sirolimus therapy for angiomyolipoma in tuberous …...automated batteries (cantab; ref. 27), and...

Cancer Therapy: Clinical

Sirolimus Therapy for Angiomyolipoma in Tuberous Sclerosisand Sporadic Lymphangioleiomyomatosis: A Phase 2 Trial

D. Mark Davies1, Petrus J. de Vries2,3, Simon R. Johnson6, Deborah L. McCartney1,3, Jane A. Cox8,Andreas L. Serra12, Peter C. Watson4, Christopher J. Howe5, Tim Doyle9, Kate Pointon10,Justin J. Cross11, Anne E. Tattersfield7, J. Chris Kingswood8, and Julian R. Sampson1

AbstractPurpose: Renal angiomyolipomas are a frequent manifestation of tuberous sclerosis and sporadic

lymphangioleiomyomatosis (LAM). These disorders are associated with mutations of TSC1 or TSC2 that

lead to overactivation of mTOR complex 1 (mTORC1), suggesting an opportunity for targeted therapy by

using mTORC1 inhibitors. This study investigated the efficacy and safety of the mTORC1 inhibitor

sirolimus for treatment of renal angiomyolipomas in patients with these disorders.

Experimental Design: In this multicenter phase 2 nonrandomized open label trial, 16 patients with

tuberous sclerosis or sporadic LAM and renal angiomyolipoma(s) were treated with oral sirolimus for up to

2 years. Steady-state blood levels were 3 to 10 ng/mL. The primary outcome was change in size of renal

angiomyolipomas measured by MRI and assessed by Response Evaluation Criteria in Solid Tumors

(RECIST) criteria. Secondary outcomes included safety, neurocognitive function, and pulmonary function.

Results: The response rate, by RECIST criteria, was 50%. Summated angiomyolipoma diameters were

reduced in all 16 patients and by 30% or more in eight (all from the per protocol group of 10). Forty-one of

48 angiomyolipomas were smaller at the last measurement than at baseline. Most shrinkage occurred during

the first year of treatment. There was little change in pulmonary function. Recall memory improved in seven of

eight patients with tuberous sclerosis. Adverse events were consistent with the known toxicities of sirolimus.

Conclusions: This study showed sustained regression of renal angiomyolipomas in patients with

tuberous sclerosis or sporadic LAM receiving 2 years of sirolimus treatment. Possible effects on pulmonary

function and neurocognition require further investigation.Clin Cancer Res; 17(12); 4071–81.�2011 AACR.

Introduction

Tuberous sclerosis is an autosomal dominant disordercaused by mutations in either TSC1 or TSC2 (1). It ischaracterized by development of tumors in many organs,

including the kidneys and brain, and by a range of neu-ropsychiatric manifestations that include seizures, cogni-tive impairments, and autism (1). Renal angiomyolipomasaffect up to 80% of patients causing morbidity and mor-tality because of hemorrhage and renal insufficiency. Theydo not regress spontaneously and current treatmentsinclude embolization and nephrectomy (2, 3). Lymphan-gioleiomyomatosis (LAM) is the main pulmonary mani-festation and occurs almost exclusively in females. Itcan lead to respiratory failure as a consequence of prolif-eration of smooth muscle–like cells (LAM cells) and cysticdegeneration and may require lung transplantation (4, 5).LAM also occurs as a rare sporadic disorder associatedwith acquired mutations of TSC2 in women without tuber-ous sclerosis, 40% of whom also have renal angiomyoli-pomas. Identical TSC2 mutations have been identified intheir angiomyolipomas and pulmonary LAM cells indicat-ing a clonal origin and ametastasis-like disease process (6).The neuropsychiatric manifestations of tuberous sclerosisare ranked by families and caregivers among the mostsignificant problems (7). Forty percent to 50% of patientshave intellectual disability, but even those with normalintellectual abilities may have specific neurocognitiveproblems, including deficits in executive function andmemory (8, 9).

Authors' Affiliations: 1Institute of Medical Genetics, School of Medicine,Cardiff University; 2Neurodevelopmental Service, Cambridgeshire &Peterborough NHS Foundation Trust, Peterborough; 3DevelopmentalPsychiatry Section, 4MRCCognition and Brain Sciences Unit and 5Depart-ment of Biochemistry, University of Cambridge; 6Division of Therapeuticsand Molecular Medicine, Nottingham NIHR Respiratory BiomedicalResearch Unit and 7Division of Respiratory Medicine, University of Not-tingham; 8Renal Unit and 9Department of Radiology, Royal Sussex CountyHospital, Brighton; 10Department of Radiology, Nottingham UniversityHospital; 11Department of Radiology, Addenbrooke's Hospital, Cam-bridge, United Kingdom; and 12Division of Nephrology, University Hospi-tal, Zurich, Switzerland.

Note: Supplementary data for this article are available at Clinical CancerResearch Online (http://clincancerres.aacrjournals.org/).

D.M. Davies, P.J. de Vries, and S.R. Johnson contributed equally to thiswork.

Corresponding Author: Julian R. Sampson, Institute of Medical Genetics,School of Medicine, Cardiff University, Cardiff CF14 4XN, United Kingdom.Phone: 44-2920-746412; Fax: 44-2920-746551;E-mail: [email protected]

doi: 10.1158/1078-0432.CCR-11-0445

�2011 American Association for Cancer Research.

