- Email: [email protected]
Measuring success in clinical gene therapy research
MOLECULAR
M E D I C I N E T O D A Y , J U N E 1996
in an asynchronous, irregular, and incremental mechanism, euphemistically known as the scientific method. Ultimate success in clinical research is the elusive 'cure'. But in progress towards that goal, success is also | measured first as the 'absence of doing harm', and then by various stages of efficacy. Perhaps only in the case of smallpox has medicine achieved total 'victory'; it is now exactly 200 years since Jenner's first clinical trial. Recently, gene therapy has come under criticism either because it has not produced any cures so far, or because it is claimed to be more 'hype than hope'. Journalists might be excusedfor such 'rash judgments', but some of this bad press has come from researchers involved in genetic medicine itself1'2, The motivations of the latter are not entirely clear, but this is a highly competitive field where good reasons can mask the real ones. Their public pronouncements, however, serve to divide the scientific community and confuse the general public. Although we would like it to be the case, there is not one defined end point at which success can be declared for clinical research, just as for laboratory research. Rather, there are a series of points during the scientific process of discovery that leads to completion of a research program, and this will always be followed by longterm evaluation and monitoring of efficacy and effectiveness.The results of the early clinical trials in gene therapy have provided clear evidence of such incrementalsuccess,encouragingmanyother scientists throughout the academic community worldwide to press ahead with the development of their own clinical gene therapy experiments. I believe part of this contentious situation has arisen from the paradigm shift (or maybe shifts) that gene therapy represents. Within the field, the concept of gene therapy ~as moved from one or more techniques aimed at correcting congenital disorders by transforming hemopoietic stem cells, to many means of gene delivery, in a variety of cell types, for use in virtually any type of clinical dis:~34
order. This broadened view has led to greater opportunities for clinical experimentation, and the knowledge gained so far has been useful whether the clinical results are positive or indefinite. This is the nature of all pioneering medicine. For some people, the successes in gene therapy so far have not been the ones anticipated under the old paradigm, and have caused confusion and consternation among those people whose expectations have not evolved with the field 1. It took 20 years from the first attempt at gene therapyin humansto the first successfulexperiment in 1990. During that period of time, technological advancesled to several new gene-transfersystems, the mapping of hundreds of disease-associated genes, and improved methods for ex vivo cell growth. Therefore, once the first approved gene therapy clinical trial began in 1990, it had a reservoir of knowledge to draw upon. Since then, there has been an average of 22 trials approved per year with therapeutic intent in the USA alone3. The increasing number of trials is important because a large quantity of new data will be generated. Few trials are identical, yet together they probably raise expectations unrealistically in the scientific community and the general public, but this may be unavoidable in the face, say, of the rising mortali~, caused by cancer or the spread of AIDS. Predicting how quickly gene therapy will become part of medicine's standard armamentarium necessarily falls into the realm of speculation given the wide spectrum of potential applications, and is not the critical issue. The assessmentof the
successful transition of gene therapy into the clinic must be based on information gleaned through clinical experimentation- an incremental process. At this stage in the development of gene therapy, no other measures are appropriate. Not only are clinical gene therapy research programs already providing important information, but in 1996 we are completing phase II of a three-part process. A brief review of the history of the clinical development of gene therapy supports this conclusion.
