CD98 regulates the phosphorylation of HER2 and a bispecific anti-HER2/CD98 antibody inhibits the growth signal of human breast cancer cells.

Human epidermal growth factor receptor (HER) family proteins are currently major targets of therapeutic monoclonal antibodies against various epithelial cancers. However, the resistance of cancer cells to HER family-targeted therapies, which may be caused by cancer heterogeneity and persistent HER phosphorylation, often reduces overall therapeutic effects. We herein showed that a newly discovered molecular complex between CD98 and HER2 affected HER function and cancer cell growth. The immunoprecipitation of the HER2 or HER3 protein from lysates of SKBR3 breast cancer (BrCa) cells revealed the HER2-CD98 or HER3-CD98 complex. The knockdown of CD98 by small interfering RNAs inhibited the phosphorylation of HER2 in SKBR3 cells. A bispecific antibody (BsAb) that recognized the HER2 and CD98 proteins was constructed from a humanized anti-HER2 (SER4) IgG and an anti-CD98 (HBJ127) single chain variable fragment, and this BsAb significantly inhibited the cell growth of SKBR3 cells. Prior to the inhibition of AKT phosphorylation, BsAb inhibited the phosphorylation of HER2, however, significant inhibition of HER2 phosphorylation was not observed in anti-HER2 pertuzumab, trastuzumab, SER4 or anti-CD98 HBJ127 in SKBR3 cells. The dual targeting of HER2 and CD98 has potential as a new therapeutic strategy for BrCa.

Rapid Isolation of Extracellular Vesicles Using a Hydrophilic Porous Silica Gel-Based Size-Exclusion Chromatography Column.

Extracellular vesicles (EVs) are nanoscale lipid bilayer vesicles released by almost all cell types and can be found in biological fluids, such as blood and urine. EVs play an important role in various physiological and pathological processes via cell-cell communication, highlighting their potential applications as diagnostic markers for diseases and therapeutic drug delivery carriers. Although various methods have been developed for the isolation of EVs from biological fluids, most of them exhibit major limitations, including low purity, long processing times, and high cost. In this study, we developed a size-exclusion chromatography (SEC) column device using hydrophilic porous silica gel (PSG). Owing to the resistance to pressure of the device, a rapid system for EV isolation was developed by connecting it to a flash liquid chromatography system furnished with a UV detector and a fraction collector. This system can be used for the real-time monitoring of eluted EVs by UV absorption without further analysis and separation of high-purity EVs from urine samples with high durability, reusability, and reproducibility. In addition, there were no significant differences between the PSG column- and conventional SEC column-isolated EVs in the proteome profiles and cellular uptake activities, suggesting the good quality of the EVs isolated by the PSG column. These findings suggest that the PSG column device offers an effective and rapid method for the isolation of intact EVs from biological fluids.

Metabolomics of small extracellular vesicles derived from isocitrate dehydrogenase 1-mutant HCT116 cells collected by semi-automated size exclusion chromatography.

Cancer-derived small extracellular vesicles (sEVs) are multifunctional particles with a lipid bilayer structure that are involved in cancer progression, such as malignant proliferation, distant metastasis, and cancer immunity evasion. The separation protocol used to isolate sEVs is an important process and thus, several have been developed, including ultracentrifugation (UC), size exclusion chromatography (SEC), and affinity purification using antibodies against sEV surface antigens. However, the effects of different separation methods on sEV components have not been adequately examined. Here, we developed a semi-automated system for collecting sEVs by combining SEC and preparative high-performance liquid chromatography and applied it to metabolome analysis. The developed SEC system could recover sEVs more efficiently and non-destructively than UC, suggesting that it is an appropriate recovery method for metabolic analysis and reflects biological conditions. Furthermore, using the developed SEC system, we performed metabolome analysis of sEVs from isocitrate dehydrogenase 1 (IDH)-mutated human colon HCT116 cells, which produce the oncogenic metabolite, 2-hydroxyglutaric acid (2-HG). IDH1-mutated HCT116 cells released significantly more sEVs than wild-type (WT) cells. The metabolomic profiles of IDH1 mutant and WT cells showed distinct differences between the cells and their sEVs. Notably, in IDH mutant cells, large amounts of 2-HG were detected not only in cells, but also in sEVs. These results indicate that the SEC system we developed has wide potential applications in sEVs research.