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Mutation of TSC1 or TSC2 leads to overactivation ofsignaling via the mTOR complex 1 (mTORC1), a regulatorof cell growth (10). Angiomyolipomas exhibit biallelic TSC1or TSC2 mutation and mTORC1 pathway activation (2). Inthe central nervous system, mTORC1 regulates neuronaldifferentiation, synaptic plasticity, and the encoding of spa-tial and auditorymemory (11, 12). Sirolimus (rapamycin), apotent mTORC1 inhibitor, reversed renal tumor growth,abnormal synaptic plasticity, and deficits in spatial recallmemory in rodent models of tuberous sclerosis (13–15).Clinical studies indicate that sirolimus also reduces the size ofkidney and brain tumors in patients with tuberous sclerosis(16–18). In a phase 1 to 2 trial in 20 patients with tuberoussclerosis or sporadic LAM, renal angiomyolipoma volumeswere reduced by approximately 50% after 12 months ofsirolimus treatment but the tumors increased in size whentreatment was stopped (19). In an interim report of thecurrent trial, we also showed size reduction of angiomyoli-pomas in patients who had been treated for up to 12months(20). We have now evaluated the longer term efficacy andsafety of sirolimus treatment for angiomyolipoma in adultswith tuberous sclerosis or sporadic LAM. We alsomonitoredaspects of neurocognitive function and, in those with LAM,assessed changes in lung function.

Methods

EligibilityPatientswith a definite diagnosis of tuberous sclerosis (21)

or sporadic LAM (22)were eligible if they were 18 to 65 yearsof age, had at least 1 renal angiomyolipoma of 2 cm ormorein longest diameter, and an estimated glomerular filtration

rate (GFR; ref. 23) of at least 40 mL/min. The major exclu-sion criteria were angiomyolipoma embolization within6 months or bleeding within 12 months, urinary proteinexcretion of more than 1 g/d, inability to walk 100 meterson the flat, requirement for continuous supplemental oxy-gen, or an intelligence quotient (IQ) of less than 70.

Trial designThis trial was a prospective multicenter, phase 2 study

(ClinicalTrials.gov number, NCT00490789) conducted atthe University Hospital of Wales (United Kingdom), Not-tingham City Hospital (United Kingdom), Royal SussexHospital (United Kingdom), and University Hospital, Zur-ich (Switzerland) from October 2005 through Septem-ber 2009 in accordance with the United KingdomMedicines for Human Use Regulations 2004, the Declara-tion of Helsinki, the Good Clinical Practice guidelines ofthe International Conference of Harmonization, and localregulatory requirements. The Thames Valley Multi-CentreResearch Ethics Committee approved the protocol, and allpatients gave written informed consent.

Data collectionAt baseline, angiomyolipomas were visualized by abdo-

minal MRI without contrast media and measured. Up to5 angiomyolipomas with longest diameters of 2.0 cm ormore were selected in each kidney as target lesions andtheir longest diameters were summated for each patient.Angiomyolipomas with a longest diameter of less than2.0 cm were recorded as nontarget lesions. Oral liquid siro-limuswas initiated at adailydoseof 0.5mg/m2body-surface-area, and the dosage was adjusted to achieve steady-statelevels between 3 and 6 ng/mL. Follow-up visits were at 3weeks and at 2, 4, 6, 9, 12, 18, and 24months after initiation,and abdominalMRIwas repeated at 2, 6, 12, and 24months.If the longest diameter of all target lesions was not reducedby at least 10% at 2 months, the sirolimus dose was increas-ed to achieve a steady-state level of 6 to 10 ng/mL.

At each visit, blood creatinine, lipids, liver enzymes, andhematologic parameters were measured, and protein andcreatinine were measured in spot urine. GFR was estimatedwith the MDRD equation (23). Steady-state levels of sir-olimus were determined by liquid chromatography–massspectrometry from whole blood. Adverse events were clas-sified according to the National Cancer Institute CommonTerminology Criteria for Adverse Events version 3.0.

ImagingTransaxial unenhanced abdominal MRI scans were done

with1.5Tesla systems. Scanswere acquiredwithT1-weightedfast spoiled gradient echo and T2-weighted fast spin echoprotocols and reviewed by a single radiologist (T.D.). CranialMRI was undertaken at baseline and, in patients with tuber-ous sclerosis, at 12 months by using 1.5 Tesla systems. AxialT2-weighted images, axial FLAIR images, and coronal 3DSPGR were obtained and reviewed by a single neuroradiol-ogist (J.J.C). Computed tomography (CT) of the lungs wasdone at baseline and at 24 months in patients with LAM.

Translational Relevance

Inhibition of the mTOR complex 1 (mTORC1) isimportant in the treatment of a diverse and growing rangeof tumors. However, inmost cases, the tumors concernedare highly heterogeneous at a molecular genetic level andtumor responses are unpredictable. By contrast, renalangiomyolipomas thatdevelop inpatientswith the inher-ited disorder tuberous sclerosis and the related sporadicdisorder lymphangioleiomyomatosis are specificallyassociated with inactivation of the TSC1 or TSC2 tumorsuppressor genes whose products are key regulators ofmTORC1. In this phase II trial 16 patients with thesedisorders and renal angiomyolipomas were treated for 2years with the mTORC1 inhibitor sirolimus. Angiomyo-lipoma burdenwas reduced in all patients and a responseby Response Evaluation Criteria in Solid Tumors wasobserved in 50% overall and in 80% of the per protocolgroup. Our study suggests that mTORC1 inhibition maybe an effective strategy for treating angiomyolipoma inthese rare disorders. It is also the first to report onneurocognitive changes in patients with these disorderstreated with an mTORC1 inhibitor.