Phase I (1970-1990) Stansfield Rogers, Senior Scientist in the Biology Division of the Oak Ridge National Laboratory, and colleagues in Germany attempted the first genetic therapy in 1970 (Ref. 4). They injected Shope papilloma virus into two children suffering from arginase deficiency, as the virus was known to synthesizearginase.The experimentwas based upon animal studies that suggested that the direct injection of the infectious virus could successfully deliver the gene encoding arginase and decrease plasma arginine levels. However, neither clinical benefit nor adverse reaction was observed. Two attempts were made at ex vivogene transfer in nemopoieticcells during the 1980s. The first, in 1980, employed a calcium phosphate precipitation method to transfer the ~-globin gene into the bone marrow stem cells of two children with thalassemias.This controversialexperimentdemonstrated the persistence of the transferred gene for up to nine months (using Southern blot detection), showing for the first time that the transfer of
Copyright ©1996 Elsevier Science Ltd. All rights reserved. 1357 - 4310/96/$15.00
PII: S1357-4310(96)20018-8
MOLECLILAR
MED~CINI
i 'IFODA'~', JLJNE
E,~ou~d
11996
recombinant DNA molec,Jlesinto humans can be safely achieved. This protocol was successful in altering the intended target-cell population and documenting the long-term survival of the genetically altered cells, confirmingthe results from prec~inicalanimal studies; but, as expected from the techniques used, no therapeuticbenefit was seen in either patient. Another gene transfer protocol using ex vivo manipulation of hemopoietic lineages was done at the National Institutes of Health(Bethesda,MD, USA)in 1989(Ref. 6). T cells derived from tumors were marked ex vivo using a murine retroviral vector, and returned intravenously to the patient. This experiment showed that a replication-incompetent recombinant virus could safely deliver a gene ex vivo, as predicted by animal studies. The only broad conclusion that could be drawn from these three trials, which took place over a period of 20 years, was that relative safety was achievable. The greatest concern regarding the transfer of genes into humans has been, and continues to be, its safety. Clinicaland laboratoryscientistsalike are very concerned about possible adverse reactions related to the transfer of viruses and recombinant DNA molecules into humans. Safety concerns led to a temporary hold on recombinantDNA research in the USA 20 years ago. As a result, the scientific community established the Recombinant DNA Advisory Committee (RAC) to develop guidelines and monitorrecombinantDNA activity.The fact that no adverse reactions were noted in the first three clinical studies of gene therapy (two of which had used viruses) was highly significant; especially in comparisonto the (in somewayscomparable)advent of allogeneic bone marrow transplantation, where safety issues limited progression for many years. However, these three trials were just the beginning. Large numbers of patients and gene transfer systems would need to be studied before firm conclusions could be drawn about the overall safety of gene therapy. None the less, the initial safe application of gene transfer into humans provided an ethical and scientificbasisfor the necessaryexpansion of treatment of larger numbers of patients before any attemptsat achievingclinicalefficacycould be hopedfor. In this way,the historyof gene therapy development as a scientific process is no different from that of other types of medicaltherapies, such as antibiotics and bone marrow transplantation. Phase II (1990-1996) g new phase in gene therapy research began in 1990 when the first gene therapy experiment with the possibility of producing a clinical benefit was initiated7. This trial revealed no adverse reactions to the geneticallycorrectedcells, persistence of gene-marked cells in vivo, the development of new immunologic functions and improved clinical status. With the successful regulatory approval,
implementationand the encouragingresults of this first trial, other investigatorswere moved to initiate other types of clinical trials for gene therapy. As further positive results followed, such as the marking of tumor cells in marrow transplants,persistent
~p
expression of the gene for the low-densitytipoprotein receptor in the liver and regression of tumor deposits in some patients, enthusiasm for clinical trials continuedto grow worldwide. By March 1996, more than 200 additionalclinicalgene markingand
Blaese, R.M. et al. (1995) T lymphocyte=~lireeted gene therapy for ADA-~CID: initia~ ~'iai results after 4 years, Science 270, 475-480 Bordignon, C. et aL (1995) Gene therapy in peripheral bmoodlymphocy~esand bone marrow for ADA-immunodefic[ent patients, Science 270, 470-475 Brenner, M.K. et aL (1993) Gone marking to determ~e whether autologous marrow hifmion restores tongoterm haematopoiesh in cancer patients, Lancet 342, 1134-1137 Brenner, M.K. et alp (1993) Gone=marking to trace origin of relapse after autologoas bonemarrow transplantation, Lancet 341, 85-86 Caplen, N.J. et al. (1995) Liposomeomediated CFTR gone transfer to the nasal epithelium of patienis with cystic fibrosis, Nat. Med. 1, 39--46 Cl3~stal,R.G. et al. (1994) Admimstration of an adenovirus containing the human CFTR cDNA to the respiratory tract of individuah with cystic fibrosis, Nat. Genet. 8, 42-51 Deisseroth, A.B. etal. (1994) Genetic marldng shows that Ph÷cells present in autologous transplants of chronic myelogenous leukemia (CI'~L) cont~bute to relapse after autologous bone marrow in CIV~, Blood 83, 3068-3076 Dunbar, C.E. et al. (1995) Re~'ovh'allymarked CD34-enrich~ peripheral bI~KIand bone mar~w cells con~bute to long-term engraftment after antologous transplantation, Blood 85, 3048-3057 Grossman, M. et al. (1994) Successful ex vivo gene therapy directed to liver in a patient with f a t a l hypercbolesterolemia, Nat. Genet. 