Klaus Schieweck

Highly selective inhibition of estrogen biosynthesis by CGS 20267, a new non-steroidal aromatase inhibitor

CGS 20267 is a new non-steroidal compound which potently inhibits aromatase in vitro (IC50 of 11.5 nM) and in vivo (ED50 of 1-3 micrograms/kg p.o.), CGS 20267 maximally inhibits estradiol production in vitro in LH-stimulated hamster ovarian tissue at 0.1 microM with an IC50 of 0.02 microM and does not significantly affect progesterone production up to 350 microM. In ACTH-stimulated rat adrenal tissue in vitro, aldosterone production was inhibited with an IC50 of 210 microM (10,000 times higher than the IC50 for estradiol production); no significant effect on corticosterone production was seen at 350 microM. In vivo, in ACTH-treated rats, CGS 20267 does not affect plasma levels of corticosterone or aldosterone at a dose of 4 mg/kg p.o. (1000 times higher than the ED50 for aromatase inhibition in vivo). In adult female rats, a 14-day treatment with 1 mg/kg p.o. daily, completely interrupts ovarian cyclicity and suppresses uterine weight to that seen 14 days after ovariectomy. In adult female rats bearing estrogen-dependent DMBA-induced mammary tumors, 0.1 mg/kg p.o. given daily for 42 days caused almost complete regression of tumors present at the start of treatment. Thus compared to each other, CGS 16949A and CGS 20267 are both highly potent in inhibiting estrogen biosynthesis in vitro and in vivo. The striking difference between them is that unlike CGS 16949A, CGS 20267 does not affect adrenal steroidogenesis in vitro or in vivo, at concentrations and doses several orders of magnitude higher than those required to inhibit estrogen biosynthesis.

Inhibition of estrogen biosynthesis and its consequences on gonadotrophin secretion in the male

Of the gonadal steroids in the male, testosterone is the most important regulator of gonadotrophin secretion. However, whether testosterone affects gonadotrophin secretion directly or whether it must first be aromatized to estrogens is controversial. We have reported extensively on the endocrine and anti-tumor effects of the non-steroidal aromatase inhibitors CGS 16949A and CGS 20267 in adult female rats. In these animals, both inhibitors potently and selectively inhibit estrogen biosynthesis. Thus these agents can be effectively used in studying estrogen-dependent processes. CGS 16949A was administered for 14 days to adult male rats, over a dose range which in females suppresses estradiol and elevates LH. In male rats a suppression of estradiol was seen, however, there was no significant effect on either serum LH or on the weights of androgen-dependent organs. CGS 16949A, when administered to healthy men at a dose of 1 mg b.i.d. for 10 days, causes a significant fall in plasma estradiol and significant elevations of plasma FSH and testosterone. Dose-dependent suppression of serum estradiol and an increase in serum testosterone and LH are seen after administration of single oral doses of CGS 20267. These results indicate that in the male rat, inhibition of aromatization of testosterone to estrogens does not influence gonadotrophin secretion whereas in men the negative feedback exerted by testosterone on gonadotrophin secretion is dependent on the aromatization of testosterone to estrogens.

Novel aromatase inhibitors

Aminoglutethimide (AG), an inhibitor of the aromatase enzyme, inhibits the biosynthesis of estrogens and displays well-documented anti-tumor efficacy in breast-cancer. However, this efficacy is accompanied by a relative lack of specificity in inhibiting aromatase and moderate tolerability. We report on two new non-steroidal aromatase inhibitors (CGS 16949A and CGS 18320B) which are more potent, selective and efficacious in their inhibition of aromatase than AG. Both compounds inhibit aromatase more potently in vitro and in vivo (over 400 and 1000 times respectively) than AG. They are both more selective in their inhibition of aromatase with CGS 18320B showing an improved selectively over CGS 16949A. When administered to adult female rats, both compounds elicit responses in serum hormones similar to those seen after ovariectomy. The duration of action of CGS 18320B, however, appears to be longer than that of CGS 16949A. CGS 18320B and CGS 16949A cause almost complete regression of DMBA-induced mammary tumors in adult female rats and almost completely suppress the appearance of new tumors. Thus CGS 16949A and CGS 18320B represent significant advances in the search for novel aromatase inhibitors which are more potent, selective and efficacious than aminoglutethimide.

