Clinical Trials of Copolymer I in
`Multiple Sclerosis
`
`M. B. BORNSTElN." A. l. MILLER." s. SLAGLE.‘
`R. ARNON.’ M. st-:LA,” AND D. TEITELBAUM”
`
`“The Albert Einstein College of Medicine
`Bronx. New York 10461
`
`” The Weizmann Institute
`Rehovot, Israel
`
`INTRODUCTION
`
`The understanding of the pathogenetic mechanisms involved in multiple sclerosis
`(MS) and the search for an efl'ective treatment have been intimately associated with
`the laboratory model. experimental allergic encephalomyelitis (EAE). The validity of
`EAE as such a model system has been clearly and convincingly presented by Paterson
`in a series of scholarly publications.” Our own work in tissue culture’ also served to
`relate MS. as a naturally occurring disorder, to its laboratory counterpart, EAE.
`Organotypic cultures of mammalian CNS tissue respond with identical patterns of
`demyelination,‘ swollen myelin sheaths.’ and eventual “sclerosis” when exposed to
`serum from EAE (whole white matter)-alfected animals and MS patients. The
`demyelinating elfect is not produced by these EAE sera on cultured peripheral nerve
`which, nevertheless, are responsive to serum from animals with experimental allergic
`neuritis.""' The cultures also demonstrated the capacity of mammalian CNS to
`remyelinate after being demyelinated by antisera.” These laboratory demonstrations
`provided further support for the extension to MS patients of therapeutic possibilities
`arising from animal studies.
`The synthetic polypeptide, copolymer I (COP l), was prepared from alanine.
`glutamic acid, lysine. and tyrosine (TABLE I) as one of a series of compounds which,
`alone or in combination with various lipids, might simulate the ability of myelin basic
`protein (MBP) to induce EAE (Sela. personal communication). None of the prepara-
`tions proved to be encephalitogenic,
`i.e.. capable of inducing EAE, but some.
`particularly COP I. did suppress EAE in animals challenged with either whole white
`matter or MBP in complete Freund‘s adjuvant. The numerous laboratory investiga-
`tions of the elfectiveness of COP l in EAB, involving mice. rats. guinea pigs. rabbits.
`monkeys. chimpanzees and baboons are of particular interest to the clinical trials and
`have recently been reviewed by Arnon and Teitelbaum." In addition, extensive
`laboratory studies failed to demonstrate any toxicological or other undesirable side
`reactions in experimental animals exposed to COP I under a variety of testing
`situations (A. Meshorer. personal communication). Finally, Abramsky et al." lirst
`examined COP I for its effect on three patients with acute disseminated encephalo-
`myelitis (ADE) and four with terminal MS. The three ADE patients recovered rapidly
`and completely. The MS patients may have demonstrated slight improvements. What
`is more important in these first clinical studies was the absence of any significant
`undesirable side reactions.
`This report presents the data derived from a preliminary trial of the efiectiveness of
`copolymer I in patients with the exacerbating-remitting (ER) and chronic progressive
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`(C P) types of MS. it will also describe the extensions of that study to pilot trials in each
`group of patients. their organizational aspects and present position.
`A preliminary trial is
`
`. conducted for the purpose of establishing dosages. studying toxicity. and obtaining a
`.
`.
`lead as to the possible efficacy of a new treatment which may be a new investigative
`drug .
`.
`. Different dosages with different schedules . . . are tried on a few patients who are
`very closely monitored for toxic reactions. For the assessment of therapeutic dosages. the
`patient with MS will serve as his own control. Therefore. the physician-investigator should
`be well acquainted with the medical history and past clinical course of MS in each
`patient .
`. . in most instances. it will not be necessary to involve more than perhaps l0
`patients in a given preliminary trial. If the preliminary trial brings forth evidence of
`therapeutic elficacy and little or no evidence of serious toxicity. it would be reasonable to
`move onto the next stage of investigation. the pilot trial."
