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Delivery of anticancer drugs into tumor cores comprised of malignant cancer cells can result in potent therapeutic effects. However, conventional nanoparticle-based drug delivery systems used for cancer therapy often exhibit inefficient tumor-penetrating properties, thus, suggesting the need to improve the functional design of such systems. Herein, we focus on the interactions between cancer cells and the extracellular matrix (ECM) and demonstrate that liposomes modified with slightly acidic pH-sensitive peptide (SAPSp-lipo) can penetrate in vivo tumor tissue and in vitro spheroids comprised of cancer cells and ECM. We previously reported SAPSp-lipo, tumor microenvironment-sensitive liposomes, are effectively delivered to tumor tissue (Hama et al. J Control Release 2015, 206, 67-74). Compared with conventional liposomes, SAPSp-lipo could be delivered to deeper regions within both spheroids and tumor tissues. Furthermore, tumor penetration was found to be promoted at regions where actin depolymerization was induced by SAPSp-lipo and inhibited by the polymerization of actin. In addition, SAPSp-lipo attenuated the interaction between cancer cells and ECM, contributing to the penetration of SAPSp-lipo. These results suggest that SAPSp-lipo penetrates tumors via the interspace route and is accompanied by actin depolymerization. Taken together, SAPSp-lipo demonstrates potential as a novel tumor-penetrable drug carrier for induction of therapeutic effects against malignant cells that comprise tumor cores.

We have developed a novel drug delivery system (DDS) using an antitumor agent, α-tocopheryl succinate (TS). TS has attracted attention as a unique anti-cancer drug for its ability to induce apoptosis in various cancer cells. Furthermore, TS itself readily forms nanovesicles (TS-NVs) and is a prospective tool for use as an antitumor DDS. However, TS-NVs are unstable for encapsulating drugs and passive targeting delivery to tumor tissue via enhanced permeation and retention effect. Therefore, to improve the stability of vesicles, we developed a novel nanovesicle consisting of TS and egg phosphatidylcholine (TS-EPC-NVs). The stability of vesicles of TS-EPC-NVs was significantly higher than that of TS-NVs. As a result, the in vivo antitumor activity of TS-EPC-NVs was more potent than that of TS-NVs. The enhanced antitumor activity of TS-EPC-NVs was found to be due to its effective intratumoral distribution. Moreover, the in vitro anticancer efficiency of TS-EPC-NVs increased seven-fold. We suggest that the improvement is due to homogenous cellular uptake and enhanced cytosolic delivery of the nanovesicles via alteration of intracellular trafficking. Furthermore, TS-EPC-NVs encapsulating siRNA showed significant knockdown efficiency. In summary, TS-EPC-NVs represent a novel and attractive drug delivery system. The system shows antitumor activity of the encapsulated drug and the carrier itself.

Based on the ratios of (-)-epigallocatechin-3-gallate (EGCG) and caffeine (CAF) levels found in commercial tea drinks, EGCG and CAF were co-administered to human volunteers at various EGCG/CAF ratios, and plasma EGCG was determined by high performance liquid chromatography with chemiluminescence detection. As for the results, in plasma taken after ingestion of a beverage containing 95 mg of EGCG alone, the area under the plasma EGCG concentration-time curve (AUC) was 857 ngxh/ml. A higher AUC (1,370 ngxh/ml) was observed when subjects ingested a beverage containing EGCG (95 mg) and a low amount of CAF (40 mg). In the case of ingestion of a beverage containing EGCG (95 mg) and a high amount of CAF (180 mg), the AUC tended to be somewhat higher (1,165 ngxh/ml), but not significantly so, compared with the beverage with EGCG alone. These findings (modulation of plasma EGCG level by CAF) provide ideas for modulating the bioavailability of tea catechins, which can be applied to tea-related drinks and foods.

