Significant Differences In EFS Distribution Research Articles

Initial testing (stage 1) of SGI‐1776, a PIM1 kinase inhibitor, by the pediatric preclinical testing program

The PIM kinase inhibitor, SGI-1776, was tested against the PPTP in vitro (1.0 nM-10 µM) and in vivo panels (148 mg/kg daily × 5 days for 3 weeks). SGI-1776 exhibited cytotoxic activity in vitro with a median relative IC(50) of 3.1 µM. SGI-1776 induced significant differences in EFS distribution in vivo in 9 of 31 solid tumor xenografts and in 1 of 8 of the evaluable ALL xenografts. SGI-1776 induced tumor growth inhibition meeting criteria for intermediate EFS T/C activity in 1 of 39 evaluable models. In contrast, SGI-1776 induced complete responses of subcutaneous MV4;11 (B myeloid leukemia).

Initial testing of the investigational NEDD8‐activating enzyme inhibitor MLN4924 by the pediatric preclinical testing program

MLN4924 is an investigational first-in-class small molecule inhibitor of NEDD8-activating enzyme (NAE). NAE is an essential component of the NEDD8 conjugation pathway, controlling the activity of a subset of ubiquitin-proteasome system (UPS) E3 ligases, multiprotein complexes that transfer ubiquitin molecules to substrate proteins. MLN4924 was tested against the PPTP in vitro panel using 96-hour exposure time at concentrations ranging from 1.0 nM to 10 µM. It was tested in vivo at a dose of 100 mg/kg [66 mg/kg for the acute lymphoblastic leukemia (ALL) xenografts] administered orally twice daily × 5 days. Treatment duration was 3 weeks. The median relative IC(50) for MLN4924 against the PPTP cell lines was 143 nM, (range: 15-678 nM) with that for the Ewing panel being significantly lower (31 nM). MLN4924 induced significant differences in EFS distribution compared to control in 20 of 34 (59%) evaluable solid tumor xenografts. MLN4924 induced intermediate activity (EFS T/C values >2) in 9 of the 33 evaluable xenografts (27%), including 4 of 4 glioblastoma xenografts, 2 of 3 Wilm's tumor xenografts, 2 of 5 rhabdomyosarcoma xenografts, and 1 of 4 neuroblastoma xenografts. For the ALL panel, 5 of 8 evaluable xenografts showed intermediate activity for the EFS T/C measure. MLN4924 did not induce objective responses in the PPTP solid tumor or ALL panels. MLN4924 showed potent activity in vitro and in vivo showed tumor growth inhibitory activity against a subset of the PPTP solid tumor and ALL xenografts.

Initial testing of JNJ‐26854165 (Serdemetan) by the pediatric preclinical testing program

JNJ-26854165 was originally developed as an activator of p53 capable of inducing apoptosis in cancer cell lines. In vitro, JNJ-26854165 demonstrated cytotoxic activity. The ALL cell line panel had a significantly lower median IC(50) (0.85 µM) than the remaining cell lines. In vivo JNJ-26854165 induced significant differences in EFS distribution compared to control in 18 of 37 solid tumors and in 5 of 7 of the evaluable ALL xenografts. Objective responses were observed in 4 of 37 solid tumor xenografts, and 2 of 7 ALL xenografts achieved PR or CR. Responses were noted in xenografts with both mutant and wild-type p53.

Initial testing (stage 1) of LCL161, a SMAC mimetic, by the pediatric preclinical testing program

LCL161, a SMAC mimetic, was tested against the PPTP in vitro panel (1.0 nM to 10.0 µM) and the PPTP in vivo panels (30 or 75 mg/kg [solid tumors] or 100 mg/kg [ALL]) administered orally twice in a week. LCL161 showed a median relative IC(50) value of >10 µM, being more potent against several leukemia and lymphoma lines. In vivo LCL161 induced significant differences in EFS distribution in approximately one-third of solid tumor xenografts (osteosarcoma and glioblastoma), but not in ALL xenografts. No objective tumor responses were observed. In vivo LCL161 demonstrated limited single agent activity against the pediatric preclinical models studied.

