An integrative analysis reveals functional targets of GATA6 transcriptional regulation in gastric cancer.

Lineage-restricted transcription factors (TFs) are frequently mutated or overexpressed in cancer and contribute toward malignant behaviors; however, the molecular bases of their oncogenic properties are largely unknown. As TF activities are difficult to inhibit directly with small molecules, the genes and pathways they regulate might represent more tractable targets for drug therapy. We studied GATA6, a TF gene that is frequently amplified or overexpressed in gastric, esophageal and pancreatic adenocarcinomas. GATA6-overexpressing gastric cancer cell lines cluster in gene expression space, separate from non-overexpressing lines. This expression clustering signifies a shared pathogenic group of genes that GATA6 may regulate through direct cis-element binding. We used chromatin immunoprecipitation and sequencing (ChIP-seq) to identify GATA6-bound genes and considered TF occupancy in relation to genes that respond to GATA6 depletion in cell lines and track with GATA6 mRNA (synexpression groups) in primary gastric cancers. Among other cellular functions, GATA6-occupied genes control apoptosis and govern the M-phase of the cell cycle. Depletion of GATA6 reduced the levels of the latter transcripts and arrested cells in G2 and M phases of the cell cycle. Synexpression in human tumor samples identified likely direct transcriptional targets substantially better than consideration only of transcripts that respond to GATA6 loss in cultured cells. Candidate target genes responded to the loss of GATA6 or its homolog GATA4 and even more to the depletion of both proteins. Many GATA6-dependent genes lacked nearby binding sites but several strongly dependent, synexpressed and GATA6-bound genes encode TFs such as MYC, HES1, RARB and CDX2. Thus, many downstream effects occur indirectly through other TFs and GATA6 activity in gastric cancer is partially redundant with GATA4. This integrative analysis of locus occupancy, gene dependency and synexpression provides a functional signature of GATA6-overexpressing gastric cancers, revealing both limits and new therapeutic directions for a challenging and frequently fatal disease.

PAK1 is a breast cancer oncogene that coordinately activates MAPK and MET signaling.

Activating mutations in the RAS family or BRAF frequently occur in many types of human cancers but are rarely detected in breast tumors. However, activation of the RAS-RAF-MEK-ERK MAPK pathway is commonly observed in human breast cancers, suggesting that other genetic alterations lead to activation of this signaling pathway. To identify breast cancer oncogenes that activate the MAPK pathway, we screened a library of human kinases for their ability to induce anchorage-independent growth in a derivative of immortalized human mammary epithelial cells (HMLE). We identified p21-activated kinase 1 (PAK1) as a kinase that permitted HMLE cells to form anchorage-independent colonies. PAK1 is amplified in several human cancer types, including 30--33% of breast tumor samples and cancer cell lines. The kinase activity of PAK1 is necessary for PAK1-induced transformation. Moreover, we show that PAK1 simultaneously activates MAPK and MET signaling; the latter via inhibition of merlin. Disruption of these activities inhibits PAK1-driven anchorage-independent growth. These observations establish PAK1 amplification as an alternative mechanism for MAPK activation in human breast cancer and credential PAK1 as a breast cancer oncogene that coordinately regulates multiple signaling pathways, the cooperation of which leads to malignant transformation.

TTF1 expression in non-small cell lung carcinoma: association with TTF1 gene amplification and improved survival.

Acquired chromosomal aberrations play an important role in tumour development and progression. Such genetic alterations occur in a significant proportion of non-small cell lung carcinomas (NSCLCs) and include amplification of 14q13.3, which contains the TTF1 gene. We asked whether TTF1 amplification is associated with increased TTF1 protein expression in NSCLCs, and whether TTF1 is associated with clinicopathological features, including patient survival. We used a FISH assay and quantitative immunohistochemical staining to interrogate a population-based cohort of 538 NSCLCs from Swiss patients for TTF1 amplification and protein expression. We found TTF1 amplification in approximately 13% of adenocarcinomas (ACs) and in approximately 9% of squamous cell carcinomas (SCCs) and TTF1 amplification was associated with increased TTF1 protein expression. High-level TTF1 expression was significantly associated with smaller tumour size, female gender and longer overall survival only among ACs (median survival 82 versus 28 months; p = 0.002). On multivariate analysis, high TTF1 expression was an independent predictor of favourable prognosis in patients with AC [hazard ratio, 0.56 (95% CI 0.38-0.83); p = 0.008]. We conclude that TTF1 amplification is a mechanism of high-level TTF1 expression in a subset of NSCLCs. When expressed at high levels, this routinely used diagnostic marker is also an independent biomarker of favourable prognosis in AC.

The major lung cancer-derived mutants of ERBB2 are oncogenic and are associated with sensitivity to the irreversible EGFR/ERBB2 inhibitor HKI-272.

Mutations in the ERBB2 gene were recently found in approximately 2% of primary non-small cell lung cancer (NSCLC) specimens; however, little is known about the functional consequences and the relevance to responsiveness to targeted drugs for most of these mutations. Here, we show that the major lung cancer-derived ERBB2 mutants, including the most frequent mutation, A775insYVMA, lead to oncogenic transformation in a cellular assay. Murine cells transformed with these mutants were relatively resistant to the reversible epidermal growth factor receptor (EGFR) inhibitor erlotinib, resembling the resistant phenotype found in cells carrying the homologous mutations in exon 20 of EGFR. However, the same cells were highly sensitive to the irreversible dual-specificity EGFR/ERBB2 kinase inhibitor HKI-272, as were those overexpressing wild-type ERBB2. Finally, the NSCLC cell line, Calu-3, overexpressing wild-type ERBB2 owing to a high-level amplification of the ERBB2 gene were highly sensitive to HKI-272. These results provide a rationale for treatment of patients with ERBB2-mutant or ERBB2-amplified lung tumors with HKI-272.

