Ligand Release Research Articles

SÍNTESE, FOTORREATIVIDADE E INTERAÇÃO COM DNA DE UM NOVO COMPLEXO DE RUTÊNIO PROTÓTIPO PARA O DESENVOLVIMENTO DE AGENTES FOTOQUIMIOTERAPÊUTICOS

SYNTHESIS, PHOTOREACTIVITY AND INTERACTION WITH DNA OF A NEW PROTOTYPE RUTHENIUM COMPLEX FOR THE DEVELOPMENT OF PHOTOCHEMOTHERAPEUTIC AGENTS. A new ruthenium polypyridine complex, cis-[Ru(bpy)2(1- meimN)(4-pyCHO)](PF6)2 (WP1), where 1-meimN and pyCHO are the ligands 1-methylimidazole and 4-pyridinecarboxaldehyde, respectively, was prepared as a prototype for use in phototherapeutic process. This compound was characterized by electronic absorption in UV-Visible, FTIR, 1H and 13C NMR, cyclic voltammetry, besides DFT that supported its molecular structure. The compound WP1 was photoreactive, where blue or green light promoted the release of 4-pyCHO ligand. Additionally, this compound was able to interact with DNA, mainly through electrostatic interactions showing photoinduced nuclease activity. There is also photoproduction of superoxide ions using blue or green light suggesting a route for DNA damage. These promising results opened new opportunities for the potential use of this system and analogues as phototherapeutic agents, which is an ongoing study in our lab.

Pivalate complexes of copper(ii) with aliphatic amines as potential precursors for depositing nanomaterials from the gas phase

New copper(ii) complexes [Cu2(RNH2)2(μ-O2CtBu)4]n can be promising for the vapour deposition methods (CVD and FEBID). The pivalate-amine ligand system allowed multinuclear single crystals to grow through the partial release of the secondary ligand.

Antibacterial efficacy of copper-based metal-organic frameworks against Escherichia coli and Lactobacillus.

The widespread and excessive use of antimicrobial drugs has resulted in a concerning rise in bacterial resistance, leading to a risk of untreatable infections. The aim of this study was to formulate a robust and efficient antibacterial treatment to address this challenge. Previous work focused on the effectiveness of the Cu-BTC metal-organic framework (MOF; BTC stands for 1,3,5-benzenetricarboxylate) in combatting various bacterial strains. Herein, we compare the antibacterial properties of Cu-BTC with our newly designed Cu-GA MOF, consisting of copper ions bridged by deprotonated gallate ligands (H2gal2-), against Escherichia coli (E. coli) and Lactobacillus bacteria. Cu-GA was synthesized hydrothermally from copper salt and naturally derived gallic acid (H4gal) and characterized for antibacterial evaluation. The gradual breakdown of Cu(H2gal) resulted in a significant antibacterial effect that is due to the release of copper ions and gallate ligands from the framework. Both copper MOFs were nontoxic to bacteria at low concentrations and growth was completely inhibited at high concentrations when treated with Cu-BTC (1500 μg for E. coli and 1700 μg for Lactobacillus) and Cu-GA (2000 μg for both bacterial strains). Furthermore, our agarose gel electrophoresis results indicate that both MOFs could disrupt bacterial cell membranes, hindering the synthesis of DNA. These findings confirm the antibacterial properties of Cu-BTC and the successful internalization of Cu2+ ions and gallic acid by bacteria from the Cu-GA MOF framework, suggesting the potential for a sustained and effective therapeutic approach against pathogenic microorganisms.

Adrenodoxin allosterically alters human cytochrome P450 11B enzymes to accelerate substrate binding and decelerate release.