The characteristics of multi-directional step distance and the association between stepping laterality and walking ability of patients with stroke.

[Purpose] The purpose of this study was to determine the effect of stepping limb and step direction on step distance and the association of step distance and stepping laterality in step difference with walking ability and motor dysfunction. [Subjects and Methods] The subjects were thirty-nine patients with chronic hemiparesis as a result of stroke, who performed the MSL (Maximum Step Length) test along with tests of motor impairment, gait speed and Functional Ambulation Category. The MSL test is a clinical test of stepping distance in which participants step to the front, side, and back. The subjects were classified into three groups according to the stepping laterality in front step distance. [Results] Step distance did not differ across stepping limbs but did differ across step directions. Front step distance was significantly longer than side and back step distance. Participants with forward paretic step length shorter than forward non-paretic step length had significantly higher walking ability than participants with symmetric forward step length or forward paretic step length longer than forward non-paretic step length [Conclusion] Patients with stroke have characteristic step distances in each direction. Adequate weight shift toward the paretic limb when stepping with the non-paretic limb is associated with walking ability.

Association of performance of standing turns with physical impairments and walking ability in patients with hemiparetic stroke.

[Purpose] The effect of turn direction and relation between turn performance and walking ability in patients with hemiparetic stroke is not clear. The purpose of this study was to determine the effect of turn direction on the performance of standing turns and to examine the relations between turn performance and walking ability in patients with hemiparetic stroke. [Subject and Methods] The participants were 38 outpatients with chronic hemiparesis due to stroke. Turn performance was evaluated using the time and number of steps required to complete a 360° standing turn, and was evaluated for turns toward the paretic side and the non-paretic side. Walking ability was assessed using gait speed in the 10-m walk test, the Timed Up and Go test, and the Functional Ambulation Category. [Results] Thirty-six participants were analyzed, and the time needed for turns and number of steps were similar for turns to the paretic and non-paretic sides. The time needed for turns was correlated walking ability. A turn time of 10.0 s distinguished FAC 5 (independent ambulation in the community) from FAC ≤4 with a sensitivity of 0.94 and specificity of 0.85. [Conclusion] The performance of standing turns was not affected by the turning direction and was closely correlated with walking ability.

Significant differences of brain blood flow in patients with chronic low back pain and acute low back pain detected by brain SPECT.

BACKGROUND: The aim of this study was to examine and compare the areas of brain blood flow in patients with chronic low back pain (CLBP) without structural abnormality and acute low back pain (ALBP) with lumber disc herniation (LDH). Functional neuroimaging studies provide evidence of abnormalities in the regional cerebral blood flow during low back pain. Recent studies have shown that CLBP is associated with plastic, pathophysiological changes in the brain. However, there has been no report yet statistically or by neuro-images on the compared brain single photon-emission computed tomography (SPECT) findings between CLBP and ALBP patients. METHODS: The subjects comprised 14 patients, 7 CLBP and 7 ALBP patients. The CLBP group included the patients who had no or minor structural abnormality in the lumbar spine on magnetic resonance imaging (MRI) and met the criteria for a classification of "pain disorder" (chronic) according to the Diagnostic and Statistical Manual of Mental Disorders, 4th Edition, Text Revision. The ALBP group included the patients who had symptoms within 3 months of onset and LDH revealed by MRI. All patients were assessed using brain SPECT. We then performed a two-tailed view analysis using the easy Z score imaging system, determined the mean Z scores, and performed vBSEE software (Fujifilm RI Pharma, Tokyo, Japan) for both CLBP and ALBP patients. RESULTS: The CLBP group showed significantly reduced blood flow in the bilateral prefrontal cortex of the frontal lobe and increased blood flow in the bilateral posterior lobe of the cerebellum. CONCLUSIONS: SPECT images and statistical analyses revealed the brain blood flow alterations in the patients with ALBP and CLBP. These results may suggest that the dysfunction of the prefrontal cortex could lead to the appearance of unconscious pain behavior controlled by the cerebellum in the patients with CLBP.

Primary malignant mesothelioma of the appendix.

We report a case of primary malignant mesothelioma of the appendix. A 35-year-old man without any history of asbestos exposure was admitted to our hospital for further examination following the discovery of multiple liver tumors, an ileocecal tumor, and abdominal lymph node swelling. An ultrasound-guided liver tumor biopsy revealed malignant mesothelioma. Despite receiving systemic chemotherapy, he died 3 months after the initial diagnosis. At autopsy, a diagnosis of multiple organ metastases from a malignant biphasic mesothelioma of the appendix was made. To our knowledge, this is only the second reported case of primary malignant mesothelioma of the appendix.