`
`CONDUCT OF THE PRELIMINARY TRIAL
`
`Sixteen MS patients participated in the preliminary trial. They represented a
`broad spectrum of neurological
`involvement ranging from those of the chronic
`
`‘man: I. Composition of Copolymer 1'
`
`Amino Acid
`alanine
`glutamic acid
`lysine
`tyrosine
`
`N—Carboxyanhydride
`Used for Reaction
`alanine
`bcnzyl glutamate
`N-trifluoroaeetyl-lysine
`tyrosine
`
`‘Molecular weight: 23.000.
`
`Amount Used in
`the Reaction
`(mM)
`
`8.6
`6.0
`l4.0
`3.0
`
`75
`23
`52
`I4
`
`Molar Ratio of
`Amino Acid
`in Copolymer
`6.0
`1.9
`4.1
`1.0
`
`progressive type. some of whom were essentially bed or wheelchair bound. to those of
`the exacerbating-remitting type who were fully active and employed between attacks.
`There were four of the ER type and twelve of the CP type. All patients had been well
`known to the principal investigator (M BB) for years prior to their entry into the study.
`Some had participated as volunteers in earlier clinical and laboratory studies whereas
`others had been unsuccessfully tried on immunosuppressant therapy.
`The preliminary trial was conducted as an open study.“ All patients were given the
`COP l and all knew they were receiving it. The evaluating neurologist (AM) was also
`aware that this was an open study and that all patients were being treated. The initial
`dosage schedule was suggested by the group at the Weizmann Institute on the basis of
`their previous studies with laboratory animals.“ such as nonhuman primates. as well as
`the brief trial that was performed by Dr. Oded Abramsky." Thus, it was planned to
`prepare the COP I at a concentration of 5 mg per ml of sterile saline solution. This was
`to be given to each patient intramuscularly five times a week for the first three weeks.
`three times a week for the next three weeks. twice a week for the next three weeks. and.
`finally. once a week for the balance of a six-month period, at which time we originally
`planned to terminate the trial.
`At the time of introduction into the study, the first five patients were hospitalized
`at the General Clinical Research Center of the Albert Einstein College of Medicine.
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`They were examined and evaluated by Dr. Miller. samples of peripheral blood and
`cerebrospinal fluid were taken, and the copolymer injections were started. In the
`beginning, the patients were hospitalized during the first three weeks of treatment.
`because we had no knowledge as to whether or not there would be any significant local
`or systemic effects in patients who had multiple sclerosis. We did not. however. note
`any undesirable side reactions of any significance and soon found it unnecessary to
`keep the participants in the hospital for any period longer than was prudent following
`the lumbar puncture. usually 24 to 48 hours. The patients were seen, however, on an
`outpatient basis at
`the Clinical Research Center and their neurological status
`reevaluated at various times during the course of the following months.
`The specific aims of the preliminary trial were to determine the following: (I) Did
`COP I produce any apparent significant or undesirable side reactions? (2) Did COP I
`produce any apparent desirable effects? (3) Could a dosage schedule be established for
`further (pilot) trials should they appear to be warranted?
`
`RESULTS OF THE PRELIMINARY TRIAL
`
`During the institution of the copolymer treatment. many patients reported and. in
`fact, demonstrated early improvements in various neurological functions. As time went
`on, however, and the dosage of COP I was reduced as originally planned, these early
`improvements disappeared and most patients returned to their previous neurological
`status and continued their chronic progressive course. Over the period of the next
`months, the dosage was gradually increased in an effort to determine whether or not
`the previously observed effect was dose related. By the end of the first eighteen-month
`period. those patients who were still on the copolymer were receiving 20 mg a day in l
`ml of saline. 7 days a week. Three patients are still on this schedule, some 3-4 years
`later. This is the dosage currently being used in the pilot study.
