To facilitate the identification of candidate molecular biomarkers that are linked to the pathogenesis of hepatocellular carcinoma (HCC), we investigated protein-expression profiles of 146 tissue specimens including 67 pairs of tumors and adjacent non-tumors resected from HCC patients as well as 12 normal livers by 2-DE. Among the 1800 spots displayed in the liver proteome, a total of 90 protein species were found to be significantly different between the three groups (P < 0.05). Three of the top candidate markers up-regulated in HCC, with high receiver operating characteristic (ROC) curves, were identified by MS/MS analysis and belonged to the chaperone members: heat-shock protein (Hsp)27, Hsp70 and glucose-regulated protein (GRP)78. Over-expression of these chaperone proteins in HCC tissues was confirmed by Western blotting and immunohistochemistry. In correlation with clinico-pathological parameters, expression of Hsp27 was linked to alpha-fetoprotein level (P = 0.007) whereas up-regulation of GRP78 was associated with tumor venous infiltration (P = 0.035). No significant association of Hsp70 with any pathologic features was observed. The present HCC proteome analysis revealed that in response to the stressful cancerous microenvironment, tumor cells strived to increase the expression of chaperone proteins for cyto-protective function and to enhance tumor growth and metastasis.

A stable gold(III)-phosphine complex [(C^N^C) 2 Au 2 (m-dppp)](CF 3 SO 3 ) 2 [Au3, HC^N^CH ¼ 2,6diphenylpyridine; dppp ¼ bis(diphenylphosphino)propane] displays potent in vitro cytotoxicity towards various cancers with sub-micromolar range cytotoxic IC 50 values, and is significantly more potent than its structural and iso-electronic platinum(II) analog [(C^N^N) 2 Pt 2 (m-dppp)](CF 3 SO 3 ) 2 (HC^N^N ¼ 6-phenyl-2,2 0 -bipyridine) and gold(III)-carbene complexes. Complex Au3 displays promising inhibition on tumor growth in animal models, and its acute and sub-chronic toxicities have been examined in mice and beagle dogs. Transcriptomic and connectivity map analyses have revealed that the transcriptional profile of Au3 is similar to those of inhibitors of thioredoxin reductase (TrxR) and inducers of endoplasmic reticulum (ER) stress. As we found that Au3 is also a nanomolar inhibitor of TrxR, a model of ER stress-induced cell death mediated by inhibition of TrxR is proposed. The transcriptomic analysis also leads to the identification of TRAIL, a ligand for death receptor 5 (DR5), as a synergistic agent of the anti-tumor activity of Au3. Collectively, our results demonstrate that the gold(III) complex Au3 effectively inhibits tumor growth in vivo, and displays promising cytotoxicity towards cancer cells in association with the inhibition of TrxR, induction of ER stress and also a death-receptor-dependent apoptotic pathway.

Spermatogenesis is a tightly regulated process leading to the development of spermatozoa. To elucidate the molecular spermatogenic mechanisms, we identified an acrosome-specific gene AEP1 in spermatids, which is located in rat chromosome 17p14 with a transcript size of 3,091 bp encoding a signal peptide, zinc finger-like motif, coiled-coil region, several predicted glycosylation and phosphorylation sites. Northern blot and RT-PCR analyses revealed the restricted expression of AEP1 to the testis only. In postnatal rat testes, AEP1 mRNA became detectable from postnatal 25 dpp (round spermatids) and onwards. By using in situ hybridization (ISH) and flow cytometry-fluorescent ISH, only the haploid spermatids yielded the positive AEP1 signal. Immunohistochemistry showed that AEP1 was expressed in the acrosomal cap of late-staged germ cells in rat testis, and co-localized with the acrosomal marker, peanut agglutinin. The spatial expression of AEP1 immunoreactivity in testis was conserved among diverse mammalian species (rat, pig, monkey, human). To further study its roles in spermatogenesis, we showed AEP1 and beta-actin was associated together in complex by co-immunoprecipitation in adult germ cells and by immunofluorescence assay in isolated spermatozoon. In human testes diagnosed with hypospermatogenesis, lower expression of AEP1 was observed, whereas there was no detectable signal in undescended testes. In short, AEP1 is an evolutionary-conserved acrosome-specific gene and likely functions in acrosome-cap formation.