6, 335-341 Grossman, M. etal. (1995) ApUot study ofex vivo gone therapy for homozygons familial hypercholesterolaenfia, Nat. Med. 1, 1148-1154 Hoogerbrugge, P.M. et aL (1996) Bone marrow gene transfer in three patients with adena~ine deanfinase deficiency, Gene Ther. 3, 179--183 Knowles, M.R. et al. (1995) A controli~ study of adenoviral-vector-mediated gone transfer in the nasal epithelium of patients with cystic fibrosis, New Engl. J. Med. 333,823-831 Kohn, D.B. et al. (1995) Engraftment of gene-modified umbilical cord blood ceRs in neonates with adenosine deaminase deficiency, Nat. Med. 1, 1017-1023 McElvaney, N.G. and Crystal, R.G. (1995) IL-6 release and airway administration of human CFTR eDNA adenovirus vector, Nat. Med. 1,182-184 Merrouche,Y. et al. (I 995) Cliulcai application of retrovh'algone transfer in oncoiogy: results of a French study with tumor infiltrating lympbocytes transduced with the gone of resh~mce to neomycin, J. Clin. OncoL 13, 410-418 Nabel, E.G. et al. (1994) Safety and toxicity of catheter delivery to the pulmonary vascnlature in a patient with metastatic melanoma, Hum. Gene Thor. 5, 1089-1094 Nabel, G.J. et al. (I993) Direct gone transfer with DNA-liposome complexes in melanoma: expression, biologic activity,and lack of toxicity in bomam, Proc. NatlAcad. Sci. USA 90,11307-11311 Riddell, S.R. et al. (1996) T-cell mediated rejection of gone-modified HIV-specific cytotoxic T lymphocytes in HIV.infected patients, Nat. Med. 2, 216-223 Rill, D.R. et al. (1994) Direct demonstration that autoingous bone marrow transplantation for solid tumors can return a multiplicity of tumorigenic cells, Blood 84, 380-383 Rooney, C.M. et al. (1995) Use of gone-modified virus.specific T.|ympbocytes to control Epstein-Burr virus.related lympboproliferafion, Lancet 345, 9-13 Sobol, R.E. et ai. (1995) Interleulfin-2 gone therapy in a patient with glioblastoma, Gene Ther. 2, 164-167 Woffendin,C. et al. (1996) Expression of a protective gone prolongs survival o f t cells in human immunodeficicocy vh'us.infected patients, Proc. NatlAcad. Sci. USA 93, 2889-2894 Zabner, J. et al. (1993) Adenovirus-mecfiated gone transfer transiently corrects the chloride transport defect in nasal epithelia of patients with cystic fibrosis, Cell 75, 207-216
D
235
iii!!!i
.... MOLECULAR
MEDICINE
J U N E 1996
TODAY,
Opinion therapy trials were approved worldwide. Clinical results have been published on 23 of them (see Box 1). As gene therapy comprises not one therapy, but many, some clinical protocols are only asking basicquestionsabout safety of the transferredgenes and the delivery system, while others are also attemptingto define the parameters for successful therapeuticapplicationof these moleculartherapies. The diversityof these investigationsshouldexpedite new understandings of the critical issues for ac~vancingthe application of gene therapy to patients. As more patients are being treated, the number of adverse reactions has remained very low, with none reported to be directly related to gene transfer itself. The major clinical side effects have been related to in vivo transfer of adenoviral vectors and retroviral-vector producer cells. Improvements in the delivery methods for each of these systems has markedly diminished the number and severity of the adverse reactions. Several trials have demonstrated some evidence of clinical benefit, despite the fact that they are Phase I toxicity trials. While we are all eager to see unequivocal evidence of sustained clinical benefit, this will require the combined talent and cooperation of laboratory and clinical scientists. We are currently in a phase of development, encompassing a cycle between the laboratory and the clinic, that will produce the fundamental improvements required for a substantial advancement in gene therapy. As laboratory-based and clinical researchers continue to move products into the clinic, these experimental therapies will derive informationthat will move developmentback into the laboratory for further refinement before returning for clinical evaluation. The molecular redesign of adenoviral vectors is a good example of this symbiotic relationship between 'benchside' and 'bedside' research. Those who profess that the laboratory or the clinic alone can more quickly advance the science of gene therapy are arguing against the full tide of experience of medical research since Pasteur's time. Many of the critical factors relating to gene expression and gene delivery can only be adequately assessed in humans (as was the case for novel therapeutics such as antibiotics and novel procedures such as bone marrowtransplantation).Historyrecordsthat any new paradigmthat crashes the old ones will always produce critics that hope to prevent, retard or cushion the changes that new ideas representto them. This led the great physicist Max Planck to observe that 'An important scientific innovation rarely makes its way by gradually winning over and ~nverting its opponents...Whatdoes happenis that its opponents gradually die out and that the growing generation is familiar with the idea from the beginning'a. As those who understand pharmaceutical development will recognize, the early clinical results in gene therapy are a logical extension of the m
236
developmentalyears precedingtheir approval.Those individuals who received the first doses of penicillin and the first bone marrow transplants died because information such as the appropriate dosing, formulation and duration of therapy had yet to be learned. Gene therapists have already proved that the current gene delivery systems are safe and that evidenceof clinical benefit can be seen in some patients. This successful beginning has nearly completed the foundation for the next phase of clinical studies, in which directed therapies with refined delivery systems will try to achieve actual efficacy in new groups of patients who are more likely to respondto the therapy than those 'high-risk' patients enrolled in the earliest safety experiments.
