[CANCER RESEARCH57, 2325—2330,June 15, 19971

Advances in Brief

Multidrug Resistance Protein (MRP) Expression in Retinoblastoma Correlates

with the Rare Failure of Chemotherapy despite Cyclosporine for Reversal

of P-Glycoprotein'

Helen S. L. Chan,2 Ying Lu, Thomas M. Grogan, George Haddad, David R. Hipfner, Susan P. C. Cole,Roger G. Deeley, Victor Ling, and Brenda L. GallieDivisions ofllematology/Oncology, Immunology & Cancer Research, Hospitalfor Sick Children, 555 University Avenue, Toronto, Ontario, M5G 1X8 Canada (H. S. L C., G. H.];Depart@nents ofPediatrics [H. S. L C., G. H.], Ophthalmology [I'. L, B. L G.], and Molecular and Medical Genetics (B. L. G.], University of Toronto, Toronto, Ontario, MSS 1A8

Canada; British Columbia Cancer Agency, University of British Columbia, 601 West 10th Avenue, Vancouver. British Columbia, V5Z 113 Canada (V. LI; Cancer ResearchLaboratories, Queen‘sUniversity, Botterell Hall, Kingston, Ontario, K7L 3N6 Canada (D. R. H., S. P. C. C., R. G. DI; and Department of Pathology, Arizona Health ScienceCenter, College ofMedicine, University ofArizona, Tucson, Arizona 85724 (T. M. G.]

Abstract

Failure of chemotherapy associated with expression of the multidrugresistance protein p170 frequently occurs in retinoblastoma (RB). Despiteusing cyclosporine, which inhibits p170 and improves our chemotherapyresults, rare failures occur. In nonmetastatic primarily enucleated RBs,we show expression ofpl7O in 3 of 18 samples and expression of multidrugresistance protein (MRP), the second protein associated with resistance tochemotherapy, in 1 of 18 samples. All three RBs that failed chemotherapywithout cyclosporine expressed MRP with p170. All three RBs that wereenucleated immediately when chemotherapy failed despite the addition ofcyclosporine expressed only MRP. One RB enucleated 2 years after failing

chemotherapy with cyclosporine, despite radiation and salvage chemotherapy, expressed both p170 and MRP. Two metastatic RBs that expressed both p170 and MRP at diagnosis and at recurrence failed chemotherapy without cyclosporine, whereas one metastatic RB that expressedneither protein was cured by chemotherapy without cyclosporine. MRPmay result in failure of chemotherapy despite the elimination of p170-expressing clones by cyclosporine.

Introduction

The 190-kDa MRP@confers a pattern of drug resistance in vitro thatis similar to that of the multidrug resistance P-glycoprotein geneproduct, p!70 (1—5).Because many studies have found that thepresence of p!70 is strongly associated with failure of chemotherapyin neuroblastoma, sarcoma, leukemia, lymphoma, and myeloma (6—10),trialsthattestwhethercyclosporineor verapamil,inhibitorsofp170 in vitro, can improve the results of chemotherapy (8—10)areongoing. The role of MRP in the failure of chemotherapy remainspoorly defined.

After myeloid leukemia patients had failed cyclosporine-modulatedchemotherapy (8), p170 was reported to be undetectable, suggestingthat the p170-expressing tumor cells had been eliminated. If failure of

Received2/17/97;accepted4/30/97.The costs of publication of this article were defrayed in part by the payment of page

charges. This article must therefore be hereby marked advertisement in accordance with18 U.S.C. Section 1734 solely to indicate this fact.

I Supported by grants from the National Cancer Institute of Canada (to H. S. L. C.,V. L, B. L G., R. G. D., and S. P. C. C.); Medical Research Council of Canada (toH. S. L. C., S. P. C. C., R. G. D., V. L., and B. L. G.); Elsa U. Pardee Foundation (toH. S. L. C.); Sandoz Canada, Inc., Hospital for Sick Children Pediatric Consultants andResearch Institute, Atkinson Charitable Foundation (to H. S. L. C.); Canadian GeneticDiseases Network; Retinoblastoma Family Association; and Royal Arch Masons ofCanada (to B. L. G.); and by NIH USPHS Grant CA37130 (to V. L.) and National CancerInstitute Grants CA32102 and CA17094 (to T. M. G.). S. P. C. C. is a Senior Scientist ofthe Ontario Cancer Treatment and Research Foundation. R. G. D. is the Stauffer ResearchProfessor of Queen's University. B. L. G. is a Distinguished Scientist of the MedicalResearch Council of Canada.

