Somatic gain-of-function mutations in members of the RAS subfamily of small guanosine triphosphatases are found in > 30% of all human cancers. We recently described a syndrome of chronic nonmalignant lymphadenopathy, splenomegaly, and autoimmunity associated with a mutation in NRAS affecting hematopoietic cells, and initially we classified the disease as a variant of the autoimmune lymphoproliferative syndrome. Here, we demonstrate that somatic mutations in the related KRAS gene can also be associated with a nonmalignant syndrome of autoimmunity and breakdown of leukocyte homeostasis. The activating KRAS mutation impaired cytokine withdrawal-induced T-cell apoptosis through the suppression of the proapoptotic protein BCL-2 interacting mediator of cell death and facilitated proliferation through p27 kip1 down-regulation. These defects could be corrected in vitro by mitogen-activated protein kinase/ extracellular signal-regulated kinase kinase 1 or phosphatidyl inositol-3 kinase inhibition. We suggest the use of the term RAS-associated autoimmune leukoproliferative disease to differentiate this disorder from autoimmune lymphoproliferative syndrome. (Blood. 2010;117(10): 2883-2886) IntroductionThe autoimmune lymphoproliferative syndrome (ALPS) is characterized by childhood onset chronic lymphadenopathy, splenomegaly, multilineage cytopenias secondary to sequestration and autoimmune destruction, and an increased risk of B-cell lymphoma. 1 Laboratory findings include polyclonal hypergammaglobulinemia and expansion of a unique population of circulating T-cell receptor ␣␤ ϩ B220 ϩ CD4 Ϫ CD8 Ϫ T (␣␤ ϩ -DNT) lymphocytes. 2,3 Most patients with ALPS harbor heterozygous autosomaldominant germline mutations in FAS, with somatic FAS mutations representing the second most common genetic cause. [4][5][6][7] Germline mutations in the genes encoding FAS ligand and caspase 10 have been identified in a small minority of patients. [8][9][10][11][12] In our cohort, approximately one-third of the patients with ALPS have an undetermined genetic basis. In addition, there is a group of genetically undetermined ALPS-like patients without ␣␤ ϩ -DNT cell elevation.We recently reported one person among these latter patients with a syndrome of lymphoproliferation, autoimmunity, and minimally increased ␣␤ ϩ -DNT cells caused by a somatic mutation in the NRAS gene, resulting in defective lymphocyte apoptosis. 13 Here, we demonstrate that somatic mutations in the homologous KRAS gene can also be associated with a syndrome consisting of autoimmune phenomena and dysregulated leukocyte homeostasis, with normal ␣␤ ϩ -DNT cells. The activating KRAS mutation, like the previously described NRAS mutation, impaired intrinsic T-cell apoptosis through the suppression of the proapoptotic protein BCL-2 interacting mediator of cell death (BIM) and facilitated cellular proliferation by repression of p27 kip1 . Methods Cells and treatmentsAll patients were studied at the National Institutes of Health (NIH) under protocols approved by the institutional review b...

The regenerative capacity of the central nervous system must be optimized to promote repair following traumatic brain injury (TBI) and may differ with the site and form of damage. Sonic hedgehog (Shh) maintains neural stem cells and promotes oligodendrogenesis. We examined whether Shh signaling contributes to neuroblast (doublecortin) or oligodendrocyte progenitor (neural/glial antigen 2 [NG2]) responses in two distinct TBI models. Shh-responsive cells were heritably labeled in vivo using Gli1-CreERT2;R26-YFP bitransgenic mice with tamoxifen administration on Days 2 and 3 post-TBI. Injury to the cerebral cortex was produced with mild controlled cortical impact. Yellow fluorescent protein (YFP) cells decreased in cortical lesions. Total YFP cells increased in the subventricular zone (SVZ), indicating Shh pathway activation in SVZ cells, including doublecortin-labeled neuroblasts. The alternate TBI model produced traumatic axonal injury in the corpus callosum. YFP cells decreased within the SVZ and were rarely double labeled as NG2 progenitors. NG2 progenitors increased in the cortex, with a similar pattern in the corpus callosum. To further test the potential of NG2 progenitors to respond through Shh signaling, Smoothened agonist was microinjected into the corpus callosum to activate Shh signaling. YFP cells and NG2 progenitors increased in the SVZ but were not double labeled. This result indicates that either direct Smoothened activation in NG2 progenitors does not signal through Gli1 or that Smoothened agonist acts indirectly to increase NG2 progenitors. Therefore, in all conditions, neuroblasts exhibited differential Shh pathway utilization compared with oligodendrocyte progenitors. Notably, cortical versus white matter damage from TBI produced opposite responses of Shh-activated cells within the SVZ.