Two human mitochondrial membrane CYP11B enzymes play a pivotal role in steroidogenesis. CYP11B1 generates the major glucocorticoid cortisol, while CYP11B2 catalysis yields the primary mineralocorticoid aldosterone. Catalysis by both requires electron delivery by a soluble iron-sulfur adrenodoxin redox partner. However recent studies have shown that adrenodoxin/CYP11B interaction alone allosterically increases substrate and inhibitor affinity as exhibited by decreased dissociation constant (K d) values. The current study moves beyond such equilibrium studies, by defining adrenodoxin effects on the rates of P450 ligand binding and release separately. Stopped-flow data clearly demonstrate that adrenodoxin interaction with the P450 proximal surfaces increases ligand binding in both P450 CYP11B active sites by increasing the on rate constant and decreasing the off rate constant. As substrate entry and exit from the sequestered P450 active site requires conformational changes on the distal side of the P450 enzyme, a likely explanation is that adrenodoxin binding allosterically modulates CYP11B conformational changes. The 93% identical CYP11B enzymes can bind and hydroxylate each other's native substrates differing only by a hydroxyl. However, CYP11B1 exhibits monophasic substrate binding and CYP11B2 biphasic substrate binding, even when the substrates are swapped. This indicates that small differences in amino acid sequence between human CYP11B1 and CYP11B2 enzymes are more functionally important in ligand binding and could suggest avenues for more selective inhibition of these drug targets. Both protein/protein interactions and protein/substrate interactions are most likely to act by modulating CYP11B conformational dynamics.

Channel Modeling for Receptors Reversible Reactive Receiver With Continuous Release of Ligands

Channel Modeling for Receptors Reversible Reactive Receiver With Continuous Release of Ligands

"Catch and release" of the CpN3 ligand using cobalt: dissociation, protonation, and C-H bond thermochemistry.

The coordination chemistry of an amine-rich CpN3 ligand has been explored with cobalt. We demonstrate that in the presence of NaCo(CO)4, the cationic precursor [CpN3]+ yields the complex CpN3CoI(CO)2. While 2e- oxidation generates new CoIII complexes such as [CpN3Co(NCMe)3]2+ and CpN3CoI2(CO), subsequent ligand loss is facile, generating free [CpN3]+ or the protonated dication [CpN3H]2+. We have structurally characterized both these ligand release products via single crystal X-ray diffraction and obtained thermochemical C-H bond strengths via experiment and density functional theory (DFT). Upon reversible 1e- reduction, the radical cation [CpN3H]˙+ has a weak C-H BDFE of 52 kcal mol-1 in acetonitrile. Mechanistic analysis shows that [CpN3H]˙+ undergoes radical-radical disproportionation in the absence of exogenous H-atom acceptors, which is supported by deuterium isotope labelling experiments. Structural comparison of these organic molecules shows a high degree of iminium-like electron delocalization over the C-N bonds connected to the central five-membered ring.

Site-selective template-directed synthesis of antibody Fc conjugates with concomitant ligand release.

Template-directed methods are emerging as some of the most effective means to conjugate payloads at selective sites of monoclonal antibodies (mAbs). We have previously reported a method based on an engineered Fc-III reactive peptide to conjugate a radionuclide chelator to K317 of antibodies with the concomitant release of the Fc-III peptide ligand. Here, our method was redesigned to target two lysines proximal to the Fc-III binding site, K248 and K439. Using energy minimization predictions and a semi-combinatorial synthesis approach, we sampled multiple Fc-III amino acid substituents of A3, H5, L6 and E8, which were then converted into Fc-III reactive conjugates. Middle-down MS/MS subunit analysis of the resulting trastuzumab conjugates revealed that K248 and K439 can be selectively targeted using the Fc-III reactive variants L6Dap, L6Orn, L6Y and A3K or A3hK, respectively. Across all variants tested, L6Orn-carbonate appeared to be the best candidate, yielding a degree and yield of conjugation of almost 2 and 100% for a broad array of payloads including radionuclide chelators, fluorescent dyes, click-chemistry reagents, pre-targeted imaging reagents, and some cytotoxic small molecules. Furthermore, L6Orn carbonate appeared to yield similar conjugation results across multiple IgG subtypes. In vivo proof of concept was achieved by conjugation of NODAGA to the PD1/PD-L1 immune checkpoint inhibitor antibody atezolizumab, followed by PET imaging of PD-L1 expression in mice bearing PD-L1 expressing tumor xenograft using radiolabeled [64Cu]Cu-atezolizumab.

