Topoisomerase II[4]

In a pH-dependent manner, aldoxorubicin hydrochloride (INNO-206) (0.27 to 2.16 μM) decreases the proliferation of multiple myeloma cells and prevents blood vessel formation[1].

Aldoxorubicin hydrochloride (INNO-206) (10.8 mg/kg, iv) is well tolerated; 90% of mice with the LAGκ-1A tumor survived until the study's conclusion[1]. On days 28, it displays noticeably lower tumor sizes and IgG levels. Aldoxorubicin hydrochloride (INNO-206) can cause tumor regressions in breast cancer, small cell lung cancer, and sarcoma in a phase I research [2]. It also exhibits an excellent safety profile at doses up to 260 mg/mL doxorubicin equivalents. In models of mouse renal cell cancer and breast carcinoma xenografts, aldoxorubicin hydrochloride (INNO-206) exhibits better efficacy than doxorubicin[3].

Cell viability assay[1]
Cells were seeded at 1 × 105 cells/100 μL/well in 96-well plates in RPMI-1640 media with FBS for 24 hours before treatment. Cells were cultured in the presence of medium, INNO-206 or doxorubicin for 48 hours. Next, cell viability was quantified using the CellTiter 96 AQueous Non-Radioactive Cell Proliferation Assay (Promega). Each well was treated with MTS for 1 to 4 hours, after which absorbance at 490 nm was recorded using a 96-well plate reader. The quantity of formazan product as measured is directly proportional to the number of living cells. Data graphed are means ± SEM using 3 replicates per data point.

The anti–multiple myeloma effect of INNO-206 at different pH levels on multiple myeloma cell proliferation using multiple myeloma cell lines with the MTS assay and antiangiogenic activity using the chorioallantoic membrane/feather bud assay were determined. The anti–multiple myeloma effects and toxicity of INNO-206 were also compared with conventional doxorubicin and PEGylated liposomal doxorubicin (PLD) alone, and in combination with bortezomib, using our multiple myeloma xenograft models[1].

INNO-206 stock solutions (5.4 mg/mL) were prepared using 50% ethanol and 50% water and further diluted in sterile water. [1]
For the LAGκ-1A experiment, INNO-206 was administered to SCID mice at 10.8 mg/kg (doxorubicin equivalent dose of 8.0 mg/kg) once weekly. Mice were treated with conventional doxorubicin at 4.0 and 8.0 mg/kg once weekly. For the LAGκ-2 experiment, INNO-206 was administered once weekly (W) at doses of 2.7 and 5.4 mg/kg, or on 3 consecutive days (W-F) weekly at doses of 0.9 and 1.8 mg/kg. Bortezomib was administered twice weekly (W, F) at a dose of 0.5 mg/kg. Doxorubicin was administered to SCID mice at 2, 4, and 8 mg/kg, and PLD was administered to SCID mice at 2 mg/kg once weekly. Each drug was administered i.v. in a volume of 100 μL.[1]

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The (6-maleimidocaproyl)hydrazone derivative of doxorubicin (INNO-206) is an albumin-binding prodrug of doxorubicin with acid-sensitive properties that is being assessed clinically. The prodrug binds rapidly to circulating serum albumin and releases doxorubicin selectively at the tumor site. This novel mechanism may provide enhanced antitumor activity of doxorubicin while improving the overall toxicity profile. Preclinically, INNO-206 has shown superior activity over doxorubicin in a murine renal cell carcinoma model and in breast carcinoma xenograft models. In this work, we compared the antitumor activity of INNO-206 and doxorubicin at their respective maximum tolerated doses in three additional xenograft models (breast carcinoma 3366, ovarian carcinoma A2780, and small cell lung cancer H209) as well as in an orthotopic pancreas carcinoma model (AsPC-1). INNO-206 showed more potent antitumor efficacy than free doxorubicin in all tumor models and is thus a promising clinical candidate for treating a broad range of solid tumors[3].

[1]. Cell penetrating peptides fused to a thermally targeted biopolymer drug carrier improve the delivery and antitumor efficacy of an acid-sensitive doxorubicin derivative. Int J Pharm. 2012 Oct 15;436(1-2):825-32.

