Breast cancer inhibiting diastereomeric diacetato[1,2-bis(4-fluorophenyl)ethylenediamine]platinum(II) derivatives: synthesis and studies on the relationship between reactivity and antitumor activity

Antitumor active [1,2-bis(4-fluorophenyl)ethylenediamine]platinum(II) diastereoisomers containing acetic acid derivatives as ‘leaving groups’ (acetate: meso/rac-4F-Pt(Ac)₂; monochloroacetate: meso/rac-4F-Pt(ClAc)₂; dichloroacetate: meso/rac-4F-Pt(Cl₂Ac)₂; trichloroacetate: meso/rac-4F-Pt(Cl₃Ac)₂; glycolate: meso/rac-4F-Pt(OHAc)₂; phenylacetate: meso/rac-4F-Pt(PhAc)₂) were synthesized and characterized by IR and ¹H NMR spectroscopy. In all complexes except meso/rac-4F-Pt(PhAc)₂, which exist as [meso/rac-4F-PtPhAc]⁺PhAc⁻, both carboxylic acid residues are coordinated to platinum. Kinetic studies on the reaction behavior of the title compounds with nucleophiles were performed by using iodide as nucleophile. The studies show that the new complexes react with nucleophiles predominantly via the ‘solvent path’ (i.e. via the reactive intermediates = Pt(X)(OH₂)⁺ and =Pt(OH₂)₂²⁺. Therefore the rates of reactions in which the reactive species are formed affect the antitumor activity of the complexes as well as their inactivation by bionucleophiles during the transport to the tumor. The extent of accumulation in the tumor cell, too, influences the antitumor activity of a complex. The rate constants are discussed in view of the activities of the respective complexes on the human MCF-7 breast cancer cell line. From the title compounds the Cl₂Ac and Cl₃Ac derivatives do not come close to the standard cisplatin, neither in chemical reactivity nor in biological activity. Meso/rac-4F-Pt(Ac)₂ and meso/rac-4F-Pt(ClAc)₂, respectively, show similar hydrolysis rates but lower antitumor activities than cisplatin, presumably due to a reduced drug uptake by the tumor cell. Meso/rac-4F-Pt(PhAc)₂ compare well with their standard carboplatin in respect to both properties. Other than the remaining, poorly water soluble title compounds, meso/rac-4F-Pt(OHAc)₂ equal their standard cisplatin in terms of water solubility and antitumor activity rac-4F-Pt(OHAc)₂ > meso-4F--Pt(OHAc)₂). However, they are markedly faster hydrolyzed than cisplatin. By use of rac-4F-Pt(Ac)₂ as an example it was confirmed that, in contrast to the parent compound rac-4F-PtCl₂, the new complex type is also active under in vivo conditions owing to its markedly lower reactivity (mainly due to the lack of a direct substitution by strong nucleophiles), which entails a reduced inactivation of the drug on its way to the tumor. The in vitro testing on tumor cell lines combined with the evaluation of the water solubility and with kinetic studies on the reaction with nucleophiles is a useful method for the preselection of potent platinum complexes deserving further thorough in vitro and in vivo investigations.     

Synthesis, crystal structure and magnetic properties of the first structurally characterized 1,2-dithiocroconato-containing Cu(II) complex, [Cu(bpca)(H2O)]2[Cu(1,2-dtcr)2]·2H2O

