MicroRNAs as novel players in skin development, homeostasis and disease.

MicroRNAs (miRNAs) are a group of newly discovered small (∼19-24 nucleotides), noncoding RNAs that modulate gene expression by interacting with the 3' untranslated region of the corresponding target gene messenger RNA (mRNA). miRNAs have been estimated to regulate more than one-third of protein-encoding mRNAs. As a consequence, cellular protein expression and a large number of biological processes are influenced by miRNA-mediated post-transcriptional regulation of gene expression. The severe phenotype of mice lacking key enzymes of the miRNA biogenesis pathway (Dgcr8 and Dicer) in the skin confirmed the essential function of miRNAs in this tissue. In addition, a growing number of reports has identified miRNAs as regulators of the morphogenesis and homeostasis of the skin and its appendages, and miRNA deregulation was shown to be associated or even causally related to several skin diseases. Profiling studies have identified numerous differentially regulated miRNAs associated with physiological (e.g. keratinocyte differentiation) and pathological (e.g. psoriasis, melanoma) processes. These data bear enormous potential for further studies. Because of the easy accessibility of the skin, it is plausible to anticipate that, once efficient and safe methods for the topical delivery of substances mimicking or modulating miRNA activity become available, skin diseases will be among the first to be approached with miRNA-based therapies. This review article gives a short introduction to miRNA biology and summarizes and discusses existing evidence for a role of these molecules in the skin.

Betacellulin overexpression in transgenic mice improves glucose tolerance and enhances insulin secretion by isolated islets in vitro.

Betacellulin (BTC), a ligand of the epidermal growth factor receptor, has been shown to promote growth and differentiation of pancreatic beta-cells and to improve glucose metabolism in experimental diabetic rodent models. We employed transgenic mice (BTC-tg) to investigate the effects of long-term BTC overabundance on islet structure and glucose metabolism. Expression of BTC is increased in transgenic islets, which show normal structure and distribution of the different endocrine cell types, without pathological alterations. BTC-tg mice exhibit lower fasted glucose levels and improved glucose tolerance associated with increased glucose-induced insulin secretion. Surprisingly, quantitative stereological analyses revealed that, in spite of increased cell proliferation, the islet and beta-cell volumes were unchanged in BTC-tg mice, suggesting enhanced cell turnover. Insulin secretion in vitro was significantly higher in transgenic islets in medium containing high glucose (11.2 or 16.7mM) as compared to control islets. Our results demonstrate that long-term BTC overabundance does not alter pancreatic islet structure and beta-cell mass, but enhances glucose-induced insulin secretion in vivo as well as in vitro.

A highly sensitive model for quantification of in vivo tumor angiogenesis induced by alginate-encapsulated tumor cells.

A remarkable approach to a specific tumor angiogenesis model in vivo is the use of alginate implants encapsulating tumor cells. However, this previously reported approach has often been questioned because of doubts regarding the relevance of hemoglobin at the alginate implant as a parameter of vascularization. In the present investigation, we examined whether or not the use of the blood pool agents FITC-dextran of high molecular weight would significantly improve the determination of vascularization at the alginate implant. In our experiments, we found a rapid distribution of FITC-dextran within the blood circulation of mice after i.v. bolus injection. The amount of FITC-dextran within alginate implants strongly correlated with the number of LL2 carcinoma cells or B16/F10 cells encapsulated. Even a low number of 10(3) cells per alginate implant led to a significantly increased accumulation of FITC-dextran. A more than 10-fold stimulation above that of controls was found with alginate implants containing 10(4) LL2 or B16/F10 tumor cells. Using the investigational compound AGM-1470 in different treatment schedules, we found that quantification of alginate implant anglogenesis with FITC-dextran is a sensitive method for the determination of angiogenesis inhibition. In conclusion, our results demonstrated that the use of FITC-dextran enables highly sensitive, quantitative measurement of blood vessel formation by alginate implants.