"Lineage addiction" in human cancer: lessons from integrated genomics.

Genome-era advances in the field of oncology endorse the notion that many tumors may prove vulnerable to targeted therapeutic avenues once their salient molecular alterations are elucidated. Accomplishing this requires both detailed genomic characterization and the ability to identify in situ the critical dependencies operant within individual tumors. To this end, DNA microarray platforms such as high-density single-nucleotide polymorphism (SNP) arrays enable large-scale cancer genome characterization, including copy number and loss-of-heterozygosity analyses at high resolution. Clustering analyses of SNP array data from a large collection of tumor samples and cell lines suggest that certain copy number alterations correlate strongly with the tissue of origin. Such lineage-restricted alterations may harbor novel cancer genes directing genesis or progression of tumors from distinct tissue types. We have explored this notion through combined analysis of genome-scale data sets from the NCI60 cancer cell line collection. Here, several melanoma cell lines clustered on the basis of increased dosage at a region of chromosome 3p containing the master melanocyte regulator MITF. Combined analysis of gene expression data and additional functional studies established MITF as an amplified oncogene in melanoma. MITF may therefore represent a nodal point within a critical lineage survival pathway operant in a subset of melanomas. These findings suggest that, like oncogene addiction, "lineage addiction" may represent a fundamental tumor survival mechanism with important therapeutic implications.

Evidence for two types of subunits in the bacterioferritin of Magnetospirillum magnetotacticum.

In order to investigate the role of bacterioferritin (Bfr) in the biomineralization of magnetite by microorganisms, we have cloned and sequenced the bfr genes from M. magnetotacticum. The organism has two bfr genes that overlap by one nucleotide. Both encode putative protein products of 18 kDa, the expected size for Bfr subunits, and show a strong similarity to other Bfr subunit proteins. By scanning the DNA sequence databases, we found that a limited number of other organisms, including N. gonorrhea, P. aeruginosa, and Synechocystis PCC6803, also have two bfr genes. When the sequences of a number of microbial Bfrs are compared with each other, they fall into two distinct types with the organisms mentioned above having one of each type. Differences in heme- and metal-binding sites and ferroxidase activities of the two types of subunits are discussed.

Effects of one-clip, one-kidney hypertension in chronically catheterized pregnant ewes.

1. Hypertension secondary to renal disease was studied in non-pregnant and pregnant ewes to determine whether there were any changes in arterial pressure and the distribution of cardiac output and, in particular, whether uteroplacental blood flow was affected. 2. In six non-pregnant, chronically catheterized, uninephrectomized ewes, a reduction in renal blood flow (RBF) to 40-50% of control caused hypertension within 3 h. This was maintained for as long as RBF was reduced (72 h) and returned to control 24 h after the occluder around the renal artery was released. When this experiment was repeated in 16 uninephrectomized pregnant ewes (118-134 days gestation) hypertension occurred within 3 h and was sustained for as long as RBF was reduced (between 24 and 72 h). Arterial pressure returned to control within 24-72 h of restoring RBF. 3. Compared with non-pregnant ewes, pregnant ewes had similar arterial pressures, higher cardiac outputs (CO; P < 0.001) and heart rates (HR; P < 0.001), lower total peripheral resistances (TPR; P < 0.001) and similar blood flows to brain, ovary, pancreas, kidney and spleen. Splenic vascular resistance (VR) was greater (P = 0.006), gut blood flow was greater (P < 0.05) and gut VR was less (P < 0.05). Myoendometrial blood flow/g was greater (P < 0.005) and myoendometrial VR was less (P = 0.006). 4. In pregnant sheep with renal clip hypertension, there was no change in CO and HR, but TPR increased (P < 0.01), as did plasma renin activity. Gut, brain, pancreatic and myoendometrial VR were increased as long as RBF was reduced; in addition, myoendometrial VR remained high for the rest of the experiment. Placental blood flow was unchanged at 3 h; 24-72 h later it was reduced (P < 0.05) and remained low. Placental VR was increased 24-72 h after RBF was restored when ewes were again normotensive. 5. Thus, one-clip, one-kidney renal hypertension in the pregnant ewe was due to increased TPR associated with a fall in uteroplacental blood flow that persisted even when RBF was restored and ewes were normotensive. This reduction in uteroplacental blood flow could account for the high foetal morbidity and mortality that occurs in pregnant women with renovascular hypertension.

Aminoglycoside nephrotoxicity following single-dose cystoscopy prophylaxis.

OBJECTIVE: To report a case of gentamicin-associated acute tubular necrosis following single-dose exposure. SETTING: Central-city, university-affiliated teaching hospital. PATIENT: An 82-year-old man, admitted for evaluation of shortness of breath, appetite reduction with weight loss, and stable renal insufficiency, developed urinary retention, which prompted a urologic evaluation followed by a cystoscopy. INTERVENTIONS: A single 1.6-mg/kg (actual body weight) dose of gentamicin was administered as prophylaxis prior to this procedure. RESULTS: A significant decline in renal function was observed 48 hours following gentamicin administration. A nephrology evaluation excluded pre- and postrenal causes and attributed this episode of nonoliguric acute tubular necrosis to a single dose of gentamicin. CONCLUSIONS: Although not consistent with currently proposed mechanisms and reported onset of aminoglycoside-induced nephrotoxicity, an etiology other than that concluded by this case report appears unlikely.