Aberrant activation of the canonical Wnt/β-catenin pathway occurs in almost all colorectal cancers and contributes to their growth, invasion and survival. Although dysregulated β-catenin activity drives colon tumorigenesis, additional genetic perturbations are required to elaborate fully malignant disease. To identify genes that both modulate β-catenin activity and are essential for colon cancer cell proliferation, we conducted two loss-of-function screens in human colon cancer cells and compared genes identified in these screens with an analysis of copy-number alterations in colon cancer specimens. One of these genes, CDK8, which encodes a member of the mediator complex, is located at 13q12.13, a region of recurrent copy number gain in a substantial fraction of colon cancers. Suppression of CDK8 expression inhibited proliferation in colon cancer cells characterized by high levels of CDK8 and β-catenin hyperactivity. CDK8 kinase activity was necessary for β-catenin driven transformation and expression of several β-catenin transcriptional targets. Together these observations suggest that therapeutic interventions targeting CDK8 may confer clinical benefit in β-catenin-driven malignancies.Correspondence and Requests for materials should be addressed to W.C.H. (Email: william_hahn@dfci.harvard.edu).. The Wnt/β-catenin pathway is implicated in over 90% of colon cancers and in a fraction of other human malignancies. Loss of the tumor suppressor APC or activating CTNNB1 (β-catenin) mutations results in constitutive activity of the β-catenin-T cell factor (TCF) transcriptional complex, which drives adenoma formation 1,2 . Although mutations in TP53 or K-RAS cooperate with dysregulated β-catenin signaling to program a fully malignant phenotype 3 , these mutations are found in less than half of β-catenin-driven colon cancers 4 . NIH Public AccessTo identify oncogenes that modulate β-catenin-dependent transcription and regulate colon cancer cell proliferation, we conducted two RNAi-based loss-of-function screens. We engineered DLD1 colon cancer cells, which harbor APC deletions and depend on β-catenin for proliferation 5 , to stably express "TOPFLASH" β-catenin-luciferase and "FOPFLASH" mutant-Renilla reporter constructs 6,7 (DLD1 Rep ). Suppression of β-catenin expression in DLD1 Rep cells by three β-catenin-specific short hairpin RNAs (shRNA) markedly reduced the TOPFLASH/FOPFLASH ratio (Fig. 1a), confirming that reporter activity requires β-catenin expression. We then screened DLD1 Rep cells with a shRNA library containing 4849 shRNAs that target 1000 genes, including 95% of the human kinome 6 . We found 34 genes whose expression was necessary for β-catenin activity, including two known β-catenin regulators, CSNK1G3 8 and CSNK1E 9 ( Fig. 1b and Supplementary Table 1).In parallel, we performed an arrayed, kinase-enriched shRNA screen in another β-catenindependent colon cancer cell line, HCT116, to identify genes essential for cancer cell proliferation. We identified 166 candidate genes necessary for proliferatio...

Alterations in the Wnt/b-catenin pathway define a key event in the pathogenesis of colon cancer. We have recently shown that CDK8, the gene encoding a cyclin-dependent kinase (CDK) component of the Mediator complex, acts as a colon cancer oncogene that is necessary for b-catenin activity. Here, we tested the hypothesis that colorectal cancers with CDK8 expression have distinct clinical, prognostic and molecular attributes. Among 470 colorectal cancers identified in 2 prospective cohort studies, CDK8 expression was detected in 329 (70%) tumors by immunohistochemistry. Cox proportional hazards model and backward stepwise elimination were used to compute hazard ratio (HR) of deaths according to CDK8 status, initially adjusted for various patient and molecular features, including b-catenin, p53, p21, p27 (CDK inhibitors), cyclin D1, fatty acid synthase (FASN), cyclooxygenase-2 (COX-2), microsatellite instability (MSI), CpG island methylator phenotype (CIMP), LINE-1 methylation, and mutations in KRAS, BRAF and PIK3CA. CDK8 expression in colorectal cancer was independently associated with b-catenin activation (p 5 0.0002), female gender (p < 0.0001) and FASN overexpression (p 5 0.0003). Among colon cancer patients, CDK8 expression significantly increased colon cancer-specific mortality in both univariate analysis [HR 1.70; 95% confidence interval (CI), 1.03-2.83; p 5 0.039] and multivariate analysis (adjusted HR 2.05; 95% CI, 1.18-3.56; p 5 0.011) that was adjusted for potential confounders including b-catenin, COX-2, FASN, LINE-1 hypomethylation, CIMP and MSI. CDK8 expression was unrelated with clinical outcome among rectal cancer patients. These data support a potential link between CDK8 and b-catenin, and suggest that CDK8 may identify a subset of colon cancer patients with a poor prognosis.

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A 44-year-old man is presented here with 14 years of chronic purulent sinusitis, a chronic fungal rash of the scrotum, and chronic pelvic pain. Treatment with antifungal therapy resulted in symptom improvement, however he was unable to establish an effective long-term treatment regimen, resulting in debilitating symptoms. He had undergone extensive work-up without identifying a clear underlying etiology, although Candida species were cultured from the prostatic fluid. 100 genes involved in the cellular immune response were sequenced and a missense mutation was identified in the Ras-binding domain of PI3Kγ. PI3Kγ is a crucial signaling element in leukotaxis and other leukocyte functions. We hypothesize that his mutation led to his chronic infections and pelvic pain.

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The Rabbit Listened is a children's picture book that demonstrates in few words and even fewer pages the healing power of unconditional support. The book tells the story of Taylor, a child with curly black hair wearing a striped teal and white onesie. Taylor builds a pyramidal tower from wooden blocks and looks on with pride. Out of nowhere, a swarm of black birds swoop down, crash into the tower, and reduce it to rubble. Taylor crouches in a corner, overwhelmed by the feelings of loss. A parade of animal friends approaches Taylor to offer counsel. The chicken clucks apologies and encourages Taylor to talk. The bear advises Taylor to growl and shout in anger. The elephant trumpets that it can fix and rebuild the tower. The hyena suggests laughing about it, the ostrich recommends pretending it did not happen. The kangaroo sees the rubble as a mess and proposes to throw it away. The snake hisses, advising Taylor to topple someone else's tower. Taylor does not feel like doing anything and the parade of counsel leaves with disdain. Taylor remains slouched in the corner. As Taylor languishes, a cotton-tailed rabbit hops closer and closer until Taylor feels the warmth of its fur. They initially sit in silence until Taylor invites the rabbit to stay. The rabbit listens as Taylor talks, shouts, remembers, laughs, hides under a box, throws everything away, and fantasizes about destroying someone else's prized creation. Undaunted, the rabbit settles in. Over time, Taylor dreams of building again, to which the rabbit pounces in excitement. This story conveys a lesson to those mourning a loss and to those comforting others experiencing a loss. This is done through the use of tender illustrations and simple language. The author's use of a unisex name, her depiction of a child of nebulous gender, and her use of an olive skin tone potentially belonging to various ethnicities is clever and inclusive. This depiction enables children and parents of all backgrounds to relate to Taylor and the experience of loss. The illustrations of Taylor and the rabbit provoke warm feelings within the reader, sparking a desire to nuzzle and cuddle with both. The author illustrates two approaches to comforting others and their impact on the bereaved. The counsel of animals instructs Taylor on how to feel and how to process the loss. Taylor oscillates between sadness and discomfort when hearing the advice. Like many bereaved persons, Taylor has neither the space nor the support to express true feelings.