[2]. INNO-206, the (6-maleimidocaproyl hydrazone derivative of doxorubicin), shows superior antitumor efficacy compared to doxorubicin in different tumor xenograft models and in an orthotopic pancreas carcinoma model. Invest New Drugs. 2010 F.

[3]. Anti-Myeloma Effects of the Novel Anthracycline Derivative INNO-206. Clin Cancer Res.2012 18; 3856.

[4]. Kratz, F. INNO-206 (DOXO-EMCH), an Albumin-Binding Prodrug of Doxorubicin Under Development for Phase II Studies. Current Bioactive Compounds, 2011, 7(1): 33-38(6).

Aldolrubicin is an antitumor drug and an albumin-bound prodrug of doxorubicin. Aldolrubicin is a 6-maleimide hexanoyl hydrazone derivative prodrug of the anthracycline antibiotic doxorubicin (DOXO-EMCH) and possesses antitumor activity. After intravenous injection, aldolrubicin selectively binds to cysteine-34 of albumin via its maleimide moiety. In the acidic environment of the tumor, the acid-sensitive hydrazone linker breaks, releasing doxorubicin from the albumin carrier. Once inside the cell, it intercalates into DNA, inhibiting DNA synthesis and inducing apoptosis. Due to the high metabolic turnover, rapid angiogenesis, rich vascularity, and impaired lymphatic drainage of solid tumors, albumin tends to accumulate in them. This passive accumulation within the tumor may enhance the therapeutic effect of doxorubicin while minimizing systemic toxicity. Mechanism of action: INNO-206 is a (6-maleimide hexanoyl) hydrazone derivative of doxorubicin. INNO-206 is an doxorubicin prodrug that binds to endogenous albumin after administration. The bound doxorubicin is released through the cleavage of acid-sensitive linkers in the acidic environment of tumor cells. In preclinical models, INNO-206 demonstrated superior antitumor efficacy and toxicity compared to doxorubicin. Doxorubicin has shown efficacy, particularly in combination therapy for multiple myeloma; however, its side effects limit its application. INNO-206 is an albumin-bound doxorubicin prodrug that is released from albumin under acidic conditions. Since INNO-206 has not been previously evaluated in any hematologic malignancies, we investigated its anti-multiple myeloma activity. Experimental Design: We used the MTS assay to detect the effect of INNO-206 on the proliferation of multiple myeloma cells at different pH values and the chorioallantoic membrane/feather bud assay to detect its anti-angiogenic activity. Furthermore, using a multiple myeloma xenograft model, we compared the anti-multiple myeloma effects and toxicities of INNO-206 with those of conventional doxorubicin, pegylated liposomal doxorubicin (PLD) monotherapy, and combination therapy with bortezomib. Results: INNO-206 inhibited angiogenesis and reduced the growth of multiple myeloma cells in a pH-dependent manner. INNO-206 alone produced significant anti-multiple myeloma effects in vivo at doses comparable to the toxic doses of doxorubicin; the anti-multiple myeloma effect of INNO-206 in combination with bortezomib was enhanced compared to either drug alone. Conversely, all mice treated with bortezomib in combination with doxorubicin or PLD died. Conclusion: These results indicate that INNO-206 has anti-multiple myeloma effects both in vitro and in vivo, and can enhance the antitumor activity of bortezomib. These results suggest that INNO-206 may provide an anthracycline for patients with multiple myeloma, and that INNO-206 can be safely administered at higher doses compared to free doxorubicin, thus achieving superior efficacy compared to anthracyclines currently used to treat this B-cell malignancy. [1] Clin Cancer Res; 18(14); 3856–67. ©2012 AACR.