2To whom requests for reprints should be addressed. Phone: (416) 813-5872; Fax:(416) 813-5327.

3The abbreviations used are: MRP, multidrug resistance protein; RB, retinoblastoma.

chemotherapy in these patients was due to resistance to cyclosporine,p170 should continue to be expressed. Alternative mechanisms ofdrug resistance, such as MRP, might account for failure of chemo

therapy despite the use of inhibitors of p170.MRP is a member of the same AlP-dependent transmembrane

transporter superfamily as p170 (3, 5). Transfection of MRP cDNAconfers a similar pattern of resistance to important cytotoxic anthracyclines, Vinca alkaloids, antibiotics, and epipodophyllotoxins (3—5).Furthermore, deletion of the MRP gene (l6pl3.l) on the invertedchromosome 16 of myelomonocytic leukemia correlated with a better

outcome, suggesting that allelic loss of MRP might confer increaseddrug sensitivity (11). Because MRP is not blocked by the present

inhibitors of p170 (3—5),it is important to establish whether MRP isthe cause of failure to reverse multidrug resistance.

Until recently, chemotherapy rarely cured intraocular RB, whichwas often successfully treated by radiation, incurring a 30% risk ofradiation-induced secondary cancers by 40 years in children withgerm-line RB] gene mutations (12). We have found that p170 expres

sion occurs more frequently in RB that failed therapy than in untreatedRB (13), suggesting that overexpression ofpl7O could cause failure ofchemotherapy in RB, as in the other previously reported cancers(6—10).

We have recently shown that chemotherapy with cyclosporine butwithout radiation had a high cure rate for intraocular RB: 91% ofpreviously untreated tumors remained relapse-free, and 70% of thosetreated with chemotherapy and cyclosporine after having relapsedwithout cyclosponne were salvaged at a median follow-up of 2.8years (14, 15). Even for those with the worst prognosis, eyes with RBthat had seeded into the vitreous, the cure rate without radiation was88%, much better than that previously reported with radiation (16),with the same chemotherapy without cyclosporine (17), or with thesame chemotherapy without cyclosporine but with radiation (18). Wenow report that MRP expression is also associated with the rarefailures of chemotherapy in RB despite the use of cyclosporine.

Patients and Methods

Patients. Since 1991,we have recruited31 consecutivebilateralnonmetastaticRB patients,ages 15days—5.3years, into a carboplatin-vincristine-teniposidetrialwith cyclosporine but no radiation, approved by our research ethics board. The

patient characteristicsand protocolshave been reported elsewhere (14, 15). Irrevocably damaged eyes were enucleated at diagnosis. We treated 40 eyes of 31patientswith chemotherapywith cyclosporine,consolidatedwith laser with/without cryotherapy.In the precyclosporineera (before 1991),we treated 19nonmetastatic RB patients with vincristine-teniposideor vincristine-doxorubicin-cyclophosphamideprotocolswith/withoutradiationand treated3 metastaticRB patientswith radiation and vincristine-doxorubicin-cyclophosphamide or 8-in-i protocols

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MRP IN RB THAT FAILED CHEMOTHERAPY

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Table 1Nonmetastatic RB treated in the precyclosponfl era (before 1991)by p170and MRP expression before and aftertherapyPatient

eye―Initial

therapy―Therapy at relapseVision

OutcomeBefore

therapy

p170 MRPAfter

relapse

p170MRPSurgRTDrugsRTDrugsSurg1

2

3

4

5

6

7

8ros

LOD

[@OSL ODEOSL ODroSL ODroDLOS

(0DLOS

EOSL0D

ODLosT

T

TT

TT

TT

TN

TT

TT

TTE

E

E

E

E

EE

E

EEVT

Eye VTDCP

Eye VTDC

Eye VT

EyeE

E

ESees

5.7yrs

Blind9.7 yrs

Blind13.6 yrs

Blind9.2 yrs

Sees2.Syrs

Blindl1.lyrs

Sees6.3yrs

Blind8.lyrs-

-

— +

— —

— —

— —

— —

— —

— —

— —

- -+

+

++

+

+

MRP IN RB THAT FAILED CHEMOTHERAPY

a OD, tight eye; OS, left eye; T, eye with tumor; N, normal eye.

b p cisplatin or carboplatin; V. vincristine; T, teniposide or etoposide; D, doxorubicin; C, cyclophosphamide or iphosphamide; RT, radiation; Surg, surgery; E, enucleation.