Signaling-inactive epidermal growth factor receptor/ligand complexes in intact carcinoma cells by quinazoline tyrosine kinase inhibitors.

Several inhibitors of EGF receptor (EGFR) tyrosine kinase activity have been developed that compete with ATP at its binding site such as the quinazolines PD 153035 and ZD 1839 or the 4,5-dianilino-phthalimides DAPH1 and DAPH2. When tested on human A431 cells, the quinazolines completely blocked EGF-induced receptor phosphorylation at 100 nM, whereas it was inhibited by DAPH1 and DAPH2 by only 20% at 3 microM. Quinazoline-treated A431 as well as tumor cells expressing less EGFR (A549, MDA MB 231, and T47D) bound 3- to 6-fold more (125)I-labeled EGF than untreated intact control cells. Scatchard analysis revealed the disappearance of low- and high-affinity EGFR on A431 cells upon PD 153035 treatment. A single receptor class of intermediate ligand binding affinity emerged and its number corresponded to the sum of the two classes. DAPH1 and DAPH2 did not change ligand binding properties of EGFR. PD 153035 exerted the most potent effects on EGF binding to A431 or on inhibiting EGF-stimulated growth of rat MTLn3 cells at low ligand concentrations. Cross-linking of EGFR on PD 153035-treated A431 cells indicated the formation of inactive dimers that further increased upon addition of EGF. Chemical cross-linking of (125)I-labeled EGF to PD 153035-treated A431 cells revealed increased binding to monomeric and dimeric EGFR. Thus, the quinazolines sequestered EGFR plus the ligand into inactive receptor/ligand complexes. This novel mode of action of quinazoline tyrosine kinase inhibitors may be the basis for their extraordinary potency especially in conditions when the ligand is present in limiting amounts.

PTK787/ZK 222584, a novel and potent inhibitor of vascular endothelial growth factor receptor tyrosine kinases, impairs vascular endothelial growth factor-induced responses and tumor growth after oral administration.

PTK787/ZK 222584 (1-[4-chloroanilino]-4-[4-pyridylmethyl] phthalazine succinate) is a potent inhibitor of vascular endothelial growth factor (VEGF) receptor tyrosine kinases, active in the submicromolar range. It also inhibits other class III kinases, such as the platelet-derived growth factor (PDGF) receptor beta tyrosine kinase, c-Kit, and c-Fms, but at higher concentrations. It is not active against kinases from other receptor families, such as epidermal growth factor receptor, fibroblast growth factor receptor-1, c-Met, and Tie-2, or intracellular kinases such as c-Src, c-Abl, and protein kinase C-alpha. PTK787/ZK 222584 inhibits VEGF-induced autophosphorylation of kinase insert domain-containing receptor (KDR), endothelial cell proliferation, migration, and survival in the nanomolar range in cell-based assays. In concentrations up to 1 microM, PTK787/ZK 222584 does not have any cytotoxic or antiproliferative effect on cells that do not express VEGF receptors. After oral dosing (50 mg/kg) to mice, plasma concentrations of PTK787/ZK 222584 remain above 1 microM for more than 8 h. PTK787/ZK 222584 induces dose-dependent inhibition of VEGF and PDGF-induced angiogenesis in a growth factor implant model, as well as a tumor cell-driven angiogenesis model after once-daily oral dosing (25-100 mg/kg). In the same dose range, it also inhibits the growth of several human carcinomas, grown s.c. in nude mice, as well as a murine renal carcinoma and its metastases in a syngeneic, orthotopic model. Histological examination of tumors revealed inhibition of microvessel formation in the interior of the tumor. PTK787/ZK 222584 is very well tolerated and does not impair wound healing. It also does not have any significant effects on circulating blood cells or bone marrow leukocytes as a single agent or impair hematopoetic recovery after concomitant cytotoxic anti-cancer agent challenge. This novel compound has therapeutic potential for the treatment of solid tumors and other diseases where angiogenesis plays an important role.