Davies et al.

Clin Cancer Res; 17(12) June 15, 2011 Clinical Cancer Research4072




Images were obtained during full inspiration by using a lowdose protocol and reviewed by a single radiologist (K.P.).

Pulmonary function testingForced expiratory volume in 1 second (FEV1), forced

vital capacity (FVC), and diffusing capacity of the lung forcarbon monoxide (DLco) were measured at baseline andat 4, 6, 12, and 24months in a lung function laboratory inall patients with LAM and expressed as percentage ofpredicted values (24) to allow for change over time.

Neurocognitive testingNeurocognitive tests were undertaken at baseline, 4 and

12 months in patients with English as their first language.IQ was determined for eligibility by using the NationalAdult Reading Test (25). Immediate recall memory wastested by using the list learning, story, and complex figuretests from the Adult Memory and Information ProcessingBattery (26); immediate recognition memory by using thespatial recognition memory and pattern recognition mem-ory subtests from the Cambridge Neuropsychological TestAutomated Batteries (CANTAB; ref. 27), and executivefunctions by using the self-ordered spatial working mem-ory task (SWM), the stockings of Cambridge (SoC) plan-ning task, and the intra-dimensional extra-dimensional(IDED) set-shifting task, from CANTAB (27).

Statistical analysisThe primary outcome was the number of patients in

whom renal angiomyolipomas responded to sirolimustherapy. Response was evaluated by the Response Evalua-tion Criteria in Solid Tumors (RECIST; ref. 28) that define a"complete response" as the disappearance of all lesions, a"partial response" as decrease of at least 30% in the sum ofthe longest diameters of target lesions without progressionof nontarget lesions, "progressive disease" as a 20%increase in the sum of the longest diameters of targetlesions compared with the smallest value recorded sincetreatment started or progression of nontarget lesions orappearance of 1 or more new lesions, and "stable disease"as neither response nor progression. In this trial, a responsewas defined as a complete or partial response that occurredat any time during the trial.We aimed initially to recruit 15 patients with sporadic

LAM and 15 with tuberous sclerosis but because ofslow recruitment we modified our plan to recruitingat least 14 patients in total. The minimum sample sizeof 14 was based on Fleming’s single stage design (29). p1(the response rate below which the treatment wouldnot be studied further) was set at 0.1 and p2 (the responserate above which the treatment would be studied further)at 0.4. We set a (the probability of concluding thatthe response rate is greater than p1 when that is false)at 0.05 and b (the probability of concluding that theresponse rate is less than p2 when that is false) at 0.1. Theprimary analysis was based on all patients.The per protocol population included all patients in

the study apart from those who received less than

4 months treatment at the higher sirolimus blood levelof 6 to 10 ng/mL, unless a response had been achieved.

Annual rate of change in FEV1, FVC, and DLco was deter-mined by linear regression.

For neurocognitive outcomes, we used published UKpercentile bands for age and sex for each measure to derivesummary scores (30) for each neurocognitive domain,(recall memory, recognition memory, and executive func-tion).

Results

Patients and protocol completionSixteen patients, 13 female and 3 male, were enrolled.

Ten had tuberous sclerosis (including 3 with LAM) and6 had sporadic LAM with no skin signs of tuberoussclerosis or manifestations of tuberous sclerosis on brainMRI scan. Four had undergone previous unilateralnephrectomy. Among the 12 patients with 2 kidneysthe target angiomyolipomas (i.e., those of 2 cm or morein longest diameter) were bilateral in 6. During the trial,1 patient with severe sporadic LAM died from a respira-tory infection, 1 withdrew for lung transplantation, 1for elective surgery for angiomyolipoma, 2 because ofprotocol deviations, and 1 because of likely sirolimus-related peripheral edema. Ten patients completedthe 24-month trial (Fig. 1). At 2 months, 4 patientswere dose escalated to steady-state blood levels of 6 to10 ng/mL. Others were maintained at 3 to 6 ng/mLwithout further dose escalation.

Response of angiomyolipomasThe overall response rate by the RECIST criteria (28)

was 50% (8 of 16) and in the per protocol group it was80% (8 of 10). All were partial responses that occurred in4 of 10 patients with tuberous sclerosis and 4 of 6 withsporadic LAM. At 24 months, a partial response waspresent in 4 of 10 patients (40%) remaining in the trial.Moreover, in every patient the sum of the longest dia-meters of target angiomyolipomas was reduced through-out the trial compared with baseline (Fig. 2 andSupplementary Table S1). Of 23 target angiomyolipomasevaluated at 24 months, 21 were smaller and 2 wereunchanged (Table 1). Of all 48 target angiomyolipomasevaluated during the trial, 41 (85%) were smaller at thelast measurement than at baseline, 2 were unchanged,and 5 were larger (including 4 from 1 patient, TSC4, and a1-mm increase in the fifth; Table 1). Angiomyolipomasshrank most quickly early in the trial (Fig. 2). For angio-myolipomas measured at baseline and at 12 and 24months, there was a mean reduction in longest diameterof 7.3 mm at 12 months but a further mean reduction ofonly 0.7 mm at 24 months (Table 1).