Modification with polyethylene glycol (PEG) is currently considered an important strategy for anti-cancer drug delivery, because PEGylated-nanoparticles would be effectively delivered to tumor tissue by enhanced permeation and retention effects. However, PEGylation suppresses the cellular uptake of nanoparticles (NPs) to target cells (known as the PEG dilemma). Here, we propose a novel strategy, namely conferring a pathological environment-sensitive property of nanoparticles for overcoming the PEG dilemma. Specifically, although nanoparticles have an overall negative surface charge to avoid interactions with biogenic substances in blood circulation, inversion of surface charge (to positive) at the pH of the tumor microenvironment may allow the nanoparticles to be taken up by cancer cells. To prove this concept, charge-invertible nanoparticles modified with novel slightly acidic pH-sensitive peptide (SAPSP-NPs) were developed. The negatively-charged SAPSP-NPs were delivered to tumor tissue, and were successfully taken up by cancer cells upon inversion of the surface charge to positive at intratumoral pH. SAPSP-NPs may serve as an alternative carrier to the PEGylated NP for anti-cancer drug delivery.

Further isolation and examination of fusicoccane hydrocarbons biosynthetically related to fusicoccin from Phomopsis amygdali allowed us to identify new fungal diterpene hydrocarbons of fusicoccadiene and araneosene. These were assigned as (+)-fusicocca-3(16),10(14)-diene, and (+)-beta- and (+)-delta-araneosenes. These findings led to the experimental clarification of the structures of the biosynthetic hydrocarbon intermediates presumed earlier.

The extraction behavior of MnII, FeIII, CoII, NiII, CuII, ZnII, GaIII, and CdII was studied with a new chelating extractant, 5-(4-nitrophenylazo)-7-(4-ethyl-1-methyloctyl)-8-quinolinol (HNEQ), in chloroform. HNEQ exhibits a much higher extractability for CoII, CuII, ZnII, and CdII than 8-quinolinol. Especially, CuII was quantitatively extracted with 0.001mol dm-3 of HNEQ at such a low pH as 2.0 within 15min, and was readily separated from the other metal(II, III) ions studied. From equilibrium studies, the extracted CuII-HNEQ chelate was ascertained to be Cu(NEQ)2. The extraction constant (logKex) was determined to be 4.25, which was much larger than those for 7-(4-ethyl-1-methyloctyl)-8-quinolinol (Kelex 100) and 8-quinolinol.

A rare case of metastatic malignant melanoma of unknown primary origin to the right inguinal lymph nodes was reported. Although the melanoma was largely developed dimensioned 20×15×7cm, after extensive chemotherapy combined with radiotherapy and hyperthermia, we were able to excise the melanoma from the inguinal area. However, after 7 months it recurred. In addition 11 patients with malignant melanoma including the present case experienced by us in the Department of Dermatology, Saitama Medical Center, Saitama Medical School, between 1986 and 1991, were summarized and discussed.

ATM (Ataxia Telangiectasia Mutated) protein kinase plays a crucial role in cellular DNA damage responses. The inhibition of ATM leads to an abolishment of one such signal pathways termed as “checkpoints”. It is expected that the discovery of checkpoint inhibitor will be an effective assistance of anti-cancer therapy. Recent study reported that Wip1, a magnesium dependent protein phosphatase, de-phosphorylates molecules involved DNA damage checkpoint response such ATM (Ser1981). On the other hand, Squalene (SQ) has been thought to assist anti-cancer therapies such as chemo and/or radiation therapy though there is few available data which refer to its detailed mechanisms. We previously showed that SQ enhances the γ-irradiation (IR) sensitivity through the induction of Wip1 in A549 adenocarcinoma cells. It was indicated that IR induced DNA damage checkpoint was abrogated by SQ pre-treatment, and led to the shrinkage of tumor in vivo. Here, we report SQ sensitizes chemotherapy and an advanced radiation therapy, heavy ion (HI) exposure, in mice using allo/xeno-graft model. Mouse Colon-26 and human MKN45 cells (5 × 106 cells/ mouse) were transplanted into Balb/c and SCID mice, respectively. The tumor volumes were estimated by measuring the growing diameters of tumor tissue every day. SQ (200μl, oral administration) was challenged after day-8 and later days. Chemotherapy using DOX (2 mg/kg, i.p.) was conducted in same days with SQ. One-time exposure of HI (Carbon, 290MeV) was performed on the day-8 in NIRS (National Institute of Radiological Sciences, Japan). Marked change of body weight was not observed between the treatments. DOX and HI prevented the tumor growth. SQ itself inhibited tumor growth slightly. The administration of SQ significantly improved the efficacy of DOX and HI on tumor growth. Wip1 protein expression in tumor tissue was detected by western blot analysis, and was elevated by the oral administration of SQ. These results were confirmed in xenograft model using MKN45 cells and SCID mice. Taken together these findings, SQ sensitized chemo/radio therapy through the inhibition of DNA damage checkpoint activity by Wip1. Further in-depth study is required to determine the clinical feasibility of SQ as a sensitizing agent for anti-cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3873.

