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Stanford announces new Center for Definitive and Curative Medicine July 2017

To help accelerate development of the iPS-derived tissue regeneration product in our lab, Tony helped to co-found the Center for Definitve and Curative Medicine with Maria Grazia Roncarolo and Matthew Porteus--"one dose is a cure".  Stem cell and gene therapy hold enormous promise to cure conditions with well-defined genetic causes by engineering cells to treat disease or altering a patient’s personal DNA to “fix” an abnormality. To bring these new stem cell and gene therapies to their patients, Stanford Medicine has announced the opening of the Stanford Center for Definitive and Curative Medicine, a joint initiative of the Stanford University School of Medicine, Stanford Health Care and Stanford Children’s Health.

The center provides the organizational and physical infrastructure to support investigator-initiated clinical translational studies on stem cell and gene therapy from initial discovery through completion of clinical proof-of-concept studies. Stanford Medicine is in a unique position to develop the CDCM because of its outstanding expertise in disease pathophysiology, cell and stem cell biology, and an optimal and collaborative environment between the medical school and the hospitals.

New transcription factor inference model reveals initiation and maturation networks for stratified epidermal differentiation

Tissue development results from lineage-specific transcription factors (TF) programming a dynamic chromatin landscape through progressive cell fate transitions.  We interrogate the epigenomic landscape during epidermal differentiation and, in collaboration with Wing Wong's lab, create an inference network that ranks the coordinate effects of TF-accessible regulatory element-target gene expression triplets on lineage commitment. We discover two critical transition periods: surface ectoderm initiation and keratinocyte maturation, and identify TFAP2C and p63 as lineage initiation and maturation factors, respectively. Surprisingly, we find that TFAP2C, and not p63, is sufficient to initiate surface ectoderm differentiation, with TFAP2C-initiated progenitor cells capable of maturing into functional keratinocytes. Mechanistically, TFAP2C primes the surface ectoderm chromatin landscape and induces p63 expression and binding sites, thus allowing maturation factor p63 to positively auto-regulate its expression and close a subset of the TFAP2C-initiated early program. Our work provides a general framework to infer TF networks controlling chromatin transitions that will facilitate future regenerative medicine advances.

Tumors "Roll the Genetic Dice" to generate drug resistance

Genomic analysis by our group and others have revealed that BCCs are typically diploid and carry a high frequency of non-silent single nucleotide variants (SNVs) compared to other cutaneous and non-cutaneous tumors. Given their high mutational load, how these variants confer selective tumor growth without deleterious effects remains poorly understood. We previously identified and functionally validated nine SMO mutations that drive the majority of drug resistance in BCC through two distinct mechanisms that maintain HH signaling in the presence of drug: induction of constitutive activity or disruption of ligand binding (see below). However, SMO mutations with unclear function are frequently found across many HH and non-HH dependent cancers with drug-resistant BCCs bearing the highest rate of recurrent mutations at 66%. Using a sensitive functional assay, our results reveal a surprising frequency of neutral and inactivating SMO variants in our drug-resistant BCC tumor population. Our data supports a model where tumors have an inherent advantage to generate as many genetically diverse clones as possible to cultivate a competitive environment that forces clonal competition as a way to grow. This ability to “roll the genetic dice” creates many mutations in key genes like SMO that would have activating, neutral, or negative effects on the cell. However, a small percentage of clones fortunate enough to contain activating mutations would continue to divide and contribute to a larger fraction of the tumor mass.

Basal Cell Carcinomas in Mice Overexpressing Sonic Hedgehog Ligand

The Scott lab found that mutations in the PTCH1 tumor suppressor cause Basal cell nevus syndrome (Gorlin's), a disorder where patients develop hundreds of BCCs and other hedgehog-dependent cancers.  Gene regulatory relationships defined in the fruit fly suggest that SHH overproduction should mimic PTCH1 loss if SHH is sufficient to drive tumorigenesis.  We show that SHH overexpression is sufficient to induce BCCs in mice and reinforce the unique dependence of BCCs on the hedgehog pathway for growth. We went on, in collaboration with Paul Khavari, to show a similar finding in human skin using SHH-retroviral infected, regenerated human epidermis. Fan, Oro, Scott and Khavari Nat Med 3:788-92 1997.

Mtss1 / Src Kinase Signaling Cassette Regulates Ciliogenesis

The signal strength of development morphogens and oncogenes plays a critical role in both morphogenic patterning and tumor growth and resistance.  An emerging mediator of signal reception is the microtubule-based organelle, the primary cilium. The small 2 mm sensory structure provides directional signal specificity by concentrating receptor subtypes and downstream signaling cascades into a highly polarized structure.  The primary cilium provides critical regulation of the Hedgehog pathway, regulating the interactions of Patched, Smoothened, and Gli in a ligand-dependent manner. We have identified a novel Shh-induced cytoskeletal gene Missing-in-Metastasis (MTSS1 or MIM) and have shown MTSS1 is required for Shh signaling in BCC cells. MTSS1 localizes to the basal body of the primary cilium and regulates vesicle movement and ciliary length by antagonizes the activity of Src-family kinases and actin regulators like Cortactin.  MTSS1 overexpression can increase Gli1 activity.  We have identified MTSS1-interacting proteins and found that the polarity pathway including Atypical Protein Kinase C, also localizes to the basal body, and are currently investigating the role of aPKC and other MTSS1-interacting proteins in ciliary function and tumor progression.

Fly MTSS1 regulates directional migration through Src Kinase mediated endocytosis

Although directed cellular migration facilitates the coordinated movement of cells during development and repair, the mechanisms regulating such migration remain poorly understood. MTSS1 is a defining member of the inverse Bin/Amphiphysin/Rvs domain (I-BAR) subfamily of lipid binding, cytoskeletal regulators whose levels are altered in a number of cancers. We  provide the first genetic evidence that an I-BAR protein regulates directed cell migration in vivo. Drosophila MIM (MTSS1) (dmim) is involved in Drosophila border cell migration (see normal migration left below marked with moesin-GFP), with loss of dmim function resulting in a lack of directional movement by the border cell cluster (right, below). In vivo endocytosis assays combined with genetic analyses demonstrate that the dmim product regulates directed cell movement by inhibiting endocytosis and antagonizing the activities of the Src kinase/ CD2- associated protein/cortactin complex in these cells. These studies demonstrate that DMIM antagonizes pro-endocytic components to facilitate polarity and localized guidance cue sensing during directional cell migration.