Initial testing (Stage 1) of AT13387, an HSP90 inhibitor, by the pediatric preclinical testing program

AT13387, a non-geldanamycin inhibitor of heat-shock protein 90 (HSP90), was tested against the PPTP in vitro panel (1.0 nM to 10 µM) and against the PPTP in vivo panels (40 or 60 mg/kg) administered orally twice weekly. In vitro AT13387 showed a median EC(50) value of 41 nM and exhibited activity consistent with a cytotoxic effect. In vivo AT13387 induced significant differences in EFS distribution compared to controls in 17% evaluable solid tumor xenografts, but in none of the ALL xenografts. No objective tumor responses were observed. In vivo AT13387 demonstrated only modest single agent activity.

Initial testing (stage 1) of the polyamine analog PG11047 by the pediatric preclinical testing program

PG11047 is a novel conformationally restricted analog of the natural polyamine, spermine that lowers cellular endogenous polyamine levels and competitively inhibits natural polyamine functions leading to cancer cell growth inhibition. The activity of PG11047 was evaluated against the PPTP's in vitro and in vivo panels. PG11047 was evaluated against the PPTP in vitro panel using 96 hr exposure at concentrations ranging from 10 nM to 100 µM. It was tested against the PPTP in vivo panels at a dose of 100 mg/kg administered by the intraperitoneal route weekly for 6 weeks. In vitro PG11047 demonstrated a concentration-response pattern consistent with cytostatic activity. The median EC(50) for PG11047 was 71 nM. Cell lines of the Ewing sarcoma panel had a lower median EC(50) value compared to the remaining cell lines in the panel, while cell lines of the neuroblastoma panel had a higher median EC(50) value. In vivo PG11047 induced significant differences in EFS distribution compared to control in 5 of 32 (15.6%) of the evaluable solid tumor xenografts and in 0 of 7 (0%) of the evaluable ALL xenografts. The single case of tumor regression occurred in an ependymoma xenograft. Further pediatric development of PG11047 will require better defining a target population and identifying combinations for which there is a tumor-selective cytotoxic effect. The regression observed for an ependymoma xenograft and the exquisite sensitivity of some Ewing sarcoma cell lines to the antiproliferative effects of PG11047 provide leads for further preclinical investigations.

Initial testing of lenalidomide by the pediatric preclinical testing program.

Lenalidomide, a novel immunomodulatory agent, is reported to modulate stem cell differentiation, and have direct antiproliferative activity as well as inhibit inflammation and hyperalgesia. On the basis of this varied pharmacological profile, lenalidomide is under investigation as a treatment for a range of oncologic indications. Lenalidomide was evaluated against the PPTP in vitro panel using 96-hr exposure at concentrations ranging from 1 nM to 10 µM. It was tested against the PPTP in vivo panels at a dose of 30 mg/kg administered orally (PO) once daily for a planned for 6 weeks. In vitro activity was not observed at concentrations up to 10 µM. Lenalidomide was well tolerated, and induced significant differences in EFS distribution compared to control in 7 of 37 (18.9%) of the evaluable solid tumor xenografts and in 0 of 8 (0%) of the evaluable ALL xenografts. The best response in the solid tumor panel was PD2 [progressive disease with growth delay (EFS T/C > 1.5)], observed in 4 of 37 (10.8%) solid tumor xenografts. A single ALL xenograft showed a PD2 response. Direct antiproliferative effects of lenalidomide were not observed in vitro. In vivo lenalidomide demonstrated low activity against tumors in immune-deficient mice. Our results suggest that lenalidomide's utility in the pediatric clinical setting may depend upon its ability to induce antitumor activity through effects on host immune and stromal cells rather than through direct effects on tumor cells.

