Src Kinase Is Biphosphorylated at Y416/Y527 and Activates the CUB-Domain Containing Protein 1/Protein Kinase C δ Pathway in a Subset of Triple-Negative Breast Cancers.

Targeted therapeutics are needed for triple-negative breast cancer (TNBC). In this study, we investigated the activation of Src family of cytoplasmic tyrosine kinases (SFKs) and two SFK substrates-CUB-domain containing protein 1 (CDCP1) and protein kinase C δ (PKCδ)-in 56 formalin-fixed, paraffin-embedded (FFPE) TNBCs. Expression of SFK phosphorylated at Y416 (SFK_pY416+) in tumor cells was strongly associated with phosphorylation of CDCP1 and PKCδ (CDCP1_ pY743+ and PKCδ_pY311+), as assessed by immunohistochemistry, indicating increased SFK activity in situ. To enable biochemical analysis, protein extraction from FFPE tissue was optimized. Cleaved CDCP1 isoform (70 kDa) was expressed to a varying degree in all samples but only phosphorylated in TNBC tumor cells that expressed SFK_pY416. Interestingly, active SFK was found to be biphosphorylated (SFK_pY416+/pY527+). Biphosphorylated active SFK was observed more frequently in forkhead box protein A1 (FOXA1)- TNBCs. In addition, in SFK_pY416- samples, FOXA1+ TNBC tended to be SFK_pY527+ (classic inactive SFK), and FOXA1- TNBC tended to be SFK_pY527- (SFK poised for activation). Strong SFK_pY416 staining was also observed in tumor-infiltrating lymphocytes in a subset of TNBCs with high tumor-infiltrating lymphocyte content. This report will facilitate protein biochemical analysis of FFPE tumor samples and justifies the development of therapies targeting the SFK/CDCP1/PKCδ pathway for TNBC treatment.

Synchronous recurrence of concurrent colon adenocarcinoma and dedifferentiated liposarcoma.

A 62-year-old man presented with concurrent sigmoid colon adenocarcinoma and small bowel mesenteric dedifferentiated liposarcoma. Following surgical resection of the colon cancer, complete excision of the mesenteric sarcoma and adjuvant folinic acid, fluorouracil and oxaliplatin (FOLFOX) chemotherapy, the patient demonstrated no radiological evidence of disease for more than 2 years. The patient then developed synchronous recurrence of both cancers: the colon cancer metastasised to the liver and a pelvic lymph node, and the liposarcoma recurred in the original location. The patient underwent additional chemotherapy with complete response of the metastatic colon cancer and stable disease for the liposarcoma. The recurrent mesenteric tumour was subsequently resected. Although concurrent cancers have been reported, this unique case of synchronous recurrence raises interesting hypotheses regarding host-tumour interaction and immune surveillance.

A Pilot Study of Ultrasound-Guided Cryoablation of Invasive Ductal Carcinomas up to 15 mm With MRI Follow-Up and Subsequent Surgical Resection.

OBJECTIVE: The purpose of this study was to evaluate the effectiveness of ultrasound-guided cryoablation in treating small invasive ductal carcinoma and to assess the role of contrast-enhanced (CE) MRI in determining the outcome of cryoablation. SUBJECTS AND METHODS: Twenty consecutive participants with invasive ductal carcinomas up to 15 mm, with limited or no ductal carcinoma in situ (DCIS), underwent ultrasound-guided cryoablation. Preablation mammography, ultrasound, and CE-MRI were performed to assess eligibility. Clinical status was evaluated at 1 day, 7-10 days, and 2 weeks after ablation. CE-MRI was performed 25-40 days after ablation, followed by surgical resection within 5 days. RESULTS: Ultrasound-guided cryoablation was uniformly technically successful, and postablation clinical status was good to excellent in all participants. Cryoablation was not clinically successful in 15% (three of 20 patients). Three participants had residual cancer at the periphery of the cryoablation site. Two participants had viable nonmalignant tissue within the central zone of cryoablation-induced necrosis. Postablation CE-MRI had a sensitivity of 0% (0/3) and specificity of 88% (15/17). The predictive value of negative findings on CE-MRI was 83% (15/18). Correlations between cancer characteristics, cryoablation procedural variables, postablation CE-MRI findings, and surgical specimen features were not statistically significant. There were also no significant differences in participants with or without residual cancer. CONCLUSION: In our pilot experience, ultrasound-guided cryoablation of invasive ductal carcinomas up to 15 mm has a clinical failure rate of 15% but is technically feasible and well tolerated by patients. The majority of cryoablation failures are manifest as DCIS outside the cryoablation field. Postablation CE-MRI does not reliably predict cryoablation outcome.

Identification of 12 or more lymph nodes in resected colon cancer specimens as an indicator of quality performance.

BACKGROUND: : Identification of > or =12 lymph nodes in resected colon cancer specimens has been endorsed as a quality indicator. METHODS: : The Hoag Hospital cancer registry was used to identify patients diagnosed with colon cancer. The proportion of colon cancer specimens for which > or =12 lymph nodes were identified was determined by anatomic location, stage of disease, patient age, and operating surgeon. Survival was correlated with stage and with whether > or =12 lymph nodes were identified. RESULTS: : Pathology procedural changes in 1998 were associated with an increase in the average number of lymph nodes identified from 8.0 to 14.5 (P < .0001); therefore, analysis was limited to 574 patients who underwent surgical resection of colon adenocarcinoma during 1998 to 2005. Identification of > or =12 lymph nodes varied from 57% to 83% by 7 anatomic locations (P < .0001), from 65% to 75% by 5 age cohorts (P = .027), from 59% to 73% by 4 general stages of disease (P = .004), and from 53% to 80% among 12 surgeons who performed at least 17 resections (P = .014). The proportion of resections in which > or =12 lymph nodes were identified was higher for 3 colorectal fellowship-trained surgeons compared with the other 9 surgeons (77% vs 63%, P = .0007), and with 30 surgeons who each performed <10 resections (77% vs 51%, P < .0001). Identification of > or =12 lymph nodes was associated with better survival for patients with stage I (P = .016) and stage II (P = .021) disease. CONCLUSIONS: : Anatomic location, colorectal surgical training, and case volume were strongly correlated with the number of lymph nodes identified. Cancer 2009. (c) 2009 American Cancer Society.