Elucidating Dynamics of Adenylate Kinase from Enzyme Opening to Ligand Release.

This study explores ligand-driven conformational changes in adenylate kinase (AK), which is known for its open-to-close conformational transitions upon ligand binding and release. By utilizing string free energy simulations, we determine the free energy profiles for both enzyme opening and ligand release and compare them with profiles from the apoenzyme. Results reveal a three-step ligand release process, which initiates with the opening of the adenosine triphosphate-binding subdomain (ATP lid), followed by ligand release and concomitant opening of the adenosine monophosphate-binding subdomain (AMP lid). The ligands then transition to nonspecific positions before complete dissociation. In these processes, the first step is energetically driven by ATP lid opening, whereas the second step is driven by ATP release. In contrast, the AMP lid opening and its ligand release make minor contributions to the total free energy for enzyme opening. Regarding the ligand binding mechanism, our results suggest that AMP lid closure occurs via an induced-fit mechanism triggered by AMP binding, whereas ATP lid closure follows conformational selection. This difference in the closure mechanisms provides an explanation with implications for the debate on ligand-driven conformational changes of AK. Additionally, we determine an X-ray structure of an AK variant that exhibits significant rearrangements in the stacking of catalytic arginines, explaining its reduced catalytic activity. In the context of apoenzyme opening, the sequence of events is different. Here, the AMP lid opens first while the ATP lid remains closed, and the free energy associated with ATP lid opening varies with orientation, aligning with the reported AK opening and closing rate heterogeneity. Finally, this study, in conjunction with our previous research, provides a comprehensive view of the intricate interplay between various structural elements, ligands, and catalytic residues that collectively contribute to the robust catalytic power of the enzyme.

A first estimation of the role of penguin guano on copper cycling and organic speciation in Antarctic coastal waters

Cu is a vital micronutrient, but free Cu ions (Cu2+) in seawater, even at subnanomolar concentrations, can impede phytoplankton growth. Natural Cu complexation with organic ligands regulates Cu acquisition and, in most instances, reduces Cu2+ concentrations below toxic thresholds. Along the Antarctic coast, the sources and sinks of Cu and its associated ligands remain poorly defined. Despite the high productivity in the area, there are no studies on the role of trophic transfer in Cu cycling.This study explores penguin guano release of Cu and Cu ligands and its potential in neutralizing copper toxicity along the Antarctic coast. We collected guano in a Chinstrap penguin nesting location in the West coast of Deception Island and extracted its components into aqueous solution imitating natural processes. Copper concentration in guano was 0.4 mg (dry weight g)−1 constituting a potential toxic threat and showed biomagnification with respect to krill. Surface seawater samples collected from various locations varying in penguin activity, were analyzed to assess the potential influence of guano on the area. Visual examination and elevated levels of Al suggested that a substantial portion of guano was lithogenic. Consequently, only a modest 16 % of the total Cu present in guano could be extracted using mechanical methods. Notably, the concentrations of the extracted organic ligands were approximately 23 times higher than the concentrations of the extracted Cu. This significant presence of ligands effectively nullifies any potential toxicity that could have arisen from free Cu2+ ions. Guano ligands' conditional stability constants were lower than those in surface seawater suggesting phytoplankton exudation was the main ligand source in the area. Overall, guano acts as a key node for Cu cycling in coastal Antarctic waters but its deleterious potential is neutralized by ligands from krill digestion and the high background concentration of phytoplankton exudates.

Einbau von (Bioaktiven) Äquatorialen Liganden in Platin(IV)-Komplexe.

