Saturday, November 14, 2009



"Snail1[or Snai1] gene contributes to FGFR3 receptor signal transduction, which contributes to chondrodysplasias (achondroplasia (ACH), thanatophoric dysplasia (TD) and hypochondroplasia (HCH)). The exemplary embodiments identify Snail1 as a therapeutic and diagnostic target for chondrodysplasia, as well as the use of inhibitors thereof as drugs for the treatment of these diseases."

"Chondrocytes and osteoblasts have a mesenchymal origin, whereas osteoclasts arise from the haematopoietic lineage."

"the long-bone shortening phenotype [of being FGFR3 transgenic] is due to a disorganization and shortening of the proliferative chondrocyte columns and to delayed differentiation. Defects in the proliferative chondrocytes are due to the activation of Stat1, which is responsible for the induction of cell cycle inhibitor p21, and delayed differentiation is due to the activation of the MAPK signaling cascade, which causes a reduction in the area of hypertrophic chondrocytes, in both animal and human models. On the contrary, the de-activation of Fgfr3 in mice causes prolonged endochondral growth, resulting in a "long-bone" phenotype, which is accompanied by an expansion of the proliferative chondrocyte region in the growth plate. All these data give FGFR3 a significant role as a negative regulator of the proliferation of chondrocytes. This inhibition of proliferation by the FGF pathway is unique to chondrocytes and is mediated by transcription factor STAT1, which increases the expression of cell cycle inhibitor p21, the final agent responsible for the interruption of the proliferation induced by this signaling pathway. [p21] levels can be considered to be a reflection of the activation of the FGFR3-mediated signaling pathway in the growth plate. "

"Snail1 [is] a direct repressor of type II collagen, which is characteristic of proliferative chondrocytes, and disappears when the latter cease to proliferate and differentiate into hypertrophic chondrocytes, the cell population that expresses type X collagen. In a completely independent context, it was observed that the presence of Snail attenuated the proliferation of epithelial cells in culture and evolved with an increase in p21 levels and an increase in the phosphorylation of ERK1 and ERK2."

"The use of natiuretic peptide CNP inhibits the FGF-mediated MAPK activation in the growth plate"

"Snail1 is sufficient for FGFR3 signaling in the bone."

"Exemplary inhibitory compounds include, without limitation, antisense oligonucleotides described in, without limitation, US Patent Publication No. US20060003956 and Kajita et al., 2004 or siRNAs that inhibit the expression of Snail1 such as those described in without limitation, Peinado et al., 2005 and Tripathi et al., 2005 all of which are incorporated by reference herein. Additionally, any published nucleotide sequences or those published in the future that inhibit the expression of Snail1 are incorporated as embodiments herein, as potentially useful therapeutic compounds for the preparation of drugs designed to treat a chondrodysplasia process. Gene inhibition techniques and, more specifically, transport of compounds including, without limitation, antisense oligonucleotides, siRNA, ribozymes or aptamers--can be performed using, without limitation, liposomes, nanoparticles or other carriers that increase the success rate of the transfer to the interior of the cell, in one exemplary embodiment the cell nucleus (see, without limitation, Lu and Woodle, 2005 and Hawker and Wooley, 2005 which are incorporated by reference herein). In principle, Snail1 mRNA translation inhibitors can be used which bind both to the encoding region and/or the non-encoding region, for example, in front of the 3' non-encoding area. "

Compensatory regulation of the Snai1 and Snai2 genes during chondrogenesis.

"Several genes involved in cartilage and bone development have been identified as target genes for the Snail family of zinc finger transcriptional repressors, and a gain-of-function study has demonstrated that upregulation of Snai1 activity in mouse long bones caused a reduction in bone length. The Snai1 and Snai2[also known as slug] genes function redundantly during embryonic long bone development in mice. Deletion of the Snai2 gene, or limb bud-specific conditional deletion of the Snai1 gene, did not result in obvious defects in the skeleton. However, limb bud-specific Snai1 deletion on a Snai2 null genetic background resulted in substantial defects in the long bones of the limbs. Long bones of the Snai1/Snai2 double mutants exhibited defects in chondrocyte morphology and organization, inhibited trabecular bone formation and delayed ossification. Chondrocyte proliferation was markedly reduced, and transcript levels of genes encoding cell cycle regulators, such as p21(Waf1/Cip1) , were strikingly upregulated in the Snai1/Snai2 double mutants, suggesting that during chondrogenesis Snail family proteins act to control cell proliferation by mediating expression of cell cycle regulators. Snai2 transcript levels were increased in Snai1 mutant femurs, while Snai1 transcript levels were increased in Snai2 mutant femurs. In addition, in the mutant femurs the Snai1 and Snai2 genes compensated for each other's loss not only quantitatively, but also by expanding their expression into the other genes' normal expression domains. The Snai1 and Snai2 genes transcriptionally compensate temporally, spatially, and quantitatively for each other's loss, and demonstrate an essential role for Snail family genes during chondrogenesis in mice."

"mean bone length of femurs of the Snai1/Snai2 DM embryos was reduced approximately 20%, whether the measurements were taken of total bone length, or the length of trabecular bone"

"in Snai1/Snai2 DM growth plates the pattern of well-aligned columnar chondrocytes was disorganized, and chondrocyte morphology was altered. Proliferating chondrocytes in Snai1/Snai2 DM growth plates were more compact, and exhibited an elliptical shape with a higher chondrocyte to lacuna ratio"

"statistically significant increase in the length of the hypertrophic chondrocyte zone in Snai1/Snai2 DM growth plates"

"Snai1 and Snai2 genes are required to maintain the high rate of chondrocyte proliferation in the rapidly growing long bone"

"Cdkn1a RNA expression was approximately six-fold higher in Snai1/Snai2 DM femurs than in littermate controls"

"several markers were upregulated in Snai1/Snai2 DM femurs, including Col1a1, Col2a1, Col10a1, Sox9, and Acan"<-all these genes were upregulated by LSJL.

"genes encoding the matrix metalloproteases Mmp9 and Mmp13 were downregulated"

"Progression into chondrocyte prehypertropy and hypertrophy appeared to be delayed in Snai1/Snai2 DM growth plates. At E14.5, expression of both Col10a1, a hypertrophic chondrocyte marker, and Indian hedgehog (Ihh), a prehypertrophic chondrocyte marker, was markedly delayed in double mutant femurs"

"Blood vessel formation and penetration into the trabecular region of the femur was observed in both double mutant and controls"

"Despite the increase in Trp53 expression [in double mutants], we did not observe increased chondrocyte cell death in these embryos"

Snai1 mutant knockout mice had a slight non-signficiant increase in bone length.  At E16.5, Snai mutant knockout mice had a slight non-significant increase in Col10A1 and MMP9 levels.

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