The EGFR-HER2 module: a stem cell approach to understanding a prime target and driver of solid tumors.

The epidermal growth factor receptor (EGFR) and a coreceptor denoted HER2/ERBB2 are frequently overexpressed or mutated in solid tumors, such as carcinomas and gliomas. In line with driver roles, cancer drugs intercepting EGFR or HER2 currently outnumber therapies targeting other hubs of signal transduction. To explain the roles for EGFR and HER2 as prime drivers and targets, we take lessons from invertebrates and refer to homeostatic regulation of several mammalian tissues. The model we infer ascribes to the EGFR-HER2 module pivotal functions in rapid clonal expansion of progenitors called transient amplifying cells (TACs). Accordingly, TACs of tumors suffer from replication stress, and hence accumulate mutations. In addition, several lines of evidence propose that in response to EGF and related mitogens, TACs might undergo dedifferentiation into tissue stem cells, which might enable entry of oncogenic mutations into the stem cell compartment. According to this view, antibodies or kinase inhibitors targeting EGFR-HER2 effectively retard some solid tumors because they arrest mutation-enriched TACs and possibly inhibit their dedifferentiation. Deeper understanding of the EGFR-HER2 module and relations between cancer stem cells and TACs will enhance our ability to control a broad spectrum of human malignancies.

The target cell of transformation is distinct from the leukemia stem cell in murine CALM/AF10 leukemia models.

The CALM/AF10 fusion gene is found in various hematological malignancies including acute myeloid leukemia (AML), T-cell acute lymphoblastic leukemia and malignant lymphoma. We have previously identified the leukemia stem cell (LSC) in a CALM/AF10-driven murine bone marrow transplant AML model as B220+ lymphoid cells with B-cell characteristics. To identify the target cell for leukemic transformation or 'cell of origin of leukemia' (COL) in non-disturbed steady-state hematopoiesis, we inserted the CALM/AF10 fusion gene preceded by a loxP-flanked transcriptional stop cassette into the Rosa26 locus. Vav-Cre-induced panhematopoietic expression of the CALM/AF10 fusion gene led to acute leukemia with a median latency of 12 months. Mice expressing CALM/AF10 in the B-lymphoid compartment using Mb1-Cre or CD19-Cre inducer lines did not develop leukemia. Leukemias had a predominantly myeloid phenotype but showed coexpression of the B-cell marker B220, and had clonal B-cell receptor rearrangements. Using whole-exome sequencing, we identified an average of two to three additional mutations per leukemia, including activating mutations in known oncogenes such as FLT3 and PTPN11. Our results show that the COL for CALM/AF10 leukemia is a stem or early progenitor cell and not a cell of B-cell lineage with a phenotype similar to that of the LSC in CALM/AF10+ leukemia.

Antiangiogenic chemotherapeutic agents: characterization in comparison to their tumor growth inhibition in human renal cell carcinoma models.

The mechanism of action of anticancer chemotherapeutic agents is mainly thought to be due to a direct inhibition of tumor cell proliferation. The enhanced endothelial cell proliferation rate in tumor specimens raised the question of whether therapeutic effects of chemotherapeutic agents might be at least partially attributed to inhibition of tumor angiogenesis. In the present study, we investigated the potential effects of chemotherapeutic agents on human renal carcinoma angiogenesis with the alginate implantation model in mice. For the first time, we also compared results from the angiogenesis model with the inhibitory effects on growth of s.c. xenografts in nude mice. Vincristine and bleomycin exerted strong inhibition of tumor angiogenesis in both carcinoma lines close to the level of the standard antiangiogenic agent O-chloroacetyl-carbamyl-fumagillol (AGM-1470; T/C 22%). Adriamycin reduced angiogenesis of Caki-2 cells (T/C 33%) but had no effect on Caki-1 angiogenesis (T/C 137%). Etoposide and 5-fluorouracil reduced Caki-1 tumor angiogenesis but had no effect on Caki-2. Despite antiangiogenic effects in both carcinoma lines, vincristine, bleomycin, and AGM-1470 significantly reduced only the growth of fast-growing Caki-1 s.c. xenografts but not the slow-growing Caki-2. Antivascular effects by bleomycin and AGM-1470 were also shown by a decrease of microvessel density in nude mouse xenografts. Our findings suggest that chemotherapeutic agents may exert inhibition of tumor angiogenesis, which could be exploitable by combination therapy of fast-growing tumors. The resistance of the slow-growing Caki-2 carcinoma against acute angiogenesis inhibition indicates a need for well-tolerated angiogenesis inhibitors. Our results also suggest the use of fast-growing s.c. xenografts for demonstrating growth inhibition by antiangiogenic compounds. Further characterization of antiangiogenic compounds considered for clinical application should, however, have its focus on slow-growing tumors, which are not accessible for most therapeutic strategies.