Phase IH (1997-) Despite their differences in opinion and understanding, critics and proponents, scientists and the general public all generally agree that gene therapy holds enormous potentia/for improving human health. The question is when will the widespread clinical application and attendant benefit to humans come to fruition? Such realizationdepends upon substantial advances in knowledge about gene transfer and regulation. Furthermore, gene therapy depends heavily on other areas of clinical research. It is clear that the genotype of tumor cells can have a tremendous impact on the antitumor response to chemotherapeutic drugs9. Likewise,tumor genotype will also influence the magnitude of the antitumorefficacyof cancer gene therapeutics1°. Testingtumors to determine whether they are forming gap junctions between cells before selecting the gene for herpes simplex thymidine kinase as a gene therapeutic, for example, may be expected to facilitateclinically important antitumor responses. It would be extremely pessimistic to predict that, in conjunctionwith the further development of DNAbased diagnostics, the next five years would yield anything less than enormous advances in gene therapy systems and real benefits to patients. By no means is gene therapy the supreme answer to humar; illness. Preventing disease still stands foremost. Yet genetic medicine, with all its attendingideas such as molecular repairof damage, implantation of new traits, improved efficiency of gene transduction and control of gene expression, represents a new threshold in the potential that humans possess for health. Through that entrance pass all other aspirations and achievements.
clinical trials versus complete optimization in animal models; established an efficient regulatory system that ensures safety; and shown that gene therapy can be beneficial to patients. While gene therapy research has had undeniable success in its two decades of clinical history, doubt and faith continue to struggle with each other. Doubt might always have the upper hand because it is inherent in the scientific method (and perhaps cynicism is in fashion). But faith is also an active motivator for scientific endeavor. Max Pianck noted that point too: 'Anybody who has been seriously engaged in scientific work of any kind realizes that over the entrance to the gates of the temple of science are written the words: Ye must have faith. It is a quality which the scientist cannot dispense with'11. References 1 Friedman,T. (1996) Human gene therapy - an immature genie but certainly out of the botlJe, Nat. Med. 2, 144-147 2 Touchette,N. (1996)Gene therapy: not ready for prime time, Nat. Med. 2, 7-8 3 Culver, K.W. (1996) Gene Therapy: a Primer for Physicians, MaryAnn Liebert 4 Rogers,S. (1976)Reflections on issues posed by recombinant DHA molecule technology, II, Ann. New YorkAcad. Sci. 265, 66-70 5 Cline,M.J. (1985)Perspectives for gene therapy: inserting newgeneticinfo~ation into mammalian celia by physical techniques and vira~ vectors, PharmacoL Ther.29, 69-92 6 Rosenberg, S.A. et al. (1990) Gene transfer into humans - immunotherapy of patients with advancod me!anoma, using tumor-infiltrating iymphocytesmodifiedby retroviralgonetransduction, New Engl. J. Med. 323, 570-578
7 Blaese,R.M.,Culver,K.W.andAnderson,W.F.(1990) The ADA human gene therapy clinical protocol, Hum. Gene Ther. 1,331-362 8 Planck, M. (1936) The Philosophy of Physics, C.
Allen & Unwin 9 Wahl, A.F. etal. (1996) Loss of normal p53 function confers sensitization to taxol by increasing G2/M arrest and apoptosis, Nat. Med. 2, 72-79 10 Fick, J. et al. (1995) The extent of heterocellular communication mediated by gap junctions is predictiveof bystandertumor cytotoxicityin vitro, Proc. NatlAcad. Sci. USA 92, 11071-11075 11 Planck, M. (1932) Where is science going? W.W.
Norton & Co.
Concluding remarks There have been undeniable successes in gene therapy research over the past 20 years. We have already: redefined gene therapy as a potential treatment for both acquired and congenital disorders; developed gene transfer systems that are clearly far less dangerous than many standard therapies; redrawn the line on transition into
.
