Current Eye Research

Suramin treatment suppresses induction of experimental autoimmune uveoretinitis (EAU) in rodents

Gil Sartani, Phyllis B. Silver, Gideon Strassmann', Chi-Chao Chan and Rachel R. Caspi

Laboratory of Immunology, National Eye Institute, NIH, Bethesda, MD and 'Otsuka America Pharmaceuticals, Inc., Rockville, MD, USA

Abstract The experimental drug suramin has been shown to possess several immunosuppressive properties. In this study we investi- gated the effect of suramin on the development of experimental autoimmune uveoretinitis (EAU) in mice and in rats. EAU was induced either by active immunization with a uveitogenic protein or peptide, or by the adoptive transfer of a uveitogenic T cell line. The development of EAU was assessed by clinical evaluation as well as by histopathology. Immunological responses were measured by delayed type hypersensitivity (DTH), lymphocyte proliferation, and serum antibody levels to the immunizing antigen. Suramin treatment was most effective in suppressing EAU when started concurrently with immunization (afferent). Treatment was less effective in sup- pressing disease when first administered 7 days after immuniza- tion or when given to animals that received an adoptive transfer of uveitogenic T cells (efferent). The effect of suramin on DTH and lymphocyte proliferation roughly paralleled its effect on EAU. Aferent treatment of mice with suramin completely suppressed anti-IRBP antibody titers. Interestingly, animals receiving efferent treatment had unreduced IgM levels but little or no IgG, suggesting prevention of the IgM-to-IgG switch. Depressed in vitro lymphocyte proliferative responses in animals treated with suramin during the afferent stage suggested that the suppressive effect on disease was due at least in part to an inhibition of antigen priming. Our results suggest that suramin merits further investigation as a potential treatment for some types of uveitis. Curr. Eye Res. 14: 887- 896, 1995.

Key words: autoimmune disease; experimental autoimmune uveoretinitis (EAU); mouse; rat; suramin; uveitis

Correspondence: Rachel R. Caspi, Laboratory of Immunology, National Eye Institute, NIH, Building 10, Room 10N218, 10 CENTER DR MSC 1858, Bethesda, MD 20892, USA

Introduction Experimental autoimmune uveoretinitis (EAU), induced in rodents by immunization with purified retinal proteins, serves as a model for several immune-mediated diseases that affect the posterior segment of the eye in humans (1, 2) . Some of these diseases are part of a generalized systemic syndrome ( e g Sarcoidosis, Behcet's disease and Vogt-Koyanagi-Harada syndrome), whereas others are confined primarily to the eye (e.g. sympathetic ophthalmia and birdshot retinochoroidopathy) (3). Various immunosuppressive and antiinflammatory treat- ments are used clinically to control uveitic diseases. However, there is no single treatment that is effective in all cases, and none of the treatments are free of side effects. Thus, there is a need to identify additional compounds that might constitute potentially useful treatments, either as standalone therapies or as adjuncts to other treatment modalities.

The mouse and rat models have been widely used as templates to evaluate the efficacy of novel treatment strategies. Although neither model by itself represents all the full spectrum of human uveitis, each can represent specific types of disease and each offers different advantages. In mice, the course of disease is variable, and can be controlled by the immunization dose. High doses of antigen tend to result in a more acute disease course, whereas low doses tend to result in a chronic- relapsing course (4). In rats, the disease is of the acute form irrespective of antigen dose, however, the availability of highly pathogenic T cell lines permits to evaluate treatment effects under conditions of 'clean' efferent-type disease, where already primed cells are introduced directly into the animal and the immunization stage is entirely avoided.

The experimental drug suramin, evaluated here, is a sulfated naphthylurea that was originally described as an antifilarial and antitrypanosomal agent (5). More recently, suramin was found to interfere with the binding of several growth factors, including IGF- 1, PDGF, EGF and TGF-P, to their correspond- ing cell surface receptors (68). Primarily for these reasons, suramin is currently being investigated in the clinic for its effects on several cancers that are refractory to conventional

Received on December 16, 1994; accepted on July 11, 1995

0 Oxford University Press

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.