The stable prostacyclin analogue Cicaprost inhibits metastasis to lungs and lymph nodes in the 13762NF MTLn3 rat mammary carcinoma.

Prostacyclin and its stable analogues have been shown to interfere specifically with certain steps of the metastatic cascade. The antimetastatic activity of the stable prostacyclin analogue Cicaprost (Schering AG) on haematogenous metastasis in a series of tumours in rats and mice has been well established. In order to test the effect of Cicaprost on lymphogenous metastasis we chose the metastatic cell clone MTLn3 derived from the 13762NF rat mammary carcinoma. The effect of Cicaprost on prevention of lung metastasis, lymph node metastasis and primary tumour growth was investigated. Cicaprost given in daily doses of 0.01, 0.03 and 0.1 mg/kg orally, reduced the number of lung metastases in a dose-dependent manner. Whereas the median number of lung metastases in the controls was greater than 1000, Cicaprost at a dose of 0.1 mg/kg reduced the number of lung metastases to between 11 and 100. The weight of the ipsilateral axillary lymph nodes was diminished by Cicaprost to 30-50% of controls. Moreover, metastasis to the contralateral axillary lymph node was completely inhibited by Cicaprost at all three doses tested. Cicaprost did not influence the growth rate of the MTLn3 cell clone implanted into the mammary fat pad or the weight of the primary tumour at the end of treatment. In conclusion, in addition to its dose-dependent effect on haematogenous metastasis, Cicaprost strongly inhibits lymph node metastasis.

Insulin-like growth factor-binding protein-4 inhibits growth of the thymus in transgenic mice.

Numerous in vitro studies have demonstrated that IGF-binding protein (IGFBP)-4 is a consistent inhibitor of IGF actions. In order to investigate the functions of IGFBP-4 in vivo, transgenic mice were generated by microinjection of a transgene, in which the murine Igfbp4 cDNA is driven by the H-2K(b) promoter, and followed by a splicing cassette and polyadenylation signal of the human beta-globin gene. Transgene mRNA was expressed ubiquitously, and elevated IGFBP-4 protein was detected in the spleen, thymus, kidney and lung of transgenic mice. The activities of serum IGFBPs were not changed in transgenic mice. Immunohistochemical studies revealed transgene expression predominantly in the thymic medulla and red pulp of the spleen. Body weight and the weights of the spleen, kidney and lung of transgenic mice were not different from controls. In contrast, the thymus of transgenic mice showed a significantly reduced weight and cortex volume. In transgenic thymus and spleen, cell proliferation was inhibited and apoptosis was stimulated. Transgenic mice showed normal T- and B-cell development and normal basal plasma immunoglobulin levels. In conclusion, overexpression of IGFBP-4 inhibits growth of the thymus. IGFBP-4 excess inhibits cell proliferation and stimulates apoptosis in lymphoid tissues, but does not affect lymphocyte development. These findings suggest that IGFBP-4 is a potential growth inhibitor of lymphoid tissues.