.
.
.
.
.
.
.
M E D I C I N E T O D A Y , J U N E 1996
in an asynchronous, irregular, and incremental mechanism, euphemistically known as the scientific method. Ultimate success in clinical research is the elusive 'cure'. But in progress towards that goal, success is also | measured first as the 'absence of doing harm', and then by various stages of efficacy. Perhaps only in the case of smallpox has medicine achieved total 'victory'; it is now exactly 200 years since Jenner's first clinical trial. Recently, gene therapy has come under criticism either because it has not produced any cures so far, or because it is claimed to be more 'hype than hope'. Journalists might be excusedfor such 'rash judgments', but some of this bad press has come from researchers involved in genetic medicine itself1'2, The motivations of the latter are not entirely clear, but this is a highly competitive field where good reasons can mask the real ones. Their public pronouncements, however, serve to divide the scientific community and confuse the general public. Although we would like it to be the case, there is not one defined end point at which success can be declared for clinical research, just as for laboratory research. Rather, there are a series of points during the scientific process of discovery that leads to completion of a research program, and this will always be followed by longterm evaluation and monitoring of efficacy and effectiveness.The results of the early clinical trials in gene therapy have provided clear evidence of such incrementalsuccess,encouragingmanyother scientists throughout the academic community worldwide to press ahead with the development of their own clinical gene therapy experiments. I believe part of this contentious situation has arisen from the paradigm shift (or maybe shifts) that gene therapy represents. Within the field, the concept of gene therapy ~as moved from one or more techniques aimed at correcting congenital disorders by transforming hemopoietic stem cells, to many means of gene delivery, in a variety of cell types, for use in virtually any type of clinical dis:~34
order. This broadened view has led to greater opportunities for clinical experimentation, and the knowledge gained so far has been useful whether the clinical results are positive or indefinite. This is the nature of all pioneering medicine. For some people, the successes in gene therapy so far have not been the ones anticipated under the old paradigm, and have caused confusion and consternation among those people whose expectations have not evolved with the field 1. It took 20 years from the first attempt at gene therapyin humansto the first successfulexperiment in 1990. During that period of time, technological advancesled to several new gene-transfersystems, the mapping of hundreds of disease-associated genes, and improved methods for ex vivo cell growth. Therefore, once the first approved gene therapy clinical trial began in 1990, it had a reservoir of knowledge to draw upon. Since then, there has been an average of 22 trials approved per year with therapeutic intent in the USA alone3. The increasing number of trials is important because a large quantity of new data will be generated. Few trials are identical, yet together they probably raise expectations unrealistically in the scientific community and the general public, but this may be unavoidable in the face, say, of the rising mortali~, caused by cancer or the spread of AIDS. Predicting how quickly gene therapy will become part of medicine's standard armamentarium necessarily falls into the realm of speculation given the wide spectrum of potential applications, and is not the critical issue. The assessmentof the
successful transition of gene therapy into the clinic must be based on information gleaned through clinical experimentation- an incremental process. At this stage in the development of gene therapy, no other measures are appropriate. Not only are clinical gene therapy research programs already providing important information, but in 1996 we are completing phase II of a three-part process. A brief review of the history of the clinical development of gene therapy supports this conclusion.
