An Official Journal
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`.. ti ~ovem er 2000 o Volume 6 0 Supplement
`-"? 1P. 4467s-4597s o ISSN 1078-0432
`c,i
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`
`West-Ward Exhibit 1083
`Raymond NCI 2000 Abs #414
`Page 001
`
`

`

`Clinical Cancer Research 0 Volume 6 o November 2000 (Supplement)
`
`Poster Session 17
`
`4549s
`
`observed also. Other adverse events, generally mild-moderate in severity, oc(cid:173)
`curred over a broad range of doses. Toxicities include asthenia, cutaneous
`toxicity, mucositis, and hypertriglyceridemia. In 17 pts receiving 0.75 to 3.~2
`mg/m2/d, CCl-779 exhibited little accumulation from cycle to cycle, preferential
`binding to RBCs, dose-related increase in AUG, and mean t'h of 32.6 h.
`Preliminary evidence of antitumor activity has been.observed, with 1 PR (non(cid:173)
`small cell lung cancer) and minor responses in other tumor types. The safety
`profile and antitumor activity observed to date. associated with plasma con(cid:173)
`centrations at which biological activity was observed in vitro, are encouraging.
`
`414 CCl-779, an ester analogue of rapamycin that interacts with PTEN/
`Pl3 kinase pathways: A phase I study utilizing a weekly intravenous sched(cid:173)
`ule. E. Raymond, J. Alexandre, H. Depenbrcck, N. Ady Vago, S. Faivre, A.
`l.ahr-Randak, E. Materman, J. Boni, S. Abbas. E. Angevin, B. Escudier, J.P.
`Armand. /nstitut Gustave Roussy, Villejuif, France; Onkologische Tageskllnik &
`Wyeth Ayerst Research I Genetics Institute, Munich, Germany.
`Background. CCl-779 inhibits mTOR, thus the phosphorylation of elF4E(cid:173)
`BP1 and p7056 kinases, prevents elF4E to initiate protein synthesis and the
`phosphorylation of the ribosomal protein S6 required for the translation of
`mRNAs. Patients and Methods. CCI-779 was given as a weekly 30-min
`infusion in patients (pts) with advanced tumors using the modified CAM. Re(cid:173)
`sults. 18pts (lvVF: 12/6) received: 7.5 (1pt), 15 (2pts), 22.5 (1pt), 34 (3pts), 45
`(4pts), 60 (1pt), 80 (1pt), 110 (1pt), 165 (lpt) and 220 mg/m2/week (3~ts). DLT
`was observed in only 1 pt; MTD has not been reached. No prolonged 1mmuno(cid:173)
`suppression has been induced. Grade (Gr) 1-2 skin toxicity was observ~:
`dryness with mild itching (6pts), eczema-like lesions (2pts), ~ub-acute .u.rticana
`(2pts), and aseptic folliculitis (11 pts). Gr1-2 and Gr-3 mucos1t1s/stomat1bs were
`observed in 10pts and 1 pt, respectively. All pts receiving ;z: 8 doses expen(cid:173)
`enced Gr-1 nail changes. Thrombocytopenia was observed in 9pts; 2pts with
`G-3 at 34 and 45 mg/m2/week. Leukopenia was reported in 4pts and anemia in
`7pts. Asymptomatic increases of triglyceride and cholesterol levels were ob(cid:173)
`served in 9pts and 5pts, respectively. A reversible decrease in testosterone
`concentrations with increased levels of LH/FSH were observed in 5/9 men
`receiving «4 doses at dosages :;;::15mg/m2/week. Pharmacokinetic analysis
`from 12pts (doses: 7.5-60 mg/m2/week) indicates that CCl-779 Cmax increased
`linearly but AUG increased sub-proportionately. Clearance and volume of dis(cid:173)
`tribution at steady state increased with increasing dose. Mean half-life was
`about 20hrs. Of the 16pts evaluable for anti-tumor activity, 3pts had a partial
`response (renal cell carcinoma with lung metastases; neuro-endocrine tumor
`with hepatic metastases and breast cancer with liver, lymph node and Pl'.n(cid:173)
`orbital metastases). Conclusion. Current data show that CCl-779 has promis(cid:173)
`ing activity and mild-moderate toxicity over a broad range of doses.
`
`415 Effect of the proteasome inhibitor PS-341 on cell cycle progression
`and bcl-2: A potentially unique mechanism of action. R. Perez-Solert, YH
`Lingt. B Ngt, J Adams", P Elliott·, L Liebest. Kaplan Cancer Center, tNew York
`University School of Medicine, New York, NY, and •Millennium Pharmaceuticals,
`Cambridge, MA.