2-Alkyl-substituted 1,1-bis(4-acetoxyphenyl)-2-phenylethenes. Estrogen receptor affinity, estrogenic and antiestrogenic properties, and mammary tumor inhibiting activity.

1,1-Bis(4-acetoxyphenyl)-2-phenylethenes that are substituted with H, CH3, C2H5, n-C3H7, i-C3H7, or CH2CF3 in position 2 were synthesized in order to study the influence of the alkyl side chain on estradiol receptor affinity, estrogenic and antiestrogenic properties, and inhibition of the hormone-dependent MXT mammary carcinoma of the mouse. Furthermore, the double bond of 1,1-bis(4-acetoxyphenyl)-2-phenylbut-1-ene was hydrogenated or epoxidated to yield the corresponding ethane and oxirane derivative. Compounds 14 (R = H), 15 (R = CH3), and 16 (R = C2H5) had the best binding affinities. Lengthening the side chain, hydrogenation, or epoxidation decreased the RBA values. In the immature mouse assay, 15 (R = CH3) and 19 (R = CH2CF3) had the highest uterotrophic activity. There was no correlation between receptor affinity and estrogenic properties. Compounds 14 (R = H), 17 (R = n-C3H7), the ethane 20, and the oxirane 21 had some antiuterotrophic activity in a low dosage. The MXT tumor was best inhibited by compounds 15 (R = CH3), 16 (R = C2H5), and 18 (R = i-C3H7) without significant elevation of the uterine weight determined at the end of the experiment. The antitumor effect of 15, 16, and 18 was significantly better than that of tamoxifen. In this series, a certain estrogenic potency in the immature mouse test seems to be necessary for a good antitumor activity, as all compounds with antiuterotrophic and low uterotrophic properties did not exert any significant tumor-inhibiting effect.

2-Phenylindenes: development of a new mammary tumor inhibiting antiestrogen by combination of estrogenic side effect lowering structural elements.

A new antiestrogenic, mammary tumor inhibiting 2-phenylindene was developed by the use of structural elements that we have shown to decrease estrogenic side effects but to increase antiestrogenic activity and retain the antitumor effect of certain stilbenes. The new 2-phenylindenes were synthesized from their corresponding methoxy-substituted 3,4-diphenylhexane-3,4-diols by cyclization with acetyl chloride and acetic anhydride and subsequent ether cleavage and acetylation. In this series, the 2-phenylindene derivative (compound 13) with a 5,6,3',4'-tetraacetoxy and a 1-methyl-3-ethyl substitution had the highest affinity for the estrogen receptor, the strongest antiestrogenic effect, and the lowest estrogenic effect. This compound was superior to the 2-phenylindenes with 5,3'-diacetoxy substitution or 1,1-dimethyl and 3-isopropyl moieties, respectively. Compound 13 exhibited a strong, significant inhibiting effect on the growth of the hormone-dependent MXT mouse mammary tumor without estrogenic side effects.

Indolo[2,1-a]isoquinolines. Syntheses, steroid hormone receptor binding affinities, and cytostatic activity.

A number of acetoxy-substituted 5,6-dihydroindolo[2,1-a]isoquinolines were synthesized and tested for binding affinity for steroid hormone receptors. All of the derivatives bind to the estrogen receptor with RBA values ranging from 1.5 to 17 (17 beta-estradiol = 100). Some of them show binding affinities for the androgen receptor as well. In the mouse uterine weight test, the tetracycles proved to be weak estrogens with partial antagonistic activity. All of the compounds were tested in vitro for cytostatic activity with hormone-independent MDA-MB 231 and hormone-dependent MCF-7 breast cancer cells. A cytostatic effect was found in both cell lines. The comparison of results exhibited a stronger inhibitory effect on MCF-7 cells only for compounds with high binding affinity for the estrogen receptor. For those derivatives, it can be assumed that the growth inhibition is partly mediated by the estrogen receptor.

Catechol estrogens of the 1,1,2-triphenylbut-1-ene type: relationship between structure, estradiol receptor affinity, estrogenic and antiestrogenic properties, and mammary tumor inhibiting activities.

