Topoisomerase I

9-Aminocamptothecin cytotoxicity rises with increasing drug concentrations and longer exposure times in human breast (MCF-7), bladder (MGH-U1), and colon (HT-29) cancer cell lines. Unless the concentration of 9-Aminocamptothecin surpasses a threshold of 2.7 nm, minimal cell killing is also observed[1]. After 96 hours of medication exposure, PC-3, PC-3M, DU145, and LNCaP cells are inhibited by 9-Aminocamptothecin with IC50 values of 34.1, 10, 6.5, and 8.9 nM, respectively[2].

9-Aminocamptothecin (9-Amino-CPT) suppresses tumor growth at the lowest oral dose of 0.35 mg/kg/day; tumor regression is caused by s.c. administration (4 mg/kg/week) and higher oral doses of 0.75 and 1 mg/kg/day. All dosages of 9-Aminocamptothecin are well tolerated, and no group has experienced toxic death or weight loss greater than 10%[2]. A full remission is induced in 55% of SCID mice engrafted with human myeloid leukemia by 9-Aminocamptothecin. Both the intravenous and oral routes work equally well. The evaluation of 9-Aminocamptothecin as an antileukemic agent in a phase I trial involving AML patients is supported by the results obtained from this pre-clinical model[3].

The clonogenic assay is used to evaluate the cytotoxicity of 9-amino-CPT (9-amino-20(S)-camptothecin). 100–250 cells are inoculated in triplicate onto 60 15-mm dishes holding 5 mL of medium after exponentially growing cells are resuspended in media and the number of cells is ascertained using an electronic counter. 5 μL of 9-Aminocamptothecin stock solutions are added to the dishes to reach final concentrations of 137, 274 nM, 0.27, 1.37, 2.74, 13.7, 27.4, and 0.27 nM after an overnight incubation. Fresh medium is added to the dishes and the medium is removed by aspiration after 4, 8, 12, 24, 48, 72, and 240 hours of exposure. The ratio of colons in the drug-treated sample to those in the control (DMSO vehicle-treated) sample is used to calculate the percentage of survival at each drug concentration and exposure time[1].

Mice: On the seventh day following the KBM-3 cell inoculation, 9-Aminocamptothecin treatment is initiated. The following treatment is given to five groups of five mice each, with an average weight of 22 g, four days a week for three weeks: 1) PBS is IV injected into group 1 control mice; 2) group 2 mice receive 1.33 mg/kg Group 3 mice are given 1.33 mg/kg of 9-Aminocamptothecin IV. 4) Group 4 mice receive 2.0 mg/kg of 9-Aminocamptothecin IV; 5) Group 5 mice receive 2.0 mg/kg of 9-Aminocamptothecin orally by gavage. Gavage of 9-Aminocamptothecin orally[3]. The intravenous (i.v.) injection of the human acute myelogenous leukemia cell line KBM-3 into severe combined immune deficient (SCID) mice results in disseminated multi-organ human disease involvement in these animals which leads to their death over a defined period of time. We utilized this model of human leukemia to investigate the in vivo therapeutic efficacy of the topoisomerase I inhibitor 9-aminocamptothecin (9-AC) given by two different routes. Mice injected with KBM-3 were divided into five groups. Group 1 received only diluent and served as control. The four remaining groups were treated with 9-AC four days a week for three consecutive weeks as follows: group 2 received 1.33 mg/kg/dose, i.v.; group 3, 1.33 mg/kg/dose, orally (p.o.); group 4, 2.0 mg/kg/dose i.v. and group 5, 2.0 mg/kg/dose p.o.. All animals in the control group died from disseminated human leukemia by day 64 from grafting, with a median survival of 59 days. Eleven out of 20 treated mice survived with no evidence of disease and were sacrificed at the termination of the experiment on day 128. PCR-assisted tissue analysis for the presence of human DNA showed no evidence of human leukemia. In conclusion, 9-AC is an active agent in SCID mice engrafted with human myelogenous leukemia and should be explored in phase I-II trials. Oral and intravenous routes are equally effective.[3]

[1]. Pharmacological determinants of 9-aminocamptothecin cytotoxicity. Clin Cancer Res. 2001 Jan;7(1):168-74.

[2]. 9-Aminocamptothecin: a topoisomerase I inhibitor with preclinical activity in prostate cancer. Clin Cancer Res. 1997 Feb;3(2):287-94.

[3]. Activity of oral and intravenous 9-aminocamptothecin in SCID mice engrafted with human leukemia. Leuk Lymphoma. 1998 Dec;32(1-2):159-64.

9-Aminocamptothecin is a pyranoindolizinoquinoline.
Aminocamptothecin has been used in trials studying the treatment of Lymphoma, Gastric Cancer, Ovarian Cancer, Esophageal Cancer, and Ovarian Neoplasms, among others.
Aminocamptothecin is a water-insoluble camptothecin derivative. Aminocamptothecin binds to the nuclear enzyme topoisomerase I, thereby inhibiting repair of single-strand DNA breakages. Because the terminal lactone ring of aminocamptothecin required for the agent's antitumor activity spontaneously opens under physiological conditions to an inactive carboxy form, the drug must be administered over an extended period of time to achieve effective cytotoxicity. (NCI04)
See also: 10-Aminocamptothecin (annotation moved to).