`PS-341 is a proteasome inhibitor currently in Phase I clinical evaluation. We
`studied the effects of PS-341 on cell cycle progression and related events in
`human NSCLC H460 (p53 wild type) and H358 (p53 null) cells. Exposure to 0.1
`µM PS-341 for 6 h resulted in a marked accumulation of cells at G21M: This
`blockade was associated with a 6-10 fold time-dependent accumulation of
`cyc!ins A and Band a 10-fold elevation of cyctins A and B kinase activities as
`assessed by 32 P-1-ATP incorporation into histone-H1. In addition, bcl-2 phos(cid:173)
`phorylation, a marker of mitotic arrest, was detected as early as 3 h after
`exposure to PS341. More impor:antly. a 25 kDa bcl-2 degradation product was
`detected as early as 12 h after exposure to PS341. This degradation product
`appeared specific for proteasome inhibition since it was observed with the
`proteasome inhibitors MG132 and PSI but not with the chemotherapeutic
`agents paclitaxel, vinblastine, camptothecin, etoposide, and cisplatin, or th~
`PKG inhibitor staurosporine. In addition. it was not caspase-dependent since 1t
`was observed in the presence of caspase inhibitors and appeared to localize in
`the triton X-100 insoluble cellular fraction. In view of the ability of PS341 to
`induce arrest at G2/M we then studied in vitro cytotoxicity of the combination of
`PS341 and the antitubulin agent docetaxel against H460 and H358 cells. Cells
`were treated concomitantly with PS341 (0.1 µM or 0.5JLM) and docetaxel {0.1 to
`4µM) for 48 h. An additive cytotoxic effect was observed with the combin'."tion
`0.5µM PS341 and 0.5 and 1 µM docetaxel. In conclusion. our results 10d1cate ·
`that PS341 induces unique changes in bcl-2 that appear to be specific for
`proteasome inhibition. The functional consequences of these bcl-2 changes
`and their potential relaticnship with the demonstrated ability of this agent to
`retain its cytotoxicity against bcl-2 transfected cells is being investigated.
`
`416 A phase I phannacodynamic study of the proteasome inhibitor
`PS-341. J.P. Thomas, A. Adie~ C. Ehrlichman, P. Geiger, A. Haas, R. Arzooma(cid:173)
`nian, D. Alberti, R. Marnocha. K. Binger, J. Volkman, C. Feierabend, K. Tuts~h,
`J. Adams, P. Eliot and G. Wilding. University of Wisconsin Comprehensive
`Cancer Center, Madison, WI, Mayo Clinic, Rochester, MN and Millenium Phar(cid:173)
`maceuticals, Gambridge, MA.
`The ubiquitin-proteasome pathway is the principal enzymatic degradation path(cid:173)
`Wff.J for most intracellular proteins including those involved in cell cycle regulat10~,
`apoptosls and angiogenesis. PS-341 is a dipeptide boronic acid compound that IS
`a potent inhibi1or of the 20S/26S proteasome. Proteasome inhibition affects such
`
`diverse metabolic processes including stabilization of ceil cycle regulatory proteins
`and inhibition of NF·KB activation. PS-341 has broad activity including MOR and
`Bcl-2 overexpressing cancer cell lines. In vivo PS-341 inhibits the growth of a
`number of tumors including the HT-29, NCl-H23 and PC-3 models. Toxicity was
`seen in preclinical models when the proteasome was inhibited by greater than
`80%. We are conducting a phase I trial of PS-341 in patients with advanced
`refractory cancers. PS-341 is administered intravenously twice weekly for 4 weeks
`followed by a two week break. Dose levels of 0.5, 0.9, 1.25 and 1.50 ~!112 have
`been explored. A total. of 9 patients have been treated at the UW. Tox1c1ties seen
`have included rash, fatigue and thrombocytopenia. A MTD has not yet been
`reached. Proteasome inhibition by PS-341 has been monitored by measuring 205
`proteasome activity in whole blood samples using a ftourogenic peptide substrate.
`20S proteasome inhibition in this study measured 1 hour after PS-341 administra(cid:173)
`tion correlates highly with PS-341 dose. We are achieving levels of proteasome
`inhibition (> 60%) associated with anti-tumor activity in the preclinical models. We
`have also examined patient peripheral blood mononuclear cells to determine
`whether levels of proteasome inhibition achieved in this study may be associated
`with accumulation of ubiquitinated proteins. Cell lysates were analyzed by Western
`blot for ubiquitin protein conjugates. Up to a 3 fold increase in ubiquitinated
`proteins were seen in some patients. peaking at 5 hours after PS-341 admini(cid:173)
`stration.
`
`417 Proteasome inhibition by PS-341: A phase I study, A Hamilton1, JP
`Eder, A Pavlick', JW Clark3, A Chachoua 1, DP Ryan3 , K Farrell', H Wasserstrom 1,
`• 'NYUSch. of Med., 2Dana
`L Liebes1
`, JWright", P Elliott5,JAdams5 andFMuggia1
`Farber Can. Inst., 3Mass. General Hosp., 4CTEP NCl, 5Millennium Pharm.
