Serum cytokeratin 19 fragment 21-1 and carcinoembryonic antigen combination assay as a biomarker of tumour progression and treatment response in extramammary Paget disease.

BACKGROUND: Extramammary Paget disease (EMPD) is a rare intraepithelial adenocarcinoma affecting the genitals and axillary regions. As metastasis of these tumours is itself rare, solid disease management strategies have not been established. Serum carcinoembryonic antigen (CEA) and cytokeratin 19 fragment 21-1 (CYFRA 21-1) levels have been identified as candidate biomarkers for tumour progression in EMPD. OBJECTIVES: To determine the accuracy of and the correlation between these markers in patients with EMPD. METHODS: Serum CEA and CYFRA 21-1 levels were examined in 30 patients with EMPD treated at Keio University Hospital, and compared against clinical information. Both assays were performed at the time of diagnosis, during the postoperative observation period, and following systemic treatment in those with confirmed metastasis. Serum levels were then correlated with tumour progression status and treatment responses. RESULTS: Normal levels for both assays were observed in all 11 patients with primary localized disease (100%). In patients with metastatic disease the CEA positivity rate was 79% (15 of 19 patients), with a rate of 63% (12 of 19 patients) for CYFRA 21-1. Changes in CEA and CYFRA 21-1 levels were statistically independent; however, using a combined view, elevated levels of either marker improved the positivity rate to 95% (18 of 19 patients). Use of both markers also correlated well with treatment responses. CONCLUSIONS: The combination of CEA and CYFRA 21-1 is useful for predicting metastasis and treatment response in patients with EMPD, especially in those who only have elevation of a single marker. What's already known about this topic? Serum levels of carcinoembryonic antigen (CEA) and cytokine 19 fragment 21-1 (CYFRA 21-1) have been shown to be elevated in patients with extramammary Paget disease (EMPD). Elevation of serum CEA levels is associated with tumour progression of EMPD. A single small study reported that serum CYFRA 21-1 levels are elevated in patients with EMPD with lymph node metastasis. What does this study add? Serum CEA and CYFRA 21-1 were present in 79% and 63% of 19 cases of metastatic EMPD, respectively. Elevations of CEA and CYFRA 21-1 were statistically independent. CEA and CYFRA 21-1 combination assays were positive in 95% of cases of metastatic EMPD. What is the translational message? Combination assays with CEA and CYFRA 21-1 are useful for monitoring treatment response in patients with metastatic EMPD, particularly in those with elevation of either marker.

Serous cystic neoplasm of the pancreas: a multinational study of 2622 patients under the auspices of the International Association of Pancreatology and European Pancreatic Club (European Study Group on Cystic Tumors of the Pancreas).

OBJECTIVES: Serous cystic neoplasm (SCN) is a cystic neoplasm of the pancreas whose natural history is poorly known. The purpose of the study was to attempt to describe the natural history of SCN, including the specific mortality. DESIGN: Retrospective multinational study including SCN diagnosed between 1990 and 2014. RESULTS: 2622 patients were included. Seventy-four per cent were women, and median age at diagnosis was 58 years (16-99). Patients presented with non-specific abdominal pain (27%), pancreaticobiliary symptoms (9%), diabetes mellitus (5%), other symptoms (4%) and/or were asymptomatic (61%). Fifty-two per cent of patients were operated on during the first year after diagnosis (median size: 40 mm (2-200)), 9% had resection beyond 1 year of follow-up (3 years (1-20), size at diagnosis: 25 mm (4-140)) and 39% had no surgery (3.6 years (1-23), 25.5 mm (1-200)). Surgical indications were (not exclusive) uncertain diagnosis (60%), symptoms (23%), size increase (12%), large size (6%) and adjacent organ compression (5%). In patients followed beyond 1 year (n=1271), size increased in 37% (growth rate: 4 mm/year), was stable in 57% and decreased in 6%. Three serous cystadenocarcinomas were recorded. Postoperative mortality was 0.6% (n=10), and SCN's related mortality was 0.1% (n=1). CONCLUSIONS: After a 3-year follow-up, clinical relevant symptoms occurred in a very small proportion of patients and size slowly increased in less than half. Surgical treatment should be proposed only for diagnosis remaining uncertain after complete workup, significant and related symptoms or exceptionally when exists concern with malignancy. This study supports an initial conservative management in the majority of patients with SCN. TRIAL REGISTRATION NUMBER: IRB 00006477.

Differential gene expression screening between parental and highly metastatic pancreatic cancer variants using a DNA microarray.