Lung functionLung function in patients with sporadic and tuberous

sclerosis-associated LAM showed wide variation at base-line and fell slightly in most patients during the trial (Fig. 3

Sirolimus Therapy for Angiomyolipoma

www.aacrjournals.org Clin Cancer Res; 17(12) June 15, 2011 4073




and Supplementary Table S2). For the 5 patients withmeasurements over 2 years the mean � SD FEV1 declinedby 76 � 52 mL/y, mean FVC by 55 � 94 mL/y and meanDLco by �0.49 � 0.55 mL/min/mmHg/y.

Four patients (LAM1, LAM2, LAM4, and TSC1) had serialmeasurements of FEV1 before enrollment for 16, 132, 42,and 106 months, respectively. Mean annual changes inFEV1 for these patients before and during the trial were�172 and �94, �122 and �89, �90 and þ10, and �49and �69 mL/yr, respectively. There were no clinicallyrelevant changes in lung CT appearances during the trial.

Neurocognitive functionThe mean � SD IQ of patients with tuberous sclerosis

was 107� 12 and of sporadic LAM patients 105� 15. Fourpatients with tuberous sclerosis were taking antiepilepticdrugs during the trial, but none reported seizures during

3 were not enrolled2 were ineligible1 declined to participate

19 patients with tuberous sclerosis or LAM screened

Sirolimus therapy, 16 started

10 had angiomyolipomasmeasured at 24 months

11 had neurocognitiveassessment at 12 months

13 had neurocognitiveassessment at 4 months

15 had angiomyolipomasmeasured at 2 months

13 had angiomyolipomasmeasured at 6 months*

12 had angipomyolipomasmeasured at 12 months

9 with LAM hadpulmonary functionassessed at baseline

8 had pulmonary functionassessed at 4 months

6 had pulmonary functionassessed at 6 months#

6 had pulmonary functionassessed at 12 months#

5 had pulmonary functionassessed at 24 months

16 had angiomyolipomasmeasured at baseline

14 had neurocognitiveassessment at baseline

16 patients enrolled6 had LAM only3 had tuberous sclerosis and LAM7 had tuberous sclerosis only

1 withdrew

1 withdrew

1 withdrew1 died

1 withdrew

1 withdrew

Figure 1. Flow diagramsummarizing patients andassessments. *One patient whowas still in the study was unable toundergo MRI of the kidneys at the2-month assessment and anotherat the 6-month assessmentbecause of inter-current illness.#, one patient was unable toundergo pulmonary function testsat 6 and at 12 months because ofinter-current illness.

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Figure 2. Changes in angiomyolipoma burden during sirolimus therapy.Renal angiomyolipomas were measured at baseline and at 2, 6, 12, and24 months by MRI. The sum of the longest diameters of all targetangiomyolipomas in each patient was calculated at each time point andthe percentage reduction calculated by comparison with the baselinevalue. Each solid line shows change in angiomyolipoma burden in onepatient. The top dashed line represents the baseline value and the bottomdashed line 70% of the baseline value. Patients in whom the sum of thelongest diameters of target angiomyolipomas fell to or below 70% of thebaseline value are defined as partial responders by the RECIST criteria.

Davies et al.





Table 1. Longest diameters of individual target angiomyolipomas

Patient Angiomyolipoma Baseline 2 mo 6 mo 12 mo 24 mo

LAM1 R1 2.2 1.4 ND 2.0 DeceasedR2 2.6 2.7 ND 2.3 DeceasedR3 2.3 2.3 ND 1.9

LAM2 R1 2.8 2.5 1.4 1.2 1.0R2 2.0 2.0 1.9 1.6 1.4

LAM3 R1 2.2 1.9 1.9 1.9 1.3LAM4a L1 2.8 2.5 2.8 2.4 2.2

L2 2.0 1.9 1.6 1.3 0.8LAM5 R1 2.3 2.2 WithdrewLAM6 L1 5.6 3.4 Withdrew

L2 2.7 1.7 WithdrewL3 2.6 1.8 Withdrew

TSC1 (LAM) R1 2.4 1.9 1.9 1.9 2.0R2 2.2 1.4 1.4 1.3 1.3

TSC2 (LAM) R1 3.2 2.6 2.1 2.3 WithdrewR2 2.2 1.8 1.4 1.6 WithdrewR3 2.2 2.1 2.0 2.0 WithdrewL1 3.8 3.4 2.8 2.4 Withdrew

TSC3 R1 2.4 2.2 2.1 WithdrewR2 2.4 1.9 1.9 WithdrewL1 5.4 4.6 4.3 WithdrewL2 2.2 2.4 1.9 WithdrewL3 2.7 2.6 2.8 Withdrew

TSC4 R1 14.1 12.8 13.3 WithdrewR2 2.7 2.7 2.5 WithdrewR3 3.4 3.3 3.7 WithdrewR4 3.2 4.2 4.2 WithdrewR5 5.4 5.4 6.4 WithdrewL1 10.6 10.2 8.3 WithdrewL2 4.4 4.5 4.8 WithdrewL3 3.6 3.5 3.4 WithdrewL4 3.4 3.3 3.2 Withdrew

