• KSBB Journal
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• v.15 no.5
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• pp.428-433
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• 2000

Cell cultures of Camptotheca acuminata, which is known to produce the anticancer indole alkaloid camptothecin and its dervatives, were made to enhance the productivity of camptothecin. In suspension cultures, the maximum cell growth rate in exponential growth phase was $0.269day^{-1}$ which was correlated to 2.58days of cell doubling time. The production of camptothecin was non-growth associated. The camptothecin production was the highest at 11th day form inoculation and then it decreased. Various elicitors were applied to enhance the production of camptothecin. Both of jasmonic acid and cellulase increased the production of camptothecin. The optimal dosing time of elicitor was the beginning of the cultures. The combination of two elicitors was more effective to produce camptothecin than single applications. $20.42{\times}10^{-3}mg/\ell$ of camptothecin was obtained with combined application of two elicitors in two days.

• KSBB Journal
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• v.15 no.6
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• pp.609-614
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• 2000

Suspension cultures of Camptotheca acuminata, which is known to produce the anticancer indole alkaloid camptothecin and its derivatives, were made to increase camptothecin production. The capability of camptothecin production in suspended cells is decreased by repeated subculturein. Aggregated cells produced more camptothecin than single cells. Optimal cell aggregation was achieved in hybrid medium supplemented with 4% sucrose. Aggregated cells in hybrid medium with 4% sucrose produced $18.04{\times}10^{-4} mg/L$ of camptothecin. The control of shaking speeds was effective at inducing cell aggregation and camptothecin production. A shaking speed of 100 rpm was found optimum to increase the cell aggregation with a camptothecin production of $19.4{\times}10^{-4} mg/L$.

• The Korean Journal of Physiology and Pharmacology
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• v.13 no.2
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• pp.115-121
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• 2009

Functional defects in mitochondria are involved in the induction of cell death in cancer cells. We assessed the toxic effect of camptothecin against the human cervical and uterine tumor cell line SiHa with respect to the mitochondria-mediated cell death process, and examined the combined effect of camptothecin and anticancer drugs. Camptothecin caused apoptosis in SiHa cells by inducing mitochondrial membrane permeability changes that lead to the loss of mitochondrial membrane potential, decreased Bcl-2 levels, cytochrome c release, caspase-3 activation, formation of reactive oxygen species and depletion of GSH. Combination of camptothecin with other anticancer drugs (carboplatin, paclitaxel, doxorubicin and mitomycin c) or signaling inhibitors (farnesyltransferase inhibitor and ERK inhibitor) did not enhance the camptothecin-induced cell death and caspase-3 activation. These results suggest that camptothecin may cause cell death in SiHa cells by inducing changes in mitochondrial membrane permeability, which leads to cytochrome c release and activation of caspase-3. This effect is also associated with increased formation of reactive oxygen species and depletion of GSH. Combination with other anticancer drugs (or signaling inhibitors) does not appear to increase the anti-tumor effect of camptothecin against SiHa cells, but rather may reduce it. Combination of camptothecin with other anticancer drugs does not seem to provide a benefit in the treatment of cervical and uterine cancer compared with camptothecin monotherapy.

• Journal of Microbiology and Biotechnology
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• v.8 no.6
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• pp.631-638
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• 1998

Studies were made to elucidate the cell growth and the production of camptothecin and its derivatives in cell cultures of Camptotheca acuminata. High resolution HPLC chromatograms to analyze camptothecin and 10-hydroxycamptothecin in lactone and carboxylate forms were obtained with a fluorescence detector. Calli inductions were optimized with the young stem of explant on Schenk and Hildebrandt (SH) medium supplemented with 5 mg/l $\alpha$-naphthaleneacetic acid (NAA), 0.2 mg/l 6-benzylamino purine (BAP), 2.0% sucrose, and 0.5% agar. The hybrid medium, a mixture of SH and Murashige and Skoog (MS) salts, was developed for homogeneous suspension cultures without large cell aggregates. The optimum phytohormone concentrations for successful suspension cultures were 1.0mg/l of 2,4-D and 0.5 mg/l of kinetin. The highest growth in suspension cultures was observed when 49.7% (w/w) of the cells was composed of small aggregates which were below 0.1 mm in diameter. Time course changes of cell growth and camptothecin production showed that camptothecin accumulation was started at the end of the growth phase and the maximum content was obtained 10 days after inoculation. Yeast extract elicitor increased camptothecin accumulation 4 times. Methyl jasmonate and jasmonic acid also increased camptothecin production 6 and 11 times, respectively.

• Biotechnology and Bioprocess Engineering:BBE
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• v.3 no.2
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• pp.91-95
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• 1998

Camptothecin productoin was increased with elicitors, methyl jasmonate, jasmonic acid, yeast extract elicitor, and ferulic acid in suspension cultures of Camptotheca acuminata. Jasmonic acid was found to be the most efficient elicitor. Camptothecin production increased 11 times by using the optimum dosing concentration of jasmonic acid which was 50 ${\mu}$M. The kinetics of camptothecin accumulation in response to the treatment with jasmocin acid showed that the comptothecin accumulation reached the maximum value at 4 days after jasmonic acid dosing and then a rapid decrease in camptothecin accumulation was observed.