Initial testing (stage 1) of the mTOR kinase inhibitor AZD8055 by the pediatric preclinical testing program

AZD8055 is a small molecule ATP-competitive inhibitor of the serine/threonine kinase mTOR that regulates cap-dependent translation through the mTORC1 complex and Akt activation through the mTORC2 complex. Procedures AZD8055 was tested against the PPTP in vitro panel at concentrations ranging from 1.0 nM to 10 µM and against the PPTP in vivo panels at a dose of 20 mg/kg administered orally daily x 7 for 4 weeks. In vitro the median relative IC(50) for AZD8055 against the PPTP cell lines was 24.7 nM. Relative I/O values >0% (consistent with a cytostatic effect) were observed in 8 cell lines and 15 cell lines showed Relative I/O values ranging from -4.7 to -92.2% (consistent with varying degrees of cytotoxic activity). In vivo AZD8055 induced significant differences in EFS distribution compared to controls in 23 of 36 (64%) evaluable solid tumor xenografts, and 1 of 6 evaluable ALL xenografts. Intermediate activity for the time to event activity measure (EFS T/C >2) was observed in 5 of 32 (16%) solid tumor xenografts evaluable. The best response was stable disease. PD2 (progressive disease with growth delay) was observed in 20 of 36 (55.6%) evaluable solid tumor xenografts. AZD8055 significantly inhibited 4E-BP1, S6, and Akt phosphorylation following day 1 and day 4 dosing, but suppression of mTORC1 or mTORC2 signaling did not predict tumor sensitivity. AZD8055 demonstrated broad activity in vitro, but at the dose and schedule studied demonstrated limited activity in vivo against the PPTP solid tumor and ALL panels.

Initial testing (stage 1) of the IGF‐1 receptor inhibitor BMS‐754807 by the pediatric preclinical testing program

BMS-754807 is a small molecule ATP-competitive inhibitor of the type-1 insulin-like growth factor receptor currently in phase 1 clinical trials. BMS-754807 was tested against the Pediatric Preclinical Testing Program (PPTP) in vitro panel at concentrations ranging from 1.0 nM to 10 µM and was tested against the PPTP in vivo panels at a dose of 25 mg/kg administered orally BID for 6 days, repeated for 6 weeks. In vitro BMS-754807 showed a median EC(50) value of 0.62 µM against the PPTP cell lines. The median EC(50) for the four Ewing sarcoma cell lines was less than that for the remaining PPTP cell lines (0.19 µM vs. 0.78 µM, P = 0.0470). In vivo BMS-754807 induced significant differences in EFS distribution compared to controls in 18 of 32 evaluable solid tumor xenografts (56%) tested, but in none of the ALL xenografts studied. Criteria for intermediate activity for the time to event activity measure (EFS T/C > 2) were met in 7 of 27 solid tumor xenografts evaluable for this measure. The best response was PD2 (progressive disease with growth delay), which was observed in 18 of 32 solid tumor xenografts. PD2 responses were most commonly observed in the rhabdomyosarcoma, neuroblastoma, osteosarcoma, Ewing sarcoma, and Wilms tumor panels. BMS-754807 activity in vitro is consistent with a specific IGF-1R effect that has half-maximal response in the 0.1 µM range and that is observed in a minority of the PPTP cell lines. In vivo intermediate activity was most commonly observed in the neuroblastoma and rhabdomyosarcoma panels.

Initial testing (stage 1) of AZD6244 (ARRY‐142886) by the pediatric preclinical testing program