TSC5a R1 2.3 1.9 1.8 1.8 1.6R2 4.5 3.8 2.8 2.8 2.9

TSC6 R1 2.4 ND 2.3 1.8 1.6L1 2.8 ND 2.3 1.7 2.1L2 2.4 ND 2.6 1.9 1.9

TSC7a R1 2.5 2.4 2.2 1.8 1.8R2 3.8 2.8 2.7 2.8 3.2R3 3.3 2.4 2.3 2.1 2.1R4 2.9 2.2 2.4 2.1 1.9

TSC8 (LAM) R1 3.9 2.3 2.6 2.5 2.5L1 4.3 3.1 3.6 4.2 4.3

TSC9a R1 2.0 1.7 1.7 1.9 1.7R2 4.2 3.5 3.8 3.2 3.2

TSC10 R1 2.9 2.8 2.8 2.2 2.0L1 3.2 2.9 2.7 2.5 2.5L2 3.3 3.5 3.3 3.4 3.3

NOTE: Target angiomyolipomas in the right (R) and/or left (L) kidney in each patient were visualized by MRI at baseline and at 2, 6, 12,and 24months and the longest diameter of each angiomyolipomawasmeasured. Values are given in cm. ND¼MRI not done becauseof intercurrent illness. TSC1-TSC10 ¼ patients with tuberous sclerosis. LAM1-LAM6 ¼ patients with sporadic LAM. TSC (LAM)indicates patients with tuberous sclerosis and LAM. aThese patients had previous unilateral nephrectomy for angiomyolipoma.






the trial or in the preceding year. Baseline cranial MRI scansin patients with tuberous sclerosis showed cortical tubersand subependymal nodules, but no subependymal giantcell astrocytomas and no changes were seen at 12 months.

Increased immediate recall memory scores were seen in 7of 8 tested patients with tuberous sclerosis. By contrast,immediate recognition memory scores fell in 5 and noneshowed an increase. Executive function scores increased in5 of 8 patients with tuberous sclerosis. Changes in neuro-cognitive scores were also observed in patients with spora-dic LAM (Fig. 4 and Supplementary Table S3).

Adverse eventsThe most common adverse events were oral mucositis

(6 of 16 patients), respiratory infections (5 patients) and

proteinuria (5 patients; Table 2). Seven serious adverseevents occurred of which 3 were possibly related tosirolimus. One patient with severe sporadic LAM diedfollowing a respiratory infection and 2 further patientswith LAM were hospitalized, 1 with pharyngitis and 1 witha chest infection. The 4 remaining serious adverse eventswere categorized as not related or unlikely to be relatedto sirolimus: a fractured tibia and fibula, chest pain ofunknown cause, urinary obstruction secondary to an ovar-ian cyst, and musculoskeletal back pain.

Discussion

This trial determined the response of renal angiomyoli-pomas in patients with tuberous sclerosis or sporadic LAMto 2 years of sirolimus treatment. A tumor response byRECIST criteria (28) was observed in 8 of 16 patientsoverall and 8 of 10 in the per protocol group. Of 23angiomyolipomas evaluated at 24months, 21 were smallerand 2 were unchanged. Although all target angiomyolipo-mas shrank inmost patients, one (TSC4) who had themosttarget lesions and who withdrew at 12 months, showedheterogeneity of tumor response, with 5 of 9 angiomyoli-pomas shrinking and 4 growing. One previous trialreported by Bissler and colleagues investigated sirolimustreatment for angiomyolipomas in patients with tuberoussclerosis or sporadic LAM and found a mean reduction inangiomyolipoma volume of 47% at 12months (19). In thecurrent trial, angiomyolipomas measured at 12 monthsshowed amean reduction in their longest diameters of 25%compared with baseline, equivalent to a volume reductionof 60% if a spherical shape is assumed. Although 12 of16 patients in the current trial were maintained at troughblood levels of 3 to 6 ng/mL and the other 4 at 6 to 10 ng/mL, all but one of the patients in the trial by Bissler andcolleagues were escalated to trough blood levels of 10 to15 ng/mL. The current trial shows that sirolimus levels atthe bottom end of the immunosuppressive range are effec-tive in reducing angiomyolipoma size in tuberous sclerosisor sporadic LAM.

In the trial reported by Bissler and colleagues, angiomyo-lipoma volumes increased after sirolimus therapy was with-drawn at 12 months, returning from 53% to 86% of thebaseline value by 24 months (19). In the current trial, themean longest diameter of angiomyolipomas measured atboth 12 and 24months was 2.19 and 2.11 cm, respectively,indicating that tumor response was maintained by conti-nuation of therapy, but little further shrinkage occurredduring the second year of treatment. Long-term sirolimustherapy may be needed to prevent tumor regrowth inpatients with tuberous sclerosis or LAM and this could beacceptable, as it is in organ transplant recipients.

In recent studies of LAM, themean rate of decline in FEV1

has ranged from 75 to 118 mL/y (31–33). During this trial,it was 76 mL/yr in the 5 patients who had measurementsmore than 2 years and 49 mL/y in 7 patients with measure-ments over 1 year or more. We did not observe a significantimprovement in lung function during sirolimus therapy as

LAM1

LAM2

LAM3

LAM4

LAM5

LAM6

TSC1(L)

TSC2(L)

TSC8(L)

FEV1

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DLCO

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Figure 3. Pulmonary function studies in patients with sporadic or tuberoussclerosis-associated LAM. FEV1, FVC, and DLco are shown aspercentages of the predicted value for women of equivalent age andheight (24).

Davies et al.