888 G. Srrrtuni et al.

therapy. The inhibitory capacity of the drug is not restricted to growth factors. I t has recently been shown that suramin blocks the binding of several cytokines, including IFN-y, IL- I , 11,-2 and IL-6 to their corresponding cell surface receptors (9- 12). Furthermore, this receptor blocking activity may also operate in vivo; amelioration of experimental cancer cachexia by suramin has been attributed inhibition of the interaction of IL-6 with its receptor (9). Because a number of these cytokines have been implicated in uveitis (13) we decided to evaluate suramin as a potential treatment modality for EAU.

In this study we show that suramin treatment suppresses the development of EAU in mice and in rats, and present evidence that the effect on EAU can be attributed to inhibition of the induction stage of discase.

Materials and methods Animals

Female B 10.A mice were purchased from Frederick Cancer Research facility, Frederick, MD. Female Lewis rats were purchased from Harlan Sprague Dawley, Indianapolis, IN. All animals were housed under specific pathogen-free conditions, were given water and laboratory chow (Purina) and were used at 6-8 weeks of age. Animals were treated in accordance with Institutional guidelines and the ARVO Resolution on Use of Animals in Research.

Antigens and adjuvants

IRBP, prepared from bovine retinas as reported previously, by Concanavalin A (Con A) Sepharose affinity chromatography and fast performance liquid chromatography (FPLC) (14), was kindly provided by Dr B. Wiggert, LMOD, NEI. Antigen preparations were aliquoted and stored at -20°C until use. Purified Bordetella pertussis toxin (PTX) was purchased from Sigma, St Louis, MO. Complete Freud’s adjuvant (CFA) was purchased from DIFCO (Detroit, MI) and was supplemented with additional Mycobuctrriurn tuberculosis strain H37RA to a final concentration of 2.5 nig/ml. Peptide 35, representing the major pathogenic epitope of human S-Ag for Lewis rats (residues 34 1-360) was synthesized using FMOC chemistry (Nuros, San-Jose, CA) ( IS).

EAU induction

The disease was induced using the following procedures: a) Active immunization: Mice were immunized on day 0 with SO pg or with 100 pg of IRBP in 0.2 ml emulsion in CFA ( I : 1 v/v) divided among three subcutaneous sites: 100 p1 in the base of the tail; SO pI in each thigh. All mice received 0.5 or 1.0 pg PTX intraperitoneally (i.p.). Rats were immunized on day 0 with 5.0 pg of peptide 35 in 0. I ml in CFA ( I : 1 v/v) in,jected into one hind footpad. b) Adoptive transfer: The uveitogenic SP-35 T cell line was derived and maintained in culture as described previously ( 1 6). Before transfer cells were activated with 2 pg/ml of peptide 35 in the presence of syngeneic antigen-presenting cells for 48 h. The activated

lymphocytes were washed and injected i.p. into naive Lewis recipients (0.5X lo6 cells/rat in 1.0 ml volume). Mice were sacrificed 2 1 days after immunization (approximately 7 days after the onset of EAU in control animals). Rats were sacrificed 7 days after inflammatory disease was first observed in the anterior chamber of the eye in control animals.

Treatment

Suramin (m.w. 1429) was supplied by NCI, NIH. Animals immunized on day 0 were treated with 100 mg/Kg of suramin, or with PBS for control, injected i.p. once weekly. Treatment groups were: suramin on days 0 and 7, PBS on day 14 (afferent treatment); PBS on day 0, suramin on days 7 and 14 (efferent treatment) and PBS on days 0, 7, and 14 (control). Adoptively transferred rats received a single injection of 33 mg/Kg or 100 mg/Kg of suramin i.p. one day before cell transfer (d - 1 ) . This dose range and administration schedule were chosen on the basis of our previous experience with using suramin in vivo (9). Due to the low clearance of suramin (serum half-life of 40-50 days) only one or two administrations of the drug appeared to be sufficient to maintain therapeutic levels.