AZD6244 (ARRY-142886) is a potent small molecule inhibitor of MEK1/2 that is in phase 2 clinical development. AZD6244 was tested against the Pediatric Preclinical Testing Program (PPTP) in vitro panel (1 nM-10 microM). In vivo AZD6244 was tested at a dose of 100 mg/kg administered orally twice daily 5 days per week for 6 weeks. Subsequently, AZD6244 was evaluated against two juvenile pilocytic astrocytoma (JPA) xenografts using once and twice daily dosing schedules. Phosphorylation of ERK1/2 was used as a surrogate for in vivo inhibition of MEK1/2 was determined by immunoblotting. At the highest concentration used in vitro (10 microM) AZD6244 only inhibited growth by 50% in 5 of the 23 cell lines. Against the in vivo tumor panels, AZD6244 induced significant differences in EFS distribution in 10 of 37 (27%) solid tumor models and 0 of 6 acute lymphoblastic leukemia (ALL) models. There were no objective responses. Pharmacodynamic studies indicated at this dose and schedule AZD6244 completely inhibited ERK1/2 phosphorylation. AZD6244 was evaluated against two JPA xenografts, BT-35 (wild-type BRAF) and BT-40 (mutant [V600E] BRAF). BT-40 xenografts were highly sensitive to AZD6244, whereas BT-35 xenografts progressed on AZD6244 treatment. At the dose and schedule of administration used, AZD6244 as a single agent had limited in vitro and in vivo activity against the PPTP tumor panels despite inhibition of MEK1/2 activity. However, AZD6244 was highly active against BT-40 JPA xenografts that harbor constitutively activated BRAF, causing complete regressions.

Abstract 5265: Pediatric preclinical testing program (PPTP) stage 1 evaluation of BMS-754807 IGF-1 receptor inhibitor

Abstract Background: Signaling through the type-1 insulin-like growth factor receptor (IGF-1R) is involved in autocrine or paracrine growth of many tumor types including childhood malignancies, and provides a strong anti-apoptotic signal that induces resistance to many forms of cellular stress. BMS-754807 is a potent inhibitor of IGF-1R and the insulin receptor that has entered phase 1 clinical trials. Methods: The PPTP includes a molecularly characterized in vitro panel of cell lines (n=27) and in vivo panel of xenografts (n=61) representing common types of childhood solid tumors and ALL. BMS-754807 was tested against the PPTP in vitro panel at concentrations from 1.0 nM to 10 μM and results were compared to those obtained with the anti-IGF-1R antibody mAb391 (50 μg/ml). In vivo testing used a dose of 25 mg/kg BID administered daily × 6 × 6 weeks by oral gavage. In vivo antitumor activity was primarily assessed by using response criteria modeled after the clinical setting and by using a time to event measure based on the median EFS of treated and control lines (intermediate activity required EFS T/C > 2, and high activity additionally required a net reduction in median tumor volume at the end of the experiment). Results: The median EC50 for BMS-754807 against the in vitro panel was 0.62 μM (range, 0.07 - 4.96 μM). The median BMS-754807 EC50 value for the 5 cell lines with the greatest response to mAb391 was 0.12 μM compared to 1.1 μM for the 11 cell lines with the least response to mAb391 (p=0.0009). These results are consistent with a specific IGF-1R effect that has half-maximal response in the 0.1 μM range and with a non-IGF-1R effect that shows half-maximal response at approximately 1 μM. The mortality rate among treated mice was 6.5%, and 39 of 45 xenograft models were evaluable for efficacy. BMS-754807 induced significant differences in EFS distribution compared to controls in 18 of 32 evaluable solid tumor xenografts (56%) tested, but in none of the ALL xenografts studied. Criteria for intermediate activity for the time to event activity measure (EFS T/C > 2) were met in 7 of 27 solid tumor xenografts and were most commonly observed in the neuroblastoma (3 of 6) and rhabdomyosarcoma (2 of 6) panels. Objective responses (i.e., tumor regression) were not observed for any xenografts. The best response was PD2 (progressive disease with growth delay), which was observed in two or more xenografts in the rhabdomyosarcoma, neuroblastoma, osteosarcoma, Ewing, and Wilms tumor panels. Conclusions: BMS-754807 showed broad tumor growth inhibition activity against the PPTP in vivo preclinical models. Future studies will focus on defining how pharmacokinetic and pharmacodynamic effects of BMS-754807 relate to tumor sensitivity and to evaluating combinations of BMS-754807 with standard cytotoxic agents. (Supported by NCI NO1CM42216) Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5265.

