Rachel Courtney

Selective CDK4/6 inhibition with tumor responses by PD0332991 in patients with mantle cell lymphoma

Mantle cell lymphoma (MCL) carries an unfavorable prognosis and requires new treatment strategies. The associated t(11:14) translocation results in enhanced cyclin D1 expression and cyclin D1-dependent kinase activity to promote cell-cycle progression. A pharmacodynamic study of the selective CDK4/6 inhibitor PD0332991 was conducted in 17 patients with relapsed disease, using 2-deoxy-2-[(18)F]fluoro-D-glucose (FDG) and 3-deoxy-3[(18)F]fluorothymidine (FLT) positron emission tomography (PET) to study tumor metabolism and proliferation, respectively, in concert with pre- and on-treatment lymph node biopsies to assess retinoblastoma protein (Rb) phosphorylation and markers of proliferation and apoptosis. Substantial reductions in the summed FLT-PET maximal standard uptake value (SUV(max)), as well as in Rb phosphorylation and Ki-67 expression, occurred after 3 weeks in most patients, with significant correlations among these end points. Five patients achieved progression-free survival time of > 1 year (range, 14.9-30.1+ months), with 1 complete and 2 partial responses (18% objective response rate; 90% confidence interval, 5%-40%). These patients demonstrated > 70%, > 90%, and ≥ 87.5% reductions in summed FLT SUV(max) and expression of phospho-Rb and Ki67, respectively, parameters necessary but not sufficient for long-term disease control. The results of the present study confirm CDK4/6 inhibition by PD0332991 at a well-tolerated dose and schedule and suggest clinical benefit in a subset of MCL patients. This study is registered at www.clinicaltrials.gov under identifier NCT00420056.

Treatment with PF-04449913, an oral smoothened antagonist, in patients with myeloid malignancies: a phase 1 safety and pharmacokinetics study

BACKGROUND Activation of the Hedgehog signalling pathway contributes to cancer progression and the development of myeloid leukaemia stem cell therapeutic resistance. We aimed to identify the maximum tolerated dose (MTD) and the recommended phase 2 dose of the selective Hedgehog antagonist PF-04449913 in myeloid malignancies. METHODS We undertook an open-label, dose-finding, standard 3+3 design phase 1 study of PF-04449913 in adult patients with acute myeloid leukaemia, chronic myeloid leukaemia, chronic myelomonocytic leukaemia, myelodysplastic syndrome, or myelofibrosis who were refractory, resistant, or intolerant to previous treatments, at three centres in the USA and one in Italy. Patients who had newly diagnosed, untreated disease were included if they were not eligible for standard treatment options or if standard treatments were not deemed appropriate. Patients received PF-04449913 once daily continuously until disease progression, unacceptable toxic effects, or patient withdrawal for up to 12 28-day cycles. Additional cycles were given if patients showed evidence of clinical benefit. The starting dose was 5 mg and was increased by 100% until the first dose-limiting toxic effect (DLT) and by 50% thereafter, in keeping with a 3+3 clinical trial statistical design. The primary endpoint was first-cycle DLTs. Secondary endpoints were safety, tolerability, pharmacokinetics, pharmacodynamics, and preliminary clinical activity. This trial is registered with ClinicalTrials.gov, number NCT00953758. FINDINGS Between March 24, 2010, and Sept 7, 2012, 47 patients were enrolled and included in the study: 28 with acute myeloid leukaemia, six with myelodysplastic syndrome, five with chronic myeloid leukaemia (two with chronic-phase and three with blast-phase disease), one with chronic myelomonocytic leukaemia, and seven with myelofibrosis. Patients received PF-04449913 once daily at 5 mg (n=3), 10 mg (n=3), 20 mg (n=4), 40 mg (n=4), 80 mg (n=8), 120 mg (n=3), 180 mg (n=3), 270 mg (n=5), 400 mg (n=9), and 600 mg (n=5). Two patients experienced DLTs (one each in the 80 mg and 600 mg dose groups). The MTD for PF-04449913 was established to be 400 mg once daily. Of the 47 patients enrolled, 28 (60%) experienced treatment-related adverse events, three of which were grade 4 in severity. The most common treatment-related adverse events included dysgeusia (13 [28%] patients), decreased appetite (nine [19%]), and alopecia (seven [15%]). None of the 15 deaths reported were treatment related. Pharmacokinetics seemed to be dose proportional. The mean half-life was 23·9 h (SD 14·0) in the MTD group. Some suggestion of clinical activity was noted in 23 (49%) of 47 patients with haematological malignancies. Based on these results, the recommended phase 2 dose was 200 mg or lower once daily. INTERPRETATION Based on these findings, PF-04449913 is being tested in phase 2 studies in patients with myelodysplastic syndrome, acute myeloid leukaemia, and myelofibrosis. FUNDING Pfizer.

