H U M A N G E N E T H E R A P Y 1:327-329 (1990) Mary Ann Liebert, Inc., Publishers
The A D A
Human
Gene
Therapy Clinical Protocol
ORIGINAL COVERING MEMO Date: February 23, 1990 From: R. Michael Blaese, M.D., Chief, Cellular Immunology Section, N C I W . French Anderson, M.D., Chief, Molecular Hematology Branch, N H L B I Subject: Submission of A D A H u m a n Gene Therapy Clinical Protocol To: N I H IBC Attached is our human gene therapy clinical protocol that we are submitting for IRB/IBC/RAC/FDA approval. Title: Treatment of Severe Combined Immunodeficiency Disease (SCID) Due to Adenosine Deaminase ( A D A ) Deficiency with Autologous Lymphocytes Transduced with a H u m a n A D A Gene. Principal Investigator: R. Michael Blaese, M.D., Chief, Cellular Immunology Section (CIS), Metabolism Branch, N C I Associate Investigators: W . French Anderson, M.D., Chief, Molecular Hematology Branch, N H L B I Kenneth W . Culver, M.D., Medical Staff Fellow, CIS, Metabolism Branch, N C I Steven A. Rosenberg, M.D., Ph.D., Chief of Surgery, N C I /. Background A. Preclinical Data D o c u m e n t O n April 24, 1987, w e submitted a "Human Gene Therapy Preclinical Data Document" to the H u m a n Gene Therapy Subcommittee ofthe R A C in response to the February 25, 1986, m e m o of Dr. William J. Gartland (then Director of O R D A ) in which he requested that investigators supply preclinical data to the Subcommittee for "information purposes and part of an education process for the Subcommittee and submitters." In that document (copies of which are on file with O R D A ) w e proposed to treat adenosine deaminase ( A D A ) deficiency by inserting a normal human A D A gene into the bone marrow cells of infants suffering from severe A D A deficiency. Each ofthe questions raised in the "Points to Consider" document was addressed. The "protocol" was reviewed over a seven-month period by a dozen outside reviewers. O n December 7, 1987, a formal public meeting of the Subcommittee took place in which the protocol was evaluated. The conclusion was that bone marrow gene therapy was not ready to be used as a first line therapy for A D A deficiency but may have a place as a "last hope" procedure. The problems were threefold (see Preface, pp. 4-5, Preclinical Data Document):
1. The number of transduced cells obtained in monkey bone marrow transplant/gene transfer models was ver low and there was no evidence that totipotent stem cells, the optimal target cells, had been transduced. 327
A D A CLINICAL P R O T O C O L M E M O
2. Even though the ADA gene was expressed efficiently in mature ADA-deficient T cells in tissue cultur there was no evidence that the gene would remain active during the differentiation of the T-cell precursor in the treated bone marrow to the final mature T cell found in the periphery. 3. The A D A vector preparation available at that time was helper virus contaminated and there were only preliminary in vivo primate data which suggested that murine helper virus might not pose a public health risk.
These questions, together with the encouraging data being obtained with a new treatment modality (viz. P E G - A D A ) and the fact that the protocol called for the treatment of infants, led to the consensus view that additional work was required before an attempt at gene therapy for A D A deficiency was appropriate. B. In Vivo Non-Human Primate Safety Studies W e therefore sought to obtain substantial additional information in order to evaluate the safety issue relating to the use of retroviral-mediated gene transfer as a clinical procedure for humans. A series of in vivo studies in monkeys was carried out to determine if infectious murine amphotropic retroviruses might pose a public health danger. The studies indicated that primates appear to be resistant to murine retroviruses and, even when intentionally infected, appear to suffer no pathology. The data are reported in the attached manuscript "Amphotropic Murine Leukemia Retrovirus is Not an Acute Pathogen for Primates." Concurrently, vector/packaging line combinations were developed which could be used to obtain large quantities of helper-free retroviral vector supernatant. Thus, the safety issues that had been a source of concern in 1987 were significantly reduced by mid-1988. C. N2ITIL Human Gene Transfer Clinical Protocol W e sought to develop a clinical protocol that would avoid several ofthe issues that were of primary concern in 1987:
1. We wanted to choose a mature readily obtained target cell in order to avoid attempting to transduce elusive totipotent stem cell. 2. W e wanted to use the retroviral vector only as a marker in order to avoid the necessity of having efficient expression. 3. W e wanted to choose an adult patient population that could give informed consent.