1,1,2-Triphenylbut-1-enes (26-35), which are substituted with one or two 3,4-diacetoxy groups or with one 3,4-diacetoxy and one 3- or 4-acetoxy group in two aromatic rings, were synthesized. The occurring E and Z isomers were isolated, and their identity was established by 1H NMR spectroscopy. A study on structure-activity relationship was carried out with regard to estradiol receptor affinity in vitro, estrogenic and antiestrogenic properties in the immature mouse, and inhibition of the hormone-dependent MXT mammary tumor of the mouse in vivo. Among the tested compounds, most of the 1,1-disubstituted 1,1,2-triphenylbut-1-enes (29, Z-30, Z,E-31) and (E)-1-(3-acetoxyphenyl)-1-phenyl-2-(3,4-diacetoxyphenyl)but- 1-ene (E-35) as well as its respective Z isomer (Z-35) exerted antiestrogenic properties. Compounds Z-30, Z,E-31, Z-35, and E-35 inhibited the growth of the hormone-dependent MXT tumor. The best antitumor effect without estrogenic side effects during therapy was shown by E-35.

Acetoxy-substituted 1,1,2-triphenylbut-1-enes with antiestrogenic and mammary tumor inhibiting properties.

1,1,2-Triphenylbut-1-enes (E- and Z-10-12), which are substituted with one p- and one m-acetoxy group in two different aromatic rings, were synthesized. The E and Z isomers were isolated, and their identity was established by 1H NMR spectroscopy. A study of the structure-activity relationship was carried out with regard to estradiol receptor affinity in vitro, estrogenic and antiestrogenic properties (mouse), inhibition of the hormone-dependent human MCF7 breast cancer cell line in vitro, and the hormone-dependent MXT mammary tumor of the mouse in vivo. Among the tested compounds, (E)- and (Z)-1-(3-acetoxyphenyl)-1-(4-acetoxyphenyl)-2-phenylbut-1-enes+ ++ (E-10 and Z-10) and (Z)-1-(3-acetoxyphenyl)-1-phenyl-2-(4-acetoxyphenyl)-but-1-ene (Z-12) proved to be partial antiestrogens, which lead to an inhibition of the MCF7 cell line. They exert a growth-inhibiting activity on the hormone-dependent MXT mammary carcinoma of the mouse. In the case of E-10 and Z-10, this effect is only slightly weaker than that of 1,1-bis(4-acetoxyphenyl)-2-phenylbut-1-ene (13) and tamoxifen. Under the applied experimental conditions, there were no significant changes of uterine weight as an indicator of estrogenic side effects.

Mammary tumor inhibiting effect of 3,3'-diacetoxy-alpha, beta-dialkylstilbenes and of related stilbene oxides.

3,3'-Diacetoxy-alpha, beta-dialkylstilbenes (alkyl = CH3 to C4H9, 1-4) 3,3'-dihydroxy-alpha, beta-diethylstilbene (5), and their corresponding stilbene oxides (6-10) were synthesized. Compounds 1-10 competitively antagonized in vitro the interaction of [3H]estradiol with its receptor. 3,3'-Diacetoxy-alpha, beta-diethylstilbene (2), 3,3'-diacetoxy-alpha, beta-diethylstilbene oxide (7), and their phenolic analogues (5 and 10) were most effective. Shortening or lengthening the alkyl side chains led to a decrease in receptor affinity. Among the stilbenes and epoxides, those with C2H5 and C3H7 groups (2, 3, 5 and 7, 8, 10) caused the strongest inhibition of the growth of a hormone-dependent postmenopausal human mammary carcinoma serially implanted in nude mice. The strong antitumor activity of 5 and 10 was confirmed by experiments on the 9,10-dimethyl-1,2-benzanthracene-induced, hormone-dependent mammary carcinoma of the Sprague-Dawley rat.