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The camptothecins are a group of anticancer agents with a unique mechanism of action: poisoning of eukaryotic DNA topoisomerase I. 9-aminocamptothecin (9-AC), a potent water-insoluble derivative of camptothecin, is currently undergoing clinical testing. The kinetics of the active derivative 9-AC lactone in cell culture media was defined, and then 9-AC cytotoxicity against human breast (MCF-7), bladder (MGH-U1), and colon (HT-29) cancer cell lines was studied. The relationship between cytotoxic effects, drug concentration, and exposure time was then explored. For all of the three cell lines, 9-AC cytotoxicity increased with both higher drug concentrations and longer exposure times. However, when the duration of exposure was less than 24 h, cytotoxicity was limited and less than 1 log of cell killing occurred, even with very high drug concentrations. Minimal cell killing was also observed unless 9-AC concentrations exceeded a threshold of 2.7 nM. No fixed relationship between the survival fraction and the area under the drug concentration-time curve could be modeled that would fit all of the three cell lines. However, data for the three cell lines from the multiple exposure time experiments were fitted very well to the pharmacodynamic model C(n)t = k (r2, 0.90-0.99), where C is the drug concentration, n is the drug concentration coefficient, and t is the exposure time. For the three cell lines, to kill 1 log of cells, 0.30 < n < 0.85, which indicated that duration of exposure was more important than concentration. Our data support the use of 9-AC by infusion for 24 h or longer in clinical studies providing target plasma concentrations can be achieved.[1]

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9-Aminocamptothecin (9-AC) is a topoisomerase I inhibitor currently being developed as an antineoplastic agent. The aim of these preclinical studies was to assess the activity of 9-AC against prostate cancer, a malignancy notoriously insensitive to most cytotoxic agents in the clinic. The activity of 9-AC was first tested in vitro against one hormone-sensitive (LNCaP) and three hormone-resistant (PC-3, PC-3M, and DU145) human prostate cancer cell lines. After 96 h of drug exposure, concentrations required to inhibit cell viability to 50% of control values (IC50s) were 34.1, 10, 6.5, and 8.9 nm for PC-3, PC-3M, DU145, and LNCaP, respectively. Because 9-AC is known to undergo rapid hydrolysis, we assayed lactone levels in tissue culture medium over 24 h and found that the half-life was 20 min, with only 15%of the drug remaining as lactone at steady state. Consequently, the IC50s calculated from a single dose of the drug may represent overestimates. Subsequently, we tested the activity of a colloidal dispersion formulation of 9-AC against PC-3 implanted into flanks of nude mice. 9-AC was given for a total of 3 weeks by daily oral gavage (excluding weekends) or by twice weekly s.c. injections. 9-AC inhibited tumor growth at the lowest oral dose (0.35 mg/kg/day), whereas higher oral doses (0.75 and 1 mg/kg/day) and s.c. administration (4 mg/kg/week) caused tumor regression. 9-AC was well tolerated at all doses, with no toxic death or weight loss of more than 10% observed in any group. Finally, we considered that the activity of 9-AC seen in the mouse xenograft model might be explained, in part, by the relatively acidic tumor microenvironment, which would favor the formation of the more potent lactone. Simultaneous determination of plasma and tumor 9-AC lactone concentrations confirmed this hypothesis. Taken together, these studies suggest that 9-AC should be submitted for clinical trials in patients with prostate cancer.[2]

C20H17N3O4

363.36668

363.121

C, 66.11; H, 4.72; N, 11.56; O, 17.61

91421-43-1

72402

Light yellow to brown solid powder

1.6±0.1 g/cm3

819.6±65.0 °C at 760 mmHg

449.5±34.3 °C

0.0±3.1 mmHg at 25°C

1.771

0.44

2

6

1

27

775

1

C([C@]1(C(=O)OCC2C(N3CC4C=C5C(N)=CC=CC5=NC=4C3=CC1=2)=O)O)C

FUXVKZWTXQUGMW-FQEVSTJZSA-N

InChI=1S/C20H17N3O4/c1-2-20(26)13-7-16-17-10(6-11-14(21)4-3-5-15(11)22-17)8-23(16)18(24)12(13)9-27-19(20)25/h3-7,26H,2,8-9,21H2,1H3/t20-/m0/s1

(19S)-8-amino-19-ethyl-19-hydroxy-17-oxa-3,13-diazapentacyclo[11.8.0.02,11.04,9.015,20]henicosa-1(21),2,4,6,8,10,15(20)-heptaene-14,18-dione

9AC; 9aminoCPT; 9-amino-20-camptothecin; 9-amino-camptothecin; 9-AC9-amino-CPT; 9-amino-20(S)-camptothecin; 9aminocamptothecin; Aminocamptothecin

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: ~3.3 mg/mL (~9.2 mM)

Solubility in Formulation 1: ≥ 0.33 mg/mL (0.91 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% 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 3.3 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 0.33 mg/mL (0.91 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 3.3 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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Solubility in Formulation 3: ≥ 0.33 mg/mL (0.91 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (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 3.3 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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