Autologous Gene Therapy for Artemis-Deficient SCID

Children with SCID generally do not survive beyond the first year of life without definitive
treatment. The most effective current cure is hematopoietic stem cell transplant (HCT) with a
human leukocyte antigen (HLA) matched sibling. While a matched sibling HCT can successfully
treat ART-SCID, fewer than 20% of affected children have such a donor, and even when a
matched sibling donor is available there is often incomplete T and B cell immune
reconstitution. ART-SCID is the most difficult type of SCID to cure by hematopoietic stem
cell transplant using alternative donors. Engraftment typically requires intensive
conditioning with high dose alkylating agents to prevent rejection and to open marrow niches.
These patients also have a high risk of developing graft versus host disease (GVHD) when
alternative donors are used. The great majority of patients have absent B cell reconstitution
and require lifelong administration of immunoglobulin infusions. Patients with ART-SCID who
do receive high doses of alkylators, especially when 2 agents are used, have poorer survival,
abnormal dental development, endocrinopathies, and short stature in comparison with children
exposed to no or limited alkylators or children with SCID types that are not associated with
a DNA repair defect. For these reasons, a safer, more effective approach to curing ART-SCID
is needed. Autologous gene-corrected hematopoeitic stem cell transplant may eliminate both
the risk of GVHD and the need for alkylators to prevent rejection.

The study design is a single-cohort, longitudinal experiment using non-randomized patients
treated once with a lentiviral vector for gene-correction of Artemis-deficient SCID after
conditioning with low-dose busulfan. No formal control group is planned for gauging safety;
rather, intensive monitoring of the initial 6 enrollees will preclude continued accrual in
the presence of safety signals, and long-term safety will be monitored for 15 years. Bone
marrow stem cells will be harvested from participants who weigh ≤7.5 kilograms or have failed
cytokine mobilization previously, and cytokine-mobilized peripheral blood stem cells will be
harvested from participants weighing >7.5 kilograms. CD34 cells will be isolated using the
CliniMACS® CD34 Reagent System cell sorter device. After a back-up untransduced cell graft
has been cryopreserved, the remaining cells will be transduced with the AProArt lentiviral
vector. These transduced cells will then be cryopreserved, and aliquots of the cells will
undergo safety testing and be reserved for potency evaluation. All patients will receive
busulfan conditioning targeted over 2 days to achieve a cumulative area under the curve (AUC)
of 20 mg*hr/L (an ablative cumulative AUC is 60-90mg*hr/L). Following the infusion of
AProArt-transduced cells, patients will be evaluated at 2, 4, 6, 8, 16, and 24 weeks for
evidence of gene transduced peripheral blood mononuclear cells and when possible cell
lineages including T, B, NK and granulocyte/myeloid cells. If there is no evidence of gene
transduced cells at 6 weeks (42 days) post infusion, a decision will be made regarding
further therapy.

After day 42 post-transplant, recipients will be followed for toxicity and durable
reconstitution of T and B cell immunity. Immune reconstitution of T cells will be monitored
on a regular basis. If the absolute neutrophil count is < 200/µl or platelets < 20,000/µl on
3 independent determinations after day 42 post infusion of transduced cells, the patient may
receive infusion of the back-up cells or an allogeneic hematopoeitic stem cell transplant.
Patients who were neutropenic prior to conditioning (SCID-related neutropenia) but responsive
to granulocyte-colony stimulating factor (GCSF) will not be considered to have failed,
provided the absolute neutrophil count can be maintained above >500/µl with GCSF.

After day 42, patients will be assessed weekly through 16 weeks post-transplant, monthly
through month 6 post-transplant, and then 3 monthly through month 12. They will then be
assessed at 6 monthly intervals during years 2-5 and annually through year 15. Study
follow-up will include completion of Quality of Life questionnaires and administration of
neurodevelopmental testing.