The first crystal and molecular structure of a transition metal complex containing 1,2-dithiocroconate (1,2-dtcr, dianion of 1,2-dimercaptocylopent-1-ene-3,4,5-trione), [Cu(bpca)(H₂O)]₂[Cu(1,2-dtcr)₂]·2H₂O (where bpca is the bis(2-pyrdidylcarbonyl)amide anion), has been determined by single crystal X-ray diffraction methods. The compound crystallizesin the monoclinic syste, space group P2₁/c, with a = 11.661(3), b = 20.255(6), c = 8.265(3) Å, ß = 107.26(2)° and Z = 2. The structure is formally built of [Cu(1,2-dtcr)₂]²⁻ and [Cu(bpca)(H₂O)]⁺ ions and water of hydration. The copper atom of the anion is situated at a crystallographic inversion centre, bonded to four sulfur atoms in a planar, approximately square arrangement. In the cation the copper equatorial plane is formed by the three nitrogen atoms of the bpca ligand and a water oxygen atom. In addition there is a very weak axial bond to one of the sulfur atoms of a 1,2-dtcr ligand in the anion. Through these latter weak bonds each anion is connected to, and sandwiched between, two cations, resulting in neutral, trinuclear, centrosymmetric formula units. The triple-decker molecules are arranged in stacks along the crystallographic a-axis creating close contacts between the terminal copper atoms and bpca groups of the neighbouring molecules. This intermolecular interaction is, however, too weak to define the structure as a chain compound. The distance between adjacent copper atoms within the trinuclear unit is 4.189(1) Å, while the shortest intra-stack metal-metal separation between terminal copper atoms is 5.281(1) Å. Variable-temperature magnetic susceptibility measurements in the temperature r.2–140 K reveal that a Curie law is followed; with three non-interacting copper(II) ions in the formula unit.     

Polynuclear copper(II) complexes with μ2-1,1-azide bridges. Structural and magnetic properties

Two novel, weakly antiferromagnetically coupled, tetranuclear copper(II) complexes [Cu₄(PAP)₂(μ₂-1,1-N₃)₂(μ₂-1,3-N₃)₂(μ₂-CH₃OH)₂(N₃)₄ (1) (PAP = 1,4-bis-(2′-pyridylamino)phthalazine) and [Cu₄(PAP3Me)₂ (μ₂-1,1-N₃)₂(μ₂-1,3-N₃)₂(H₂O)₂(NO₂)₂]- (NO₃)₂ (2) (PAP3Me = 1,4-bis-(3′-methyl-2′-pyridyl)aminophthalazine) contain a unique structural with two μ₂-1,1-azide intramolecular bridges, and two μ₂-1,3-azide intermolecular bridges linking pairs of copper(II) centers. Four terminal azide groups complete the five-coordinate structures in 1, while two terminal waters and two nitrates complete the coordination spheres in 2. The dinuclear complexes [Cu₂(PPD)(μ₂-1,1-N₃)(N₃)₂(CF₃SO₃)]CH₃OH) (3) and [Cu₂(PPD)(μ₂-1,1-N₃)(N₃)₂(H₂O)(ClO₄)] (4) (PPD = 3,6-bis-(1′-pyrazolyl)pyridazine) contain pairs of copper centers with intramolecular μ₂-1,1-azid and pyridazine bridges, and exhibit strong antiferromagnetic coupling. A one-dimensional chain structure in 3 occurs through intermolecular μ₂-1,1-azide bridging interactions. Intramolecular Cu-N₃-Cu bridge angles in 1 and 2 are small (107.9 and 109.4°, respectively), but very large in 3 and 4 (122.5 and 123.2°, respectively), in keeping with the magnetic properties. 2 crystallizes in the monoclinic system, space group C2/c with a = 26.71(1), b = 13.51(3), c = 16.84(1) Å, β = 117.35(3)° and R = 0.070, R_w = 0.050. 3 crystallizes in the monoclinic system, space group P2₁/c with a = 8.42(1), b = 20.808(9), c = 12.615(4) Å, β = 102.95(5)° and R = 0.045, R_w = 0.039. 4crystallizes in the triclinic system, space group P1, with a = 10.253(3), b = 12.338(5), c = 8.072(4) Å, = 100.65(4), β = 101.93(3), γ = 87.82(3)° and R = 0.038, R_w = 0.036 . The magnetic properties of 1 and 2 indicate the presence of weak net antiferromagnetic exchange, as indicated by the presence of a low temperature maximum in χ_m (80 K (1), 65 K (2)), but the data do not fit the Bleaney-Bowers equation unless the exchange integral is treated as a temperature dependent term. A similar situation has been observed for other related compounds, and various approaches to the problem will be discussed. Magnetically 3 and 4 are well described by the Bleaney-Bowers equation, exhibiting very strong antiferromagnetic exchange (− 2J = 768(24) cm⁻¹ (3); − 2J = 829(11) cm⁻¹ (4)).     