AbstractPlatin(IV)‐Prodrugs sind aufgrund ihrer erhöhten Tumorselektivität und geringeren Nebenwirkungen äußerst interessante Alternativen zu Platin(II)‐Antitumortherapeutika. Im Gegensatz zur gängigen Theorie haben wir kürzlich beobachtet, dass äquatoriale Liganden von z. B. Oxaliplatin(IV)‐Komplexen unter Bildung von [(DACH)Pt(OHeq)2(OAcax)2] hydrolysiert werden können. In der hier vorgestellten Arbeit untersuchten wir die Reaktivität und synthetische Verwendbarkeit dieses Komplexes, als Vorstufe für die Entwicklung neuartiger Platin(IV)‐Komplexe, welche mit herkömmlichen Methoden nicht zugänglich sind. Tatsächlich war es möglich die äquatorialen Hydroxidoliganden z. B. durch ein oder zwei monodentate Biotin‐Liganden, die unter Standardmethoden oxidiert werden würden, zu ersetzen. Die gebildeten Komplexe erwiesen sich als sehr stabil und zeigten auch nach der Reduktion eine langsame Ligandenfreisetzung, eine ideale Eigenschaft für lang zirkulierende zielgerichtete Strategien. Daraufhin wurden zwei Platin(IV)‐Komplexe mit äquatorialen Maleimiden, für die Bindung an Serumalbumin als natürlichen Nanocarrier, synthetisiert. Die Komplexe zeigten im Vergleich zu Oxaliplatin eine stark verlängerte Plasmahalbwertszeit und eine deutlich verbesserte Antitumoraktivität in vivo. Zusammenfassend ermöglicht diese neu entwickelte Syntheseplattform den einfachen und gezielten Einbau äquatorialer Liganden in Platin(IV)‐Komplexe. Des Weiteren können verschiedene (bioaktive) Einheiten koordiniert werden, wodurch sogar zielgerichtete dreifach‐wirksame Platin(IV)‐Prodrugs mit nur einem Platinzentrum möglich wären.

Insertion of (Bioactive) Equatorial Ligands into Platinum(IV) Complexes.

Platinum(IV) prodrugs are highly interesting alternatives to platinum(II) anticancer therapeutics due to their increased tumor selectivity and reduced side effects. In contrast to the established theory, we recently observed that the equatorial ligand(s) of e.g. oxaliplatin(IV) complexes can be hydrolyzed with formation of [(DACH)Pt(OHeq )2 (OAcax )2 ]. In the work presented here, we investigated the reactivity and synthetic usability of this complex to be exploited as a precursor for the development of novel platinum(IV) complexes, not able to be synthesized by conventional protocols. Indeed, we could substitute the equatorial hydroxido ligand(s) e.g. by one or two monodentate biotin ligands (which would be oxidized under standard methods). The formed complexes turned out to be very stable with slow ligand release after reduction, ideal for long-circulating tumor-targeting strategies. Therefore, two platinum(IV) complexes with equatorial maleimides, capable of exploiting serum albumin as a natural nanocarrier, were synthesized as well. The complexes showed massively prolonged plasma half-life and distinctly improved anticancer activity in vivo compared to oxaliplatin. Taken together, the newly developed synthetic platform allows the simple and specific insertion of equatorial ligands into platinum(IV) complexes. This will enable the attachment of three different (bioactive) moieties generating targeted triple-action platinum(IV) prodrugs within one single platinum complex.

A Platinum(II) Boron-dipyrromethene Complex for Cellular Imaging and Mitochondria-targeted Photodynamic Therapy in Red Light.

Cisplatin-derived platinum(II) complexes [Pt(NH3 )2 (pacac)](NO3 ) (1, DPP-Pt) and [Pt(NH3 )2 (Acac-RB)](NO3 ) (2, Acacplatin-RB), where Hpacac is 1,3-diphenyl-1,3-propanedione and HAcac-RB is a red-light active distyryl-BODIPY-appended acetylacetone ligand, are prepared, characterized and their photodynamic therapy (PDT) activity studied (RB abbreviated for red-light BODIPY). Complex 2 displayed an intense absorption band at λ=652 nm (ϵ=7.3×104 M-1 cm-1 ) and 601 nm (ϵ=3.1×104 M-1 cm-1 ) in 1 : 1 DMSO-DPBS (Dulbecco's Phosphate Buffered Saline). Its emission profile includes a broad maximum at ~673 nm (λex =630 nm). The fluorescence quantum yield (ΦF ) of HAcac-RB and 2 are 0.19 and 0.07, respectively. Dichlorodihydrofluorescein diacetate and 1,3-diphenylisobenzofuran assay of complex 2 indicated photogeneration of singlet oxygen (ΦΔ : 0.36) as reactive oxygen species (ROS). Light irradiation caused only minor extent of ligand release forming chemo-active cisplatin analogue. The complex showed ~70-100 fold enhancement in cytotoxicity on light exposure in A549 lung cancer cells and MDA-MB-231 multidrug resistant breast cancer cells, giving half maximal inhibitory concentration (IC50 ) of 0.9-1.8 μM. Confocal imaging showed its mitochondrial localization and complex 2 exhibited anti-metastasis properties. Immunostaining of β-tubulin and Annexin V-FITC/propidium iodide staining displayed complex 2 induced photo-selective microtubule rupture and cellular apoptosis, respectively.