(cisplatin, procarbazine, i,3-bis(2-chloroethyi)-l-nitrosourea,vincristine, l-@3-D-arabinofuranosyicytosine, cyclophosphamide, hydroxyurea, and methylprednisolone).

Because there is no safe way to biopsy intraocular RB without riskingsystemic spread, we have been able to test for p170 and MRP only at

therapeutic failure when the eye is enucleated. Furthermore, RB expression ofp170 or MRP in the primarily enucleated eye does not necessarily representthat of the separate tumors in the retained eye undergoing chemotherapy (13).However, we did biopsy metastatic RB assaying for p170 and MRP bothbefore and after therapy.

Multilayer Immunoperoxidase. The expression of p170 (6, 7) and MRP(19) was scored by sensitive assays in seven samples from seven intraocularRB patients and seven samples from three metastatic RB patients that weretreated with chemotherapy with/without cyclosponne. Eighteen samples from16 nonmetastatic RB patients enucieated before any chemotherapy served ascontrols. We stained the formalin-fixed samples three times with two monoclonal antibodies directed against separate p170 epitopes (C494, specific forthe MDRJ gene conferring multidrug resistance, and C219, recognizing MDRJand the multidrug resistance-unrelated MDR3 gene) or separate MRP epitopes(QCRL1 and QCRL3 are MRP-specific) and with isotype-matched negativecontrols of these antibodies (I, 2). QCRL1 does not cross-react with p170, nordoes C219 cross-react with MRP (2). Two observers, masked to the identity ofsamples and to each other's scores, interpreted the results; the final interpretation was based on the consensus score. We scored samples with no p170-

positive or MRP-positive tumor cells as negative, grading those with p170-positive tumor cells from 1+ to 5+, and grading those with MRP-positivetumor cells from 1+ to 3+ . We used appropriate controls with p170 and MRPthat have been quantified by immunoblot, including the sensitive and the 8-,16-,64-, 510-, 1000-fold resistant vincristine-selected ovarian cancer lines thatstained negative or 1+ to 5 + for p170 and the sensitive and the 2—3-,4—12-,and 50—100-foldresistant doxorubicin-selected small cell lung cancer orMRP-transfected HeLa lines that stained negative or 1+ to 3 + for MRP (3—7,19, 20).

Results

Overall, we detected MRP in 12 of 30 samples (4 of 22 at diagnosisand 8 of 8 at relapse) and p170 in 13 of 32 samples (7 of 23 at

diagnosis and 6 of 9 at relapse). There was considerable heterogeneityin the patterns and expression levels of p170 and MRP in differentsamples obtained at the same or different time points for each patientand between different patients (Fig. 1, A—H).All IgG control sampleswere negative. The observers were unanimous in interpreting whetheror not each sample was positive for p170 or MRP, varying only intheir scoring of the precise degree of positivity. We observed focal ordiffuse p!70 staining in the plasma membrane and Golgi, and MRPwas observed in the plasma membrane and cytoplasmic endocytoticvesicles. The normal retina did not express p170 or MRP.

Both p170 and MRP were found more commonly at relapse than atdiagnosis and found more often in metastatic than in nonmetastaticRB. For the 25 samples from 23 nonmetastatic RB patients, we foundMRP in 7 of 7 samples at intraocular relapse but in only 1 of 18samples at diagnosis and found p170 in 4 of 7 samples at intraocularrelapse but in only 3 of 18 samples at diagnosis (Tables 1 and 2).However, for the three metastatic RB patients, we found both MRPand p170 present in two of two samples at relapse and in two of threesamples at diagnosis.

In the precyciosporine era (before 1991 ), 22 eyes of 19 bilateral RBpatients that did not require enucleation at diagnosis were treated withchemotherapy with/without radiation; 32% were cured (6 of 19 newlydiagnosed patients and 1 of 3 previously relapsed patients) at a medianfollow-up of 5.6 years (14) and thus could not be biopsied for p170and MRP. Fifteen eyes failed therapy, 13 by 1 year and 2 by 2 years.Six of these failures occurred before 1991 : three were salvaged byirradiation, but three were enucleated, all of which showed coexpression of MRP with p170 (Table 1). The nine eyes that failed after 1991were recruited into the cyclosporine trial.