Influence of alkyl-chain fluorination on the action of mammary tumor inhibiting 2,3-bis(hydroxyphenyl)butanes and 2,3-bis(hydroxyphenyl)but-2-enes.

trans-1,2-Bis(trifluoromethyl)-1,2-bis(4- and 3-hydroxyphenyl)ethenes 2 and 4 were prepared by reductive coupling (TiCl4/Zn/pyridine) of the methoxy-substituted alpha, alpha, alpha-trifluoroacetophenones, separation of the resulting cis- and trans-stilbene derivatives, and ether cleavage with BBr3. The cis-stilbenes were catalytically hydrogenated to give meso-1,1,1,4,4,4-hexafluoro-2,3-bis(4- and 3-hydroxyphenyl)butanes 6 and 8. Compounds 2, 4, 6, and 8 showed 2- to 10-fold increased binding affinities for the estradiol receptor (E2R) and enhanced estrogenicity in the uterine weight test of the immature mouse compared to their unfluorinated analogues. Compound 8 exhibited a 46% inhibition of the estrone-stimulated uterine growth. Antitumor activity was evaluated with use of the transplantable, hormone-dependent MXT mammary tumor of the BD2F1 mouse. All compounds showed tumor growth inhibitory activity corresponding to their RBA values. The most interesting compound 8 led to a significant inhibition of the tumor growth on the DMBA-induced hormone-dependent mammary carcinoma of the Sprague-Dawley rat.

1,1,2-triphenylbut-1-enes: relationship between structure, estradiol receptor affinity, and mammary tumor inhibiting properties.

1,1,2-Triphenylbut-1-enes, which are substituted with acetoxy groups on one, two, or three aromatic rings in the para and/or meta positions, were synthesized. The identity of the occurring E and Z isomers were established by 1H NMR spectroscopy. A study on structure-activity relationships was carried out with regard to estradiol receptor affinity and to inhibiting effects on the growth of a postmenopausal human mammary carcinoma implanted in nude mice. The para-substituted compounds generally exhibited a higher receptor affinity and a better antitumor activity than the corresponding meta-substituted ones. The E isomers were superior to the respective Z isomers in those two properties. The tumor-inhibiting effect of the mono- and disubstituted compounds was better than that of the trisubstituted ones. Except for the trisubstituted compounds, they all show a good correlation between estradiol receptor affinity and antitumor activity. One of the compounds was also tested on the 9,10-dimethylbenz[a]-anthracene-induced, hormone-dependent mammary carcinoma of the Sprague-Dawley rat, and the results corresponded to those obtained in the xenograft tumor.

Ring-substituted [1,2-bis(4-hydroxyphenyl)ethylenediamine]dichloroplatinum (II) complexes: compounds with a selective effect on the hormone-dependent mammary carcinoma.

[1,2-Bis(4-hydroxyphenyl)ethylenediamine]dichloroplatinum (II) complexes with one substituent in the 2-position (CH3, CF3, F, Cl, Br, I: meso- and d,l-1-PtCl2, meso-(3-5)-PtCl2, meso-(7 and 8)-PtCl2) or two substituents in the 2,6-positions (CH3, Cl: meso-2-PtCl2, meso- and d,l-6-PtCl2) in both benzene rings were synthesized and tested for estrogenic and cytotoxic activities. Two complexes (meso-6-PtCl2 and meso-7-PtCl2) possess both effects. In comparative tests on estrogen receptor positive and negative mammary tumors in cell culture (MCF 7, ER+ and MDA-MB 231, ER-) and in animals (MXT, ER+ and MXT, ER-, mouse), meso-6-PtCl2 shows a selective effect on the estrogen receptor positive mammary carcinoma. A further increase of efficacy was achieved with the water-soluble (sulfato)platinum(II) derivative (meso-6-PtSO4). On the DMBA-induced hormone dependent mammary carcinoma of the SD rat, meso-6-PtSO4 is significantly more active than its ligand (meso-6) and cisplatin.