An independent Data Safety Monitoring Board (DSMB) will be appointed for safety monitoring of
this trial. The DSMB will review all data for safety on a regular schedule, based on numbers
of enrolled subjects and will also conduct special urgent review of any protocol related
Serious Adverse Events (SAE). As the trial is initiated, the DSMB will review results of each
of the first 3 cases prior to proceeding with subsequent patients.

Inclusion Criteria:

- ≥2.0 months of age at initiation of busulfan conditioning

- Diagnosis of typical or leaky ART-SCID:

Newly diagnosed ART-SCID patients must have:

- Artemis deficiency; AND

- CD3 count < 300 autologous cells/µL (typical ART-SCID) OR spontaneous maternal
chimerism, OR CD3 count >300/µL but with restricted T cell receptor Vb diversity,
defined as 18/24 or fewer polyclonal families.

AND - CD45 cell response to mitogens (PHA) < 50% of the lower limit of normal range for the
lab (leaky ART-SCID).

Patients diagnosed with ART-SCID per the criteria above who have failed an allogeneic
transplant (including an HLA matched sibling transplant) may participate if they meet the
criteria below:

- Are at least 3 months post allogeneic hematopoeitic stem cell transplant without evidence
of engraftment of allogeneic donor cells (excluding maternal cells)

OR are engrafted but have at least 2 of the following 4 conditions:

- Declining CD3 donor chimerism with at least 3 evaluations separated by at least 1
month prior to time of enrollment OR < 5% overall donor chimerism in blood and marrow
at ≥3 months post transplant.

- Incompletely reconstituted T cell immunity at ≥6 months (1 of the following 2):

- CD4 < 200/μL AND CD45 cell PHA < 50% of the lower limit of normal for lab;

- CD4 CD45RA < 20% of total CD4 cells OR T cell receptor Vb diversity is
restricted, defined as 18/24 or fewer polyclonal families.

- No donor B cells OR lack of B cell function (immunoglobulin M isohemagglutinins <
1:8 (not blood type AB) AND immunoglobulin A (IgA) or IgM values below reference
range for age AND if not receiving intravenous immunoglobulin (IVIG), no
protective level of antibody to tetanus immunization x2).

- Clinical manifestations consistent with persistent T and B cell immunodeficiency
e.g., chronic infection including norovirus, cytomegalovirus, human herpes virus
6; OR acute or recurrent infection (e.g., PJP), bronchiectasis, chronic
sinusitis.

AND

- Have no prior exposure to high dose busulfan (≥10 mg/kg total dose or average
cumulative exposure of ≥40 mg*hr/L). If the total cumulative AUC including previous
busulfan exposure plus the dose to be administered in this protocol is predicted to be
≤60 mg*hr/L, then patient would be eligible providing other criteria are satisfied.

- No medically eligible HLA-identical sibling with a normal immune system who could
serve as an allogeneic bone marrow donor (applies to newly diagnosed patients only).

Written informed consent according to guidelines of the Institutional Review Board (IRB).

Exclusion Criteria:

- Liver function tests (aspartate aminotransferase, alanine transaminase, gamma-glutamyl
transferase) > three times the upper limit of normal for lab and/or total bilirubin
>1.50 mg/dl at the time of planned initiation of busulfan conditioning.

- Prior history of veno-occlusive disease (Sinusoidal obstruction syndrome) of the
liver.

- Medically eligible HLA-matched sibling (applies to newly diagnosed patients only).

- Evidence of HIV infection by polymerase chain reaction or p24 antigen testing.

- Unable to tolerate general anesthesia and/or marrow harvest or peripheral blood stem
cell collection (apheresis) or insertion of central venous catheter.

- Presence of a medical condition indicating that survival is predicted to be less than
4 months, such as the requirement for mechanical ventilation, severe failure of a
major organ system, or evidence of a serious, progressive infection that is refractory
to medical therapy.

- Pregnancy

- A social situation indicating that the family may not be able to comply with protocol
procedures and recommended medical care and follow-up.

- Other conditions which in the opinion of the Principal Investigator and/or
co-investigators, contra-indicate the infusion of transduced cells or study
participation.