Initial testing of a monoclonal antibody (IMC‐A12) against IGF‐1R by the pediatric preclinical testing program

Many childhood malignancies including sarcomas, neuroblastoma, and Wilms tumor show the presence of both, active, type-1-insulin-like growth factor receptor (IGF-1R), and the autocrine production of its ligands IGF-1/IGF-2. IMC-A12 is a fully human IgG1 antibody that prevents ligand binding to the IGF-1R. IMC-A12 was evaluated against the 23 cell lines of the Pediatric Preclinical Testing Program (PPTP) in vitro panel using 96 hr exposure at concentrations ranging from 0.01 nM to 0.1 microM. IMC-A12 was tested in vivo at a dose of 1 mg/mouse administered intraperitoneally twice weekly for 6 weeks. In vitro, IMC-A12-induced T/C values <50% in only three cell lines, a rhabdomyosarcoma cell line (Rh41) and two Ewing sarcoma cell lines (TC-71 and CHLA-9). In vivo, IMC-A12 induced significant differences in EFS distribution compared to control in 24 of 34 (71%) evaluable solid tumor xenografts. Using the PPTP "time to event" activity measure, IMC-A12 induced intermediate (n = 13) or high (n = 1) activity in 33 xenografts evaluable for this activity measure, including 6 of 6 rhabdomyosarcoma xenografts, 3 of 5 osteosarcoma xenografts, 2 of 5 neuroblastoma xenografts, and 1 of 5 Ewing sarcoma xenografts. The only objective response observed was observed in a rhabdomyosarcoma xenograft (Rh28) that achieved a maintained complete response. IMC-A12 demonstrated broad antitumor activity against the PPTP's in vivo solid tumor panels, with the activity primarily being tumor growth inhibition rather than tumor regression. IMC-A12 showed its greatest activity in vivo against the PPTP's rhabdomyosarcoma xenografts.

Abstract C57: Pediatric Preclinical Testing Program (PPTP) Stage 1 evaluation of AZD8055 an inhibitor of mTOR Kinase

Abstract Background: The serine/threonine kinase mTOR in the mTORC1 complex plays an important role in regulating cap-dependent translation and in the mTORC2 complex is required to fully activate Akt signaling. AZD8055 is a small molecule being evaluated in phase 1 clinical trials that selectively inhibits mTOR kinase in both complexes. The activity of AZD8055 was evaluated against the in vitro and in vivo panels of the Pediatric Preclinical testing Program (PPTP). Methods: The PPTP includes a molecularly characterized in vitro panel of cell lines (n=27) and in vivo panel of xenografts (n=61) representing most of the common types of childhood solid tumors and childhood ALL. AZD8055 was tested against the PPTP in vitro panel at concentrations ranging from 1.0 nM to 10 µM to determine the IC50 (concentration at which T/C% = 50%) and the minimum T/C%, and it was tested against the PPTP in vivo panel at a dose of 20 mg/kg daily by oral gavage for 4 consecutive weeks. Three measures of antitumor activity were used: 1) response criteria modeled after the clinical setting; 2) treated to control (T/C) tumor volume at day 21; and 3) a time to event (4-fold increase in tumor volume) measure based on the median EFS of treated and control lines (intermediate activity required EFS T/C &amp;gt; 2, and high activity additionally required a net reduction in median tumor volume at the end of the experiment). Results: AZD8055 potently inhibited growth of the PPTP cell lines with a median IC50 value for the in vitro panel of 31.7 nM (range 2.6 nM to &amp;gt; 10 µM) and a minimum T/C% value of 6.2% (range 0.1% to 62.3%). AZD8055 was well tolerated in vivo (4.3% lethality), with only 3 of 45 tested xenograft models considered non-evaluable because of toxicity. AZD8055 induced significant differences in EFS distribution compared to control in 23 of 36 (64%) of the solid tumor xenografts and in 1 of 6 (17%) of the ALL xenografts. Criteria for intermediate activity for the time to event activity measure (i.e., EFS T/C &amp;gt; 2) were met in 5 of 32 (16%) solid tumor xenografts evaluable for this measure and were observed in the rhabdomyosarcoma panel (3 of 6) and Ewing sarcoma panel (2 of 5). Tumor regression was not observed at the 20 mg/kg daily dose, with the best response being stable disease, which was observed in 2 of 36 (5.6%) evaluable solid tumor xenografts. PD2 (progressive disease with growth delay) was observed in 20 of 36 (55.6%) evaluable solid tumor xenografts, including 4 of 5 Ewing sarcoma and 4 of 6 rhabdomyosarcoma xenografts. Conclusions: The activity observed for AZD8055 against the PPTP preclinical models is modest, although a subset of PPTP xenografts in the rhabdomyosarcoma and Ewing sarcoma panels showed greater tumor growth inhibition compared to other solid tumor xenografts. Future studies will focus on defining how the pharmacokinetics and the pharmacodynamic effects of AZD8055 relate to tumor sensitivity and on evaluating combinations of AZD8055 with standard cytotoxic agents. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C57.