Stevioside enhances apoptosis induced by serum deprivation in PC12 cells.

In the laboratory, using a PC12 cell system, studies have been conducted on the effects of various chemicals on apoptosis, as it is considered to be an essential part of normal development, maintenance, and defense in organisms. Stevioside is a natural sweetener extracted from the leaves of Stevia rebaudiana. Since it is widely used as a sugar replacement, it was decided to evaluate the toxicological effects of low concentrations of stevioside on apoptosis induced by serum deprivation using the PC12 cell system. It was found that based on data from DNA electrophoresis and TUNEL signal assays stevioside enhanced apoptosis induced by serum deprivation. This enhancement was caused by increased expression of Bax and of cytochrome c released into the cytosol. These findings suggest that stevioside affects the regulation of the normal apoptotic condition. Further investigation will be needed to clarify the detailed mechanism of the enhancement due to the treatment with stevioside.

Diethyl phthalate enhances apoptosis induced by serum deprivation in PC12 cells.

In this study, to examine the mechanism of diethyl phthalate toxicity to cells, the effects of diethyl phthalate on apoptosis in a PC12 cell system were investigated by assaying apoptotic factors such as caspase-3, Bax, cytochrome c and DNA damage. Diethyl phthalate was shown to enhance the apoptosis induced by serum deprivation according to the results of DNA electrophoresis and TUNEL signal assays, although it could not induce apoptosis itself in the cells. This enhancement was thought to be because of an increase in caspase-3-like activity. In addition, the expression of bax and contents of cytochrome c in the cytosol showed a tendency to increase the cells exposed to diethyl phthalate. These results indicated that diethyl phthalate, a potential endocrine disrupter, affects the apoptotic system in PC12 cells. Diethyl phthalate may enhance oxidative stress such as that induced by reactive oxygen species in PC12 cells.

A novel anti-human HB-EGF monoclonal antibody with multiple antitumor mechanisms against ovarian cancer cells.

PURPOSE: Heparin-binding epidermal growth factor (EGF)-like growth factor (HB-EGF) is a member of the EGF family and plays a pivotal role in tumor progression in ovarian cancer. We developed an anti-HB-EGF monoclonal antibody (mAb) and investigated its antitumor activities in vitro and in vivo to evaluate its potential as a therapeutic antibody against ovarian cancer. EXPERIMENTAL DESIGN: We prepared mAbs from HB-EGF null mice immunized with recombinant human soluble HB-EGF and evaluated their binding and neutralizing activity against HB-EGF. Next, we generated a mouse-human chimeric antibody and examined its in vitro and in vivo antitumor activities. RESULTS: Two murine anti-HB-EGF mAbs were developed, and one of them, KM3566, was revealed to have a high binding reactivity for membrane-anchored HB-EGF (pro-HB-EGF) expressed on the cell surface, as well as neutralizing activity against growth promoting activity of soluble HB-EGF. The mouse-human chimeric counterpart for KM3566 (cKM3566) induced dose-dependent antibody-dependent cellular cytotoxicity (ADCC) against cancer cells expressing HB-EGF in vitro, and significantly inhibited tumor growth in severe combined immunodeficient mice inoculated with MCAS or ES-2 human ovarian cancer cells. CONCLUSIONS: A novel anti-HB-EGF chimeric antibody, cKM3566, with two antitumor mechanisms, neutralization and ADCC, exhibits potent in vivo antitumor activity. These results indicate that cKM3566 is a promising antiovarian cancer therapeutic antibody.

Involvement of mitoKATP channel in protective mechanisms of cerebral ischemic tolerance.