Establishment and characterization of a human primary prostate carcinoma cell line, HH870.

BACKGROUND: Development of new therapeutic modalities for human prostate carcinoma has been impeded by a lack of adequate in vitro and in vivo models. Most in vitro studies have been carried out using a limited number of human prostate cancer cell lines that are mostly derived from metastatic tumors sites or are immortalized. METHODS: Characterization of the prostate cancer cell line, HH870, included description of morphology, determination of doubling time, response to androgens, immunocytochemistry, and immunoblotting of proteins known to be associated with prostate carcinoma, karyotyping, fluorescence in situ hybridization (FISH), DNA profiling, and growth as xenograft in athymic rodents. RESULTS: HH870 expresses various epithelial marker antigens that correlate with known basic immunostaining profiles of prostate adenocarcinoma, although the cell line does not express PSA, PSMA, or PAP. HH870 exhibits complex chromosomal abnormalities and harbors no immortalizing HPV, BKV, JCV, and SV40 DNA. CONCLUSIONS: We report the successful establishment and characterization of a new long-term primary human prostate tumor cell line HH870.

A plasminogen-like protein selectively degrades stearoyl-CoA desaturase in liver microsomes.

Stearoyl-CoA desaturase (SCD) is an integral membrane protein of the endoplasmic reticulum that is rapidly and selectively degraded when isolated liver microsomes are incubated at 37 degrees C. We previously reported the purification of a 90-kDa microsomal protein with SCD protease activity and characterized the inhibitor sensitivity of the protease. Here we show that the 90-kDa protein is a microsomal form of plasminogen (Pg) and that the purified SCD protease contains a spectrum of plasmin-like derivatives. The 90-kDa protein was identified as Pg by mass spectrometry of its tryptic peptides. The purified SCD protease reacted with Pg antibody, and immunoblotting demonstrated enrichment of Pg by the purification procedure established for the SCD protease. Analysis of microsomes by zymography demonstrated a single band of proteolytic activity at 70-kDa corresponding to the mobility of Pg in nonreduced polyacrylamide gels. When microsomes were incubated at 37 degrees C prior to zymography, an intense band of proteolytic activity developed at 30-kDa. The purified SCD protease displayed a spectrum of proteolytic bands ranging from 70 to 30 kDa. Degradation of SCD by the purified protease and by microsomes was inhibited by bdellin, a plasmin inhibitor from the medicinal leech Hirudo medicinalis. To explore the role of Pg in the degradation of SCD in vivo, we examined SCD expression and degradation in microsomes isolated from Pg-deficient (Pg-/-) mice. Compared with microsomes from wild-type littermate control mice, liver microsomes from Pg-/- mice had significantly higher levels of SCD. Degradation of SCD in microsomes from Pg-/- mice was markedly diminished, whereas liver microsomes from control mice showed rapid SCD degradation similar to that observed in rat liver microsomes. These findings indicate that SCD is degraded by a protease related to Pg and suggest that plasmin moonlights as an intracellular protease.

A microsomal endopeptidase from liver that preferentially degrades stearoyl-CoA desaturase.

A protease was purified some 700-fold from rat liver microsomes by a combination of differential detergent solubilization, hydroxyapatite column chromatography, and gel filtration. The protease exhibits substrate selectivity for stearoyl-CoA desaturase (SCD). The purified protease rapidly degraded SCD while other microsomal proteins including cytochrome b(5) and 11beta-hydroxysteroid dehydrogenase were degraded slowly or not at all. The isolated form of the protease has an apparent molecular mass of approximately 90 kDa. Upon incubation, the 90 kDa form of the protease undergoes rapid conversion to a series of smaller proteins. This conversion is associated with a marked increase in proteolytic activity. Diisopropyl phosphofluoridate (DFP) at high concentration partially inhibited the protease activity. The [(3)H]DFP-labeled protease was detected as three protein bands of approximately 66 kDa under nonreducing conditions and a single 25 kDa band under reducing conditions. The purified protease was inhibited by dithiothreitol, suggesting the presence of an essential disulfide bond. These results further define the mechanism by which SCD is rapidly and selectively degraded in isolated liver microsomes.

Stearoyl-CoA desaturase, a short-lived protein of endoplasmic reticulum with multiple control mechanisms.

Stearoyl-CoA desaturase (SCD) is a short-lived, polytopic membrane-bound non-heme iron enzyme localized primarily in the endoplasmic reticulum. SCD is required for the biosynthesis of monounsaturated fatty acids, and plays a key role in hepatic synthesis of triglycerides and very-low-density lipoproteins. The intracellular concentration of SCD fluctuates in a wide range in response to complex and often competing hormonal and dietary factors. A combination of transcriptional regulation and rapid protein degradation produces transient elevations of SCD enzyme activity in response to physiologic demands. Dysregulation of SCD has been implicated in non-alcoholic fatty liver disease, hyperlipidemia, and obesity.