Abstract C111: Pediatric Preclinical Testing Program (PPTP): Evaluation of the MEK1/2 inhibitor AZD6244 against juvenile pilocytic astrocytoma (JPA) xenografts

Abstract Background: AZD6244 (ARRY-142886) is a potent small molecule inhibitor of MEK1/2 that is in phase 2 clinical development. Against the PPTP in vivo tumor panels, AZD6244 induced significant differences in EFS distribution in 10 of 37 (27%) solid tumor models and 0 of 6 acute lymphoblastic leukemia (ALL) models, but there were no objective responses. Recently, molecular characterization of JPAs has shown tandem duplication producing a novel fusion gene (KIAA1549-BRAF) that lacks the BRAF regulatory domain and leads to constitutive activation of BRAF. Activating point mutations in BRAF are less frequent in JPA, accounting for only 5 percent of cases. AZD6244 was evaluated against two JPA xenografts, BT-35 and BT-40, that are used for secondary testing by the PPTP. Methods: AZD6244 was dissolved in 0.5% hydroxypropyl methyl cellulose, 0.1% Polysorbate 80 and administered p.o., using a twice daily schedule (excepting weekends, for which a once daily schedule was used) for 6 weeks at a dose of 100 or 75 mg/kg. Tumors were followed for an additional 6 weeks. Three measures of antitumor activity were used: 1) response criteria modeled after the clinical setting; 2) treated to control (T/C) tumor volume at day 21; and 3) a time to event (4-fold increase in tumor volume) measure based on the median EFS of treated and control lines (intermediate activity required EFS T/C &amp;gt; 2, and high activity additionally required a net reduction in median tumor volume at the end of the experiment). Phosphorylation of ERK1/2, as a surrogate for in vivo inhibition of MEK1/2 was determined by immunoblotting. Genomic DNA from BT-35 and BT-40 was screened for BRAF mutations with primers designed to amplify exons 1–18. Purified BRAF BAC DNA was labeled with digoxigenin-11-dUTP (Roche Molecular Biochemicals, Indianapolis, IN) by nick translation and used for FISH analysis. Results: Both JPA models have additional copies of chromosome 7, but lack evidence of focal duplication of BRAF. Sequencing showed that BRAF is wild type in BT-35 but that BT-40 harbors an activating mutation [V600E]. Pharmacodynamic studies indicated that AZD6244 administered at 100 mg/kg twice-daily completely inhibited ERK1/2 phosphorylation. AZD6244 was evaluated against two JPA xenografts; BT-40 xenografts completely regressed on treatment with AZD6244, but re-grew subsequent to stopping therapy. In contrast, BT-35 xenografts progressed on AZD6244 treatment. Conclusions: The PPTP previously demonstrated that AZD6244 had limited in vivo activity against the pediatric preclinical models evaluated, despite inhibition of MEK1/2 activity. However, AZD6244 was highly active at the same dose and schedule against BT-40 JPA xenografts that harbor constitutively activated BRAF. The BT-40 xenograft model may be valuable for developing rational combinations of molecularly-targeted agents against JPAs with BRAF activation. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C111.