Little work has been performed to determine roles of mitochondrial ATP-sensitive potassium channels (mitoK(ATP)) in ischemic preconditioning (IPC) in brain. To investigate the role on cerebral IPC, we examined effect of 5-hydroxydecanoate (5-HD), a selective mitoK(ATP) blocker, and diazoxide (DZX), a selective mitoK(ATP) opener on various IPC models. An IPC model with gerbil: 2 min bilateral common carotid arteries occlusion (BLCO)+24 h recovery+5 min BLCO. 5-HD, DZX, vehicle was administered 30 min before 5 min BLCO. Seven days later, surviving CA1 neurons were counted. A focal IPC model with rat: 15 min middle cerebral artery occlusion (MCAO)+48 h recovery+90 min MCAO. Twenty-four hours before 90 min MCAO, 5-HD, DZX, or vehicle was administered. One day after 90 min MCAO, neurological symptoms and infarct volumes were evaluated. An in vitro IPC model with primary neuronal cultures: 8 min oxygen-glucose deprivation (OGD)+24 h recovery+70 min OGD. Thirty minutes before 70 min OGD, 5-HD or DZX were added. One day later, surviving neurons were counted. Mitochondrial membrane potential was also monitored. 5-HD significantly attenuated the protective effect of IPC in gerbil model, rat model, and in vitro OGD model. DZX significantly facilitated the protective effect of IPC in gerbil and rat model. The mitochondrial membranes were depolarized with IPC, and 5-HD treatment significantly reduced this effect. These results strongly suggest that mitoK(ATP) channel activation plays a key role in development of a protective mechanism of cerebral IPC.

Combination therapy with transductive anti-death FNK protein and FK506 ameliorates brain damage with focal transient ischemia in rat.

Many practical therapies have been explored as clinical applications for ischemic cerebral infarction; however, most are still insufficient to treat acute stroke. We show here a potential combination therapy in a rat focal ischemic model to improve neurological symptoms as well as to reduce infarct volumes at the maximum level. We applied protein transduction technology using artificial anti-death Bcl-xl derivative with three amino acid-substitutions (Y22F, Q26N and R165K) (FNK) protein fused with a protein-transduction-domain peptide (PTD-FNK). When PTD-FNK was administrated 1 h after initiating ischemia followed by the administration of an immunosuppressant FK506 with a 30-min time lag, infarct volumes of the total brain and cortex were markedly reduced to 27% and 14%, respectively. This procedure not only reduced the infarct volume and edema, but also markedly improved neurological symptoms. The therapeutic effect continued for at least 1 week after ischemia. FK506 inhibited the transduction of PTD-FNK in vitro, which explains the requirement of a time lag for the administration of FK506. An additional in vitro experiment showed that PTD-FNK, when administered 30 min before FK506, gave the maximal protective effect by reducing the intracellular calcium concentration. We propose that this combination therapy would provide a synergistic protective effect by both drugs, reducing adverse the effects of FK506.

Clinical features and prognoses of 23 patients with chronic granulomatous disease followed for 21 years by a single hospital in Japan.

In this paper, we examined the details of severe infections, treatment efficacies, and the prognoses of 23 Japanese patients with chronic granulomatous disease (CGD). We described the mean ages at diagnosis and follow-up, which were 2.8 years (range, 0.7-10 years) and 14.9 years (range, 0.2-28.4 years), respectively. There were three deaths, two from Aspergillus pneumonia and one from liver abscess. Eighteen of the 23 patients (78%) had a complete loss of gp91phox, and three had p22-phox and one had p67phox deficiencies. Aspergillus species were found in 45% of 174 severe infections. The mean height and weight of the 20 surviving patients were -0.8 +/- 1.3SD and -1.9 +/- 1.9SD below the means for age, respectively. Short stature and underweight (below the 10th percentile of the means) for age were seen in 22% and 17% of the patients, respectively. This growth retardation reflects the severity of the disease. At 20 years of age, there was 87% survival. Ongoing prophylaxis with trimethoprim-sulfamethoxazole (TMP-SMX) or antifungal drugs was given in 16 and 11 patients, respectively. Interferon-gamma (IFN-gamma) was given once a week to 14 patients. Four patients underwent hematopoietic stem cell transplantation (HSCT) and are currently well. There were infections observed in three of 21 identified related carriers of X-linked CGD. A carrier with a liver abscess had 5% normal neutrophils during the acute phase of infection, which returned to 40% normal neutrophils after recovery. The high survival rate in this hospital results from regular follow-up and prophylaxis with TMP-SMX and anti-fungal drugs beginning at the time of diagnosis, along with treatment with weekly IFN-gamma.

A neutralizing anti-fibroblast growth factor (FGF) 8 monoclonal antibody shows anti-tumor activity against FGF8b-expressing LNCaP xenografts in androgen-dependent and -independent conditions.