`As for undesirable side reactions. patients occasionally reported transient slight
`pain, discomfort. itching, swelling or redness at the injection site. No systemic or
`general reactions of any kind were noted or reported during the preliminary trial.
`Examinations of urine were unremarkable. Of the sixteen patients, two of the ER type
`withdrew from the study at the time of an acute attack. One later returned. Of the
`balance. all remained in the study for at least six months as originally planned. Finally.
`three patients have been maintained to this date on 20 mg daily. In general. I I of the
`I6 patients demonstrated no apparent favorable effects in that they either had an
`exacerbation during the course of the study or continued their chronic progressive
`course. On the other hand. 5 of the 16 patients have demonstrated a definite change for
`the better. (See TABLE 2.)
`Nine patients received COP I for over two years. a few for over four years. No
`patient in this group has had any significant or undesirable local or systemic side
`reaction.
`
`LABORATORY DATA
`
`Laboratory examinations have included CBC. routine urinalysis and culture and
`blood chemistry analyses (SMA 6 and I2) VDRL. CSF protein and glucose and cells.
`Except for an occasional and transient eosinophilia (reaching l6% in one instance) no
`significant changes have been noted in any of these clinical tests. There has been no
`evidence of albuminuria or other evidence of altered kidney function. No pertinent
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`BORNSTIZIN er al.: CLINICAL TIIIAIS OF COPOLYMER I
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`369
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`alteration of the patient’s serum demyelinating potency on CNS cultured tissues has
`been observed. Several sera have been examined for antibody titers against COP I. In
`general. they have not been elevated. Lymphoblast transformation in response to
`phytohemagglutinin. MBP and COP I has not occurred.
`
`EXTENSION T0 PILOT TRIAIS
`
`The question now is whether or not the demonstrated improvements were. in fact.
`due to the polypeptide. As stated by Brown er al.,'’ the aim of a pilot trial is “to
`
`TABLE 2. Results of Preliminary Trial of Copolymer l Therapy in 16 Patients with
`Multiple Sclerosis
`
`Patient Type Age Sex
`
`CF‘
`IY
`CP
`RH
`CP
`GT
`ER‘
`PP
`CP
`AT
`PM CP
`
`JP
`
`ER
`
`JW CP
`KJ
`CP
`CN
`ER
`
`46
`25
`35
`30
`23
`39
`
`39
`
`F
`M
`F
`F
`M
`F
`
`F
`
`M
`32
`F
`33
`32 M
`
`Date of
`Entry
`
`4/25/78
`5/15/78
`5/30/78
`5/30/78
`6/27/78
`7/18/78
`
`7/l8/78
`
`6/27/78
`7/31 /78
`8/7/78
`
`Date of
`Termination
`
`Results
`
`no effect
`5/27/81
`no effect
`5/29/79
`no efiect
`9/20/79
`2/5/79 (6/83) no elfect
`2/8/79
`no effect
`—
`arrested (not terminated)
`marked improvement
`withdrew at time of exacerba-
`tion
`
`l0/27/78
`
`6/5/78
`12/30/80
`5/ l /82
`
`no effect
`no effect
`cessation of characteristic
`attacks
`
`CP
`WR
`SM CP
`HW CP
`SR
`CP
`FH
`ER
`
`49
`42
`36
`38
`27
`
`M
`F
`M
`F
`F
`
`arrest (slight improvement)
`no effect
`no eflect
`no eflect
`cessation of characteristic
`attacks (not terminated)
`arrest and improvement
`—
`I l/20/78
`F
`34
`JM CP
`(not terminated)
`
`10/ 3/78
`I0/I6/78
`10/24/78
`I0/24/78
`l l /‘I/78
`
`5/ I6/82
`9/20/82
`I I/I3/78
`I/28/80
`—
`
`‘Chronic progressive.
`‘Exaeerbating-remitting.