Figure 4. Changes inneurocognitive test performancein patients with tuberous sclerosis(TSC) and sporadic LAM.Neurocognitive performance wasmeasured at baseline and at 12months (4 months in patientsTSC4 and LAM6 who withdrewbefore the 12 month assessment).Test scores were converted topercentile band scores accordingto published UK norms for age andgender, and for parallel testversions. This enabled directcomparison across measures andbetween parallel test versions.Integer scores were allocated toeach percentile band for eachmeasure used (below 5th ¼ 1;5th–9th ¼ 2, 10th–24th ¼ 3,25th–49th ¼ 4, 50th–74th ¼ 5,75th–89th ¼ 6, 90th andabove ¼ 7). Summary scoreswere then calculated for the threeneurocognitive domains ofinterest (30). A, the total immediaterecall memory score was the sumof the integer scores for listlearning, story recall, and figurerecall. B, the total immediaterecognition memory score wasthe sum of the spatial recognitionmemory and pattern recognitionmemory integer scores. C, thetotal executive function score wasthe sum of the spatial workingmemory, stockings of Cambridge,and intradimensional-extradimensional set shiftinginteger scores. All summaryand integer scores are shownin Supplementary Table S3.

A

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LAMTSC

1 2 3 4 5 6 7 8

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1 2 3 4 5 6 7 8

1 2 3 4 6LAMTSC

1 2 3 4 5 6 7 8






Table 2. Sirolimus-related adverse reactions

Category Diagnosis No. of events No. of patients Grade 1 to 2 Grade 3 to 4 Grade 5

GastrointestinalOral mucositis All 9 6 9 - -

SLAM 4 3 4 - -TLAM 3 2 3 - -TSC 2 1 2 - -

Diarrhea All 1 1 1 - -SLAM 1 1 1 - -TLAM - - - - -TSC - - - - -

Nausea All 1 1 1 - -SLAM 1 1 1 - -TLAM - - - - -TSC - - - -

InfectionUpper respiratory tractor bronchitis

All 16 5 14 1 1SLAM 16 5 14 1a 1a

TLAM - - - - -TSC - - - - -

Pharyngitis All 1 1 - 1 -SLAM 1 1 - 1a -TLAM - - - - -TSC - - - - -

Urinary tract infection All 3 3 3 - -SLAM 2 2 2 - -TLAM 1 1 1 - -TSC - - - - -

Cellulitis All 2 2 2SLAM - - - - -TLAM 1 1 1 - -TSC 1 1 1 - -

Oral cavity All 1 1 1 - -SLAM 1 1 1 - -TLAM - - - - -TSC - - - - -

MetabolicProteinuria All 5 5 5 - -

SLAM - -TLAM 1 1 1 - -TSC 4 4 4 - -

Raised creatine kinase All 3 3 3 - -SLAM - -TLAM 2 2 2 - -TSC 1 1 1 - -

Hypertriglyceridemia All 2 2 2 - -SLAM - - - - -TLAM 1 1 1 - -TSC 1 1 1 - -

Raised ALP All 1 1 1 - -SLAM - - - - -TLAM - - - - -TSC 1 1 1 - -

Hypokalemia All 1 1 1 - -SLAM 1 1 1 - -TLAM - - - - -

(Continued on the following page)

Davies et al.





Table 2. Sirolimus-related adverse reactions (cont'd )

Category Diagnosis No. of events No. of patients Grade 1 to 2 Grade 3 to 4 Grade 5

TSC - - - - -Soft tissues

Peripheral oedema All 3 3 3 - -SLAM - - - - -TLAM 1 1 1 - -TSC 2 2 2 - -

Constitutional symptomsGeneral malaise All 1 1 1 - -

SLAM 1 1 1 - -TLAM - - - - -TSC - - - - -

Fatigue All 3 3 3 - -SLAM - - - - -TLAM 2 2 2 - -TSC 1 1 1 - -

DermatologyAcneform rash All 2 2 2 - -

SLAM - - - - -TLAM - - - - -TSC 2 2 2 - -

Exacerbation of eczema All 1 1 1 - -SLAM - - - - -TLAM 1 1 1 - -TSC - - - - -

CardiacPalpitations All 2 2 2 - -

SLAM 2 2 2 - -TLAM - - - - -TSC - - - - -

NeurologyDepression All 1 1 1 - -

SLAM - - - - -TLAM - - - - -TSC 1 1 1 - -

EndocrineHypothyroidism All 1 1 1 - -

SLAM - - - - -TLAM - - - - -TSC 1 1 1 - -

OcularRetinal tear All 1 1 1 - -

SLAM - - - - -TLAM 1 1 1 - -TSC - - - - -

BloodAnemia All 1 1 1 - -

SLAM - - - - -TLAM - - - - -TSC 1 1 1 - -

NOTE: Sirolimus-related adverse reactions were those adverse events that were considered to be possibly, probably, or definitelyrelated to sirolimus. aThese events were reported as severe adverse events, defined as those that resulted in death or were life-threatening, required hospitalization, or resulted in disability. "All," All patients, n ¼ 16; "SLAM," Sporadic LAM, n ¼ 6; "TLAM,"tuberous sclerosis with LAM, n ¼ 3; "TSC," tuberous sclerosis only, n ¼ 7.






was reported by Bissler and colleagues (19) but neitherstudy was powered to evaluate lung function. A recentlyreported randomized control trial of sirolimus for thetreatment of LAMdemonstrated that sirolimusmay preventor reduce decline in lung function, consistent with ourfindings (34).