BACKGROUND: Fibroblast growth factor 8-isoform b (FGF8b) has been detected in human clinical sex-organ related cancers including hormone-refractory prostate cancer. There are, however, few relevant experimental models. A murine monoclonal anti-FGF8 antibody, KM1334, has been shown to neutralize FGF8b and inhibit the growth of androgen-dependent mouse mammary SC-3 cells in vitro and in vivo. In the present study, we evaluated the anti-tumor activity of KM1334 against androgen-dependent and -independent progression of FGF8b-expressing human prostate cancer xenografts. METHODS: FGF8b cDNA was transfected into androgen-dependent human prostate cancer cell line LNCaP, and its xenograft tumors were established subcutaneously in SCID mice with or without castration. KM1334 at the dose of 400 microg/head was injected twice weekly. RESULTS: FGF8b-expressing LNCaP cells secreted FGF8b, showed enhanced level of Erk1/2 phosphorylation, and showed more potent growth properties than mock-expressing cells in vitro and in vivo. KM1334 reduced these properties in vitro, inhibited tumorigenecity in vivo (T/C=0.33), and showed anti-tumor activity against established tumors (T/C=0.47) of FGF8b-expressing cells. FGF8b-expressing LNCaP tumors were androgen-dependent. However, they recurred as androgen-independent FGF8b positive tumors after castration. KM1334 also inhibited the growth of established FGF8b-expressing tumors in the androgen-independent states (T/C=0.47). CONCLUSIONS: These results indicate that humanized monoclonal antibodies, conserving the paratope of KM1334, are a promising candidate for therapy of FGF8b-expressing clinical prostate cancers. Follow-up studies using xenograft models with clinical FGF8b-expressing tumors are required to validate these early findings.

Brain perfusion abnormality in patients with chronic pain.

We performed single photon emission computed tomography (SPECT) of the brain in 15 patients with chronic pain (males, 7; females, 8; average age 49.1 +/- 17.9 years) and identified the locus of cerebral blood flow reduction by a new analytical method (easy Z-score Imaging System: eZIS) to clarify the functional neuroanatomical basis of chronic pain. Of the 15 patients, 6 had backache, 2 neck pain, 2 gonalgia, and 5 pain at other sites, with an average Visual analog scale of pain (VAS) value of 6.1 +/- 1.9. In comparison with a information on a data base on physically unimpaired persons, the dorsolateral prefrontal area (both sides, right dominant), medial prefrontal area (both sides), dorsal aspect of the anterior cingulate gyrus nociceptive cortex (both sides) and the lateral part of the orbitofrontal cortex (right side) were found to have blood flow reduction in the group of patients with chronic pain. As for chronic pain and its correlation with clinical features such as a depressive state, anticipation anxiety, PTSD, and conversion hysteria, the mechanism in the brain that was suggested by this study should be followed-up by functional neuroimaging studies.

Hydrogen acts as a therapeutic antioxidant by selectively reducing cytotoxic oxygen radicals.

Acute oxidative stress induced by ischemia-reperfusion or inflammation causes serious damage to tissues, and persistent oxidative stress is accepted as one of the causes of many common diseases including cancer. We show here that hydrogen (H(2)) has potential as an antioxidant in preventive and therapeutic applications. We induced acute oxidative stress in cultured cells by three independent methods. H(2) selectively reduced the hydroxyl radical, the most cytotoxic of reactive oxygen species (ROS), and effectively protected cells; however, H(2) did not react with other ROS, which possess physiological roles. We used an acute rat model in which oxidative stress damage was induced in the brain by focal ischemia and reperfusion. The inhalation of H(2) gas markedly suppressed brain injury by buffering the effects of oxidative stress. Thus H(2) can be used as an effective antioxidant therapy; owing to its ability to rapidly diffuse across membranes, it can reach and react with cytotoxic ROS and thus protect against oxidative damage.

G-CSF receptor-binding cyclic peptides designed with artificial amino-acid linkers.

Designing small molecules that mimic the receptor-binding local surface structure of large proteins such as cytokines or growth factors is fascinating and challenging. In this study, we designed cyclic peptides that reproduce the receptor-binding loop structures of G-CSF. We found it is important to select a suitable linker to join two or more discontinuous sequences and both termini of the peptide corresponding to the receptor-binding loop. Structural simulations based on the crystallographic structure of KW-2228, a stable and potent analog of human G-CSF, led us to choose 4-aminobenzoic acid (Abz) as a part of the linker. A combination of 4-Abz with beta-alanine or glycine, and disulfide bridges between cysteins or homocysteins, gave a structure suitable for receptor binding. In this structure, the side-chains of several amino acids important for the interactions with the receptor are protruding from one side of the peptide ring. This artificial peptide showed G-CSF antagonistic activity in a cell proliferation assay.

