BG & TMZ Therapy of Glioblastoma Multiforme

OBJECTIVES:

Primary

- To evaluate the feasibility of introducing and expressing P140K MGMT cDNA from a
lentiviral-based provirus in autologous hematopoietic stem cells harvested from
Glioblastoma multiforme (GBM) patients.

- To assess the safety associated with infusion of autologous hematopoietic stem cells
transduced ex vivo with a lentiviral vector expressing P140K MGMT in patients with GBM.

Secondary

- To determine whether any patients who receive P140K MGMT-transduced CD34 cells tolerate
O6-benzylguanine (BG) and dose-escalated temozolomide (TMZ) without myelosuppression.

- To evaluate the ability to detect P140K-transduced BG and TMZ-resistant hematopoietic
cells from the bone marrow and peripheral blood in patients infused with
P140K-transduced CD34 progenitors.

- To evaluate the feasibility of in vivo enrichment of P140K-expressing hematopoietic
cells by repeated treatments of BG and TMZ at doses that appear therapeutic for GBM.

- To evaluate the efficacy of various types of chemotherapy with or without radiotherapy
on conditioning the patient's bone marrow to host the transduced autologous hematopoetic
stem cells.

- To evaluate tumor response, progression-free survival, and overall survival.

OUTLINE: Patients are assigned to 1 of 3 treatment cohorts.

- Cohort 1 (LV P140K MGMT gene transfer after concurrent chemoradiotherapy): Patients
receive radiotherapy (60cGy in 30 2cGy daily doses) and TMZ 75mg/m2 /daily for 6 weeks,
cell infusion at week 7 (T0) followed by BG 120 mg/m2 intravenous infusion over 1h and
TMZ 50 mg/m2/day x 5 days, every 28 days (starting on T+28) for 6 cycles.

- Cohort 2 (LV P140K MGMT gene transfer prior to concurrent chemoradiotherapy): Patients
receive BG 120mg/m2 intravenous infusion over 1h and TMZ 400 mg/m2 one dose given on day
T-2 or T-3 days prior to cell infusion, followed within 72-96 hours by radiotherapy
(60cGy in 30 2cGy daily doses) and concurrent BG + TMZ at 50 mg/m2/day x 5 days, every
28 days,starting on T+28 for a total of 7 cycles of BG + TMZ.

- Cohort 3 (intra-patient dose escalation of TMZ in patients with evidence of P140K-marked
cells): Dose escalation of TMZ in patients with evidence of P140K marked cells in vivo
given as described above for cohort 1 or cohort 2. After completion of radiotherapy,
patients will receive BG + TMZ at 50 mg/m2/day x 5 days. Patients not experiencing any
grade 3 toxicity will be increased to the next TMZ dose level of 65 mg/m2/day x 5.
Subsequent dose escalation without grade 3 toxicity will be 80 mg/m2/day, 100 mg/m2/day,
120mg/m2/day and 140 mg/m2/day x 5. If at subsequent cycles a grade 3 or greater
hematologic toxicity occurs, the dose level for the next cycle will be reduced one
level.

Blood samples are collected periodically for replication-competent lentivirus detection and
other laboratory biomarker studies.

After completion of study therapy, patients are followed up every 2 months.

Inclusion Criteria:

- Patients with histologically confirmed, newly diagnosed, supratentorial GBM who have
undergone gross total tumor resections or near gross total resection (resection of
>90% of enhancing tumor demonstrated by MRI) are eligible up to their third
post-operative week. Patients with infratentorial disease, multifocal or
leptomeningeal disease will be excluded. In general, patients will not have > 1 cm
residual measurable or evaluable disease after surgical tumor resection.

- ECOG performance status 0-2 or Karnofsky ≥ 70.

- Patients must have received no myelosuppressive chemotherapy prior to the diagnosis of
GBM.

- Life expectancy of at least 12 weeks.

- Adequate hematologic (ANC ≥ 1,000/mm3, platelets ≥ 100,000/mm3, Hgb ≥ 9.5) , hepatic
(Bilirubin ≤ 2.0 mg/dl, AST and ALT less than or equal to 3 times upper limit of
normal, prothrombin time <1.2 times normal), and renal (Serum creatinine ≤ 2.0 mg/dl
or Creatinine Clearance ≥ 60mL/min/1.73 m2 for subjects with serum creatinine levels
above institutional normal) . These tests will be repeated within 2 weeks of treatment
with BG and TMZ, and must meet the same criteria.

- EKG without evidence of acute cardiac disease.

- Left ventricular ejection fraction (LVEF) ≥ 40

- Post-operative steroids are tapered to ≤ 24 mg decadron/d

- Patients of child-bearing potential must be using single barrier contraception

- Willingness and ability to provide informed consent.

- Patient must have all sutures removed prior to registration

- Patient must be considered to be clinically stable.

Exclusion criteria:

- Medical condition associated with immunosuppression, active infection or medical
illness which may jeopardize patient safety.

- HIV seropositivity. This exclusion is included for two reasons. First, there is
evidence of decreased marrow reserve in HIV+ patients and antiviral treatment is
associated with myelosuppression. Thus, drug treatment designed to be myelosuppressive
may be more toxic in this patient population. Second, extensive laboratory culturing
of the bone marrow and peripheral blood progenitor cells is required. No preclinical
samples which are HIV+ have been evaluated with the gene transfer modality proposed
and thus the feasibility and safety of gene transfer and selection in HIV+ samples
cannot yet be advocated. Such studies are planned so as to not preclude HIV+ patients
in later studies.

- Pregnant or lactating women. There is data to indicate that TMZ is teratogenic and
carcinogenic. Thus, its use in pregnant women would confer unnecessary risk to the
fetus.

- Patients with symptomatic pulmonary disease and other severe co-morbid conditions

- Patients with cardiac insufficiency and an LVEF of < 40%. History of acute coronary
event disease or arrhythmia within 6 months prior to enrollment

- Prior chemotherapy (including gliadel wafers) or hematopoietic cell transplantation.

- Inability to undergo repeated MRI evaluation.

- Prior diagnosis of malignant disease within a three year period with the exception of
surgically cured basal cell carcinoma or carcinoma in situ of the cervix

- Mental incapacity or psychiatric illness preventing informed consent