ObjectiveThe nature of the tumour-infiltrating leucocytes (TILs) is known to impact clinical outcome in carcinomas, including hepatocellular carcinoma (HCC). However, the role of tumour-infiltrating B cells (TIBs) remains controversial. Here, we investigate the impact of TIBs and their interaction with T cells on HCC patient prognosis.DesignTissue samples were obtained from 112 patients with HCC from Singapore, Hong Kong and Zurich and analysed using immunohistochemistry and immunofluorescence. RNA expression of CD19, CD8A, IFNG was analysed using quantitative PCR. The phenotype of freshly isolated TILs was analysed using flow cytometry. A mouse model depleted of mature B cells was used for functional study.ResultsTumour-infiltrating T cells and B cells were observed in close contact with each other and their densities are correlated with superior survival in patients with HCC. Furthermore, the density of TIBs was correlated with an enhanced expression of granzyme B and IFN-γ, as well as with reduced tumour viability defined by low expression of Ki-67, and an enhanced expression of activated caspase-3 on tumour cells. CD27 and CD40 costimulatory molecules and TILs expressing activation marker CD38 in the tumour were also correlated with patient survival. Mice depleted of mature B cells and transplanted with murine hepatoma cells showed reduced tumour control and decreased local T cell activation, further indicating the important role of B cells.ConclusionsThe close proximity of tumour-infiltrating T cells and B cells indicates a functional interaction between them that is linked to an enhanced local immune activation and contributes to better prognosis for patients with HCC.

No abstract

Aneuploidy is a key process in tumorigenesis. Dysfunction of the mitotic spindle checkpoint proteins has been implicated as a cause of aneuploidy in cells. We have previously reported that FAT10, a member of the ubiquitin-like modifier family of proteins, is overexpressed in several gastrointestinal and gynecological cancers. Here we show that FAT10 interacts with MAD2, a spindle checkpoint protein, during mitosis. Notably, we show that localization of MAD2 at the kinetochore during the prometaphase stage of the cell cycle was greatly reduced in FAT10-overexpressing cells. Furthermore, compared with parental HCT116 cells, fewer mitotic cells were observed after double thymidine-synchronized FAT10-overexpressing cells were released into nocodazole for more than 4 h. Nonetheless, when these double thymidine-treated cells were released into media, a similar number of G 1 parental and FAT10-overexpressing HCT116 cells was observed throughout the 10-h time course. Additionally, more nocodazole-treated FAT10-overexpressing cells escape mitotic controls and are multinucleate compared with parental cells. Significantly, we observed a higher degree of variability in chromosome number in cells overexpressing FAT10. Hence, our data suggest that high levels of FAT10 protein in cells lead to increased mitotic nondisjunction and chromosome instability, and this effect is mediated by an abbreviated mitotic phase and the reduction in the kinetochore localization of MAD2 during the prometaphase stage of the cell cycle. Genetic instability is an important phenomenon that underlies tumorigenesis. Chromosome instability (CIN)2 involving gains and loss of chromosomes has been found to occur in most malignancies, whereas microsatellite instability, which occurs at the nucleotide level, is less commonly observed in cancers (1). Two forms of CIN, namely structural instability and numerical instability (aneuploidy), can be observed in various tumors. Genes responsible for CIN in human cancers include those involved in the condensation of chromosomes, cohesion of sister chromatids, formation of microtubules, and kinetochore structure and function as well as mitotic "checkpoint" genes that monitor the proper progression through the cell cycle (1, 2).MAD2 (mitotic arrest-deficient 2) is a key mitotic spindle checkpoint protein whose primary role is to ensure that all of the chromosomes are properly attached to the mitotic spindle before the onset of anaphase (3). It is activated by associating with unattached kinetochores. Activated MAD2 binds to Cdc20 and prevents the anaphase-promoting complex from ubiquitylating securin. As a result, anaphase is delayed until all of the kinetochores are attached by microtubules and the chromosomes are properly aligned along the metaphase plate (4 -6). MAD2 is an essential gene, and MAD2 Ϫ/Ϫ mice die in utero (7). Loss of one allele of MAD2 has been reported to result in premature anaphase and CIN in mammalian cells (8). Dysregulation of MAD2 has been implicated in various cancers. Reduced expression of ...

Purpose: To characterize pregnane X receptor (PXR) polymorphic variants in healthy Asian populations [Chinese, Malay and Indian (n = 100 each)], and to investigate the association between PXR haplotypes and hepatic mRNA expression of PXR and its downstream target genes, CYP3A4 and ABCB1, as well as their influence on the clearance of doxorubicin in Asian breast cancer patients. Experimental Design: PXR genotyping was done by direct DNA sequencing, and PXR haplotypes and haplotype clusters were derived by expectation-maximization algorithm. Genotype-phenotype correlations were done using Mann-Whitney U test and Kruskal-Wallis test. Results: Significant interethnic variations were observed in PXR pharmacogenetics among the threeAsian ethnic groups.The expression of PXR mRNA in liver tissues harboring the PXR*1B haplotype clusters was 4-fold lower compared with the non-PXR*1B (*1A + *1C) haplotype clusters [PXR*1B versus PXR*1A ; P = 0.015; PXR*1B versus PXR*1C ; P = 0.023]. PXR*1B-bearing liver tissues were associated with significantly lower expression of CYP3A4 (PXR*1B versus non-PXR*1B, P = 0.030) and ABCB1 (PXR*1B versus non-PXR*1B, P = 0.060) compared with non^PXR*1B-bearing liver tissues. Doxorubicin clearance in breast cancer patients harboring the PXR*1B haplotypes was significantly lower compared with patients carrying the non-PXR*1B haplotypes [PXR*1B versus non-PXR*1B, CL/BSA (L h Conclusions: This study showed that PXR*1B was associated with reduced hepatic mRNA expression of PXR and its downstream targets, CYP3A4 and ABCB1. Genotype-phenotype correlates in breast cancer patients showed PXR*1B to be significantly associated with lower doxorubicin clearance, suggesting that PXR haplotype constitution could be important in influencing interindividual and interethnic variations in disposition of its putative drug substrates.

