Research Interests

In work originally supported by the Defense Advanced Research Program Administration (DARPA), we are using the CRISPR Cas13d RNAse to specifically degrade viral and host RNA and CRISPRa, in which catalytically inactivated CAS proteins are fused to transcriptional activator proteins for targeting viral infection of respiratory tract epithelial cells. A major goal is for targeting epidemic/pandemic respiratory viruses, such as influenza A virus and SARS-CoV2 for both the prophylaxis and treatment of established infection.  

As part of developing in vivo delivery systems for these anti-viral studies, we discovered that some of these systems also deliver CRISPR Cas protein/guide RNA complexes or other protein cargo to T cells. With the support of the Naddisy Foundation Professorship of Pediatric Food Allergy, Immunology, and Asthma and the Sean M Parker Center for Allergy and Asthma Research, we are currently using this approach for the development of CRISPR-based approaches to target T helper 2 (Th2) immune responses. These immune responses play an essential role in the development of severe allergies to food and environmental allergens and the development of this type of therapy has potential application to many other diseases involving the immune system.  

A long-standing interest is to understand the cellular and molecular basis for this vulnerability of the human neonate to infection with intracellular pathogens that require T helper 1 (Th1) cells [CD4 T cell producing interferon-gamma (IFN-gamma)] for effective immune control. We have previously shown that CD4 T cells of the newborn have a unique limitation in the ability to transcribe mRNAs encoding certain effector molecules, such as CD40-ligand (CD154) and IFN-gamma compared to these cells in adults. These transcriptional differences are the result of both altered upstream signal transduction events in the T cells of the fetus and young infant as well as altered chromatin accessibility. Recent RNA-Seq studies indicate that these developmental differences are the result of  fetal and young infant  T cells having a unique transcriptional program that impacts hundreds of genes and multiple regulatory RNAs. This developmental transcriptional profile is likely important for the maintenance of fetal-maternal tolerance during pregnancy.

Among the many genes that are differentially expressed by CD4 T cells,  we have focused on defining the role of increased expression of protein tyrosine kinase 7 (PTK7) by fetal and neonatal CD4 T cells in immune function. We found that PTK7, which is a member of the protein tyrosine surface receptor family originally identified in colon cancer cells, is also expressed by subpopulations of CD4 T cells in later childhood and in the adult.  PTK7 surface expression by CD4 T cells in the child and adult can be used to identify recent thymic emigrant (RTE) CD4 T cells in the peripheral blood. We are now determining the biological importance of PTK7 expression by fetal and young infant CD4 T cells and RTEs in children and adults for regulation of T-cell activation and homeostasis using both RNAi and CRISPR-based approaches. 

In work supported by the Kruzn’ for a Kure Foundation and the Little Giants Foundation we are pursuing studies to understand the disease mechanisms and possible treatments for a Schimke Immuno-Osseous Dysplasia (SIOD). SIOD is a multisystem disease that has a unique disease phenotype that includes an intrathymic developmental T-cell defect with reduced T-cell IL-7 receptor surface expression, short stature due to impaired long bone growth, nephrotic syndrome due to focal segmental glomerulosclerosis of podocytes, and neurovascular disease that affects mainly the blood vessels supplying the brain. A major medical mystery is how biallelic mutations of the SMARCAL1 gene, which is the only cause of SIOD, leads to this unique phenotype. Extensive studies in tissue culture systems has revealed that the SMARCAL1 protein plays an important role in genomic DNA replication and telomere maintenance, but the linkage between impairments of these functions and the disease phenotypes of SIOD remains obscure. In collaboration with Matthew Porteus’ laboratory, we are currently using CRISPR-mediated homologous directed repair (HDR) approaches for genomic DNA editing to generate unique induced pluripotent stem cell (IPSC) and hematopoietic stem cell (HSC) isogenic lines involving the SMARCAL1 gene, which is biallelically mutated in SIOD. Thanks to a major recent additional gift from the Kruzn’ for a Kure foundation, we are now using these cell lines for studies of SIOD disease mechanisms, the development of markers for disease activity, and identifying potential treatments for SIOD. We are also using this CRISPR IPSC and HSC HDR-editing approach for the study of other inherited monogenic disorders that result in immunodeficiency and/or immunodysregulation.   

Finally, in collaboration with Drs. Alice Bertaina and Paul Grimm, we have been involved in translational and clinical studies in which children with SIOD have received sequential  hematopoietic stem cell and a kidney transplant from the same parent. This “haplo/haplo” transplantation approach results in long-term tolerance of the kidney allografts by the immune system, allowing the discontinuation of all immunosuppressive medications. These studies have also confirmed the importance of the SMARCAL1 protein in maintaining the telomere length of HSCs and their blood-cell derivatives. We are interested in defining the importance of short telomeres in other features of the SIOD disease phenotype and in how SMARCAL1 deficiency results in short telomere length. 

Sean N. Parker Center for Allergy  and Asthma Research at Stanford University

Naddisy Foundation

Kruzn' for a Kure Foundation

Little Giants Foundation

DARPA

NIH (National Institutes of Health)

Stanford Maternal & Child Health Research Institute (MCHRI)