Artemis is a chemical in all cells in the body that is essential for the normal development of the immune system and repairing damaged DNA. Artemis deficiency (AD) causes Severe Combined Immunodeficiency (SCID-A), a “bubble baby” syndrome associated with increased sensitivity to radiation and chemotherapy. SCID-A is hard to treat with a bone marrow stem cell (SC) transplant from another person due to rejection, reactions from the graft, and toxicity from high dose chemotherapy. Gene corrected (GC) patient’s SC will minimize the risks and cure SCID-A. Our objectives are to 1) Maximize engraftment of GC SC by opening marrow space without using high dose chemotherapy; 2) Assess harmful effects after gene correction of mouse and human SC by developing safety testing suitable for clinical trial use; and 3) Demonstrate that GC human SC from SCID-A patients correct the defective immunity in animal and cell models. Using an AD mouse model we will open marrow spaces by using a genetically engineered drug which targets SC (yr 1) ± an agent which blocks marrow SC attachment (yr 2), and find the minimal effective dose of chemotherapy (year 3). We will test for toxicity using several approaches (yrs 1-3). AD SC will be corrected with our lentiviral vector (AProArt) using clinical trial conditions and cultured on special cells that support SC growth into immune cells. Immunodeficient mice will be injected with GC human SC and human SC differentiation into immune cells evaluated (yrs 2-3).
Statement of Benefit to California:
Artemis deficient Severe Combined Immunodeficiency Disease (SCID-A) results in T-B-NK+ SCID with increased sensitivity to alkylator chemotherapy, and accounts for ~10% of all SCID patients. Athabascan-speaking Native Americans have a very high incidence of SCID-A (2/5000 births) and affected children from other states are sent to UCSF for curative treatment. California has been among the leading states in instituting newborn screening for SCID and UCSF is one of the main referring hospitals for these newly-diagnosed babies. In the first year of newborn screening, 8 babies with SCID were born in CA. Currently, the only cure for this otherwise fatal disease is an allogeneic hematopoietic stem cell (HSC) transplant in which high dose alkylator chemotherapy is often necessary to overcome graft rejection and open sufficient bone marrow niches to reconstitute both T and B cell immunity. Successful gene correction of SCID-A will eliminate the need for high dose alkylator chemotherapy and significantly reduce the mortality of HSC transplantation and cost of lengthy hospitalization and long term care for the late effects due to alkylator use in these newborn babies. The approaches that are developed in this project for successful gene therapy without using high dose chemotherapy will benefit all children in California (and elsewhere) with a variety of genetic diseases who may benefit from curative cellular therapy.
Year 1Children born with Severe Combined Immunodeficiency Disease (SCID) are unable to fight common infections and often die within the first 6 months of life. The most severe form of SCID is the result of an inherited deficiency of an essential chemical in the body called Artemis (SCID-A) without which T and B cells are unable to develop. Artemis is an essential component of a process which repairs DNA (the instructions in cells which make us who we are) when it is damaged or cut during normal metabolic activities. Because of this, children with SCID-A are also more sensitive to typical chemotherapeutic drugs used for bone marrow stem cell transplant (BMT). For these reasons, if the patient’s own blood stem cells could be corrected by inserting a normal human Artemis gene into their DNA and these cells put back into the patient’s bone marrow to grow and restore normal T and B lymphocyte function, many if not all of the problems associated with BMT would be eliminated. In order to prove that the viral vector that we have constructed for clinical use this past year, that contains the human gene for Artemis, can effectively insert the Artemis gene into mouse and human cells we are using a variety of assays. We have shown that the Artemis protein is made by the gene when Artemis-deficient skin cells are "infected" with this viral vector and that these cells which previously were susceptible to radiation are now similar to normal cells. We have also established that normal human stem cells can be transduced (or "infected") with a control(non-Artemis)vector and still mature into human T cells (in mice) and when cultured outside the body on a special "feeder" layer of cells that support maturation of stem cells into B cells. We plan to repeat these experiments with our "clinical grade" viral vector using normal and Artemis-deficient human and mouse stem cells. We are also studying ways that we can open up "space" in the bone marrow to permit large numbers of donor stem cells to live and grow. For a typical bone marrow transplant this is done by pre-treating the patient (recipient) with high doses of chemotherapy that destroy the patient's own stem cells and allow the donor stem cells to stay and grow in the bone marrow. Since patients (and mice) with Artemis deficiency are unable to tolerate these high doses of chemotherapy, we are studying approaches that don't require this pre-treatment. In early experiments we have demonstrated that a drug called anti-ckit (ACK2) that attacks bone marrow stem cells in severely immunodeficient mice, also works in Artemis-deficient mice and following transplantation of highly purified normal mouse stem cells, the ACK2-treated mice begin to generate both donor T and B lymphocytes that are absent in Artemis deficiency as early as 4 weeks post transplant. The ACK2 is not a chemotherapy drug and has no other toxicity than its affect on bone marrow cells. Now that we have the clinically comparable viral vector we are planning experiments with Artemis deficient donor stem cells that will be treated with the vector so that they are no longer Artemis-deficient. With ACK2 pre-treatment we expect that these cells will grow in the bone marrow of Artemis-deficient mice and restore normal immune function. There is a preparation of ACK2 that may be used in humans but this will require further study. Therefore, we plan in the coming year to evaluate three other possible agents that are currently available for use in humans and that may also be effective in opening "space" in the marrow and allow the viral vector-treated bone marrow stem cells to grow and mature.