Computational insights of the molecular recognition between volatile molecules and odorant binding proteins from the red palm weevil Rhynchophorus ferrugineus

The red palm weevil Rhynchophorus ferrugineus (Coleoptera: Curculionidae) is one of the most harmful pests for palm trees, causing serious economic damage worldwide. The present work aims to model and study the 3D structures of highly expressed odorant binding proteins from R. ferrugineus (RferOBPs) and identify possible binding modes and ligand release mechanism by docking and molecular dynamics. Highly confident 3D structures of a total of 11 odorant binding proteins (OBPs) were obtained with AlphaFold2. All 3D RferOBPs modeled structures displayed six characteristic α-helices, except for RfeOBP7 and RfeOBP10, which had an extra terminal α-helix. Among the eleven modeled RferOBPs, RferOBP4 was highly expressed in the antennae and subsequently selected for further analyses. Molecular docking analyses demonstrated that ferruginol, α-pinene, DEET, and picaridin can favorably bind the RferOBP4 cavity with low affinity energies. Molecular dynamic simulations of RferOBP4 bound to ferruginol at different pH values showed that low pH environments dictate a structural change into an apo-state that modifies the number of tunnels where the ligand can coexist, further triggering ligand release by a pH-dependent mechanism. This is the first report concerning the modelling and study of ligand binding modes and release mechanism of R. ferrugineus OBPs. Communicated by Ramaswamy H. Sarma

Cobalt(III)-py2en systems as potential carriers of β-ketoester-based ligands

Two cobalt(III) complexes containing different β-ketoesters, namely [CoIII(L1)(py2en)](ClO4)2·H2O (1) and [CoIII(L2)(py2en)](ClO4)2 (2) (py2en = N,N′-bis(pyridin-2-ylmethyl)ethylenediamine; L1− = methylacetoacetate; L2− = ethyl 4-chloroacetoacetate) have been prepared and investigated as prototypes of bioreductive prodrugs. The presence of β-ketoester and py2en ligands in 1 and 2, as well as the perchlorate counterions, was supported by IR spectroscopy and CHN elemental analysis. The composition molecular structure of both complexes was confirmed by NMR spectroscopy and ESI mass spectrometry. Structural information was also obtained for 2via X-ray diffraction analysis. The redox properties indicate that 1 and 2 are suitable for reduction under biological conditions. Investigation of DNA-interacting suggest that 1 and 2 bind DNA via electrostatic forces. Both complexes may be employed as possible platforms for the delivery of biologically active compounds, since their reaction with ascorbic acid in PBS at pH 6.2 and 7.4 at 37°C results in the release of the β-ketoester ligands upon Co(III)/Co(II) reduction.