Phase I (1970-1990) Stansfield Rogers, Senior Scientist in the Biology Division of the Oak Ridge National Laboratory, and colleagues in Germany attempted the first genetic therapy in 1970 (Ref. 4). They injected Shope papilloma virus into two children suffering from arginase deficiency, as the virus was known to synthesizearginase.The experimentwas based upon animal studies that suggested that the direct injection of the infectious virus could successfully deliver the gene encoding arginase and decrease plasma arginine levels. However, neither clinical benefit nor adverse reaction was observed. Two attempts were made at ex vivogene transfer in nemopoieticcells during the 1980s. The first, in 1980, employed a calcium phosphate precipitation method to transfer the ~-globin gene into the bone marrow stem cells of two children with thalassemias.This controversialexperimentdemonstrated the persistence of the transferred gene for up to nine months (using Southern blot detection), showing for the first time that the transfer of
Copyright ©1996 Elsevier Science Ltd. All rights reserved. 1357 - 4310/96/$15.00
PII: S1357-4310(96)20018-8
MOLECLILAR
MED~CINI
i 'IFODA'~', JLJNE
E,~ou~d
11996
recombinant DNA molec,Jlesinto humans can be safely achieved. This protocol was successful in altering the intended target-cell population and documenting the long-term survival of the genetically altered cells, confirmingthe results from prec~inicalanimal studies; but, as expected from the techniques used, no therapeuticbenefit was seen in either patient. Another gene transfer protocol using ex vivo manipulation of hemopoietic lineages was done at the National Institutes of Health(Bethesda,MD, USA)in 1989(Ref. 6). T cells derived from tumors were marked ex vivo using a murine retroviral vector, and returned intravenously to the patient. This experiment showed that a replication-incompetent recombinant virus could safely deliver a gene ex vivo, as predicted by animal studies. The only broad conclusion that could be drawn from these three trials, which took place over a period of 20 years, was that relative safety was achievable. The greatest concern regarding the transfer of genes into humans has been, and continues to be, its safety. Clinicaland laboratoryscientistsalike are very concerned about possible adverse reactions related to the transfer of viruses and recombinant DNA molecules into humans. Safety concerns led to a temporary hold on recombinantDNA research in the USA 20 years ago. As a result, the scientific community established the Recombinant DNA Advisory Committee (RAC) to develop guidelines and monitorrecombinantDNA activity.The fact that no adverse reactions were noted in the first three clinical studies of gene therapy (two of which had used viruses) was highly significant; especially in comparisonto the (in somewayscomparable)advent of allogeneic bone marrow transplantation, where safety issues limited progression for many years. However, these three trials were just the beginning. Large numbers of patients and gene transfer systems would need to be studied before firm conclusions could be drawn about the overall safety of gene therapy. None the less, the initial safe application of gene transfer into humans provided an ethical and scientificbasisfor the necessaryexpansion of treatment of larger numbers of patients before any attemptsat achievingclinicalefficacycould be hopedfor. In this way,the historyof gene therapy development as a scientific process is no different from that of other types of medicaltherapies, such as antibiotics and bone marrow transplantation. Phase II (1990-1996) g new phase in gene therapy research began in 1990 when the first gene therapy experiment with the possibility of producing a clinical benefit was initiated7. This trial revealed no adverse reactions to the geneticallycorrectedcells, persistence of gene-marked cells in vivo, the development of new immunologic functions and improved clinical status. With the successful regulatory approval,
implementationand the encouragingresults of this first trial, other investigatorswere moved to initiate other types of clinical trials for gene therapy. As further positive results followed, such as the marking of tumor cells in marrow transplants,persistent
~p
expression of the gene for the low-densitytipoprotein receptor in the liver and regression of tumor deposits in some patients, enthusiasm for clinical trials continuedto grow worldwide. By March 1996, more than 200 additionalclinicalgene markingand
Blaese, R.M. et al. (1995) T lymphocyte=~lireeted gene therapy for ADA-~CID: initia~ ~'iai results after 4 years, Science 270, 475-480 Bordignon, C. et aL (1995) Gene therapy in peripheral bmoodlymphocy~esand bone marrow for ADA-immunodefic[ent patients, Science 270, 470-475 Brenner, M.K. et aL (1993) Gone marking to determ~e whether autologous marrow hifmion restores tongoterm haematopoiesh in cancer patients, Lancet 342, 1134-1137 Brenner, M.K. et alp (1993) Gone=marking to trace origin of relapse after autologoas bonemarrow transplantation, Lancet 341, 85-86 Caplen, N.J. et al. (1995) Liposomeomediated CFTR gone transfer to the nasal epithelium of patienis with cystic fibrosis, Nat. Med. 1, 39--46 Cl3~stal,R.G. et al. (1994) Admimstration of an adenovirus containing the human CFTR cDNA to the respiratory tract of individuah with cystic fibrosis, Nat. Genet. 8, 42-51 Deisseroth, A.B. etal. (1994) Genetic marldng shows that Ph÷cells present in autologous transplants of chronic myelogenous leukemia (CI'~L) cont~bute to relapse after autologous bone marrow in CIV~, Blood 83, 3068-3076 Dunbar, C.E. et al. (1995) Re~'ovh'allymarked CD34-enrich~ peripheral bI~KIand bone mar~w cells con~bute to long-term engraftment after antologous transplantation, Blood 85, 3048-3057 Grossman, M. et al. (1994) Successful ex vivo gene therapy directed to liver in a patient with f a t a l hypercbolesterolemia, Nat. Genet. 