Leukemia/lymphoma-related factor (LRF), a zinc-finger transcription factor encoded by Zbtb7a, is a protooncogene that regulates differentiation in diverse cell lineages, and in the CNS, its function is relatively unexplored. This study is the first to examine the role of LRF in CNS pathology. We first examined LRF expression in a murine viral model of spinal cord demyelination with clinically relevant lesion characteristics. LRF was rarely expressed in oligodendrocyte progenitors (OP) yet, was detected in nuclei of the majority of oligodendrocytes in healthy adult CNS and during remyelination. Plp/CreER T :Zbtb7a fl/fl mice were then used with cuprizone demyelination to determine the effect of LRF knockdown on oligodendrocyte repopulation and remyelination. Cuprizone was given for 6 weeks to demyelinate the corpus callosum. Tamoxifen was administered at 4, 5, or 6 weeks after the start of cuprizone. Tamoxifen-induced knockdown of LRF impaired remyelination during 3 or 6-week recovery periods after cuprizone. LRF knockdown earlier within the oligodendrocyte lineage using NG2CreER T :Zbtb7a fl/fl mice reduced myelination after 6 weeks of cuprizone.LRF knockdown from either the Plp/CreER T line or the NG2CreER T line did not significantly change OP or oligodendrocyte populations. In vitro promoter assays demonstrated the potential for LRF to regulate transcription of myelin-related genes and the notch target Hes5, which has been implicated in control of myelin formation and repair. In summary, in the oligodendrocyte lineage, LRF is expressed mainly in oligodendrocytes but is not required for oligodendrocyte repopulation of demyelinated lesions. Furthermore, LRF can modulate the extent of remyelination, potentially by contributing to interactions regulating transcription. K E Y W O R D Sdemyelination, cuprizone, notch, differentiation, oligodendrocyte progenitor

Autoimmune lymphoproliferative syndrome (ALPS) is a non-malignant disease characterized by early-onset chronic lymphadenopathy, splenomegaly, multilineage cytopenias, polyclonal hypergammaglobulinemia, expansion of circulating TCRαβ+B220+CD4-CD8- T (αβ+-DNT) lymphocytes, and an increased risk of B-cell lymphoma. Most ALPS patients have germline or somatic TNFRSF6 (FAS) mutations, and a small minority of patients have germline mutations in FAS ligand and caspase 10. We recently reported that an ALPS-like phenotype can be caused by a somatic NRAS mutation (G13D) resulting in the defective lymphocyte apoptosis. Here we demonstrate that somatic mutations in the related KRAS gene can also be associated with a non-malignant syndrome of autoimmunity and breakdown of leukocyte homeostasis. The activating KRAS mutation (G13C) impaired cytokine-withdrawal induced T cell apoptosis through the suppression of the pro-apoptotic protein BIM and facilitated proliferation through p27kip1 downregulation. These defects could be corrected by MEK1 or PI3K inhibition in vitro. We suggest the use of the term RAS-associated autoimmune leukoproliferative disease (RALD) to differentiate this disorder from ALPS. In the hematopoietic system, somatic KRAS and NRAS mutations are commonly observed in aggressive tumors such as multiple myeloma or juvenile myelomonocytic leukemia (JMML). From our reports, we think patients with somatic KRAS and NRAS mutations can follow a more benign clinical course.