`
`determine whether a treatment that ‘looked good‘ in a preliminary trial still ‘looks
`good‘ when tested under more rigorously controlled conditions.“ For this purpose a
`double-blind. placebo-controlled. randomized pilot trial was started about three years
`ago. Since another purpose of a pilot trial is to determine whether or not a full clinical
`trial
`is justilied and.
`in addition. how it may be structured.
`the following brief
`description presents the current organization of the pilot trial.
`The clearly defined objectives of the pilot trial of the ER patient are (l) whether or
`not the frequency of attacks is diflerent between the COP I and the saline-injected
`groups. and (2) whether there is a difference in the degree of disability developed after
`two years of participation in the trial.
`The patient population of 50 ER MS patients was selected from 935 volunteers to
`have at least one and preferably two attacks a year in each of the two years prior to
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`entry into the trial. to be between the ages of 20 and 35. to be ambulatory. to be
`emotionally stable. and to have had no prior treatment with immunosuppressive drugs.
`The selected patients were matched for age. sex, frequency of attacks and degree of
`disability and randomly distributed by the statistician (SS) into the placebo or COP l
`groups. All information collected is sent to the Coordinating Center where it is edited.
`coded and stored in a data base specifically designed and constructed for this study.
`The organizational structure for the pilot study includes an External Advisory
`Board. Steering Committee, Coordinating Center and Clinical Center. The External
`Advisory Board. an outside independent group which serves the trial in an advisory
`and consultative capacity. includes Dr. John Kurtzlte, Professor of Neurology and
`Community Medicine. Georgetown University School of Medicine and chief of
`Neurology Service. Veterans Administration Medical Center; Dr. William Weiss,
`chief of Biometry and Field Studies at the National Institute of Neurological and
`Communicative Disorders and Stroke. NIH; and Dr. I. Herbert Scheinbcrg. chair-
`man of the Clinical Investigation Committee at Albert Einstein College of Medicine.
`This group receives periodic reports from the Coordinating Center that describe the
`results of the trial to date and keep the members informed as to the conduct of the
`trial.
`The Steering Committee is composed of Murray Bornstein. M.D.. principal
`investigator (who is blinded); Dr. Sylvia Wassertheil-Smoller. Ph.D.. head of the
`Division of Epidemiology and Biostatistics; and Ms. Susan Slagle, M.P.H.. biostatis-
`tion. It is their function to monitor the progress of the trial on an ongoing basis as well
`as to make decisions as to the design and conduct of the trial. At the Clinical Center.
`patients are examined routinely every three months and more frequently at the time of
`exacerbations. The investigating neurologists and the principal investigator. as well as
`the patients themselves. are unaware of the treatment assignments to placebo or COP
`I. This blinding of patients and examiners is maintained through the use of a
`Coordinating Center. which is a separate unit responsible for matching. randomiza-
`tion. procedures. assignment to treatment and data management.
`The pilot trial of the ER patients will end in December l984. The code will be
`broken, the data analyzed and the results published.
`A report summarizing the status of the ER trial as of 1 January I983 was
`submitted to the External Advisory Committee. An excerpt from this report that shows
`current enrollment status of the pilot trial is presented in TABLE 3. Based on the
`recommendations from the External Advisory Committee after reviewing the report of
`January I and the fact that there have been few significant undesirable side effects or
`reactions observed or reported to date. the ER pilot trial is now being extended to
`chronic progressive (CP) patients. For this purpose. 80 participants will beentered into
`the trial. varying from 25 to 55 years of age. The conduct of the rnulticenter CP pilot
`trial differs in some respects from the ER pilot trial.
`During a pretrial observation period, each CP patient must demonstrate a
`predetermined and sustained degree of worsening. specifically, at least two units in any
`one of Kurtzke‘s eight functional groups or one unit in any two unrelated functional
`groups. Once this is demonstrated and maintained. the patient becomes eligible to
`enter the trial and will then be randomly distributed into placebo or COP I groups. The
`dosage of COP I will be 15 mg in 0.75 of bacteriostatic saline subcutaneously. twice a
`day. The placebo consists of 0.75 ml of bactcriostatic saline. Patients will be evaluated
`at l. 4, 8. l2, I6, 20 and 24 months after admission to the trial. The evaluation will be
`the same as those listed for the current ER pilot trial. The trial will of course be
`conducted in a double-blind manner.