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The (6-maleimide hexanoyl)hydrazone derivative of doxorubicin (INNO-206, formerly known as DOXO-EMCH) is a prodrug of the anticancer drug doxorubicin, which was planned to be administered intravenously in 2011 and selectively bind to cysteine-34 of endogenous albumin within minutes. Preclinical and clinical studies have shown that the albumin-bound form of INNO-206 has a larger AUC, a smaller volume of distribution, and a lower clearance rate compared to doxorubicin. Its uptake in solid tumors is mediated by the pathophysiological mechanisms of tumor tissue, characterized by angiogenesis, angiogenesis, vascular structural defects, and impaired lymphatic drainage. This prodrug contains an acid-sensitive hydrazone linker, enabling extracellular release of doxorubicin in the weakly acidic environment common in tumor tissue, or intracellular release in acidic endosomes or lysosomal compartments after tumor cells take up the albumin conjugate. In various preclinical tumor models, INNO-206 demonstrated significantly superior antitumor efficacy compared to free doxorubicin. In a Phase I study, INNO-206 showed good safety at doses up to 260 mg/m² doxorubicin equivalents. Although not the primary endpoint of this Phase I study, INNO-206 induced tumor regression in breast cancer, small cell lung cancer, and sarcoma. Phase II clinical trials for gastric cancer, pancreatic cancer, and sarcoma are planned to begin at the end of 2010. [2] Elastin-like peptides (ELPs) are thermoresponsive macromolecular carriers that passively accumulate in solid tumors and further aggregate in tumor tissue upon exposure to high temperatures. In this study, ELPs were conjugated with the anticancer drug doxorubicin (DOXO) and three different cell-penetrating peptides (CPPs) to inhibit tumor growth in mice compared to free doxorubicin. Fluorescence microscopy studies of MCF-7 breast cancer cells showed that all three different CPP-ELP-DOXO conjugates could deliver doxorubicin to the cell nucleus. All CPP-ELP-DOXO conjugates exhibited cytotoxicity at 42 °C, with IC50 values ranging from 12 to 30 μM, but the ELP carrier using SynB1 as the cell-penetrating peptide showed the lowest inherent cytotoxicity. Therefore, the antitumor efficacy of SynB1-ELP-DOXO versus doxorubicin was compared under hyperthermic conditions. Female C57BL/6 mice carrying mammary tumors from the E0771 mouse conjugate were treated with either free doxorubicin or the SynB1-ELP-DOXO conjugate, with or without local hyperthermia targeting the tumor. Under hyperthermic conditions, the tumor inhibition rate of SynB1-ELP-DOXO was twice that of an equivalent dose of free doxorubicin, making it an ideal candidate for optimizing thermoresponsive drug polymer conjugates. [4]

C37H43CLN4O13

786.251

1361563-03-2

Aldoxorubicin;1361644-26-9

10056071

Typically exists as solid at room temperature

8

15

12

55

1510

6

Cl.O([C@H]1C[C@@H]([C@@H]([C@H](C)O1)O)N)[C@@H]1C2C(=C3C(C4C(=CC=CC=4C(C3=C(C=2C[C@@](/C(/CO)=N/NC(CCCCCN2C(C=CC2=O)=O)=O)(C1)O)O)=O)OC)=O)O

NGKHWQPYPXRQTM-UKFSEGPMSA-N

InChI=1S/C37H42N4O13.ClH/c1-17-32(46)20(38)13-27(53-17)54-22-15-37(51,23(16-42)39-40-24(43)9-4-3-5-12-41-25(44)10-11-26(41)45)14-19-29(22)36(50)31-30(34(19)48)33(47)18-7-6-8-21(52-2)28(18)35(31)49;/h6-8,10-11,17,20,22,27,32,42,46,48,50-51H,3-5,9,12-16,38H2,1-2H3,(H,40,43);1H/b39-23+;/t17-,20-,22-,27-,32+,37-;/m0./s1

N-[(E)-[1-[(2S,4S)-4-[(2R,4S,5S,6S)-4-amino-5-hydroxy-6-methyloxan-2-yl]oxy-2,5,12-trihydroxy-7-methoxy-6,11-dioxo-3,4-dihydro-1H-tetracen-2-yl]-2-hydroxyethylidene]amino]-6-(2,5-dioxopyrrol-1-yl)hexanamide;hydrochloride

480998-12-7; ALDOXORUBICIN HYDROCHLORIDE; MC-DOXHZN hydrochloride; INNO-206 hydrochloride; Aldoxorubicin?HCl; 1361563-03-2; Aldoxorubicin (hydrochloride); Aldoxorubicin Hydrochloride [USAN];

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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