6, 335-341 Grossman, M. etal. (1995) ApUot study ofex vivo gone therapy for homozygons familial hypercholesterolaenfia, Nat. Med. 1, 1148-1154 Hoogerbrugge, P.M. et aL (1996) Bone marrow gene transfer in three patients with adena~ine deanfinase deficiency, Gene Ther. 3, 179--183 Knowles, M.R. et al. (1995) A controli~ study of adenoviral-vector-mediated gone transfer in the nasal epithelium of patients with cystic fibrosis, New Engl. J. Med. 333,823-831 Kohn, D.B. et al. (1995) Engraftment of gene-modified umbilical cord blood ceRs in neonates with adenosine deaminase deficiency, Nat. Med. 1, 1017-1023 McElvaney, N.G. and Crystal, R.G. (1995) IL-6 release and airway administration of human CFTR eDNA adenovirus vector, Nat. Med. 1,182-184 Merrouche,Y. et al. (I 995) Cliulcai application of retrovh'algone transfer in oncoiogy: results of a French study with tumor infiltrating lympbocytes transduced with the gone of resh~mce to neomycin, J. Clin. OncoL 13, 410-418 Nabel, E.G. et al. (1994) Safety and toxicity of catheter delivery to the pulmonary vascnlature in a patient with metastatic melanoma, Hum. Gene Thor. 5, 1089-1094 Nabel, G.J. et al. (I993) Direct gone transfer with DNA-liposome complexes in melanoma: expression, biologic activity,and lack of toxicity in bomam, Proc. NatlAcad. Sci. USA 90,11307-11311 Riddell, S.R. et al. (1996) T-cell mediated rejection of gone-modified HIV-specific cytotoxic T lymphocytes in HIV.infected patients, Nat. Med. 2, 216-223 Rill, D.R. et al. (1994) Direct demonstration that autoingous bone marrow transplantation for solid tumors can return a multiplicity of tumorigenic cells, Blood 84, 380-383 Rooney, C.M. et al. (1995) Use of gone-modified virus.specific T.|ympbocytes to control Epstein-Burr virus.related lympboproliferafion, Lancet 345, 9-13 Sobol, R.E. et ai. (1995) Interleulfin-2 gone therapy in a patient with glioblastoma, Gene Ther. 2, 164-167 Woffendin,C. et al. (1996) Expression of a protective gone prolongs survival o f t cells in human immunodeficicocy vh'us.infected patients, Proc. NatlAcad. Sci. USA 93, 2889-2894 Zabner, J. et al. (1993) Adenovirus-mecfiated gone transfer transiently corrects the chloride transport defect in nasal epithelia of patients with cystic fibrosis, Cell 75, 207-216
D
235
iii!!!i
.... MOLECULAR
MEDICINE
J U N E 1996
TODAY,
Opinion therapy trials were approved worldwide. Clinical results have been published on 23 of them (see Box 1). As gene therapy comprises not one therapy, but many, some clinical protocols are only asking basicquestionsabout safety of the transferredgenes and the delivery system, while others are also attemptingto define the parameters for successful therapeuticapplicationof these moleculartherapies. The diversityof these investigationsshouldexpedite new understandings of the critical issues for ac~vancingthe application of gene therapy to patients. As more patients are being treated, the number of adverse reactions has remained very low, with none reported to be directly related to gene transfer itself. The major clinical side effects have been related to in vivo transfer of adenoviral vectors and retroviral-vector producer cells. Improvements in the delivery methods for each of these systems has markedly diminished the number and severity of the adverse reactions. Several trials have demonstrated some evidence of clinical benefit, despite the fact that they are Phase I toxicity trials. While we are all eager to see unequivocal evidence of sustained clinical benefit, this will require the combined talent and cooperation of laboratory and clinical scientists. We are currently in a phase of development, encompassing a cycle between the laboratory and the clinic, that will produce the fundamental improvements required for a substantial advancement in gene therapy. As laboratory-based and clinical researchers continue to move products into the clinic, these experimental therapies will derive informationthat will move developmentback into the laboratory for further refinement before returning for clinical evaluation. The molecular redesign of adenoviral vectors is a good example of this symbiotic relationship between 'benchside' and 'bedside' research. Those who profess that the laboratory or the clinic alone can more quickly advance the science of gene therapy are arguing against the full tide of experience of medical research since Pasteur's time. Many of the critical factors relating to gene expression and gene delivery can only be adequately assessed in humans (as was the case for novel therapeutics such as antibiotics and novel procedures such as bone marrowtransplantation).Historyrecordsthat any new paradigmthat crashes the old ones will always produce critics that hope to prevent, retard or cushion the changes that new ideas representto them. This led the great physicist Max Planck to observe that 'An important scientific innovation rarely makes its way by gradually winning over and ~nverting its opponents...Whatdoes happenis that its opponents gradually die out and that the growing generation is familiar with the idea from the beginning'a. As those who understand pharmaceutical development will recognize, the early clinical results in gene therapy are a logical extension of the m
236
developmentalyears precedingtheir approval.Those individuals who received the first doses of penicillin and the first bone marrow transplants died because information such as the appropriate dosing, formulation and duration of therapy had yet to be learned. Gene therapists have already proved that the current gene delivery systems are safe and that evidenceof clinical benefit can be seen in some patients. This successful beginning has nearly completed the foundation for the next phase of clinical studies, in which directed therapies with refined delivery systems will try to achieve actual efficacy in new groups of patients who are more likely to respondto the therapy than those 'high-risk' patients enrolled in the earliest safety experiments.