Consequently, we established a collaboration with Dr. Steven A. Rosenberg, Chief, Surgery Branch, NCI, in which a retroviral vector (LNL6, which is a safety-modified version of N2) was used to mark tumor infiltrating lymphocytes (TIL) as a part of an adoptive immunotherapy protocol for the treatment of advanced cancer. That protocol was submitted on June 10, 1988, was extensively reviewed by the IRBs/IBC/RAC/FDA and was finally approved January 19, 1989. At present, six patients have received gene-marked cells. The results acquired to date (45 patient-months) have demonstrated no abnormalities, side effects, toxicities, or pathology due to the retroviral-mediated gene transfer procedure. Useful data are being obtained that should lead to improvements in TIL adoptive immunotherapy. //. ADA Human Gene Therapy Clinical Protocol A. Rationale Concurrently with the establishment of the N2/TIL human gene transfer clinical protocol, w e have been evaluating the possibility of using mature T lymphocytes as a primary target cell in the treatment of A D A
328
A D A CLINICAL P R O T O C O L M E M O deficiency. Our reasoning is developed in the accompanying Clinical Protocol but can be summarized as follows. There are a number of children (2-13 years old) with A D A deficiency who have some T-cell function but who are still at considerable risk for severe opportunistic infections. Prominent in this group of patients are children receiving regular enzyme replacement therapy with P E G - A D A . It should be possible to safely develop an A D A gene therapy protocol that could benefit these patients. Autologous T lymphocytes can be removed from the patient, a normal human A D A gene inserted, the cells grown up to a large number (exactly analogous to the growth of gene-marked TIL), and the ADA-corrected T lymphocytes returned to the patient. The patient can then be studied to determine if there is a beneficial result. This procedure can be carried out in several stages in order to minimize any potential risk. Initially a "Phase I" type of study would be done. Only a tiny number of gene-corrected cells would be given, and the patient studied for any ill effects. If none are found, then increasing numbers of cells would be given until a dosage is reached that should have a beneficial result. If functional ADA-corrected T lymphocytes are found in the patient's blood, then P E G - A D A can be carefully reduced to see if the autologous gene-corrected T cells can correct the patient's immune dysfunction on their own. B. Preclinical Studies W e have shown the following: 1. Autologous T lymphocytes have been isolated from ADA-deficient patients, a human A D A gene has been inserted, the ADA-corrected T cells have been grown to large numbers, and these gene-modified cells have been shown to express normal A D A levels and to successfully detoxify an in vitro deoxyadenosine challenge. 2. Autologous T lymphocytes have been isolated from a monkey, a human A D A gene has been inserted, the ADA-containing T cells have been grown to large numbers, and these gene-modified cells have been returned to the monkey. ADA-modified T cells were isolated from the monkey's blood four months after infusion. Other in vivo studies in monkeys have also demonstrated that T cells can be used successfully as a target cell for retroviral-mediated gene transfer. In addition, similar studies in mice suggest the same conclusion. Furthermore, all the data collected in the ongoing N2/TIL human gene transfer clinical protocol have demonstrated no untoward consequences of any kind related to the gene transfer procedure in man. All the safety features that were built into the N2/TIL human gene transfer clinical protocol can be used with equal effectiveness in this protocol. Specifically, the autologous T cells will be grown in vitro for a sufficiently long period to evaluate them for the presence of contaminating replication-competent virus, acquisition of autonomous growth, etc. ///. Conclusion W e feel that the accompanying Clinical Protocol should have a low level of risk (no higher than the N2/TIL protocol) and should provide valuable information that could lead to a successful treatment of A D A deficiency by means of gene therapy.