A Phase I Study of PF-04449913, an Oral Hedgehog Inhibitor, in Patients with Advanced Solid Tumors

Purpose: To estimate the maximum tolerated dose (MTD) of single-agent PF-04449913, and to evaluate safety, tolerability, pharmacokinetics, pharmacodynamics, and preliminary antitumor activity in patients with advanced tumors. Experimental Design: A 3+3 design was used in this open-label, multicenter, phase I study and dose escalation/de-escalation applied until identification of the MTD. PF-04449913 was orally administered once daily in continuous 28-day treatment cycles. The starting dose was 80 mg. Results: A total of 23 patients were enrolled; 19 were evaluable for first-cycle dose-limiting toxicity (DLT). The first-cycle DLT rate at the 640 mg dose level was 33.3%, and the MTD was estimated to be 320 mg once daily. The recommended phase II dose was not determined. PF-04449913 was generally well tolerated at doses of 80 to 320 mg once daily. The most common treatment-related adverse events (AE) were grade 1–2 dysgeusia, fatigue, decreased appetite, nausea, dizziness, dehydration, and diarrhea. Treatment-related grade 3 AEs only occurred in patients receiving PF-04449913 640 mg once daily. No treatment-related grade 4–5 AEs were reported. Pharmacokinetic analysis indicated a generally dose-proportional kinetics with biphasic elimination, supporting once-daily dosing. PF-04449913 modulated hedgehog signaling at the dose levels tested, as demonstrated by >80% downregulation of GLI1 expression in the skin of treated patients. Eight patients (34.8%) achieved stable disease; none had complete or partial response. Three patients with disease progression at enrollment had prolonged disease stabilization (≥6 months). Conclusions: The results obtained in this study support further evaluation of PF-04449913 in patients with advanced solid tumors. Clin Cancer Res; 21(5); 1044–51. ©2014 AACR.

Latanoprost systemic exposure in pediatric and adult patients with glaucoma: a phase 1, open-label study.