The reversal of spatial learning deficits by rapamycin(sirolimus) treatment in heterozygous Tsc2 mice (15) hassuggested that mTORC1 inhibitors might also improvespecific neurocognitive problems associated with tuberoussclerosis (35). The nonrandomized design and smallpatient numbers restricted the analysis of neurocognitiveoutcomes in this trial, and the patients with tuberoussclerosis were mildly affected with far fewer neurocognitiveproblems than more typical patients. Nevertheless, scoresfor recall memory that has been associated with mTORactivity (15) increased in 7 of 8 patients with tuberoussclerosis whereas those for recognition memory did not.Larger randomized control trials are warranted to deter-mine whether treatment-related changes in neurocognitionoccur with mTORC1 inhibition in patients with or withouttuberous sclerosis.

Adverse events were common, consistent with the knowntoxicities of sirolimus and mostly of low grade. However, 1patient with severe LAM died of a respiratory infectionduring this study. Because of their immunosuppressiveproperties, the risks and benefits of mTORC1 inhibitorsneed specific and careful evaluation in this setting.

Although our trial involved a small number of patientsand was noncontrolled and open label, it reports one ofthe first examples of therapeutic targeting of tumors inthe context of both a Mendelian and a sporadic disorderby inhibition of a shared signaling pathway. The highresponse rate seen in the trial underscores the potentialfor effective targeted treatment when the setting is one ofrelative molecular homogeneity.

Recruitment to the trial proved difficult, reflecting therarity of these disorders and our restrictive inclusion cri-teria, particularly for patients with tuberous sclerosis.Nonetheless, it provides evidence that renal angiomyoli-pomas in patients with tuberous sclerosis or sporadic LAMdecrease in size in response to treatment with sirolimus andthat this response is maintained by continuation of treat-ment for 2 years. Larger trials should now address whetherin patients with large tumors and/or extensive renal invol-vement, mTORC1 inhibition leads to reduction of theclinically important complications of renal angiomyoli-poma, notably hemorrhage and renal failure.

Disclosure of Potential Conflicts of Interest

J.R. Sampson and A.L. Serra received fees to attend a Wyeth MedicalAdvisory Board meeting. All other authors declared no conflicts of interest.

J.R. Sampson and D.M. Davies had full access to all of the data in thestudy and take responsibility for the integrity of the data and the accuracy ofthe data analysis. All authors contributed to writing this report and vouch forits fidelity to the trial protocol and statistical analysis plan.

Acknowledgments

We thank A. Hunt, F. Dunstan, K. Hood, and S. Lewis for planning andstatistical advice and colleagues at the study centers for pulmonary functiontesting. Wyeth provided sirolimus for this trial but did not provide financialsupport.

Grant Support

This work was supported by the Tuberous Sclerosis Association (to J.R.Sampson and P.J. de Vries), the James Tudor Foundation, and the Wales GenePark.

The costs of publication of this article were defrayed in part by thepayment of page charges. This article must therefore be hereby markedadvertisement in accordance with 18 U.S.C. Section 1734 solely to indicatethis fact.

Received February 17, 2011; revised April 1, 2011; accepted April 18,2011; published OnlineFirst April 27, 2011.

References1. Crino PB, Nathanson KL, Henske EP. The tuberous sclerosis com-

plex. N Engl J Med 2006;355:1345–56.2. Bissler JJ, Kingswood JC. Renal angiomyolipomata. Kidney Int

2004;66:924–34.3. Sooriakumaran P, Gibbs P, Coughlin G, Attard V, Elmslie F, Kings-

wood C, et al. Angiomyolipomata: challenges, solutions, and futureprospects based on over 100 cases treated. BJU Int 2010;105:101–6.

4. Johnson SR. Lymphangioleiomyomatosis. Eur Respir J 2006;27:1056–65.

5. Kpodonu J,MassadMG, Chaer RA, Caines A, Evans A, SnowNJ, et al.The US experience with lung transplantation for pulmonary lymphan-gioleiomyomatosis. J Heart Lung Transplant 2005;24:1247–53.

6. Carsillo T, Astrinidis A, Henske EP. Mutations in the tuberous sclerosiscomplex gene TSC2 are a cause of sporadic pulmonary lymphangio-leiomyomatosis. Proc Natl Acad Sci U S A 2000;97:6085–90.

7. Hunt A. Tuberous sclerosis: a survey of 97 cases. III: Family aspects.Dev Med Child Neurol 1983;25:353–7.

8. Prather P, de Vries PJ. Behavioral and cognitive aspects of tuberoussclerosis complex. J Child Neurol 2004;19:666–74.

9. Ridler K, Suckling J, Higgins NJ, de Vries PJ, Stephenson CM, BoltonPF, et al. Neuroanatomical correlates of memory deficits in tuberoussclerosis complex. Cerb Cortex 2007;17:261–71.

10. Huang J, Manning BD. The TSC1-TSC2 complex: a molecular switch-board controlling cell growth. Biochem J 2008;412:179–90.

11. Tavazoie SF, Alvarez VA, Ridenour DA, Kwiatkowski DJ, Sabatini BL.Regulation of neuronal morphology and function by the tumor sup-pressors Tsc1 and Tsc2. Nat Neurosci 2005;8:1727–34.

12. Costa-Mattioli M, Sossin WS, Klann E, Sonenberg N. Translationalcontrol of long-lasting synaptic plasticity and memory. Neuron2009;61:10–26.

13. Kenerson H, Dundon TA, Yeung RS. Effects of rapamycin in the Ekerrat model of tuberous sclerosis complex. Pediatr Res 2005;57:67–75.