Purpose: Small-molecule growth factor receptor inhibitors block cell growth in vitro and downstream signaling in vivo, but controlled trials in patients with advanced solid tumors have yielded disappointing response rates. To clarify this discrepancy, we compared the patterns of tyrosine phosphoprotein expression in human cancer cells and primary tumors.Experimental Design: Immunoaffinity chromatography, two-dimensional electrophoresis, and antiphosphotyrosine immunoblotting were combined with mass spectrometry to determine the phosphoproteomic signatures of 40 matched normal and malignant tissues from patients with breast or liver cancer. The identities and abundance of the detected tyrosine phosphoproteins were compared with those of ligand-responsive A431 cells.Results: Patterns of tyrosine-phosphorylated proteins are similar among normal tissues of the same origin but vary markedly between different tissues. Primary breast tumors exhibit a strikingly homogeneous tyrosine phosphorylation profile, whereas liver cancers display greater phosphoproteomic diversity. The main breast-tumor-specific tyrosine phosphoproteins are cytoskeletal molecules (actin, tubulin, and vimentin) and molecular chaperones (Hsp70, Hsc71, and Grp75). In contrast, control studies in ligand-stimulated A431 human cancer cells revealed an additional phosphorylated subset of promitogenic phosphoproteins (Grb2, Shc, Jnk2, phospholipase C-␥, and phosphatidylinositol 3-kinase).Conclusions: Identification of cytoskeletal and stress proteins as the most abundant tyrosine phosphoproteins in breast tumors implicates these molecules, rather than promitogenic effectors, as the prime stoichiometric substrates for kinase-inhibitory anticancer drugs in vivo. Because phosphorylated cytoskeletal proteins and chaperones mediate cell motility and apoptotic resistance, respectively, these data raise the intriguing possibility that small-molecule tyrosine kinase inhibitors may be of greatest value either as adjuvant antimetastatic/-invasive drugs or as chemo-/radiosensitizers.

Liver resection is commonly performed for solitary hepatocellular carcinoma (HCC) in well-compensated cirrhotic and noncirrhotic patients. Data concerning exacerbation of chronic hepatitis B (ECHB) post-liver resection are scant. To determine the incidence, risk factors, and clinical outcomes of ECHB in patients who underwent hepatic resection for HCC. The methods consisted of a retrospective review of consecutive patients with chronic hepatitis B virus (HBV) infection who had undergone liver resection for HCC from January 2002 to December 2004. Seventy-seven patients underwent 82 liver resections; the mean age was 58.0 +/- 12.1 years; 87% male; 20% hepatitis B e-antigen positive. Incidence of all causes of postoperative hepatitis was 25.6% (n = 21), and ECHB was 8.5% (n = 7). Both groups had their peak alanine aminotransferases, 231.0 IU/L (74-1,400) and 312 IU/L (147-1,400), respectively, observed at day 84 postresection. Three patients died as a result of ECHB within 4 months postsurgery. One- and 2-year survival rates were poorest for the ECHB group at 42.9 and 21.4%, compared with those with postoperative hepatitis due to other causes at 60.3 and 45.2% and those without postoperative hepatitis at 87.7 and 73.5% (p < 0.001). Liver resection for HCC in patients with chronic HBV infection carries a risk for ECHB, and affected patients have poorer clinical outcomes. There is a need for close monitoring of these patients preoperatively and in the early postoperative period.

Uncontrolled cell division is a hallmark of cancer. Deregulation of Wnt components has been linked to aberrant cell division by multiple mechanisms, including Wnt-mediated stabilisation of proteins signalling, which was notably observed in mitosis. Analysis of Wnt components revealed an unexpected role of B-cell CLL/lymphoma 9 (BCL9) in maintaining mitotic Wnt signalling to promote precise cell division and growth of cancer cell. Mitotic interactome analysis revealed a mechanistic role of BCL9 in inhibiting clathrin-mediated degradation of LRP6 signalosome components by interacting with clathrin and the components in Wnt destruction complex; this function was further controlled by CDK1-driven phosphorylation of BCL9 N-terminal, especially T172. Interestingly, T172 phosphorylation was correlated with cancer patient prognosis and enriched in tumours. Thus, our results revealed a novel role of BCL9 in controlling mitotic Wnt signalling to promote cell division and growth.

The prevalence of unmet psychosocial needs among cancer patients in ambulatory care is generally high. Young patients with disease recently diagnosed at advanced stage will benefit from additional support.