EAU grading and presentation of results

Eyes were collected for histopathology approximately 7 days after onset of disease (21 days in mice, 11-13 days in rats). Freshly enucleated eyes were fixed for I h in 4% phosphate- buffered glutaraldehyde and transferred into 10% phosphate- buffered formaldehyde until processing. Fixed and dehydrated tissue was embedded in methacrylate, and 4 to 6 pm sections, cut through the pupillary-optic nerve plane, were stained by standard hematoxylin and eosin. Six to 8 sections cut at different levels were examined for each eye in a masked fashion by one of us (C-C. C.), who is an ophthalmic pathologist, and the presence and extent of lesions were determined. Incidence and severity of EAU were scored on a scale of 0 to 4 in half-point increments, according to the semiquantitative grading systems described previously for mice and for rats (17, 18). Briefly, the minimal criterion for scoring an animal as positive by histopathology was inflammatory cell infiltration of the uvea and the retina. Progressively higher scores were assigned for presence of discrete lesions in the tissue (such as choroiditis, retinal vasculitis, granuloma formation, retinal folding and/or detachment, photoreceptor damage, etc.), taking into account lesion type, size and number. Severity of disease is calculated as mean grading of all eyes in the group. Incidence is shown as the number of positive out of total animals in each group.

Delayed type hypersensitivity (DTH)

Animals were anesthetized and 20 pg of the specific antigen was injected into the ear pinna in a volume of 20 pl. The other ear received PBS. Ear thickness was measured 48 h later with Spring-Loaded micrometer. Specific DTH was calculated as the difference between the antigen and the PBS-injected ear.

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.

Suramin suppresses EAU 889

Lymphocyte proliferation assay

Draining lymph nodes were collected from mice 21 days after immunization. Triplicate 0.2 ml cultures of 5X 10’ cells were cultured in 96-well round-bottom plates (Costar, Cambridge, MA, USA) in RPMl 1640 supplemented with 10XIO-s M 2- mercaptoethanol, 2 mM glutamine, 1 mM sodium pyruvate, 0.1 mM nonessential amino acids, 50 pg/ml gentamycin, and 1 .O% mouse serum. Alpha-methyl-D-mannopyranoside (a-MP, 20 pg/ml, Sigma, St Louis, MO) was added to the medium to neutralize any traces of Con A (which is used in the initial stages of IRBP purification). This concentration of a -MP had previously been determined not to affect cell proliferation. Stimulants were added to the wells to the following final concentrations: IRBP - 50 pg/ml, purified protein derivative of tuberculin (PPD, Connaught Laboratories, Toronto, Can- ada) - 20 pg/ml, and phytohemagglutinin (PHA, DIFCO) - 1.0 lg/rnl. Cells were cultured for a total of 66 h and were pulsed with I pCi/well of 3H- thymidine (New England Nuclear, Wilmington, DE, USA) during the last 18 h. The cells were harvested with a PHD harvester (Cambridge Technology, Watertown, MA, USA) and ’H-thymidine uptake was deter- mined by standard liquid scintillation. Stimulation index (SI) is calculated as the counts per min (c.p.m.> in the antigen stimulated wells divided by c.p.m. in wells containing medium only. SI 3 2 is considered as a positive response.

Antibody measurements

Blood was collected individually from each mouse. Levels of specific IgG and IgM to IRBP were measured by ELISA, essentially as described previously (19). Dilutions of the tested sera were incubated on IRBP-coated ELISA plates and the reaction was developed using secondary antibodies (goat anti- mouse IgC-HRP or goat anti-mouse IgM-HRP) obtained from Southern Biotechnology Associates, Inc. (Birmingham, Alab- ama). Antibody concentrations in pg/ml were calculated from a standard curve constructed with a reference mouse anti- IRBP serum. The concentration of IgM and IgG anti-IRBP antibodies in the reference serum had previously been deter- mined in using IgM and IgG standards, according to the protocol described elsewhere for titration of anti-TNP reference serum (20).

Statistical analysis

Parametric data (DTH) were analyzed by independent t-test. Nonparametric data (EAU scores) were analyzed by Snedecor and Cochran’s test for linear trend in proportions (21). Each mouse, not each eye, was treated as one statistical event (both eyes were averaged for analysis). Probability values of s0.05 were considered statistically significant.

Results Suramin inhibits the afferent phase of EAU

Mice were immunized with IRBP, and rats were immunized with peptide 35 of S-Ag, as described in Materials and methods.