Phase IH (1997-) Despite their differences in opinion and understanding, critics and proponents, scientists and the general public all generally agree that gene therapy holds enormous potentia/for improving human health. The question is when will the widespread clinical application and attendant benefit to humans come to fruition? Such realizationdepends upon substantial advances in knowledge about gene transfer and regulation. Furthermore, gene therapy depends heavily on other areas of clinical research. It is clear that the genotype of tumor cells can have a tremendous impact on the antitumor response to chemotherapeutic drugs9. Likewise,tumor genotype will also influence the magnitude of the antitumorefficacyof cancer gene therapeutics1°. Testingtumors to determine whether they are forming gap junctions between cells before selecting the gene for herpes simplex thymidine kinase as a gene therapeutic, for example, may be expected to facilitateclinically important antitumor responses. It would be extremely pessimistic to predict that, in conjunctionwith the further development of DNAbased diagnostics, the next five years would yield anything less than enormous advances in gene therapy systems and real benefits to patients. By no means is gene therapy the supreme answer to humar; illness. Preventing disease still stands foremost. Yet genetic medicine, with all its attendingideas such as molecular repairof damage, implantation of new traits, improved efficiency of gene transduction and control of gene expression, represents a new threshold in the potential that humans possess for health. Through that entrance pass all other aspirations and achievements.
clinical trials versus complete optimization in animal models; established an efficient regulatory system that ensures safety; and shown that gene therapy can be beneficial to patients. While gene therapy research has had undeniable success in its two decades of clinical history, doubt and faith continue to struggle with each other. Doubt might always have the upper hand because it is inherent in the scientific method (and perhaps cynicism is in fashion). But faith is also an active motivator for scientific endeavor. Max Pianck noted that point too: 'Anybody who has been seriously engaged in scientific work of any kind realizes that over the entrance to the gates of the temple of science are written the words: Ye must have faith. It is a quality which the scientist cannot dispense with'11. References 1 Friedman,T. (1996) Human gene therapy - an immature genie but certainly out of the botlJe, Nat. Med. 2, 144-147 2 Touchette,N. (1996)Gene therapy: not ready for prime time, Nat. Med. 2, 7-8 3 Culver, K.W. (1996) Gene Therapy: a Primer for Physicians, MaryAnn Liebert 4 Rogers,S. (1976)Reflections on issues posed by recombinant DHA molecule technology, II, Ann. New YorkAcad. Sci. 265, 66-70 5 Cline,M.J. (1985)Perspectives for gene therapy: inserting newgeneticinfo~ation into mammalian celia by physical techniques and vira~ vectors, PharmacoL Ther.29, 69-92 6 Rosenberg, S.A. et al. (1990) Gene transfer into humans - immunotherapy of patients with advancod me!anoma, using tumor-infiltrating iymphocytesmodifiedby retroviralgonetransduction, New Engl. J. Med. 323, 570-578
7 Blaese,R.M.,Culver,K.W.andAnderson,W.F.(1990) The ADA human gene therapy clinical protocol, Hum. Gene Ther. 1,331-362 8 Planck, M. (1936) The Philosophy of Physics, C.
Allen & Unwin 9 Wahl, A.F. etal. (1996) Loss of normal p53 function confers sensitization to taxol by increasing G2/M arrest and apoptosis, Nat. Med. 2, 72-79 10 Fick, J. et al. (1995) The extent of heterocellular communication mediated by gap junctions is predictiveof bystandertumor cytotoxicityin vitro, Proc. NatlAcad. Sci. USA 92, 11071-11075 11 Planck, M. (1932) Where is science going? W.W.
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Concluding remarks There have been undeniable successes in gene therapy research over the past 20 years. We have already: redefined gene therapy as a potential treatment for both acquired and congenital disorders; developed gene transfer systems that are clearly far less dangerous than many standard therapies; redrawn the line on transition into
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