`The central statistic for the CP patients will be arrest of progression, which
`currently is defined as a change of not more than one unit in the eight functional groups
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`over the two-year period. Any degree of improvement will also be recorded and
`reported. but is not being used to determine the overall efiectiveness of COP l. The
`essential reasons for this limitation are both theoretical and practical. Theoretically.
`COP l is thought to interfere with the immunological attack on the nervous system by
`promoting the development of antigen-specific suppressor T cells. Whether or not these
`cells can eventually be demonstrated in human subjects. the point is that an arrest of
`clinical progression can be interpreted as an arrest of the immunological attack. A
`number of factors of structural. biochemical or bioelectrical nature, many of which are
`unknown. may contribute to the degree of clinically manifest neuronal dysfunctions.
`Moreover. they are not only unknown. but also uncontrolled in this trial. Thus.
`although improvements in function will be noted and reported, only arrest of
`progression (as stated) will be used as the central statistic to evaluate the eflectiveness
`of COP I in the CP patient.
`
`TABLE 3. Pilot Trial of COP I in ER Patients‘
`Placebo
`
`Number entered
`Average age
`Sex‘
`males
`females
`Race
`white
`black/hispanic
`Kurtzke DSS‘
`0-2
`3-4
`5-6
`mean DSS
`Previous attack rate’
`attacks/year
`Time in study
`total patient mos.
`total patient years
`average no. mos. in the study
`
`25
`30.96
`
`I0
`IS
`
`I3
`5
`7
`3.2
`
`2.02
`
`412
`34.33
`18.40
`
`COP I
`
`25
`29.96
`
`I I
`I4
`
`I3
`5
`7
`3.2
`
`I .96
`
`460
`38.33
`l6.48
`
`Total
`
`50
`30.44
`
`2|
`29
`
`48
`2
`
`26
`l0
`l4
`3.2
`
`I .99
`
`872
`72.66
`I144
`
`‘Baseline characteristics of the study population and the accumulated time in the study as of I
`January I983.
`‘Matching variables.
`
`Meanwhile. on the basis of current data. the Steering Committee with the advice of
`the External Advisory Committee is beginning to plan a full, multicenter—-probably
`internationaI—clinical trial of COP I in the ER type of MS patient.
`If COP I should prove to be efl'ective in arresting the progression of MS in a
`significant number of MS patients without evidence of unacceptable side reactions,
`two further possibilities may be considered.
`COP I may represent a protein component of CNS tissue acting as an encephali-
`togen. Evidence of the possible involvement of a lipid factor. galactocercbroside. as an
`antigen has been demonstrated." Proteolipid lipoprotein (PLP) may also be considered
`a candidate in this regard. A carbohydrate moiety. e.g., myelin-associated glycoprotein
`(MAG). may have similar properties. Consequently, if COP I proves to be ellective in
`a given proportion of patients, the addition of other antigens such as lipid (GC and
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`PLP) and carbohydrate (MAG) substances may prove even more effective in altering
`the course of MS.
`Finally. if the synthetic polypeptide. COP I. proves valuable in MS as a disease
`involving an autoallergic mechanism, one can suggest that other synthetic polypeptides
`should be examined for their potential therapeutic elfects in other diseases known or
`suspected to result from similar immunological factors.
`
`REFERENCE
`
`l.
`
`2.
`
`3.
`
`4.
`
`PATERSON. P. Y. 1977. Autoimmune neurologic disease: Experimental animal systems and
`implications for multiple sclerosis. In Autoimmunity. Genetic. Immunologic. Virologic
`and Clinical Aspects. N. Talmage. Ed.:6-13-692. Academic Press. New York.