W v)

+I

Control Afferent Efferent

Treatment group

Figure I. Effect of suramin during the afferent and efferent phases of EAU in mice. Groups of 5 BI0.A mice were immunized with IRBP in CFA and PTX on day 0, and were given 100 m g K g of suramin on days 0, 7 (afferent) or 7, 14 (efferent). (A) Experiment 1: immunization was with 50 pg IRBP in CFA and 0.5 pg FTX per mouse. (B) Experiment 2: immunization was with 100 pg IRBP and I pg of PTX per mouse. Shown are average EAU scores i SE, analyzed by histopathology 21 days after immunization. Incidence is shown within the columns as positive/total animals. Statistically significant difference from control is denoted with an asterisk.

Suramin treatment was started either concurrently with immun- ization, to include the afferent (priming) stage of disease induction, or on day 7, to cover only the efferent phase of disease induction. For the purpose of these experiments, the afferent phase of disease is defined as the priming stage, and the efferent phase is considered the stage after antigen-specific effector cells have been generated. On the basis of previous studies (22), day 7 is considered to represent a boundary between these two stages. An alternative efferent-stage model of EAU was produced in rats by adoptive transfer of the highly uveitogenic T cell line SP-35. It should be taken into account that suramin given as afferent treatment persists into the

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.

890 c. Sartani et al.

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.

Suramin suppresses EAU 89 1

Figure 2. Histopathology of EAU in B1O.A mice (A) Retina of mouse that received suramin during the afferent phase. This mouse was protected from disease. Note preservation of retinal architecture. (B) Mouse that received suramin during the efferent phase. Note a few infiltrating cells in the outer segments of the photoreceptors (arrows), early granuloma formation (asterisk), and choroidal infiltration (open arrow). (C) Mouse that did not receive suramin. Note marked retinal vasculitis, (arrows), serous retinal detachment (asterisk) and damage to outer segments of photoreceptors (open arrow) (hematoxylin and eosin, Bar = 30 pm).

efferent stage of disease as well, due to the extremely low rate of body clearance of this compound (serum half life of over 40 days).

Although the exact degree of protection from disease showed variability depending on the experiment and the immunization dose, protection was consistently better in mice treated from the afferent stage than in mice in which treatment was started during the efferent stage (Figs lA, IB). Representative pathology in these treatment groups is shown in Figure 2. The same pattern was observed in Lewis rats immunized with peptide 35, in which early treatment ameliorated EAU, but treatment instituted 7 days after immunization was ineffective (Figs 3A, 4). Similarly, efferent-phase disease simulated in Lewis rats by adoptive transfer of uveitogenic lymphocytes was also largely resistant to suramin treatment (Fig. 3B).

Afferent suramin treatment reduces DTH responses

The DTH response is considered as an in vivo measure of cell-mediated immunity. Mice in which treatment was started during the afferent stage of EAU had significantly reduced DTH responses to the uveitogen. In contrast, responses of mice that received efferent treatment and were not significantly

protected from disease, did not achieve a statistically significant difference from the untreated control (Table 1). (The EAU developed by these animals is shown in Figure 1B). Thus, cellular immunity as measured by DTH paralleled the severity of disease as measured by histopathology in the two treatment regimens.

Depressed lymphocyte proliferation suggests interference with antigen priming

Draining lymph nodes of mice whose EAU scores are shown in Figure 1A were collected 21 days after immunization and were stimulated with IRBP, PPD or PHA as described in Material and methods. The results showed that afferent treat- ment with suramin strongly suppressed lymphocyte prolifera- tion to IRBP in comparison to control, and mice treated starting with the efferent stage had an intermediate response (Fig. 5) . Antigen-specific responses to PPD (a component of CFA) were affected in an essentially identical fashion (data not shown). In contrast, proliferation with PHA was only marginally affected by suramin (SI = 77.9, 42.6 and 38.4, respectively, in control mice treated with PBS, mice treated with suramin, and naive untreated animals). Because suramin was not present

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.