To clarify the difference in genes expressed in hematogenous metastasis and peritoneal dissemination, a broad analysis of differential gene expression analysis between parental cell lines and established metastatic sublines was performed. Using an oligonucleotide array (Gene Chip, Affymetrix), approximately 2,000 genes involved in cancer were analyzed for each of the cell lines. HPC-4H4 (highly metastatic lines to the liver) compared with HPC-4 (low metastatic parental lines), in which 20 overexpressed genes and 5 underexpressed genes were recognized. HPC-4P4a (highly metastatic to the peritoneum) compared with HPC-4, in which 12 overexpressed genes and 15 underexpressed genes were also recognized. Analysis of HPC-4H4 and HPC-4P4a showed comparative up-regulation of 20 genes and down-regulation of 13 in the former, HPC-4H4. Further studies are needed to validate our hypothesis that some of the resulting differentially expressed genes might be implicated in the development of metastasis in pancreatic cancer. In conclusion, this genome-wide expression analysis will help to clarify the molecular mechanisms of cancer metastasis and of the different levels of gene expression in a variety of metastatic potentials in pancreatic cancer.

Origin of the thoracic duct and pancreaticoduodenal lymphatic pathways to the para-aortic lymph nodes.

We investigated the afferent and efferent connections of the para-aortic lymph nodes (group 16 nodes) relative to the origin of the thoracic duct in 85 postmortem cadavers. The origin was usually restricted to groups 16b1-inter and -latero nodes (type I; 90.6%), regardless of whether the union of their efferents occurred at the abdominal or thoracic level. We also occasionally observed thick collecting vessels originating from the dorsal aspect of the pancreaticoduodenal region, running along the right side of and superficial to the celiac plexus and emptying into group 16b1 nodes. The thoracic duct originated occasionally not only from group 16b1 nodes but also from group 16a2 nodes (type II; 9.4%). Moreover, in all 85 specimens, the group 16a2-inter node often received afferents from the celiac plexus itself or the tight connective tissue between the plexus and diaphragmatic crus, or both. The results support the reliability of the extended D2 lymphadenectomy (D2 + group 16b1 nodes + group 16a2-inter node) for curative cancer surgery in the pancreaticoduodenal region.

Survival-factor-induced phosphorylation of Bad results in its dissociation from Bcl-x(L) but not Bcl-2.

The pro-apoptotic Bcl-2-family protein Bad heterodimerizes with Bcl-2 and Bcl-x(L) in the outer mitochondrial membranes, nullifying their anti-apoptotic activities and promoting cell death. We report that interleukin-3 (IL-3) stimulation induces Bad phosphorylation and triggers its translocation from mitochondria to cytoplasm in cells expressing Bcl-x(L) but not Bcl-2. Overexpression of Bad sensitized Bcl-x(L)-expressing FL5.12 cells to apoptosis induced by IL-3 deprivation, but had no effect on the viability of cells expressing Bcl-2. IL-3 stimulation induced Bad phosphorylation at Ser-112, impairing its binding to Bcl-x(L) and resulting in its association with 14-3-3 proteins in the cytosol. However, Ser-112 phosphorylation could not trigger Bad dissociation from mitochondria in FL5.12 cells expressing Bcl-2. In 293T cells expressing Bcl-x(L), Bad was phosphorylated at three serines, 112, 136 and 155, and was largely localized in the cytosolic fraction. In contrast, overexpression of Bcl-2 prevented phosphorylation of Bad at Ser-136 and Ser-155, sequestering this protein in the mitochondrial membranes. When the N-terminal regions of Bcl-2 and Bcl-x(L) were swapped with each other, the Bcl-x(L)(N)-Bcl-2 chimaeric protein (containing the N-terminal region of Bcl-x(L)) failed to prevent Bad phosphorylation in cells and was unable to block the cytosolic distribution of this pro-apoptotic protein. Additional experiments with the Bcl-2(N)-Bcl-x(L) chimaeric protein (containing the N-terminal region of Bcl-2) indicated that, although the N-terminal region of Bcl-2 is necessary, it is not sufficient for sequestering Bad in the mitochondrial membranes. These observations suggest that growth-factor-mediated phosphorylation of Bad contributes to the cytoprotective function of Bcl-x(L) but not Bcl-2.

The cell surface-expressed HSC70-like molecule preferentially reacts with the rat T-cell receptor Vdelta6 family.