Initial testing (stage 1) of lapatinib by the pediatric preclinical testing program

Lapatinib is a small molecule reversible tyrosine kinase inhibitor of EGFR and ErbB2 that shows in vitro and in vivo activity against a range of EGFR and ErbB2-dependent adult cancer cell lines and that has clinical efficacy against ErbB2-overexpressing breast cancer. Lapatinib was tested against the cell lines of the PPTP in vitro panel at concentrations ranging from 1.0 nM to 10.0 microM. Lapatinib was tested against the xenografts of the PPTP in vivo panels using a twice-daily oral administration schedule for 6 weeks (5 days on, 2 days off) at a dose of 160 mg/kg (320 mg/kg/day). Lapatinib pharmacokinetic parameters were determined in scid(-/-) mice. The median IC(50) value for lapatinib against the entire PPTP cell line panel was 6.84 microM (range, 2.08 to >10.0 microM). Lapatinib was well tolerated in vivo, with toxicity in only 1.5% of the treated animals. Lapatinib induced significant differences in EFS distribution compared to controls in 1 of 41 xenografts tested. No objective responses were observed in any of the solid tumor panels or in the ALL panel. Lapatinib systemic exposure was consistent with previously observed values. Lapatinib has little activity against the xenografts of the PPTP's in vivo panel, and its in vitro activity occurs at concentrations above those associated with specific EGFR/ErbB2 inhibition. These results likely reflect lack of ErbB2 overexpression in the models studied and suggest that adult and pediatric cancers may fundamentally differ in the applicability of EGFR family members as therapeutic targets.

Initial testing of aplidin by the pediatric pre‐clinical testing program

Aplidin was tested in vitro at concentrations ranging from from 0.1 nM to 1.0 microM and in vivo at a dose of 0.6 mg/kg administered intraperitoneally on an every 4 days x 3-schedule that was repeated at day 21. In vitro, Aplidin was most active against acute lymphoblastic leukemia (ALL) cell lines. In vivo, Aplidin induced significant differences in EFS distribution in 12 of 28 (43%) solid tumor models and 2 of 6 evaluable ALL models. Aplidin showed potent in vitro activity and induced significant in vivo tumor growth inhibition in some xenografts, but did not induce tumor regressions.

Initial testing of dasatinib by the pediatric preclinical testing program

Dasatinib, a dual inhibitor of the src and abl tyrosine kinases, was recently approved by the Federal Drug Administration for the treatment of imatinib mesylate-resistant chronic myeloid leukemia. Dasatinib was tested against the Pediatric Preclinical Testing Program (PPTP) in vitro panel at concentrations ranging from 0.1 nM to 1.0 microM and was tested in vivo at a dose of 50 mg/kg administered orally twice daily 5 days per week for 4 weeks for the solid tumor xenografts and once daily for the acute lymphoblastic leukemia (ALL) xenografts. Dasatinib was selectively active against the cell lines of the PPTP in vitro panel, reaching an IC(50) in 6 of the 22 lines. The most sensitive were the AML line Kasumi-1, which has a gain-of-function c-Kit mutation (Asn822Lys), and the rhabdoid tumor line CHLA-266 (IC(50) approximately 10 nM for each). In the in vivo panel, dasatinib induced significant differences in EFS distribution in 8 of 32 (25%) solid tumor models and 3 of 7 ALL models. Using the time to event activity measure, dasatinib had intermediate activity against 1 of 27 (4%) evaluable solid tumor xenografts and 3 of 7 ALL xenografts. One xenograft in the ALL panel, a Philadelphia chromosome positive (Ph(+)) ALL xenograft, demonstrated a complete response. Dasatinib was active at low nanomolar concentrations against a small subset of the PPTP's in vitro panel. Dasatinib had limited in vivo activity against the PPTP solid tumor xenografts, but was highly active against a Ph(+) ALL xenograft and also had anti-leukemia activity against two other xenografts.