In the cyclosporine era (since 1991), we consolidated the chemotherapy-cyclosporine response with laser with/without cryotherapy in40 eyes of 31 bilateral RB patients. At a median follow-up of 2.8years, we achieved a 91% cure rate in 26 of 28 newly diagnosed RBsand a 70% cure rate in 9 of 12 relapsed intraocular RBs, 9 of which

Fig. 1. Multilayer immunoperoxidase staining of RB to detect p170 with the C219 antibody (A, C, E, and G) and MRP with the QCRLI antibody (B. D, F, and H; hematoxylincounterstain, x 1200). A, absence of ph0; B, 1+ MRP in an eye that was enucleated immediately on falling cyclosporine. C, 2+ p170; D, 2+ MRP in an eye that was enucleated2 years after having failed chemotherapy with cyclosporine, despite additional radiation and topotecan. E. I + p170 (1+ MRP not shown) in an eye enucleated at diagnosis despitemetastatic disease; F, 3+ MRP (3+ p170 not shown) in the brain metastasis when therapy failed. G, absence of pl7O; H, absence of MRP in the extraocular metastasis of an 18-yearsurvivor successfully treated for bone marrow and orbital metastasis.

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Table 2 Nonmetastatic RB treated in the cvclosporin era (since 1991) by p1 70 and MRP expression before and aftertherapyInitial

therapy― Therapy at relapse Vision Before therapy After relapse

Patient eye― Surg RT Drugs RT Drugs Surg Outcome p170 MRP p170MRP9

r OD T PVT + CsA E Sees — +L Os T PVT+ CsA I.1yrs10EOS T E Blind + —LOD T Eye VT + CsA E 4.9 yrs —+Il

@OS T E Blind + —LOD T Eye PVT PVT+ CsA E 3.1yrs —+12

EOS T E Blind — —L OD T PVT + CsA Orbit Topotecan E 2.4 yrs + + ++13

EOS T E Sees — —LOD T PVT+ CsA 0.2yrs14

[OS T E Sees + —LOD T PVT + CsA 0.4yrs15

EOS T E Sees — —LOD T PVT + CsA 2.6yrs16

flOD T E Sees — —L Os T PVT+ CsA 3.4yrs17

rOD T E Sees — —L Os T PVT + CsA 4.3yrsa

OD, right eye; OS, left eye; T, eye with tumor.

b@ carboplatin; V. vincristine; T, teniposide or etoposide; CsA, cyclosporine; RT, radiation; Surg, surgery; E,enucleation.were

previously treated by us (see above), and 3 of which were (Fig. lH) were undetectable at diagnosis is an 18-year survivorafterreferredfrom elsewhere (14, 15). None of these 35 cured eyes were chemotherapy andradiation.tested

for p170 and MRP because they cannot be biopsied. Fiveeyes(twonewly diagnosed eyes and three previously relapsed eyes) failedDiscussiontherapy

despite cyclosponne, all by 1 year. One of these failures wassalvaged with radiation, but four were enucleated (Table 2). All three The present series is small because: (a) RB is a rare diseaseeyes that were enucleated immediately after failing chemotherapy occurnng only in 1 of 20,000 live births; (b) extraocular RB is evenwith cyclosporine expressed MRP (Fig. lB) but not p170 (Fig. 1A). In rarer (less than 10% of all cases); (c) less than 40% of intraocular RBone eye in which enucleation was delayed for 2 years while giving tumors are treated with chemotherapy; and (d) very few patients failradiation and the non-pl7O substrate topoisomerase I inhibitor topo- our present cyclosporine regimen to require eye enucleation, allowingtecan as salvage chemotherapy, we detected high levels of MRP (Fig. assessment of the drug resistance proteins. The fact that there are soID) with p170 (Fig. lC). The expression of p170 or MRP in the few eyes available for study is a triumph for the children, butthisprimarily

enucleated RB did not correlate with the chemotherapy makes a large scientific series impossible.response in the retained fellow eye that could not be biopsied before Immunohistochemistry is a standard method to score for MRPortreatment