We previously showed that the cell surface-expressed Mr 70,000 heat shock cognate (hsc70, a constitutively expressed member of the hsp70 family) protein-like molecule (#067 molecule) interacts with rat CD3+, CD4-, CD8-, T-cell receptor (TCR)alphabeta-, natural killer recetor-P1- T cells. This 70hsc-like molecule was also suggested to present cellular peptide antigens to these T cells. In the present study, we identified the genetic structure of the TCR by establishing T-cell hybridomas between these T cells and mouse BW5147 cells. Our data indicated that these T cells preferentially used TCRs with the Vdelta6 family. Analysis of the nucleotide sequence of the CDR3 junctional portion showed that there are substantial diversities, with insertion of seven to nine amino acid residues. These data provide indirect evidences for our hypothesis that an hsc70-like molecule could be presented together with cellular peptide antigens to particular T cells with TCR gammadelta chains. Since the expression of this hsc70-like #067 antigen on the cell surface is usually induced along with cell transformation by activated oncogenes, T cells with the TCR Vdelta6 family are likely to contribute to host resistance to tumor cells.

Primary sclerosing cholangitis successfully treated by resection of the confluence of the hepatic duct.

Primary sclerosing cholangitis (PSC) is a cholestatic disease characterized by chronic inflammatory fibrosis of the extra- and intrahepatic bile ducts. Although the prognosis of patients with PSC was believed to be poor, some patients have not experienced the expected rapid clinical progression. A 51-year-old man with PSC was initially hospitalized for jaundice. Laboratory data showed low levels of the complement components C3, C4, and CH50. Percutaneous transhepatic biliary drainage was performed. Cholangiography revealed complete obstruction of the common bile duct below the confluence of the cystic duct. The confluence of the hepatic duct was resected and it was reconstructed by hepaticojejunostomy for palliation of the obstructive jaundice. Increased thickness of the walls of the common bile duct, right hepatic bile duct, and gallbladder was observed. Histopathological examination of the resected specimen revealed periductal fibrosis, with an onion-skin-like appearance. The patient is currently doing well, approximately 7 years after the surgery, without any signs of PSC recurrence. In this extraordinary patient, the laboratory data for C3, C4, and CH50 showed a complete return to normal levels. The positive results in this patient suggest that resection of the confluence of the hepatic duct may be an effective surgical treatment for noncirrhotic PSC patients who have dominant extrahepatic strictures.

Long descending lymphatic pathway from the pancreaticoduodenal region to the para-aortic nodes: its laterality and topographical relationship with the celiac plexus.

In 6 of 15 postmortem-treated cadaveric specimens, we found macroscopically thick lymphatic collecting vessels that originated from not only the nodes along the common hepatic artery (No. 8 nodes) but also from the pancreaticoduodenal region, and which drained directly into the para-aortic nodes immediately below the left renal vein (No. 16b1-inter or -latero nodes). The collecting vessels, if they originated from the ventral (dorsal) visceral side, passed to the left (right) of the superior mesenteric and celiac arteries. Moreover, the right-side vessels (5 specimens) were classified into superficial and deep courses to the celiac plexus, whereas they were superficial in the left side (2 specimens). One of the deep (right) courses continued to the thoracic duct without any intercalated nodes. In addition, another deep route drained into the para-aortic node immediately above the left renal vein (No. 16a2-inter node). We consider that these collecting vessels form "direct descending pathways" from the relatively peripheral lymphatics in the upper abdomen toward the thoracic duct origin. The pathway seems to be a collateral, or even major drainage route, and it appears responsible for skipped metastasis of primary cancer. Since the classical, limited entity of the intestinal lymph trunk does not coincide with our pathway, it should be reconsidered. The proposed entity of the direct, long descending pathway will influence the selection and modification of lymphadenectomy methods in cancer surgery in the pancreaticoduodenal region.

Acquisition of multidrug resistance in recurrent breast cancer demonstrated by the histoculture drug response assay.