892 G. Surtuni et al.

W v)

+I

0 0 fA

3 U w Q) m Q

2

b. 0) > U

Control 1 OOrng/Kg 33mg/Kg

Treatment group

Figure 3. Effect of suramin during the afferent and efferent phases of EAU in Lewis rats. (A) Active immunization: Rats were immunized with 5.0 pg of S-Ag peptide 35 in CFA. Treatment was with 100 mg/Kg suramin or PBS during the afferent or efferent phase of disease, as described in legend to Figure I . (B) Adoptive transfer: Rats received either 33 mgkg or 100 mgkg of suramin 24 h before the adoptive transfer of 0.5 X 10" SP-35 cells. Groups were composed of 6 animals cach. Shown are average EAU scores 5 SE, by histopathology, of eyes collected 7 days after onset of clinical EAU (anterior chamber opacity). Incidence is shown within the columns as positive/total animals. Statistically significant difference from control is denoted with an asterisk.

in the culture, the level of proliferation in vitro is considered to reflect the prior in vivo effects of the drug. The unreduced response to PHA indicates that the cells are healthy and capable of proliferation to nonspecific stimuli. We thus interpret the depressed proliferation in vitro to IRBP (and PPD) as indicative of a reduced number of in vivo primed antigen- specific lymphocytes in the lymph node cell population.

The effect of suramin on anti-IRBP antibody response

Levels of specific IgG and IgM antibodies to IRBP in the 3erum of treated mice were measured in comparison to levels

of antibodies in mice of control group (Figs. 6A, 6B), as described in Materials and methods. Sera obtained from mice whose EAU scores are shown in Fig IA were assayed by ELISA. The results showed that afferent treatment with suramin strongly reduced levels of specific IgM and almost totally eliminated specific IgG in the serum. Efferent treatment did not affect the level of IgM antibodies, but strongly reduced the levels of anti-IRBP IgG antibodies, suggesting possible interference with the IgM-to-IgG switch.

Discussion In this study we examined the suppressive effect of the experimental drug suramin using an autoimmune disease model. Treatment administered during the afferent stage of EAU significantly suppressed the severity of disease (p <0.01). In contrast, treatment instituted during the efferent stage of the disease was less effective. Our results are in line with a previous study that demonstrated inhibition of experimental autoimmune encephalitis in the Lewis rat by suramin (23). In parallel to inhibiting EAU, suramin treatment downregulated the in vivo cellular responses to the uveitogen as measured by DTH, and in vitro proliferative responses of the draining lymph nodes to antigen. The observation that antigen-specific (but not mitogen-induced) proliferation was more profoundly depressed in mice undergoing early treatment, together with the observation that protection from disease was better when treatment covered the induction stage of the immune process, suggested that at least part of the protective effect of suramin was mediated by interference with antigen priming. As reported previously, expression of EAU requires participation of non- antigen specific T cells (that probably amplify the inflammatory process), and eliminating these cells or inhibiting their recruit- ment would also be expected to result in disease suppression. Both the unreduced response to mitogen in culture, and the diminished effectiveness of efferent treatment, instituted qffer priming has been allowed to take place, support the interpreta- tion that the 'nonspecific' arm of the response was not affected. It should be pointed out that an afferent mode of action does not necessarily make suramin an impractical therapy from a clinical point of view, since chronic uveitis must by its nature involve continuous recruitment and priming of immunocompe- tent cells.

The immunosuppressive and antiinflammatory effects of suramin could be related to the ability of this compound to block the interaction of proinflammatory cytokines, a number of which have been linked to ocular inflammation, with their corresponding cell surface receptors (6-9, 13). In addition, suramin was also shown to differentially affect production of lymphokines. Interestingly, production of interferon gamma was strongly suppressed, at the same time that production of IL-4 was enhanced (8, 12, 24). This raises the intriguing possibility that suramin treatment might skew a developing immune response towards the Th2 pathway, or preferentially inhibit maturation of Thl -like lymphocytes. Because recent data suggest that uveitis may be mediated by ThI-like cells (16, 25 , 26), such an effect would be compatible with the

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.

Surumin suppresses EAU 893

Figure 4. Histopathology of EAU in Lewis rats. (A) Retina from a rat that received treatment during the afferent phase. Note intact retinal architecture with inflammatory cells (arrows) in the choroid and surrounding the retinal vessel. (B) Retina of rat that did not receive suramin. Note total loss of outer retinal layers, retinal detachment (asterisk), inflammatory cells in the vitreous and choroid (arrows) (hematoxylin and eosin, Bar = 30 pm).