(Tables 1 and 2). p170 (21), because it allows single-cell detection of clinicallyrelevantWewere able to biopsy three metastatic RBs before treatment levels of p170 or MRP (6, 7, 21). This is particularly applicable toRB(Table3). Two children died of metaStatic RB despite radiation and tumors that are small and express low levels of p170 and MRP. To

intensive chemotherapy without cyclosporine. We found that before remove some of the subjectivity of immunohistochemistry, the obtreatment, these two RBs coexpressed p170 (Fig. 1E) and MRP (data servers independently scored the slides in a masked fashion.Wenot

shown) in both the eye and the metastasis (not shown), with even prefer to score p170 or MRP proteins by immunohistochemistryratherstrongerexpression at relapse (Fig. 1F). One patient with bone mar- than by other means for these reasons. MRP or MDRJ gene amplifi

row and orbital metastasis in whose RB both pl'lO (Fig. 1G) and MRP cation is rare in human tumors, and the mRNA levels do not neces

Table 3 Metastatic RB treated in the precyclosporin era (before 199!) by p1 70 and MRP expression before and aftertherapyInitial

therapyb Vision Before therapy After relapse

Surg Drugs RT Outcome p170 MRP p170MRPPatient

eye― Eye Met' Eye Met MetMet18

roD T E VDC+IT Orbit Dead + + + + +++ +++LosNI

9 r@ T E 8-in- 1 + IT Eye + CS Dead + + + + +LODN20

r OD T E VDC Orbit Sees - -Los N l8yrs

MRP IN RB THAT FAILEDCHEMOTHERAPY

a OD, right eye; OS, left eye; T, eye with tumor; N, normal eye.

b Surg, surgery; E, enucleation; V, vincristine; D, doxorubicin; C, cyclophosphamide; RT, radiation; CS, craniospinal radiation; IT, intrathecal methotrexate + cytosine

arabinoside + hydrocortisone.( Met, metastasis. Patient 18, optic nerve and orbit before therapy and brain and CSF after relapse; Patient 19, optic nerve, paranasal sinuses, brain, spine, and CSF before therapy

and brain and CSF after relapse; Patient 20, orbit and bone marrow before therapy but no detectable tumor at 18 years after therapy.

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MRPINRBTHATFAILEDCHEMOTHERAPY

sarily reflect the protein levels (4, 20). Except for the high-expressioncarcinomas (22), MRP and p170 levels in tumors are generally lowerthan in gene-amplified, drug-selected cell lines (3, 5, 22). Northernblot, RT-PCR, and immunoblot measure tumor cells pooled withnormal stroma and hematopoietic cells that might or might not showMRP or p170 expression that has been reported to occur widely innormal tissues. Some of these techniques might not be sensitiveenough to detect the low and heterogeneous MRP or p170 expressionin patient samples that is clinically important. Furthermore, the verylimited RB sample size and heterogeneity of p170 or MRP expressionpreclude anything but in situ studies.

We have previously hypothesized that the poor cure rate of RB bychemotherapy without radiation may be due to drug resistance conferred by p!7O (13). To show that p170 limits the response tochemotherapy, it is necessary to find p170 in unresponsive but not inresponsive tumors, predict the outcome of chemotherapy by prospective testing, and salvage patients with resistant tumors by usinginhibitors of p170. In many types of tumors, the presence of p170before treatment is strongly associated with poor outcome (6—10).Similar correlation is impossible to establish for intraocular RB,because these tumors cannot be safely biopsied before treatment.However, the finding of p170 in all of the unresponsive tumors but inonly a fraction of untreated tumors suggests the possibility that p170might be responsible for the resistance to chemotherapy.

Because we postulated that p170 might be rate-limiting for theresponse of RB to chemotherapy (13), we added the p170 inhibitorcyclosporine to our chemotherapy protocol. Our greater than 90%success rate in newly diagnosed RB and 70% success rate in previously treated but relapsed RB suggest that cyclosporine does enhancethe efficacy of chemotherapy ( 14, 15). Higher doses of cyclosporine,higher blood levels, and the greatest projected tissue exposure conelated with the best outcome (14, 15). Notably, poor-prognosis RB withvitreous seeding did better with chemotherapy and cyclosporine thanit did with radiation (16), with the same chemotherapy without cyclosporine (17), or with the same chemotherapy without cyclosporinebut with radiation at other centers (18). Despite significant improvement of the long-term efficacy of chemotherapy with cyclosporine(14, 15), we cannot conclusively prove that the cyclosponne acts bymodulating pl7O, because we cannot test intraocular RB tumorsbefore treatment.