Multinucleating ligands from multiple palladium(0)-catalysed cross-coupling reactions — synthesis and characterisation of a trinuclear cyclometallated ruthenium(II) complex and a hexanuclear EPR-active molybdenum complex

Two multinucleating ligands have been prepared from 1,3,5-tris(3,5-dibromophenyl)benzene by multiple Pd(0)-catalysed cross-coupling reactions. 1,3,5-Tris[3,5-bis(4-pyridylethenyl)phenyl]benzene (L¹) has six remote pyridyl moieties, each of which can coordinate a 17 valence-electron Mo(tp^*)(NO)Cl fragment (tp^* = hydrotris(3,5-dimethylpyrazolyl)borate), affording the hexanuclear complex [Cl(NO)(tp^*)Mo₆(L¹) (1). 1,3,5-Tris[3,5-bis(2-pyridyl)phenyl]benzene (L²) incorporates three potentially terdentate, cyclometallating N,C,N-donor sets, and can coordinate three Ru(tpy)²⁺ fragments (tpy = 2,2′:6′,2″-terpyridine) giving the trinuclear complex [(tpy)Ru₃(L²)][PF₆]₃ (2). Complex 1 is EPR active, with nearest-neighbour pairs of molybdenum centres displaying magnetic exchange interactions. Electrochemical studies of the two complexes suggest that there is little ground-state interaction between the metal centres in either case.     

Reaction of sulfur diimides with organoboranes — studied by multinuclear magnetic resonance in solution

Reactions between sulfur diimides R(NSN)R′ (R=R′=^tBu (1a), SiMe₃ (1b), SnMe₃ (1c); R=^tBu, R′=SnMe₃ (1d); R=SiMez₃, R′=SnMe₃ (1e)) and various organoboranes were studied, and the products were characterized by multinuclear magnetic resonance data (¹H, ¹¹B, ¹³C, ¹⁵N, ²⁹Si and ¹¹⁹Sn NMR). Tetraalkyldiboranes(6) (Et₂BH₂BEt₂ (2), dimeric 9-borobicyclo[3,3.1]nonane (3)) react with 1a and 1b by 1,3-hydroboration to give the N-sulfanyl-dialkylaminoboranes 4 and 5 which are instable with respect to eliminatio of short-lived [R---NS]. Trialkylboranes (Et₃B (8)) react only sluggishly with 1a, but more readily with 1b mainly via S-ethylation, formally a 1,2-ethyloboration, to give the diethylborylamido-imino-ethanesulfinic acid 9b decomposes slowly at room temperature via ethene elimination to give 4b, followed by further decomposition via [R---NS] elimination. The compounds 9 can be prepared independently from the reaction between the N-lithio-imino-ethanesulfinic acid amide 10 and diorganoboron halides. The molecular structure of the lithium amide 10a (R=R′=tBu) was determined by X-ray analysis as a dimer in which the four nitrogen, two sulfur and two lithium atoms adopt a boat conformation, in contrast with other known derivatives of this type. If the sulfur diimides bear at least one trimethylstannyl group (1c-e), their reactions with Et₃B (8), iPr₃B (12) or 9-iso-butyl-9-borabicyclo[3,3,1]nonane (13) lead to the novel aminoboranes 14–16. These are products of a 1,1,-organoboration, since the Me₃Sn group moves from one nitrogen atom to the other, and both the boryl and an alkyl group end up at the same nitrogen atom.     

Reactivity, variable temperature solution H NMR studies and MO calculations of dimolybdenum allenylidene complexes of the type [(η-C5H5)2Mo2(CO)4(μ,η(4e)-C=C=CR1R2)]

The reactivity, towards nucleophiles and electrophiles, of dimolybdenum allenylidene complexes of the type [Cp₂Mo₂(CO)₄(μ,η²(4e)-C=C=CR1R2)] (Cp=η⁵-C₅H₅) has been investigated. The nucleophilic attacks occur at the C_γ carbon atom, while electrophiles affec the C atom. Variable temperature solution ¹H NMR studies show a dynamic behavior of these complexes consisting of an equilibrium between two enantiomers with a symmetrical [Cp₂Mo₂(CO)₄(μ-σ,σ(2e)-C=C=CR1R2)] transition state. Extended Hückel MO calculations have been carried out on the model [Cp₂Mo₂(CO)₄(μ,η²-C=C=CH₂]. The calculated charges of the allenylidene carbon atoms suggest that the electrophilic attacks are under charge control, while the nucleophilic attacks are rather under orbital control.     