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.

894 G. Sartani et al.

0

Table 1. Effect of suramin treatment on the delayed type hypersensitivity response

Treatment DTH C SE Statistics (mmX lo-') (vs. PBS control)

Suramin, afferent 14 C 3 p < 0.03 Suramin, efferent 20 -C 3 not significant PBS 29 2 5

Groups of 5 mice were immunized with 100 pg IRBP in CFA and 1 pg PTX, and were given either afferent or efferent treatment with suramin (100 mg/ kg). DTH to IRBP was tested by ear assay as described in Materials and methods. Shown is the average difference between the IRBP-challenged and the control ear in micrometers on day 21,48 h after challenge. EAU developed by these mice is shown in Figure IB.

w v) +I

~~ 10 0

Control Aff e-rent Eff e-rent

Treatment Group

Figure 5. Effect of suramin on antigen-specific lymphocyte proliferation. Draining lymph nodes were collected from mice immunized 21 days earlier with IRBP and treated with suramin (afferent or efferent) or PBS (control). Cells were pooled within each group and were stimulated with IRBP. Shown is the mean stimulation index (SI) of triplicate cultures +- SE. Controls ranged from 664 to 1590 CPM. Differences between replicate cultures did not exceed 10% of the mean. EAU developed by these mice is shown in Figure 1A. Lymphocytes from naive mice had a stimulation index of 0.7 (not shown).

observed suppressive effects on disease and on DTH. This hypothesis is currently under investigation.

It was interesting to note that, whereas mice in which treatment was started on day 7 had normal or elevated IgM antibodies to IRBP, they had no IgG antibodies. This suggested that suramin prevented the IgM to IgG switch from taking place. The mechanism of this effect remains to be determined,

z 40

n 301 0

20

0

I - + I

'"1 0

Control Afferent Efferent

lo 1 I

500

400

1 300 3 %I n 2004

100 1: -

Corkrol Affekent Effeient Treatment group

Figure 6. Suramin treatment inhibits serum antibody formation. Sera of suramin-treated mice (whose disease scores are shown in Fig. 1A) were collected 21 days after immunization. IgM (A) and IgG (B) titers were assayed by ELISA along with appropriate standards. Shown are titers in pg/ml calculated using the standard curve.

but it is conceivable that it also might be connected to blocking of cytokine receptors. Because a role for antibodies in the development of EAU is uncertain, it is difficult to evaluate whether inhibition of the antibody switch might have contrib- uted to the diminished effectiveness of the efferent treatment,

In our experiments, mice appeared to respond better than rats to treatment with the same dose of suramin. The Lewis rat is inherently more susceptible to EAU than the BI0.A mouse, which requires a relatively high dose of antigen combined with administration of PTX in order to develop disease. Thus, what appears to be a better response to treatment in the case of the mouse may be connected to this difference in susceptibility rather than reflect a species-related phenomenon. Susceptibility to EAU is genetically controlled and within each species different strains vary in their susceptibility to the disease (27). Additional strains of both species should be studied to resolve this question.

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.

Suramin suppresses EAU 895

When considering the use of any compound as a therapeutic agent, the potential hazards must be weighed against the benefits. The side effects resulting from suramin administration to humans can be significant. Toxicity of suramin includes interference with adrenal function, impaired hematopoesis, kidney and liver toxicity and neurological symptoms (28-30). This toxicity might limit the usefulness of suramin in any but the most intractable cases of uveitis. However, lower, nontoxic, doses of suramin might be useful as an adjunct therapy to other immunosuppressive modalities, similarly to the syner- gistic effects observed between Cyclosporin A and Rapamycin that target distinct stages in the lymphocyte activation process (3 1). Finally, topical or periocular administration of suramin, which is likely to circumvent the systemic side effects, should be explored as a possible therapeutic approach.

Acknowledgements The authors wish to thank Dr Barbara Wiggert, LMOD, NEI, for her generous gift of IRBP.