A neutralizing anti-fibroblast growth factor 8 monoclonal antibody shows potent antitumor activity against androgen-dependent mouse mammary tumors in vivo.

PURPOSE: Fibroblast growth factor 8b (FGF8b) has been implicated in oncogenesis of sex hormone-related malignancies. A murine monoclonal anti-FGF8 antibody, KM1334, has been raised against a FGF8b-derived peptide and shown to neutralize FGF8b activity in an androgen-dependent mouse mammary cell line (SC-3) in vitro growth. The purpose of this study was to evaluate KM1334 as a therapeutic agent for FGF8-dependent cancer. EXPERIMENTAL DESIGN: Specificity and neutralizing activity of KM1334 were examined in vitro. In vivo therapeutic studies were done in nude mice bearing SC-3 tumors s.c. RESULTS: KM1334 recognized FGF8b and FGF8f specifically out of four human FGF8 isoforms and showed little binding to other members of FGF family. Neutralizing activity of KM1334 was confirmed by both blocking of FGF8b binding to its three receptors (FGFR2IIIc, FGFR3IIIc, and FGFR4) and FGF8b-induced phosphorylation of FGFR substrate 2alpha and extracellular signal-regulated kinase 1/2 in SC-3 cells. The in vitro inhibitory effect could be extended to in vivo tumor models, where KM1334 caused rapid regression of established SC-3 tumors in nude mice. This rapid regression of tumors after KM1334 treatment was explained by two independent mechanisms: (a) decreased DNA synthesis, as evidenced by a decrease in uptake of 5-bromo-2'-deoxyuridine, and (b) induction of apoptosis as shown by the terminal deoxynucleotidyl transferase-mediated nick end labeling assay. CONCLUSIONS: KM1334 possesses strong blocking activity in vitro and antitumor activity in vivo and therefore may be an effective therapeutic candidate for the treatment of cancers that are dependent on FGF8b signaling for growth and survival.

Effects of FK506 on the translocation of protein kinase C and CaM kinase II in the gerbil hippocampal CA1 neurons.

To explore biochemical basis for cerebroprotective effect of immunosuppressant FK506, we studied changes in subcellular distribution of protein kinase C gamma (PKC gamma) as well as calcium/calmodulin-dependent protein kinase II (CaMKII) after ischemia. Male Mongolian gerbils were subjected to 5 min forebrain ischemia. FK506 (1 or 3 mg kg-1) was administered at 1 min after recirculation, which was confirmed to be cerebroprotective by histological examination at seven days after ischemia. At the designated time points (before ischemia, 5 min ischemia, 1 and 24 h recovery), heads were frozen and samples were taken from CA1 subfield of hippocampus. Western blot analysis was carried out. Persistent translocations of PKC gamma and CaMKII to synaptosomal P2 fraction were observed in vehicle-treated group. FK506 significantly decreased levels of PKC gamma and CaMKII in P2 fraction at 24 h of recovery. The present results suggest FK506 downregulates translocated PKC gamma and CaMKII, which may contribute to its survival promoting effect after cerebral ischemia.

AuCl(3)-catalyzed benzannulation: synthesis of naphthyl ketone derivatives from o-alkynylbenzaldehydes with alkynes.

The reaction of o-alkynylbenzaldehydes 1 and alkynes 2 in the presence of a catalytic amount of AuCl3 in (CH2Cl)2 at 80 degrees C gave naphthyl ketone products in high yields. The AuCl3-catalyzed formal [4 + 2] benzannulation proceeds most probably through the coordination of the triple bond of 1 to AuCl3, the formation of benzo[c]pyrylium auric ate complex via the nucleophilic addition of the carbonyl oxygen atom, the Diels-Alder addition of alkynes 2 to the auric ate complex, and subsequent bond rearrangement. Similarly, the AuCl3-catalyzed reactions of o-alkynylacetophenone and o-alkynylbenzophenone with phenylacetylene afforded the corresponding naphthyl ketone products in good yields.