14. Lee L, Sudentas P, Donohue B, Asrican K, Worku A, Walker V, et al.Efficacy of a rapamycin analog (CCI-779) and IFN-gamma in tuberoussclerosis mouse models. Genes Chromosomes Cancer 2005;42:213–27.

15. Ehninger D, Han S, Shilyansky C, Zhou Y, Li W, Kwiatkowski DJ, et al.Reversal of learning deficits in a Tsc2þ/� mouse model of tuberoussclerosis. Nat Med 2008;14:843–8.

16. Krueger DA, Care MM, Holland K, Agricola K, Tudor C, Mangeshkar P,et al. Everolimus for sub-ependymal giant cell astrocytomas in tuber-ous sclerosis. N Engl J Med 2010;363:1801–11.

17. Wieneke R, Fackler I, Linsenmaier U, Mayer K, Licht T, Kretzler M.Antitumoural activity of rapamycin in renal angiomyolipomata asso-ciated with tuberous sclerosis complex. Am J Kid Dis 2006;48:e27–9.

Davies et al.





18. Herry I, Neukirch C, Debray MP, Mignon F, Crestani B. Dramatic effectof sirolimus on renal angiomyolipomas in a patient with tuberoussclerosis complex. Eur J Intern Med 2007;18:76–7.

19. Bissler JJ, McCormack FX, Young LR, Elwing JM, Chuck G, LeonardJM, et al. Sirolimus for angiomyolipoma in tuberous sclerosis complexor lymphangioleiomyomatosis. N Engl J Med 2008;358:140–51.

20. Davies DM, Johnson SR, Tattersfield AE, Kingswood JC, Cox JA,McCartney DL, et al. Sirolimus therapy in tuberous sclerosis orsporadic lymphangioleiomyomatosis. N Engl J Med 2008;358:200–3.

21. Roach ES, Gomez MR, Northrup H. Tuberous sclerosis complexconsensus conference: revised clinical diagnostic criteria. J ChildNeurol 1998;13:624–8.

22. Johnson SR, Cordier JF, Lazor R, Cottin V, Costabel U, Harari S, et al.European Respiratory Society guidelines for the diagnosis and man-agement of lymphangioleiomyomatosis. Eur Respir J 2010;35:14–26.

23. Levey AS, Coresh J, Greene T, Stevens LA, Zhang YL, Hendriksen S,et al. Chronic Kidney Disease Epidemiology Collaboration. Usingstandardized serum creatinine values in the modification of diet inrenal disease study equation for estimating glomerular filtration rate.Ann Intern Med 2006;145:247–54.

24. Quanjer PH, Tammeling GJ, Cotes JE, Pedersen OF, Peslin R, Yer-nault JC. Lung volumes and forced ventilatory flows. Report WorkingParty Standardization of Lung Function Tests, European Communityfor Steel and Coal. Official Statement of the European RespiratorySociety. Eur Respir J Suppl 1993;16:5–40.

25. Nelson HE. National Adult Reading Test. Windsor, UK: NFER-Nelson,1982.

26. Coughlan AK, Hollows SE. The Adult Memory and Information Pro-cessing Battery (AMIPB). Leeds, UK: AK Coughlan, 1985. (ISBN0–9510844-0–2.)

27. Cambridge Neuropsychological Test Automated Battery. Cambridge,UK: Cambridge Cognition Ltd. Available from: (http://www.camcog.com/camcog/default.asp.)

28. Therasse P, Arbuck SG, Eisenhauer EA, Wanders J, Kaplan RS,Rubinstein L, et al. New guidelines to evaluate the response totreatment in solid tumors. European Organization for Research andTreatment of Cancer, National Cancer Institute of the United States,National Cancer Institute of Canada. J Natl Cancer Inst 2000;92:205–16.

29. Fleming TR. One-sample multiple testing procedure for phase IIclinical trials. Biometrics 1982;38:143–51.

30. Wilson BA, Emslie H, Foley J, Emslie H, Groot Y, Hawkins K, et al. TheCAMPROMPT prospective memory test. London: Pearson, 2005.

31. Johnson SR, Tattersfield AE. Decline in lung function in lymphangio-leiomyomatosis: relation to menopause and progesterone treatment.Am J Respir Crit Care Med 1999;160:628–33.

32. Taveira-DaSilva AM, Stylianou MP, Hedin CJ, Hathaway O, MossJ. Decline in lung function in patients with lymphangioleiomyo-matosis treated with or without progesterone. Chest 2004;126:1867–74.

33. Urban T, Lazor R, Lacronique J, Murris M, Labrune S, Valeyre D, et al.Pulmonary lymphangioleiomyomatosis. A study of 69 patients.Groupe d’Etudes et de Recherche sur les Maladies "Orphelines"Pulmonaires (GERM"O"P). Medicine (Baltimore) 1999;78:321–37.

34. McCormack FX, Inoue Y, Singer LG, Strange C, Nakata K, et al.Efficacy and safety of sirolimus in lymphangioleiomyomatosis. N EnglJ Med 2011;364:1595–606.

35. de Vries PJ, Howe CJ. The tuberous sclerosis complex proteins – aGRIPP on cognition and neurodevelopment. Trends Mol Med 2007;13:319–26.






2011;17:4071-4081. Published OnlineFirst April 27, 2011.Clin Cancer Res D. Mark Davies, Petrus J. de Vries, Simon R. Johnson, et al. Phase 2 TrialSclerosis and Sporadic Lymphangioleiomyomatosis: A Sirolimus Therapy for Angiomyolipoma in Tuberous

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