Whether MRP is also a clinically relevant cause of failure ofchemotherapy has not been firmly established. The expression ofMRP before treatment has been shown to significantly correlate withthe outcome of treatment in neuroblastoma (23). We show here thatMRP was coexpressed with p170 in two RB patients that weremetastatic at diagnosis and later died (Table 3) and in three of threeintraocular RB patients enucleated immediately after failure of chemotherapy without cyclosporine (Table 1). These observations suggestbut do not prove that both of these drug resistance proteins might havecontributed to therapeutic failure. The three of three RBs that wereenucleated immediately after failing chemotherapy with cyclosporineexpressed MRP but not p170 (Table 2), suggesting that despite possible eradication of the p170-positive clones with cyclosporine, therapy conceivably might have still failed because of the occurrence ofMRP, which is poorly or not at all blocked by cyclosporine (5, 24).Furthermore, the one eye enucleated 2 years after having failedchemotherapy with cyclosporine despite additional radiation and topotecan showed high levels of both MRP and p170. Both a cyclosporine-resistant MRP clone and a reemergent p170-positive clonemight possibly be responsible for the failure of salvage therapy in thischild.

Most reported trials that have used 24-h infusions of cyclosporinefor up to 6 days have shown modest efficacy but considerable toxicity

from increased systemic exposure to the cytotoxins (8, 9), caused bycyclosporine alteration of drug metabolism or prolonged inhibition ofthe p170 drug-efflux pump in p170-expressing normal tissues such asthe kidney, liver, bowel, blood-brain barrier, and myeloid progenitors.Conversely, we have administered short 3-h infusions of cyclosporineon two sequential chemotherapy days with acceptable chemotoxicityand excellent dose intensity (14, 15). Consequently, the efficacy wehave observed is unlikely to be due entirely to the increased systemiccytotoxic drug exposure from cyclosporine (14, 15). However, because we have not tested the pharmacokinetics of the cytotoxins, wecannot exclude the possibility that cyclosporine may have caused asmall but important increase in the true dose intensity, without noticeably increasing the overall chemotoxicity. Alternatively, high concentrations of cyclosporine might have persisted in the eye sanctuaryto effectively inhibit p170 locally, a mechanism that is not availableto other types of tumors. Cyclosporine might also have modulatednon-p170 mechanisms of resistance, such as enhancing the responsiveness to epipodophyllotoxins even in sensitive cells (25) or suppressing the carboplatin-induced expression of c-los, c-myc, or othergenes required for repair of drug-induced DNA damage (26).

We were able to biopsy metastatic RB before treatment. The twoRBs that expressed both p170 and MRP in the intraocular and metastatic tumors before treatment were not cured by therapy and succumbed. The p170-negative and MRP-negative metastatic RB tumorin one patient was cured by chemotherapy. This very rare survival ofmetastatic RB might be due to the absence of expression of the twodrug resistance proteins.

MRP is widely expressed in normal tissues, but its physiologicalfunction is unknown (3), except possibly as a transporter for theendogenous glutathione conjugate leukotnene C4 and the conjugatedestrogen 17f3-estradiol glucuronide and xenobiotics (5). Precisely howMRP causes multidrug resistance is uncertain, but in addition toreducing total cellular drug accumulation, it may also alter the drugdistribution and sequestration into subcellular organelles (3, 5). Although the MRP gene was first cloned in a small cell lung carcinomacell line (3), it has only been studied in some human cancers (5, 23),and its clinical significance is not well-defined. If MRP is establishedas a clinically relevant cause of chemotherapy failure that is notreadily blocked by the known inhibitors of p170 (3—5),it will benecessary to develop blockers specific for MRP to solve the veryimportant clinical problem of multidrug resistance. Although RB is arare tumor in which failures are rare with our present chemotherapywith cyclosporine, it may serve as a model to gain useful informationimportant for the more common tumors.

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1997;57:2325-2330. Cancer Res   Helen S. L. Chan, Ying Lu, Thomas M. Grogan, et al.   P-GlycoproteinChemotherapy despite Cyclosporine for Reversal ofRetinoblastoma Correlates with the Rare Failure of Multidrug Resistance Protein (MRP) Expression in

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