CGP 55 847, liposome-delivered zinc(II)-phthalocyanine as a phototherapeutic agent for tumors

Zinc(II)-phthalocyanine (Zn-Pc) was chosen for development as a second-generation photosensitizer for photodynamic therapy (PDT) of tumors and for benign conditions because of its advantageous chemical and photophysical properties. Zn-Pc displayed good selectivity for malignant tissue in pharmacokinetic studies with Meth-A-sarcoma-bearing BALB/c mice when injected in a dose of 0.125 mg/kg, delivered by CGP 55 847. Intravenous doses of Zn- Pc ranging from 0.032 to 0.375 mg/kg caused tumor necrosis and, subsequently, cure of Meth-A-sarcoma-bearing mice when phototreatment was performed 48 hours after injection of CGP 55 847. Intravenous injection of Zn-Pc into hairless mice in doses ranging from 0.1 to 1.0 mg/kg caused dose- and time-dependent phototoxicity. We conclude that the promising pharmacological properties of liposomally delivered Zn-Pc, along with its advantageous chemical and photophysical properties, warrant the development of CGP 55 847 as a candidate drug for photodynamic therapy of tumors in humans.

Pharmaceutical development of CGP 55847: a liposomal Zn-phthalocyanine formulation using a controlled organic solvent dilution method

Liposomes were prepared containing zinc-phthalocyanine (ZnPc). The composition was ZnPc/POPC/OOPS (1:90:10 w/w /w). The phospholipids (PL) were dissolved in t-butanol at 50 degree(s)C under magnetic stirring and mixed with ZnPc, dissolved in NMP for two hours in an ultrasonic bath at 80 degree(s)C. This mixture (50 degree(s)C) was diluted with lactose- NaCl solution (9.475% lactose, 0.027% NaCl) at 4 degree(s)C using a dynamic mixer. The collected liposomal suspension was concentrated first, to 20 mg PL/ml suspension and then the organic solvents were removed by tangential flow filtration (CentrasetteR) against a ten fold volume of lactose-NaCl solution. After sterile filtration the liposomal suspension was freeze-dried for 24 hours.

Synthesis and biological evaluation of a series of new germanium phthalocyanines incorporated into liposomes--part II: biological evaluation

The pharmacokinetic and phototherapeutic properties of new phthalocyanines with Ge(IV) or Si(VI) as the central metal ion and cholesterol, cholestan or long-chain fatty acids residues as axial ligands to the central ion have been studied in tumor-bearing mice. The new photosensitizers were selectively taken up by and relatively quickly released from the tumors. Except for Si(IV)-Pc, which showed a comparably high selectivity for tumor versus peritumoral tissue, all Ge(IV)-Pc were less selective than liposomal Zn-Pc (CGP 55847). However, all the new compounds showed excellent phototherapeutic efficiency at very low drug and light doses in studies in Meth-A-sarcoma-bearing mice.

CGP 6809, a sugar-containing nitrosourea derivative: pharmacological and physicochemical properties.

SummaryCGP 6809 is a water-soluble nitrosourea derivative with quite distinct chemical and biological properties as compared with the well-known representatives of this class of compounds. It is related to the antibiotic streptozotocin, from which it is distinguished in the structure of the sugar moiety and the position of the methylnitrosourea residue. CGP 6809 possesses practically the same alkylating potential as streptozotocin; however, its carbamoylating activity is comparable with that of CCNU. In contrast to other nitrosourea derivatives, CGP 6809 showed relatively little activity in murine leukemias but was markedly active in solid transplantable melanomas (Harding-Passay, B16), in the 11095 prostate carcinoma, and in a substrain of Yoshida hepatoma (AH 7974) resistant to BCNU and CCNU. In the Ehrlich and Yoshida ascitic tumors complete responses were seen with no toxic death. Dose-dependent activity was found in the human lung carcinoma MBA 9812 and almost complete growth inhibition was achieved in the human melanoma WM 47 by both the oral and parenteral routes of administration. However, mammary tumor lines (Ca 755, 2661/61, R-3230AC), the Guerin-T8 uterus epithelioma, and the Rous sarcoma/S-R proved to be relatively refractory to this drug. This was also the case for the Lewis lung carcinoma implanted i. m. or s. c. However, development of lung metastases was markedly inhibited. Combination therapy using CGP 6809 with cyclophosphamide, 5-fluorouracil, or chlorambucil in the same model led to partial responses of the primary tumor as well as almost total eradication of lung metastases.

New liposome-bound Ge(IV)-phthalocyanine (CGP 55398) for photodynamic therapy of tumors: preliminary studies

A phthalocyanine derivative with two cholesterol moieties as axial ligands to the central Ge(IV) ion efficiently photosensitizes the oxidative modification of L-tryptophan. Administration of liposome-bound GePc to Balb/c mice bearing a MS-2 fibrosarcoma yields a quantitative release of the dye to serum lipoproteins, followed by a selective accumulation in the tumor as well as a low content in the skin. At 24 h after injection of 0.76 mg/kg GePc, the tumor was irradiated with 600 - 700 nm light; tumor necrosis appeared in all treated mice as a consequence of extensive damage of cellular and stromal elements.