Initial testing (stage 1) of the mTOR inhibitor rapamycin by the pediatric preclinical testing program

Rapamycin is a highly specific inhibitor of mTOR, a serine/threonine kinase that controls cap-dependent translation. Here we report the activity of rapamycin against the in vitro and in vivo panels of the Pediatric Preclinical Testing Program (PPTP). Rapamycin was tested against the in vitro panel at concentrations from 0.01 to 100 nM and was tested against the in vivo tumor panels by i.p. administration daily x 5 for 6 consecutive weeks at a dose of 5 mg/kg. Rapamycin variably inhibited growth of the cell lines in the PPTP in vitro panel, with maximal inhibition values ranging from 19% to 85% (median 49%) and a median EC(50) of 0.7 nM. Ten of 23 cell lines achieved at least 50% growth inhibition. Against the in vivo panels, rapamycin induced significant differences in EFS distribution in 27 of 36 solid tumor xenografts and in 5 of 8 ALL xenografts. Using the time to event activity measure, rapamycin had intermediate or high activity against 14 of 31 evaluable solid tumor xenografts and 5 of 8 ALL xenografts. Objective responses were observed in several panels, including: rhabdoid tumor (1PR), rhabdomyosarcoma (2PR), and osteosarcoma (1 maintained CR). Two T-cell ALL xenografts had objective responses (1PR, 1 maintained CR). Rapamycin demonstrated broad antitumor activity against the PPTP's in vivo tumor panels, with particularly noteworthy activity for selected sarcoma and ALL xenografts. Future work will evaluate the molecular characteristics of responding models and the activity of combinations of rapamycin with other anticancer agents.

Pediatric preclinical testing program (PPTP) efficacy and pharmacodynamic evaluation of the Hsp90 inhibitor 17-DMAG

3575 Background: 17-DMAG is a small-molecule inhibitor of the protein chaperone HSP90 that is being developed as an anticancer agent because of the multiple HSP90 client proteins involved in cancer cell growth and survival. Methods: The PPTP includes an in vitro panel (n=27) as well as panels of xenografts (n=61) representing most of the common types of childhood solid tumors and childhood ALL. 17-DMAG was tested against the in vitro panel at concentrations from 1 nM to 10 microM and was tested against the in vivo tumor panels by IP administration using a 50 mg/kg BID twice weekly x 6 weeks dose and schedule. The PPTP’s 3 measures of antitumor activity were used (Houghton et al. Ped Blood Cancer 2006): 1) an objective response measure; 2) treated to control (T/C) tumor volume at day 21; and 3) a time to event (EFS T/C) measure. HSP70 induction was was used as a pharmacodynamic measure of HSP90 inhibition and was determined in tumor and liver tissue at 8 and 24 hours following the second of two doses of 17-DMAG (50 mg/kg IP) administered at 12 hour intervals. Results: 17-DMAG had an EC50 of 62 nM against the PPTP’s in vitro panel, with a trend for lower EC50 values for the rhabdomyosarcoma panel (median EC50 31 nM) compared to the remaining PPTP in vitro cell lines (p=0.06) and for higher EC50 values for the neuroblastoma lines (median EC50 396 nM, p=0.01). 17-DMAG induced significant differences in EFS distribution in 15 of 30 of the solid tumor xenografts, and in 4 of 6 of the evaluable ALL xenografts. Using the time to event activity measure, 17-DMAG had intermediate or high activity against 4 of 28 evaluable solid tumor xenografts (1 of 2 rhabdoid tumor and 3 of 4 alveolar rhabdomyosarcoma). The only objective response (a PR) observed was for an alveolar rhabdomyosarcoma xenograft. HSP70 induction was observed in both liver and tumor tissue, with robust induction (up to 450% increase versus control) occurring in both responding and non-responding tumors. Conclusions: 17-DMAG produced its greatest antitumor activity against alveolar rhabdomyosarcoma xenografts. Robust HSP70 induction was observed in both responding and non-responding xenografts, suggesting that tumor-specific downstream effects of HSP90 inhibition are primary determinants of response. (Supported by NCI NO1CM42216) No significant financial relationships to disclose.