329
The A D A
Human
Gene
Therapy Clinical Protocol
ORIGINAL COVERING MEMO Date: February 23, 1990 From: R. Michael Blaese, M.D., Chief, Cellular Immunology Section, N C I W . French Anderson, M.D., Chief, Molecular Hematology Branch, N H L B I Subject: Submission of A D A H u m a n Gene Therapy Clinical Protocol To: N I H IBC Attached is our human gene therapy clinical protocol that we are submitting for IRB/IBC/RAC/FDA approval. Title: Treatment of Severe Combined Immunodeficiency Disease (SCID) Due to Adenosine Deaminase ( A D A ) Deficiency with Autologous Lymphocytes Transduced with a H u m a n A D A Gene. Principal Investigator: R. Michael Blaese, M.D., Chief, Cellular Immunology Section (CIS), Metabolism Branch, N C I Associate Investigators: W . French Anderson, M.D., Chief, Molecular Hematology Branch, N H L B I Kenneth W . Culver, M.D., Medical Staff Fellow, CIS, Metabolism Branch, N C I Steven A. Rosenberg, M.D., Ph.D., Chief of Surgery, N C I /. Background A. Preclinical Data D o c u m e n t O n April 24, 1987, w e submitted a "Human Gene Therapy Preclinical Data Document" to the H u m a n Gene Therapy Subcommittee ofthe R A C in response to the February 25, 1986, m e m o of Dr. William J. Gartland (then Director of O R D A ) in which he requested that investigators supply preclinical data to the Subcommittee for "information purposes and part of an education process for the Subcommittee and submitters." In that document (copies of which are on file with O R D A ) w e proposed to treat adenosine deaminase ( A D A ) deficiency by inserting a normal human A D A gene into the bone marrow cells of infants suffering from severe A D A deficiency. Each ofthe questions raised in the "Points to Consider" document was addressed. The "protocol" was reviewed over a seven-month period by a dozen outside reviewers. O n December 7, 1987, a formal public meeting of the Subcommittee took place in which the protocol was evaluated. The conclusion was that bone marrow gene therapy was not ready to be used as a first line therapy for A D A deficiency but may have a place as a "last hope" procedure. The problems were threefold (see Preface, pp. 4-5, Preclinical Data Document):
1. The number of transduced cells obtained in monkey bone marrow transplant/gene transfer models was ver low and there was no evidence that totipotent stem cells, the optimal target cells, had been transduced. 327
A D A CLINICAL P R O T O C O L M E M O
2. Even though the ADA gene was expressed efficiently in mature ADA-deficient T cells in tissue cultur there was no evidence that the gene would remain active during the differentiation of the T-cell precursor in the treated bone marrow to the final mature T cell found in the periphery. 3. The A D A vector preparation available at that time was helper virus contaminated and there were only preliminary in vivo primate data which suggested that murine helper virus might not pose a public health risk.
These questions, together with the encouraging data being obtained with a new treatment modality (viz. P E G - A D A ) and the fact that the protocol called for the treatment of infants, led to the consensus view that additional work was required before an attempt at gene therapy for A D A deficiency was appropriate. B. In Vivo Non-Human Primate Safety Studies W e therefore sought to obtain substantial additional information in order to evaluate the safety issue relating to the use of retroviral-mediated gene transfer as a clinical procedure for humans. A series of in vivo studies in monkeys was carried out to determine if infectious murine amphotropic retroviruses might pose a public health danger. The studies indicated that primates appear to be resistant to murine retroviruses and, even when intentionally infected, appear to suffer no pathology. The data are reported in the attached manuscript "Amphotropic Murine Leukemia Retrovirus is Not an Acute Pathogen for Primates." Concurrently, vector/packaging line combinations were developed which could be used to obtain large quantities of helper-free retroviral vector supernatant. Thus, the safety issues that had been a source of concern in 1987 were significantly reduced by mid-1988. C. N2ITIL Human Gene Transfer Clinical Protocol W e sought to develop a clinical protocol that would avoid several ofthe issues that were of primary concern in 1987:
1. We wanted to choose a mature readily obtained target cell in order to avoid attempting to transduce elusive totipotent stem cell. 2. W e wanted to use the retroviral vector only as a marker in order to avoid the necessity of having efficient expression. 3. W e wanted to choose an adult patient population that could give informed consent.
