channel numbers of DiOC2 efflux entered remission.

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Fluorescence studies of a carbocyanine dye 3, 3'-diethyloxacarbocyanine iodide (DOCI) in a series of monohydroxy alcohols reveal a monotonic increasing trend for both the fluorescence quantum yield and fluorescence lifetime with solvent viscosity. The results were interpreted in terms of retardation of the photoisomerization rate of DOCI with increasing solvent viscosity, and a definite inverse relationship between the two parameters was established. This relationship was utilized to estimate the microviscosity experienced by DOCI in supramolecular assemblies like micelles and microemulsions of several well-known surfactants and amphiphilic block copolymer. In most of the assemblies, the local environment around DOCI was characterized by rather high microviscosity, comparable to those of high alcohols between butanol and decanol. Our observations suggest that the rod-like DOCI molecule is largely embedded among the hydrophobic tails of the surfactant molecules, which exert a viscous drag strong enough to hinder its photoisomerization. The fluorescence properties of DOCI were also utilized to determine the critical micellization concentration of several surfactants in water. Interestingly, the microviscosity reported by DOCI in water-free AOT reverse micelles is significantly high, and can be explained by considering enhanced electrostatic attraction between the DOC+ cation and the anionic AOT head-groups [1].

Laboratory investigations [2]
Total hP67.6 antibody concentrations in plasma samples were determined using an enzyme-linked immunosorbent assay (ELISA). Formation of antigen-antibody CMA-676 bound to peripheral blood mononuclear cells was detected by flow microfluorimetry. Cells were incubated with biotinylated goat monoclonal anti-human IgG4, followed by avidin-fluorescein isothiocyanate. Cells incubated with avidin-fluorescein isothiocyanate alone comprised the negative control. The saturation percentage was defined as 100 times the ratio of the fluorescence intensity of patient mononuclear cells (minus the negative control) over the maximum fluorescence intensity (minus the negative control). Maximum achievable saturation was determined by incubating patient mononuclear cells from the same time point with saturating amounts of CMA-676 in vitro before the addition of the anti-human IgG4 antibody. The efflux of 3, 3'-diethyloxacarbocyanine iodide (DiOC2) from CD33-positive blast cells was measured as an indication of functional drug efflux.11 Serum samples obtained from each patient before CMA-676 administration, on day 7 after initial doses, and on days 7, 14, 21, and 28 after administration of the final dose of CMA-676 were analyzed for anti-hP67.6 (humanized mouse antibody) or anti-calicheamicin/linker immune response by ELISA. [2]

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Absorption and fluorescence spectra of the 3, 3'-diethyloxacarbocyanine iodide/DOCI solutions were measured in a Shimadzu UVPC-3200 spectrophotometer and a PerkinElmer LS55 fluorimeter, respectively. Picosecond fluorescence dynamics studies of the solutions were conducted with a time-correlated single photon counting (TCSPC) setup employing a picosecond diode laser (IBH NanoLED-07) operating at a λex = 440 nm and a repetition rate of 0.9 MHz as excitation source. The overall temporal and spectral resolution of the setup were ∼70 ps and ∼10 nm, respectively. All spectroscopic measurements were performed at 22 °C [1].

[1]. 3,3′-Diethyloxacarbocyanine iodide: A new microviscosity probe for micelles and microemulsions. Colloids and Surfaces A: Physicochemical and Engineering Aspects.
[2]. Selective ablation of acute myeloid leukemia using antibody-targeted chemotherapy: a phase I study of an anti-CD33 calicheamicin immunoconjugate. Blood. 1999 Jun 1;93(11):3678-84.

C3-oxacyanine is a C3 cyanine dye having 3-ethyl-1,3-benzoxazol-2(3H)-yl units at each end. It has a role as a fluorochrome. It is an organic iodide salt, a cyanine dye and a member of 1,3-benzoxazoles. It contains a C3-oxacyanine cation.

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Our results demonstrate that the photoisomerization rate constants of 3, 3'-diethyloxacarbocyanine iodide/DOCI in monohydroxy alcohols follows a relationship kiso ∝ 1/ηa, (a < 1) over a broad range of viscosities. We also found that kiso values in most micelles and microemulsions lies in the range of 1–3 × 109 s−1, comparable to those in the high alcohols between butanol and decanol. Evidently, the effective local viscosities experienced by DOCI in these surfactant aggregates are similar to those in high alcohols and could be calculated using the relationship connecting kiso and η in monohydroxy alcohols. Physically, the similarity is explained by assuming the rod-like DOCI molecule, including the cyanine bond, is buried among the hydrophobic tails of the surfactant molecules. Thus photoisomerization around this bond is inhibited by the friction exerted by these tails, which are structurally similar to the long hydrocarbon chains of the high alcohols. However, in AOT reverse micelles at , the viscosity reported by DOCI is exceptionally high compared to all other media. Here, the absence of water as a dielectric leads to strong electrostatic attraction between the DOC+ cation and the anionic head-groups of AOT, which further arrests the mobility of the DOCI molecule. [1]

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Leukemic blast cells express the CD33 antigen in most patients with acute myeloid leukemia (AML), but this antigen is not expressed by hematopoietic stem cells. We conducted a study to determine whether normal hematopoiesis could be restored in patients with AML by selective ablation of cells expressing the CD33 antigen. In a dose escalation study, 40 patients with relapsed or refractory CD33(+) AML were treated with an immunoconjugate (CMA-676) consisting of humanized anti-CD33 antibody linked to the potent antitumor antibiotic calicheamicin. The capacity of leukemic cells to efflux 3, 3'-diethyloxacarbocyanine iodide (DiOC2) was used to estimate pretreatment functional drug resistance. Leukemia was eliminated from the blood and marrow of 8 (20%) of the 40 patients; blood counts returned to normal in three (8%) patients. A high rate of clinical response was observed in leukemias characterized by low dye efflux in vitro. Infusions of CMA-676 were generally well tolerated, and a postinfusion syndrome of fever and chills was the most common toxic effect. Two patients who were treated at the highest dose level (9 mg/m2) were neutropenic >5 weeks after the last dose of CMA-676. These results show that an immunoconjugate targeted to CD33 can selectively ablate malignant hematopoiesis in some patients with AML. [2]

C21H21IN2O2

460.31

460.065

C, 54.80; H, 4.60; I, 27.57; N, 6.09; O, 6.95

905-96-4

6538326

Purplish red to red solid powder

278ºC (dec.)(lit.)

1.086

0

4

4

26

474

0

[I-].O1C(=CC=CC2=[N+](CC)C3C(=CC=CC=3)O2)N(CC)C2C1=CC=CC=2

FIZZUEJIOKEFFZ-UHFFFAOYSA-M

InChI=1S/C21H21N2O2.HI/c1-3-22-16-10-5-7-12-18(16)24-20(22)14-9-15-21-23(4-2)17-11-6-8-13-19(17)25-21;/h5-15H,3-4H2,1-2H3;1H/q+1;/p-1

(2Z)-3-ethyl-2-[(E)-3-(3-ethyl-1,3-benzoxazol-3-ium-2-yl)prop-2-enylidene]-1,3-benzoxazole;iodide

3,3'-Diethyloxacarbocyanine iodide; 905-96-4; C3-oxacyanine; 57441-62-0; 3,3'-Diethyloxacarbocyanine (iodide); DOCI dye; Oxacarbocyanine (C3); DiOC2(3) [3,3'-Diethyloxacarbocyanine Iodide];

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO: 20.83 mg/mL (45.25 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (4.52 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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 (Please use freshly prepared in vivo formulations for optimal results.)