Recurrent breast cancer has a very poor response rate to chemotherapy. To understand the degree of acquisition of multidrug resistance in recurrent disease, 24 recurrent breast tumors and 127 primary tumors were evaluated and compared for chemosensitivity in the histoculture drug response assay (HDRA). The evaluation rate was 98.8%. The HDRA utilizes 3-dimensional culture of human tumors on collagen-gel rafts. Doxorubicin (DXR), 5-fluorouracil (5-FU) and mitomycin C (MMC) were tested as standard agents and cisplatin (CDDP) as a candidate agent on surgical specimen of breast cancer in the HDRA. In vitro drug exposure in the HDRA was for 7 days. At the end of the assay, tumor response was assessed by the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). The mean inhibition rates of primary tumors vs. recurrent tumors were 57.9% and 38.6% for DXR (p<0.0005); 59.9% and 42.8% for MMC (p<0.01); 49.0% and 33.4% for 5-FU (p<0.01); and 34.5% and 16.0% for CDDP (p<0.005), respectively. The recurrent cases were pretreated clinically with CAF (cyclophosphamide, DXR and 5-FU), CEF (cyclophosphamide, epirubicin and 5-FU) or CMF (cyclophosphamide, methotrexate and 5-FU). In the CAF and CEF group, the HDRA sensitivity to CDDP was significantly lower in recurrent disease (p<0.005) than that of primary breast cancer suggesting that one agent can induce resistance to another. This is further suggested by the fact that 64.7% of the recurrent cases were resistant to all 4 agents tested as opposed to 27% of the primary cases and that only 5.9% of the recurrent cases were sensitive to three or more agents as opposed to 18% of the primary cases. The correlation of the HDRA results to clinical outcome in the study was 80.0% with 15 cases evaluated consisting of 5 true positives, 3 false positives, 7 true negatives and no false negatives. Thus, the HDRA gives useful clinical information, in particular for the specific individualized treatment design necessary to overcome the multidrug resistance problem of recurrent breast cancer.

High dorsal drainage routes of Spiegel's lobe.

BACKGROUND/PURPOSE: The venous drainage from Spiegel's lobe to the terminal portion of the hepatic veins has been described in the literature, but its morphology remains unclear. METHODS: We examined 42 dissected liver specimens and 38 cast specimens. RESULTS: In 8 of the 42 dissected liver specimens and 5 of the 38 cast specimens we found atypical but thick (over 3 mm) caudate veins that drained Spiegel's lobe and emptied into the terminal portion (along the most proximal 5-mm course) of the middle hepatic vein (MHV) or the inferior vena cava (IVC) near the MHV terminal (less than 10 mm from the MHV). We termed these the superior caudate vein. This vein ran upward between the caudate portal branches of the left and hilar bifurcation origins or through the territory of the left origin. The superior caudate vein, consistently coexisted with the typical vein(s). We also found several analogues of the superior caudate vein, such as the cranially shifted opening of the typical caudate vein and relatively thick proximal tributaries of the MHV from Spiegel's lobe. CONCLUSIONS: Although the superior caudate vein, if present, seemed to have a large role in the venous drainage of the lobe, its incidence seemed to be too low for clinical relevance.

Ischemic cholangitis caused by transcatheter hepatic arterial chemoembolization 10 months after resection of the extrahepatic bile duct.

We report a case of ischemic cholangitis that occurred after transcatheter hepatic arterial chemoembolization (TAE). Ten months prior to TAE the patient had undergone central bisegmentectomy for hepatocellular carcinoma with resection of the extrahepatic bile duct. Eleven days after TAE, he developed suppurative cholangitis and multiple organ failure. Prior surgical ligation of the peribiliary arteries around the extrahepatic bile duct followed by TAE was considered to have played a crucial role in the development of ischemic cholangitis. This case demonstrates the importance of blood flow from the peribiliary arteries for the survival of the biliary epithelium.

Exercise capacity of thoracotomy patients in the early postoperative period.

OBJECTIVE: We investigated the mechanism involved with the initial drop and subsequent recovery of exercise capacity in the early postoperative period of thoracotomy patients. METHODS: Sixteen patients (13 who had undergone lobectomy, 3 who had undergone pneumonectomy) underwent a routine pulmonary function test (PFT) and a cardiopulmonary exercise test preoperatively, within 14 postoperative days (POD; post-1; mean +/- SD, 9 +/- 2 POD), and after 14 POD (post-2; mean, 26 +/- 12 POD). RESULTS: After surgery on post-1, PFT results of FVC, FEV(1), and maximum ventilatory volume (MVV) significantly decreased. Oxygen uptake (VO(2)) at a venous blood lactate level of 2.2 mmol/L (La-2. 2), which was adopted as the empirical anaerobic threshold, and maximum V O(2) (VO(2)max) decreased significantly to 88.2 +/- 7.9% and 73.1 +/- 15.4% of the preoperative values, respectively. La-2.2 min ventilation (VE)/ MVV and maximum VEmax)/MVV increased significantly from 0.36 +/- 0.08 to 0. 66 +/- 0.20 and from 0.58 +/- 0.14 to 0.80 +/- 0.09, respectively. On post-2, though La-2.2 VO(2) did not change, VO(2)max improved significantly to 81.5 +/- 19.7% of the preoperative values, in association with significant increases in maximal tidal volume and VEmax, which were produced by significant increases in the PFT results. La-2.2 VE/MVV also decreased significantly to 0.49 +/- 0.13, which indicated a sufficient recovery of respiratory reserve at submaximal exercise. CONCLUSIONS: The initial drop of exercise capacity after lung resection seems to be derived from both circulatory and ventilatory limitations. Further, the subsequent recovery within 1 month seems to be produced by an improvement in ventilatory limitation, which was caused by the surgical injury to the chest wall.