Substitution reaction mechanisms of dihydrido-ruthenium(II) phosphine complexes: hydribe basicity and molecular hydrogen intermediates

Kinetic and activation parameter data for the reactions of cct-Ru(H)₂(CO)₂(PPh₃)₂ (1) (cct = cis, cis, trans) in THF with thiols, CO and PPh₃ to give cct-RuH(SR)(CO)₂(PPh₃)₂, Ru(CO)₃(PPh₃)₂ and Ru(CO)₂(PPh₃)₂, respectively, reveal a common, rate-determining step, the initial dissociation of H₂ from 1; the activated complex probably resembles the corresponding Ru(η²-H₂) species. Reaction of Ru(H)₂(dppm)₂ (2) (as a cis/trans mixture, DPPM = bis(diphenylphosphino)methane) with thiols initially generated cis- and trans- RuH(SR) (dppm)₂ with a rate that depends on both the type and concentration of thiol. The higher basicity of the hydride ligands in 2 (versus 1), which is demonstrated by deuterium exchange with CD₃OD, gives rise in the thiol reaction to an initial protonation step prior to loss of H₂. A species detected in the thiol reaction is possibly [RuH(η²-H₂ (dppm)₂]², the anticipated intermediate for this reaction and for the hydrogen exchange with alcohol. A longer reaction of 2 with PhCH₂SH gives solely cis-Ru(SCH₂Ph)₂(dppm)₂.     

WIDESPREAD PHAGOCYTOSIS OF CILIATES AND OTHER PROTISTS BY MARINE MIXOTROPHIC AND HETEROTROPHIC THECATE DINOFLAGELLATES1

An electron microscopic examination of large amorphous inclusions located in a variety of photosynthetic thecate dinoflagellates (Alexandrium ostenfeldii (Paulsen) Balech et Tangen, Gonyaulax diegensis Kofoid, Scrippsiella sp., Ceratium longipes (Bailey) Gran, and Prorocentrum micans Ehrenberg) and a nonphotosynthetic thecate species (Amylax sp.) revealed each inclusion to be a food vacuole, the majority of which were ingested ciliate prey. Recognizable features of these ciliates included linear arrays of basal bodies and cilia consistent with oligotrich polykinetid structure, characteristic macronuclei, chloroplasts (evidently kleptoplastids), cup-shaped starch plates, and cylindrical extrusomes. Three species contained (apparent) nonciliate prey: Scrippsiella sp., whose food vacuoles consistently contained unusual and complex extrusome-like cylindrical bodies having a distinctive six-lobed, multilayered structure; P. micans, which contained an unidentified encysted cell; and a single A. ostenfeldii cell, containing a Dinophysis sp. dinoflagellate cell. Several food vacuoles of ciliate origin had a red hue. This, together with the resemblance of A. ostenfeldii cells to planozygotes, suggests that similar structures previously identified as accumulation bodies may in fact be food vacuoles and that feeding may in some cases be associated with sexual processes.     

The formation and development of cellulose-synthesizing linear terminal complexes (TCs) in the plasma membrane of the marine red algaErythrocladia subintegra Rosenv.