• YAKHAK HOEJI
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• v.43 no.3
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• pp.385-390
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• 1999

Camptothecin (CPT) has been known to induce apoptosis in various cancer cell lines. To examine the intracellular apoptotic death signal initiated by CPT, we investigated the possible connection between caspase-3 activation and GSH depletion during CPT-induced apoptosis in HL-60 cells. Treatment of cells with $1{\;}{\mu}M$ CPT induced PARP cleavage accompanied by DNA fragmentation. z-VAD-fmk, a caspase-3 inhibitor, blocked the CPT-induced DNA fragmentation. Pretreatment of cells with N-acetylcysteine, a precursor of GSH biosynthesis, failed to inhibit CPT-induced PARP celavage and DNA gragmenatation. No significant changes in GSH depletion is not essential for caspase activation during CPT-induced apoptosis. We also investigated whether CPT-induced apoptosis is associated with changes of the levels of Bax and Bcl-2, two proteins involved in the control of apoptosis. Bcl-2 levels exhibited a late decrease compared with the kinetics of DNA fragmentation, whereas Bax levels increased more rapidly after CPT treatment. These results suggest that Bax plays more important role than Bcl-2 in inducing DNA fragmentation and may function upsteam of proteolytic activation of caspase-3 pathway in CPT-induced apoptosis.

• KSBB Journal
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• v.16 no.1
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• pp.66-70
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• 2001

Factors affecting the supercritical carbon dioxide extraction of camptothecin(CPT) and 10-hydroxycamptothecin(HCPT) from the dried powder of Cmptotheca acuminata were studied. Only a few amount of CPT and HCPT was extracted with pure supercritical carbon dioxide. Methanol and ethanol were efficient modifiers to extract CPT and HCPT. At $40^{\circ}C$, 250 bar, 1 mL/min flow rate, 41% of CPT and 35% of HCPT were extracted with supercritical carbon dioxide modified with 16% of methanol. The diffusion effect of HCPT on extraction efficiency was studied in this solid-fluid system. Round matrix hot-ball model assumption revealed that the value of D/$r^2$ was 0.0072 $min^{-1}$ which was higher than that of solvent extraction with methanol.

• Journal of Life Science
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• v.10 no.6
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• pp.621-629
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• 2000

Camptothecin (CPT) is an antitumor alkaloid that has been isolated from the Chinese tree, Camptotheca acuminata. The cytotoxicity of CPT has been correlated to its inhibition of DNA topoisomerase (Topo) I by stabilizing drug-enzyme-DNA “cleavable complex" resulting in DNA single-strand breaks and DNA-protein crosslinks. This studies were designed to elucidate whether CPT regulates Topo I mediated by CPT in DNAs containing c-myc protooncogene. We have conducted experiments on Topo I purification, pUC-MYC I cloning and Topo I assay using electrophoresis, quantitative RT-PCR and Northern blotting techniques. CPT ingibited the relaxation activity of Topo I in pUC19 DNA at various concentrations (1-1000 $\mu$M), while it enhanced the cleavage of Topo I in the pUC-MYC I by forming a cleavable complex at relatively high concentrations (100-1000 $\mu$M). In HL-60 cells treated with CPT, the expression of c-myc gene was decreased over that in the control group with no changes in the expression of Topo I mRNA. Our results suggest that Topo I is the target of CPT cytotoxicity but it does not affect Topo I extression, and the suppression of c-myc mRNA expression by CPT is due to c-myc damage resulted from formation of a cleavable complex with CPT. CPT.

• Journal of Pharmaceutical Investigation
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• v.31 no.2
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• pp.113-117
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• 2001

CKD602, a camptothecin derivative, is a synthetic and water-soluble anticancer agent possessing of topoisomerase I inhibiting activity. DPPC and DSPE-PEG liposomal formulations entrapped with CKD602 were developed. DSPE-PEG liposome, or PEGylated liposome, encapsulating CKD602 composed of dipalmitoylphosphatidylcholine (DPPC), cholesterol and distearoyl-N-monoethoxy poly (ethyleneglycol) succinylphosphatidylethanolamine $(DSPE-PEG_{2000})$ (22:11:2) was prepared by reverse-phase evaporation method. Formed liposomes were characterized in terms of the morphology, size and encapsulation efficiency. To elucidate the in vitro stability, PEGylated liposome was incubated in human plasma, and the adsorbed proteins onto the surface of liposomes were applied to the SDS-PAGE. In vitro cytotoxicity of CKD602 encapsulated in PEGylated liposome was studied in human cervical cancer cell line (HeLa). CKD602 in PEGylated liposome was found to be 40-fold more effective $(IC_{50}=1\;nM)$ than free CKD602 $(IC_{50}=40\;nM)$ in inhibiting the growth of HeLa cells in vitro.

• Journal of Pharmaceutical Investigation
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• v.32 no.2
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• pp.81-85
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• 2002

Three polymorphic modifications of CKD-602, water soluble derivative of camptothecin, were obtained by the recrystallization from different organic solvents and characterized by differential scanning calorimetry (DSC), thermogravimetric analysis (TGA) and X-ray powder diffraction (XRPD). The major endothermic peaks of the DSC curve of Form 1, Form 2 and Form 3 was shown at $268.71^{\circ}C$, $247.83^{\circ}C$, $244.76^{\circ}C$, respectively. Form 2 was elucidated to be an acetic acid solvate and Form 3 was elucidated to be a methanol solvate. The dissolution patterns of these three modifications were also checked in distilled water at $37{\pm}0.5^{\circ}C$, for 30 minutes. The polymorphic modifications showed differences in the dissolution rate. The dissolution rate of Form 1 was faster than that of other polymorphic modifications. When stored at different relative humidity over the period of 3 months, all of the polymorphic modifications did not undergo transformation.