`The proteasome is a multimeric protease complex that regulates cellular
`proteins by degrading ubiquinated proteins. Proteasome inhibition results in
`increased levels of a variety of key cellular proteins that may contribute to
`anti-tumor activity; IKB inhibits nuclear factor KB (NF-KB) mediated transcrip(cid:173)
`tion, p53 inhibits apoptosis, and p21 inhibits cyclin-dependent kinase (CDK)
`activity. PS-341 is a dipeptide boronic acid derivative that inhibits the protea(cid:173)
`some by stabilization of its active si1e. Animal models predicted dose limiting
`gastrointestinal toxicity at ;::80% proteasome inhibition (Pl), PS-341 was ad·
`ministered as an IV bolus on 01 &4 of a 2-week cycle. Five dose levels have
`been studied to date: 0.25mg/m2 , 0.8mg/m 2, 1mg/m2, 1.2mg/m2 and 1.45mg/
`m 2 • 19 pts have been treated: 11 M /BF. Age: median 57, range 25-78. Primary
`tumors: colorectal (3), renal (3), NSCLC (3), melanoma (2), ST sarcoma (2).
`osteosarcoma (1), lymphoma (1), prostate {1), endometrial (1), esophagus (1),
`hepatoma (1). Prior therapies: chemotherapy (17), radiotherapy {13). To~icities
`have been mild and non-specific. 1/6 pts treated at 1.2mg/m2 expenenced
`self-limiting G3 diarrhea. No objective responses have been documented. One
`pt with melanoma treated at 1 mg/m2 maintained a PR in lung with SD in skin for
`6 months. Pl was measured at 1 , 4 and 24hrs after dosing. At all dose levels,
`peak Pl was seen at 1 hr, and recovery to approximately 50% of peak Pl was
`seen at 24h. Peak mean Pl were 21 %, 54%, 48% and 59% at dose levels 1, 2,
`3 and 4 respectively. Tumor Pl at 24h in one pt was 87% and averaged 54% in
`2 biopsies at 2-3 h in another pt. Accrual is ongoing at 1.9mg/m2 , and phase ll
`studies are planned. Supported by U01 CA76642, M01 RR00096 and the Lynne
`Cohen Foundation (NY). and U01 62490 (Boston).
`
`418 Phannacodynamic evaluation of the protein kinase C (PKC) inhib(cid:173)
`itor CGP41251 (PKC412) in patients with metastatic melanoma. M. Mill(cid:173)
`ward', C. House'. L. Webster', B. Linahan', I. Olver, G. Toner'. J. Zalcberg'.
`D. Bowtell1
`• 'Peter Maccallum Cancer Institute, Melbourne, 2Royal Adelaide
`Hospital, Australia.
`PKC412 selectively inhibits PKG OC50 <1µM) and has preclinical activity as a
`cytostatic and modulator of MOR. The recommended Phase II dose is 75mg tds
`which produces potentially active trough plasma levels (1 Oµ.moVI), and suppresses
`cytokine release and lymphocyte ERK2 levels (Thavasu 1999). Patients (pts) with
`measurable metastatic melanoma and 2: 2 superficial lesions received 75mg tds;
`tumor biopsies and plasma were collected prior to and after 28 ~ys treatn:ent.
`lntra-tumoral total PKG activity was measured in cytosolic and particulate fractions
`using protamine sulphate as the substrate. Initial experiments showed addition of
`lOµM PKC412 to melanoma biopsies inhibited phosphorylation. Ability of plasma
`to modulate ex vivo intracellular daunorubicin accumulation in MOR cells was
`measured with activity of 20µg/ml valspodar {?SC833) defining 100% reversal.
`Compared to the pretreatment biopsy. cytosolic PKC activity was reduced by 7%
`to 91 % in 7i9 pts. Particulate PKG activity was reduced by 11 % to 79% in 419 pts.
`Only 1 pt had >50% inhibition in both fracticns. Tumor PKG isoform profile in 1 pt
`resistant to PKC412 (unchanged cytosolic activity and >200% increase particulate
`activity) showed an abundance of PKa;, an isoform refractory to inhibition by
`PKC412 OCso >1000µ.M). Addition of 2011g/ml PKC412 to pretreatment plasma
`produced 14%-64% (mean 40%) reversal of MDR. Plasma taken following 28
`days PKC412 showed <10% reversal in 8/8 patients. All.patients hadp~~ive
`disease. This Phase llA trial did not demonstrate con51stent target 1nh1bition or
`phannacodynamic efficacy of PKC412 in melanoma patients.
`
`Proceedings of the 2000 NCioEORTCoAACR Symposium
`
`West-Ward Exhibit 1083
`Raymond NCI 2000 Abs #414
`Page 002
`
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