References 1. Caspi, R. R. (1989) Basic mechanisms in immune-

mediated uveitic disease. In Immunology of Eye Disease, (Ed. Lightman, S. L.). pp. 61-86. MTP Press Ltd.

2. Gery, I . , Mochizuki, M. and Nussenblatt, R. B. (1986) Retinal specific antigens and immunopathogenic processes they provoke. Prog. Retinal Res. 5, 75-79.

3. Nussenblatt, R. B. and Palestine, A. G. (1989) Uveitis: Fundamentals and Clinical Practice. Year Book Medical Publishers.

4. Caspi, R. R., Chan, C. C., Wiggert, B. and Chader, G. J. ( I 990) The mouse as a model of experimental autoimmune uveoretinitis (EAU). Curr. Eye. Res. 9 (suppl.), 169-175.

5. Hawkins, F. (1940) Concentration of Bayer-205 (Germanin) in human blood and cerebrospinal fluid after treatment. Trans. R. SOC. Trap. Med. Hyg. 34, 87-91.

6. Pollak, M. and Richard, M. (1990) Suramin blockade of insulin like growth factor I stimulated proliferation of human osteosarcoma cells. J. Natl.Cancer Znst. 82,

7. Williams, L. T., Trumble, P. H., Lavin, M. F. and Sunday, E. ( 1 984) Platelet derived growth factor receptors from a high affinity state in membrane preparations, kinetics and affinity cross linking studies. J. Biol. Chem. 259, 5287-5294.

8. Mills, G., Zhang, N., May, C., Hill, M. and Chung, A. (1990) Suramin prevents binding of interleukin 2 to its cell surface receptor: a possible mechanism for immuno- suppression. Cancer Res. 50, 3036-3042.

9. Strassmann, G., Fong, M., Freter, C. E., Windsor, S., D’Alessandro, F. and Nordan, R. P. (1993) Suramin interferes with Interleukin-6 receptor binding In Vitro and inhibits Colon-26-mediated Experimental Cancer Cachexia In Vivo. J. Clin. Invest. 92, 2152-2159.

10. Strassmann, G., Graber, N., Goyert, S. M., Fong, M.,

1349-1362.

McCullers, S., Rong, G. W. and Beall, L. D. (1994) Inhibition of lipopolysaccharide and IL-1 but not of TNF- induced activation of human endothelial cells by suramin. J . Immunol. 153, 2239-2247.

11. Strassmann, G., D’Alessandro, F., Fong, M., Nordan, R. P., Nickes, P. and Chizzonite, R. (1994) Suramin blocks the binding of interleukin- 1 to its receptor and neutralizes IL- I biological activities. Int. J. Immunopharmacol. 16,

12. Czernin, S., Gessl, A., Wilfing, A., Holter, W., Trieb, K., Waldhausl, W., Vierhapper, H., Forster, 0. and Grubeck- Loebenstein, B. (1993) Suramin affects human peripheral blood mononuclear cells in vitro: inhibition of T cell growth and modulation of cytokine secretion. Int. Arch. Allergy Immunol. 101, 240-246.

13. De Vos, A. F., Hoekzema, R. and Kijlstra, A. (1992) Cytokines and uveitis, a review. Curr: Eye. Res. 11, 58 1-597.

14. Redmond, T. M., Wiggert, B., Robey, F. A., Nguyen, N. Y., Lewis, M. S., Lee, L. and Chader, G. J. (1985) Isolation and characterization of monkey interphotoreceptor retinoid-binding protein, a unique extracellular matrix component of the retina. Biochemistry, 24, 787-793.

15. Thurau, S. R., Chan, C. C., Suh, E. and Nussenblatt, R. B. (1991) Induction of oral tolerance to S-Antigen induced Experimental Autoimmune Uveitis by uveitogenic 20mer peptide. J. Autoimmun. 4, 507-5 16.

16. Savion, S., Oddo, S., Grover, S. and Caspi, R. R. (1994) Uveitogenic T lymphocytes in the rat: pathogenicity vs. lymphokine production, adhesion molecules and surface antigen expression. J. Neuroimmunol. 55, 35-44.