The catalytic reaction of cytochrome c oxidase probed by in situ gas titrations and FTIR difference spectroscopy

Cytochrome c oxidase (CcO) is a transmembrane heme‑copper metalloenzyme that catalyzes the reduction of O2 to H2O at the reducing end of the respiratory electron transport chain. To understand this reaction, we followed the conversion of CcO from Rhodobacter sphaeroides between several active-ready and carbon monoxide-inhibited states via attenuated total reflection Fourier-transform infrared (ATR FTIR) difference spectroscopy. Utilizing a novel gas titration setup, we prepared the mixed-valence, CO-inhibited R2CO state as well as the fully-reduced R4 and R4CO states and induced the “active ready” oxidized state OH. These experiments are performed in the dark yielding FTIR difference spectra exclusively triggered by exposure to O2, the natural substrate of CcO. Our data demonstrate that the presence of CO at heme a3 does not impair the catalytic oxidation of CcO when the cycle starts from the fully-reduced states. Interestingly, when starting from the R2CO state, the release of the CO ligand upon purging with inert gas yield a product that is indistinguishable from photolysis-induced states. The observed changes at heme a3 in the catalytic binuclear center (BNC) result from the loss of CO and are unrelated to electronic excitation upon illumination. Based on our experiments, we re-evaluate the assignment of marker bands that appear in time-resolved photolysis and perfusion-induced experiments on CcO.

Synergy by Ristocetin and CXCL12 in Human Platelet Activation: Divergent Regulation by Rho/Rho-Kinase and Rac.

CXCL12, belonging to the CXC chemokine family, is a weak agonist of platelet aggregation. We previously reported that the combination of CXCL12 and collagen at low doses synergistically activates platelets via not CXCR7 but CXCR4, a specific receptor for CXCL12 on the plasma membrane. Recently, we reported that not Rho/Rho kinase, but Rac is involved in the platelet aggregation induced by this combination. Ristocetin is an activator of the von Willebrand factor that interacts with glycoprotein (GP) Ib/IX/V, which generates thromboxane A2 via phospholipase A2 activation, resulting in the release of the soluble CD40 ligand (sCD40L) from human platelets. In the present study, we investigated the effects of a combination of ristocetin and CXCL12 at low doses on human platelet activation and its underlying mechanisms. Simultaneous stimulation with ristocetin and CXCL12 at subthreshold doses synergistically induce platelet aggregation. A monoclonal antibody against not CXCR7 but CXCR4 suppressed platelet aggregation induced by the combination of ristocetin and CXCL12 at low doses. This combination induces a transient increase in the levels of both GTP-binding Rho and Rac, followed by an increase in phosphorylated cofilin. The ristocetin and CXCL12-induced platelet aggregation as well as the sCD40L release were remarkably enhanced by Y27362, an inhibitor of Rho-kinase, but reduced by NSC23766, an inhibitor of the Rac-guanine nucleotide exchange factor interaction. These results strongly suggest that the combination of ristocetin and CXCL12 at low doses synergistically induces human platelet activation via Rac and that this activation is negatively regulated by the simultaneous activation of Rho/Rho-kinase.

Crystal-Liquid-Glass Transition and Near-Unity Photoluminescence Quantum Yield in Low Melting Point Hybrid Metal Halides.

Hybrid metal halides (HMHs) are a class of materials that combine extraordinary photophysical properties and excellent processability. Their chemical variability allows for the solid-liquid transition toward melt-processable HMHs. Herein, we report the design and synthesis of zero-dimensional HMHs [M(DMSO)6][SbCl6], where the isolated octahedra of [M(DMSO)6]3+ and [SbCl6]3- are alternatively aligned in the crystal structure. The luminescent center of [SbCl6]3- enables the photogeneration of self-trapped excitons, resulting in broadband photoluminescence with a large Stokes shift and a nearly 100% quantum yield. Meanwhile, the release of DMSO ligands from [M(DMSO)6]3+ is controlled by the M-O coordination and thus a low melting point of ∼90 °C is achieved for HMHs. Interestingly, the glass phase is obtained by melt quenching, with a sharp change in photoluminescence colors compared to the crystal phase of melt-processable HMHs. The robust crystal-liquid-glass transition opens a new avenue to tailoring structural disorder and optoelectronic performance in organic-inorganic materials.

"Catch-Store-Release" Strategy for the Stabilization of 4,4'-Methylene Diphenyl Diisocyanate (4,4'-MDI).