Persimmon leaves have been commonly used as traditional medicines in Asia. They have variety kinds of health benefits in such prevention of cancer, hypertension, and inflammation. However, a few of scientific evidences are reported to date. Recent study reported that the cell cycle checkpoint system is playing a pivotal role in DNA damage response, and it is expected that the discovery of checkpoint inhibitor will be an effective assistance of anti-cancer therapy. Checkpoint signaling cascades are critically modulated by ATM (Ataxia telangiectasia-mutated) and ATR (ATM-Rad3-related) protein kinases. Generally, ATM primarily responds to ionizing radiation-induced DNA double-strand breaks, whereas ATR is activated by stalled replication resulting from UV and some genotoxic drugs. In the previous study, we demonstrated that a flavonoid-enriched persimmon leaf extract (FEPLE) promoted cytotoxic effect of cancer cells by chemotherapeutic agents inhibiting the checkpoint activity, especially in ATM dependent pathway. Here, we investigated whether FEPLE inhibits checkpoint activity during DNA damage response induced by radiation such as gamma irrradiation (γIR) and heavy ion (HI) treatment. Dried persimmon leaves were prepared by a decoction for 30 min. A soluble extract was subsequently partitioned into ethyl acetate layer which is flavonoid-enriched persimmon leaves extract (FEPLE). FEPLE included eight flavonol components, non-galloylated flavonoids (kaempferol 3-O-glucoside, kaempferol 3-O-galactoside, quercetin 3-O-glucoside, quercetin 3-O-galactoside) and galloylated flavonoids (their 2″-gallated flavonol glucoside). γIR (2.5 to 20 Gy) and HI (2.5 to 20 Gy) were performed in National Institute of Radiological Sciences (NIRS) and Heavy Ion Medical Accelerator in Chiba (HIMAC), respectively. Human lung adenocarcinoma A549 cells were incubated with FEPLE (0, 1, 10, 30 μg/mL) at one hour prior to γIR or HI treatment. Intracellular levels of phosphorylation in each checkpoint molecule such as p53, SMC1 and Chk1 were determined by western blot using phospho-specific antibodies of Ser15, Ser966 and Ser345, respectively. Phosphorylation on Ser10 of histone H3 was measured by flow-cytometry as a marker of cells in mitosis. The phosphorylation of p53, SMC1 and Chk1 were increased by the treatment of γIR and HI. FEPLE reduced the phosphorylation of checkpoint proteins in both γIR and HI. Furthermore, ATM activity, estimated by the phosphorylation of Ser1981, was inhibited by the treatment of FEPLE. The pre-treatment of FEPLE significantly prevented the decrease of mitotic cells in HI exposed cells. These results indicated that DNA damage checkpoint system in γIR or HI exposed cells were abrogated by FEPLE treatment through the inhibition of ATM-dependent signaling pathway. FEPLE might be useful for clinical application by combining γIR or HI cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3875.

The synthesis of dl-steporphine (1), an aporphine-type alkaloid having a hydroxyl group at the 4-position, is described. The acetophenone derivative (4), prepared from 2 and 3 by Friedel-Crafts reaction, was condensed with aminoacetaldehyde diethyl acetal to afford the ketimine derivative (5). The product (6), from the sodium borohydride reduction of 5 or its N-methyl derivative (7) was subjected to hydrolytic cyclization in concentrated hydrochloric acid to afford stereospecifically 1-(2-bromobenzyl)-4-hydroxy-6, 7-methylenedioxy-1, 2, 3, 4-tetrahydroisoquinoline (8) or its N-methyl derivative (9), respectively, with a cis relationship between C1-H and C4-H. The stereospecificity is presumed to be due to intramolecular hydrogen bonding between the alcoholic hydroxyl group and the bromo atom. Irradiation of 9 in dilute hydrochloric acid afforded dl-steporphine (1). This is the first stereospecific synthesis of 4-hydroxyaporphine-type alkaloids, and the method should be adaptable to the synthesis of other alkaloids of this type.