17. Caspi, R. R., Roberge, F. G., Chan, C. C., Wiggert, B., Chader, G. J., Rozenszajn, L. A,, Lando, Z. and Nussenblatt, R. B. (1988) A new model of autoimmune disease: Experimental autoimmune uveoretinitis induced in mice with two different retinal antigens. J. Immunol. 140, 1490-1495.

18. Caspi, R. R., Fujino, Y., Najafian, F., Grover, S., Hansen, C. B. and Wilder, R. L. (1993) Recruitment of antigen- nonspecific cells plays a pivotal role in the pathogenesis of a T cell-mediated organ-specific autoimmune disease, experimental autoimmune uveoretinitis. J. Neuroimmunol.

19. DeKruyff, R. H., Mosmann, R. R. and Umetsu, D. T. (1990) Induction of antibody synthesis by CD4+ T cells: IL 5 is essential for induction of antigen-specific antibody responses by TH2 but not THl clones. Eur: J . Immunol.

20. Rizzo, L. V., DeKruyff, R. H., Umetsu, D. T. and Caspi, R. R. (1995) The regulation of the interaction between Thl and Th2 T cell clones in vivo. Eur: J. Immunol. 25,

21. Snedecor, G . W. and Cochran, W. G. (1967) Statistical Methods (6th Edition). pp. 248. Iowa State Univ. Press, Ames, IA.

22. Caspi, R. R., Roberge, F. G., McAllister, C . G., El-Saied, M., Kuwabara, T., Gery, I., Hanna, E. and Nussenblatt, R.

93 1-939.

47, 177-188.

20, 2219-2227.

708-7 16.

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.

896 C . Sartani et al

B. (1986) T cell lines mediating experimental autoimmune uveoretinitis (EAU) in the rat. J . Immunol. 136, 928-933.

23. Van Der Veen, R. C., Asghar, S. S., Uitdehadg, B. M. J., Vand Der Helm, H. J. and Hommes, 0. R. (1985) Suppression of the development of experimental allergic encephalomyelitis by suramin. Neuropharmacology, 24, 1139-1 142.

24. Shenoy, M., MacPherson, B. and Christadoss, P. (1992) Suramin inhibits the mixed lymphocyte reaction by suppressing lymphokine production. J. Clin. Immunol. 12, 122-1 29.

25. Rizzo, L. V., Silver, P. B., Hakim, F., Chan, C. C., Wiggert, B. and Caspi, R. R. (1993) Establishment and characterization of an IRBP-specific T cell line that induces EAU in B 10-A mice. (Abstract). Invest. Ophthalmol. Vis. Sci. 34 (Suppl.), 1143.

26. Rizzo, L. V., Silver, P. B., Gazzinelli, R. T., Chan, C. C., Wiggert, B. and Caspi, R. R. ( I 994) Expression of cytokine genes within the eye in murine EAU. (Abstract). Invest. Ophthalmol. Us. Sci. 34 (suppl.), 1862.

27. Caspi, R. R. (1992) Immunogenetic aspects of clinical and experimental uveitis. Reg. Immunol. 4, 321-330.

28. Stein, C. (1993) A novel anti neoplastic agent with multiple potential mechanisms of action. Cancer Res. 53,

29. Stein, C. A,, LaRocca, R. V., Thomas, R., McAtee, N. and Myers, C. E. (1989) Suramin: an anti cancer drug with a unique mechanism of action. J. Clin. Oncol. 7 ,

30. La Rocca, R. V., Meer, J., Gilliatt, R. W., Stein, C. A., Cassidy, J., Myers, C. E. and Dalakas, M. C. (1990) Suramin-induced polyneuropathy. Neurology, 40, 954- 960.

31. Martin, D. F., DeBarge, L. R., Nussenblatt, R. B., Chan, C. C. and Roberge, F. G. (1995) Synergistic effect of rapamycin and cyclosporin A in the treatment of experimental autoimmune uveoretinitis. J. Immunol. 154,

2239-2248.

499-508.

922-927.

Cur

r E

ye R

es D

ownl

oade

d fr

om in

form

ahea

lthca

re.c

om b

y N

yu M

edic

al C

ente

r on

10/

26/1

4Fo

r pe

rson

al u

se o

nly.