4,4'-Methylene diphenyl diisocyanate (MDI) is an industrially crucial compound, being one of the most utilized linkers in the polyurethane industry. However, its long-term stability is limited due to dimerization to form insoluble uretdione. Herein we demonstrate an organometallic "catch-store-release" concept aiming at improving long-term chemical stability of MDI. Treatment of MDI with two equivalents of selected N-heterocyclic carbenes (NHC) forms stable MDI-NHC adducts. Treatment of the adducts with CuCl forms metastable di-Cu(I) complexes, which undergo decomposition to reform MDI (up to 85%), along with Cu-NHC complexes. The yield of reformed MDI can be improved (up to 95%) by the release of the NHC ligands in the form of thiourea, which prevents subsequent MDI dimerization/polymerization by the carbenes. Furthermore, the need to separate MDI from the reaction mixture can be eliminated by the direct reaction of MDI-NHC complexes with alcohols (as model for diols), which form dicarbamate (as model for polyurethane) quantitatively.

Enlarged rete pegs with excessive accumulation of melanosomes leading to darker aging spots revealed by histomorphological measurements of internal structures of the epidermis.

Various histological studies of facial pigmented spot sites such as solar lentigo have been reported, but few studies have used quantitative indices by histomorphometric analysis of the internal structure of pigmented spot sites using non-invasive methods. In the present study, to quantitatively elucidate morphological changes in the epidermis in male, darker-pigmented spots and female, light-pigmented spots, indices that characterize the internal structure of the epidermis in pigmented spot sites were measured using in vivo confocal laser scanning microscopy (CLSM). The darkness of pigmented spots on the cheeks of 69 women and 43 men was analysed using image analysis software. The L* value was calculated from RGB values obtained from facial images. The internal structures of pigmented spots on the cheeks of 13 subjects were observed by CLSM. Various parameters were measured using CLSM images from the surface of the stratum corneum to the bottom of the dermal papillae, including the thickness of the epidermis, melanosome content, and shape of the dermal papillae. Mean ΔL* values between pigmented spots and non-pigmented areas of male subjects were significantly increased in the 40s and 50s compared with those of female subjects. Conspicuous pigmented spots increased in the 40s in male subjects and the 50s in female subjects. In CLSM observations, significant increases in the thickness of the epidermis and melanosome content were confirmed in pigmented spots compared with surrounding non-pigmented areas. In particular, melanosome content in the male subject group with dark-coloured pigmented spots increased significantly to about eight times that of non-pigmented areas, and more than double that of the male subject group with light-coloured pigmented spots. From the measurements of quantitative parameters, morphological changes in the epidermis were clearly related to the surface colour tone of pigmented spots. Darker pigmented spot sites tended to show longer rete pegs in the epidermis. Accumulation of melanosomes in epidermal basal cells could be considered to increase with the degree of elongation of rete pegs at pigmented spot sites and, thus, induce darker pigmented spots.

Recent Preclinical and Clinical Progress in Liposomal Doxorubicin.

Doxorubicin (DOX) is a potent anti-cancer agent that has garnered great interest in research due to its high efficacy despite dose-limiting toxicities. Several strategies have been exploited to enhance the efficacy and safety profile of DOX. Liposomes are the most established approach. Despite the improvement in safety properties of liposomal encapsulated DOX (in Doxil and Myocet), the efficacy is not superior to conventional DOX. Functionalized (targeted) liposomes present a more effective system to deliver DOX to the tumor. Moreover, encapsulation of DOX in pH-sensitive liposomes (PSLs) or thermo-sensitive liposomes (TSLs) combined with local heating has improved DOX accumulation in the tumor. Lyso-thermosensitive liposomal DOX (LTLD), MM-302, and C225-immunoliposomal(IL)-DOX have reached clinical trials. Further functionalized PEGylated liposomal DOX (PLD), TSLs, and PSLs have been developed and evaluated in preclinical models. Most of these formulations improved the anti-tumor activity compared to the currently available liposomal DOX. However, the fast clearance, the optimization of ligand density, stability, and release rate need more investigations. Therefore, we reviewed the latest approaches applied to deliver DOX more efficiently to the tumor, preserving the benefits obtained from FDA-approved liposomes.