`PATERSON. P. Y. I978. The demyelinating diseases: Clinical and experimental studies in
`animals and man. In Immunologic Diseases. 3d ed. M. Samter. N. Alexander. B. Rose.
`W. B. Sherman. D. W. Talmage & J. H. Vaughn. Eds.: l400—l43S. Little. Brown and
`Company. Boston.
`PATERSON. P. Y. 1979. Neurological disorders. In Medical Immunology. W. J. Irvine.
`Ed.: 36l—38l . Teviot Scientific Publications. Edinburgh.
`PATERSON. P. Y. I980. The imrnunopathology of experimental allergic encephalomyelitis.
`In The Suppression of Experimental Allergic Encephalomyelitis and Multiple Sclerosis.
`A. N. Davison & M. C. Cuzner, Eds.: ll-30. Academic Pres. New York.
`BORNSTEIN. M. B. 1963. A tissue culture approach to demyelinative disorders. Nat. Cancer
`Inst. Monogr. ll: l97—2l4.
`BORNSTEIN, M. B. I973. The immunopathology of demyelinative disorders examined in
`organotypic cultures of mammalian central nerve tissues. In Progress in Neuropathology.
`vol. 2. H. M. Zimmerman. Ed.: 69-90. Grune & Stratton. New York.
`BORNSTIEIN. M. B. & C. S. RAINE. I976. The initial structural lesion in serum: Induced
`demyelination in vilro. Lab. Invest. 35: 39l—40l.
`8. RAINE. C. S. & M. B. BORNSTEIN. I970. Experimental allergic encephalomyelitis. A light
`and electron microscope study of remyelination and ‘‘sclerosis'‘ in virro. J. Neuropath.
`Exp. Neurol. 29: 552-574.
`9. Boansrsm. M. B. 8: C. S. Rams. 1977. Multiple sclerosis and experimental allergic
`encephalomyelitis. Specific demyelination of CNS in culture. Neuropathology and
`Applied Neurobiology 3: 359-367.
`I0. RAINE. C. S. &. M. B. BORNSTEIN. I979. Experimental allergic neuritis ultrastructure of
`serum-induced myelin aberrations in PNS cultures. Lab. Invest. 40: 423-432.
`ll. ARNON. R. &. D. TEITELBAIJM. I980. Desensitization of experimental allergic encephalo-
`myelitis with synthetic peptide analogues. In The Suppression of Experimental Allergic
`Enccphalomyelitis and Multiple Sclerosis. A. N. Davison & M. L. Cuzner. Eds.: I05-
`I77. Academic Press. London, New York.
`I2. ABRAMSKY, 0., D. TEITELBAUM &. R. ARNON. I977. Ell'ect of a synthetic polypeptide
`(COP I) on patients with multiple sclerosis and with acute disseminated encepltalomyeIi-
`tis. Preliminary report. J. Neurol. Sci. 3|: 433-438.
`I3. Baown, J. R.. G. W. Baas:-:. J. F. KURTZKE, R. B. LOEWENSON, D. H. SILBERBERG &
`W. W. TOURTELLOTTE. I979. The design of clinical studies to assess therapeutic efllcacy
`in multiple sclerosis. Neurology 29(Part 2): 3-23.
`I4. Bolmsn-:IN. M. B.. A. I. MILLER, D. TEITELBAUM. R. Aanon 0. M. SELA. 1982. Treatment
`of multiple sclerosis with a synthetic polypeptide. Preliminary results. Ann. Neurol.
`ll: 3l7—3I9.
`I5. Fav. J. M.. S. WEISSBARTH. G. M. Lauasa at M. B. Boansrsm. I974. Cerebroside
`antibody inhibits sulfatide synthesis and myelination and demyelinates in cord tissue
`cultures. Science 183: 540-542.
`
`5.
`
`6.
`
`7.
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