Consequently, we established a collaboration with Dr. Steven A. Rosenberg, Chief, Surgery Branch, NCI, in which a retroviral vector (LNL6, which is a safety-modified version of N2) was used to mark tumor infiltrating lymphocytes (TIL) as a part of an adoptive immunotherapy protocol for the treatment of advanced cancer. That protocol was submitted on June 10, 1988, was extensively reviewed by the IRBs/IBC/RAC/FDA and was finally approved January 19, 1989. At present, six patients have received gene-marked cells. The results acquired to date (45 patient-months) have demonstrated no abnormalities, side effects, toxicities, or pathology due to the retroviral-mediated gene transfer procedure. Useful data are being obtained that should lead to improvements in TIL adoptive immunotherapy. //. ADA Human Gene Therapy Clinical Protocol A. Rationale Concurrently with the establishment of the N2/TIL human gene transfer clinical protocol, w e have been evaluating the possibility of using mature T lymphocytes as a primary target cell in the treatment of A D A
328
A D A CLINICAL P R O T O C O L M E M O deficiency. Our reasoning is developed in the accompanying Clinical Protocol but can be summarized as follows. There are a number of children (2-13 years old) with A D A deficiency who have some T-cell function but who are still at considerable risk for severe opportunistic infections. Prominent in this group of patients are children receiving regular enzyme replacement therapy with P E G - A D A . It should be possible to safely develop an A D A gene therapy protocol that could benefit these patients. Autologous T lymphocytes can be removed from the patient, a normal human A D A gene inserted, the cells grown up to a large number (exactly analogous to the growth of gene-marked TIL), and the ADA-corrected T lymphocytes returned to the patient. The patient can then be studied to determine if there is a beneficial result. This procedure can be carried out in several stages in order to minimize any potential risk. Initially a "Phase I" type of study would be done. Only a tiny number of gene-corrected cells would be given, and the patient studied for any ill effects. If none are found, then increasing numbers of cells would be given until a dosage is reached that should have a beneficial result. If functional ADA-corrected T lymphocytes are found in the patient's blood, then P E G - A D A can be carefully reduced to see if the autologous gene-corrected T cells can correct the patient's immune dysfunction on their own. B. Preclinical Studies W e have shown the following: 1. Autologous T lymphocytes have been isolated from ADA-deficient patients, a human A D A gene has been inserted, the ADA-corrected T cells have been grown to large numbers, and these gene-modified cells have been shown to express normal A D A levels and to successfully detoxify an in vitro deoxyadenosine challenge. 2. Autologous T lymphocytes have been isolated from a monkey, a human A D A gene has been inserted, the ADA-containing T cells have been grown to large numbers, and these gene-modified cells have been returned to the monkey. ADA-modified T cells were isolated from the monkey's blood four months after infusion. Other in vivo studies in monkeys have also demonstrated that T cells can be used successfully as a target cell for retroviral-mediated gene transfer. In addition, similar studies in mice suggest the same conclusion. Furthermore, all the data collected in the ongoing N2/TIL human gene transfer clinical protocol have demonstrated no untoward consequences of any kind related to the gene transfer procedure in man. All the safety features that were built into the N2/TIL human gene transfer clinical protocol can be used with equal effectiveness in this protocol. Specifically, the autologous T cells will be grown in vitro for a sufficiently long period to evaluate them for the presence of contaminating replication-competent virus, acquisition of autonomous growth, etc. ///. Conclusion W e feel that the accompanying Clinical Protocol should have a low level of risk (no higher than the N2/TIL protocol) and should provide valuable information that could lead to a successful treatment of A D A deficiency by means of gene therapy.
329