Summary The formation and development of linear terminal complexes (TCs), the putative cellulose synthesizing units of the red algaErythrocladia subintegra Rosenv., were investigated by a freeze etching technique using both rotary and unidirectional shadowing. The ribbon-like cellulose fibrils ofE. subintegra are 27.6 ± 0.8 nm wide and only 1–1.5 nm thick. They are synthesized by TCs which are composed of repeating transverse rows formed of four particles, the TC subunits. About 50.4 ± 1.7 subunits constitute a TC. They are apparently more strongly interconnected in transverse than in longitudinal directions. Some TC subunits can be resolved as doublets by Fourier analysis. Large globular particles (globules) seem to function as precursor units in the assembly and maturation of the TCs. They are composed of a central hole (the core) with small subunits forming a peripheral ridge and seem to represent zymogenic precursors. TC assembly is initiated after two or three gobules come into close contact with each other, swell and unfold to a nucleation unit resembling the first 2–3 transverse rows of a TC. Longitudinal elongation of the TC occurs by the unfolding of globules attached to both ends of the TC nucleation unit until the TC is completed. The typical intramembranous particles observed inErythrocladia (unidirectional shadowing) are 9.15 ± 0.13 nm in diameter, whereas those of a TC have an average diameter of 8.77 ± 0.11 nm. During cell wall synthesis membranes of vesicles originating from the Golgi apparatus and which seem to fuse with the plasma membrane contain large globules, 15–22 nm in diameter, as well as tetrads with a particle diameter of about 8 nm. The latter are assumed to be involved in the synthesis of the amorphous extracellular matrix cell wall polysaccharides. The following working model for cellulose fibril assembly inE. subintegra is suggested: (1) the ribbon-like cellulose fibril is synthesized by a single linear TC; (2) the number of glucan chains per microfibril correlates with the number of TC subunits; (3) a single subunit synthesizes 3 glucan chains which appear to stack along the 0.6 nm lattice plane; (4) lateral aggregation of the 3-mer stacks leads to the crystalline microfibril.Dedicated to Prof. Dr. Dr. h.c. Eberhard Schnepf on the occasion of his retirement     

Fate of the porphyrin cofactors during the light-dependent turnover of catalase and of the photosystem II reaction-center protein D1 in mature rye leaves

The apoprotein of the enzyme catalase (EC 1.11.1.6) was shown to exhibit a light-dependent turnover in leaves. Present results indicate that photoinactivation of the enzyme was not accompanied by a synchronous destruction and new synthesis of its heme moiety. In rye (Secale cereale L.) leaves the catalase content was not depleted in light when porphyrin synthesis was inhibited by gabaculine. Photoinactivation of purified bovine liver or rye leaf catalase in vitro was not accompanied by concomitant damage to the heme groups. Both the incorporation of -[³H]aminolevulinic acid ([³H]ALA) into catalase-heme and its apparent turnover increased with irradiance. However, the apparent half-life of the catalase-heme was much longer than that of its apoprotein. It is probable that not only degradation but also an exchange with the free heme pool contributed to the apparent turnover of radioactivity of the catalase-heme. Part of the chlorophyll (Chl) associated with photosystem II (PS II) had a preferential light-induced turnover, and repair of PS II appeared to require new Chl synthesis also in mature green rye leaves. The activity of PS II, indicated by the ratio of variable to maximal fluorescence (F_v/F_m), rapidly declined in the presence of gabaculine in light and the reaction-center proteins D1 and D2 were depleted. When segments of mature green rye leaves were labeled with [³H]ALA and incorporation into Chl-protein complexes analysed after electrophoretic separation in the presence of Deriphat, the highest radioactivity was observed in the core complex of PS II, while PS I and the light-harvesting complex of PS II (LHC II) were unlabeled. In greening etiolated leaves highest incorporation was observed in LHC II. Both the incorporation of [³H]ALA into the PS II core complex of green rye leaves and its turnover increased with irradiance. However, the apparent half-life of the PS II-bound labeled porphyrin compounds (mainly Chl) was considerably longer than that of the reaction-center protein D1 under identical conditions.Abbreviations ALA -aminolevulinic acid - CII Core complex of PS II - Chl chlorophyll - DMSO dimethyl sulfoxide - F_v/F_m ratio of variable to maximal chlorophyll fluorescence - LHC light-harvesting complex - PAR photosynthetically active radiation We thank the Deutsche Forschungsgemeinschaft for financial support. Technical assistence by B. Kramer and Ch. van Oijen is greatly appreciated. We are grateful to Dr. Johanningmeier and Dr. Godde (Lehrstuhl für Biochemie der Pflanzen, Universität Bochum, Germany) for providing antisera against the D1 and D2 proteins and Dr. M. Schmidt (Botanisches Institut, Universität Frankfurt am Main, Germany) for valuable advice. Deriphat 160 was kindly supplied by Henkel Corp., Hoboken, N.J., USA.