Human CD8 and CD4 T cell epitopes of epithelial cancer antigens.

Recent human tumor immunology research has identified several genes coding immunogenic peptides recognized by CD8 cytotoxic T lymphocytes (CTLs) in melanoma tumors. Very recently, CD4 T cell antigenic epitopes were also determined in certain melanoma tumors. The use of these peptides in conjunction with human immunotherapy could prove to be of great benefit. However, such peptides in clinically common tumors of epithelial cell origin, such as of the stomach, colon, lung, etc., have not yet been determined extensively. We describe for the first time an HLA-A31 (A*31012)-restricted natural antigenic peptide recognized by the CD8 CTL TcHST-2 of gastric signet ring cell carcinoma cell line HST-2. We also identified the HLA-DRB1*08032-restricted peptide recognized by the CD4 T cell line TcOSC-20 of squamous cell carcinoma OSC-20 derived from the oral cavity. The antigenic peptide of HST-2, designated F4.2, is composed of 10 amino acid residues with two anchor motif residues necessary for binding to HLA-A31 molecules. The synthetic F4.2 peptide enhanced the reactivity of TcHST-2 against HST-2 cells. Furthermore, introduction of an expression minigene coding F4.2 peptide to HLA-A31(+) cells conferred cytotoxic susceptibility to TcHST-2 on the cells. Some stomach cancer lines into which the HLA-A31 gene had been introduced, such as MKN28-A31-2, were lysed by TcHST-2, suggesting the presence of F4.2 peptide in at least some HLA-A31(+) stomach cancers. Furthermore, F4.2 peptide induced an F4.2 peptide-specific CTL response in at least 30-40% of HLA-A31(+) peripheral blood lymphocytes from gastric cancer patients, suggesting that F4.2 peptide could be used as a cancer vaccine for gastric tumors. The natural antigenic peptide of OSC-20 was also determined using acid extraction and biochemical separation and by mass spectrometry. Consequently, OSC-20 peptide was designated as the 6-1-5 peptide, an HLA-DRB1*08032-restricted 16-mer peptide with two possible anchor motifs. It has an amino acid sequence identical to that of human alpha-enolase, suggesting that it was derived from the processed parental alpha-enolase protein. We are presently attempting to determine the genes that code tumor rejection antigens recognized by HLA-A24- and A26-restricted T cells, including those of pulmonary and pancreatic carcinomas. The search for these antigenic peptides may lead to the identification of immunogenic peptide antigens that would be suitable for clinical use in commonly occurring epithelial cancers.

[Histoculture drug response assay on non-small cell lung cancer].

We examined the chemosensitivity of non-small cell lung cancer (NSCLC) tissues to CDDP, 5-FU, ADM, MMC, ETP and SN38 using histoculture drug response assay (HDRA). One-hundred and thirty surgical specimens from NSCLC patients who were not given preoperative chemotherapy were used. The inhibition indexes of CDDP, 5-FU, MMC, ADM, ETP and SN38 were 39.1 +/- 18.2%, 48.0 +/- 19.7%, 63.3 +/- 17.7%, 47.6 +/- 22.0%, 36.9 +/- 21.1%, and 37.9 +/- 25.2%, respectively. Inhibition indexes were above the cutoff level, i.e., 'judged sensitive,' in 40 cases (31.3%) for CDDP, 34 cases (27.4%) for 5-FU, 54 cases (44.3%) for MMC, 36 cases (33.0%) for ADM, 29 cases (29.8%) for ETP, and 34 cases (37.4%) for SN38, respectively. In almost one-third of patients, the inhibition indexes of all drugs were under cutoff levels. Correlations between in vitro chemosensitivity data and patient responses to chemotherapy were obtained from 16 evaluable patients, and a 44.4% true positive rate and a 100% true negative rate were observed. Our results with HDRA for NSCLC showed a high incidence of intrinsic multidrug resistance. HDRA may help doctors to avoid non-effective chemotherapy for NSCLC patients.