OBJECTIVE To evaluate short-term safety and steady-state systemic pharmacokinetics (PK) of latanoprost acid in pediatric subjects with glaucoma or ocular hypertension who received the adult latanoprost dose. DESIGN Phase 1, open-label, multicenter study. PARTICIPANTS Pediatric patients of 3 age groups (<3, 3-<12, and 12-<18 years) and adults (≥18 years) receiving latanoprost ophthalmic solution 0.005% once daily in 1 or both eyes for ≥2 weeks. INTERVENTION Latanoprost was administered in both eyes each morning post-screening. Subjects returned 3 to 28 days later for evaluation of plasma concentrations, withholding morning latanoprost. At the clinic, a single drop of latanoprost ophthalmic solution was instilled into both eyes. Plasma latanoprost acid concentrations were collected predose and 5, 15, 30, and 60 minutes after administration. MAIN OUTCOME MEASURES Latanoprost acid plasma exposure. RESULTS The evaluable PK analysis set included data from 39 of 47 enrolled subjects. The median peak plasma concentration value was higher in the <3-year age group (166 pg/ml) versus other groups (49, 16, and 26 pg/ml for the 3-<12-year, 12-<18-year, and ≥18-year age groups, respectively). The median area under the concentration-time curve from zero to last measurable concentration value was also higher in the <3-year age group (2716 pg/min/ml) versus other groups (588, 106, and 380 pg/min/ml for the 3-<12-year, 12-<18-year, and ≥18-year age groups, respectively). Latanoprost acid was rapidly eliminated from the blood, with plasma concentrations undetectable within 10 to 30 minutes postdose in all but the <3-year age group. There were no discontinuations or dose reductions due to adverse events or treatment-emergent adverse events. CONCLUSIONS Latanoprost acid systemic exposure was higher in younger children versus adolescents and adults, attributed primarily to lower body weight and smaller blood volume. Latanoprost acid was eliminated rapidly in all age groups and resulted in only a brief period of systemic exposure after once-daily dosing. Higher systemic exposure was not accompanied by adverse events, and on the basis of extrapolation of safety data from adults, this pilot study suggests an adequate safety margin for systemic adverse effects with use of the adult topical dose of latanoprost in pediatric patients. FINANCIAL DISCLOSURE(S) Proprietary or commercial disclosures may be found after the references.

A Phase 2, Randomized, Dose-Response Trial of Taprenepag Isopropyl (PF-04217329) Versus Latanoprost 0.005% in Open-Angle Glaucoma and Ocular Hypertension

Purpose: To evaluate the safety of escalating doses of taprenepag isopropyl (PF-04217329), a selective EP2 receptor agonist administered as a topical ophthalmic solution, versus its vehicle (Stage I), and dose-response of taprenepag isopropyl alone and in unfixed combination with latanoprost ophthalmic solution 0.005% versus latanoprost alone (Stage II). Subjects and Methods: Randomized, vehicle- and active-controlled, double-masked, two-stage, dose-finding trial in primary open-angle glaucoma (POAG) or ocular hypertension; first taprenepag isopropyl study in patients (NCT00572455). Study eye: 26 mmHg ≤ intraocular pressure (IOP) <36 mmHg at 8 am and 22 mmHg ≤ IOP <36 mmHg at 10 am, 1 pm, 4 pm. Stage I: 3 cohorts (total n = 67) received 1 drop of taprenepag isopropyl unit dose formulation qPM/eye for 14 days: low dose: 0.0025%, 0.005%, vehicle; middle dose: 0.01%, 0.015%, vehicle; high dose: 0.02%, 0.03%, vehicle. Stage II: 7 groups (total n = 250) received 1 drop of taprenepag isopropyl multidose formulation qPM/eye for 28 days: 0.005%, 0.01%, 0.015% monotherapy; each in unfixed combination with latanoprost 0.005%, or latanoprost monotherapy. Main outcomes: mean change in diurnal IOP, baseline to last visit; adverse events. Results: Stage I at Day 14: statistically significantly greater IOP reductions were observed at all taprenepag isopropyl doses versus vehicle. Stage II at Day 28: statistically significantly greater IOP reductions were observed at all doses of the unfixed combination versus latanoprost monotherapy. At least 1 treatment-emergent adverse event reported for 29/67 (43.3%) subjects in Stage I and 158/250 (63.2%) in Stage II. Conclusions: Taprenepag isopropyl significantly reduces IOP in POAG and ocular hypertension. Taprenepag isopropyl monotherapy is comparable to latanoprost 0.005% in reducing IOP. As demonstrated in this report, the activity of taprenepag isopropyl is additive to that of latanoprost 0.005%. Further studies are required to determine whether it shows similar additivity when administered with other ocular antihypertensive medications.

Phase I Study of PD 0332991, a Novel, Oral, Cyclin-D Kinase (CDK) 4/6 Inhibitor in Combination with Letrozole, for First-Line Treatment of Metastatic Post-Menopausal, Estrogen Receptor-Positive (ER+), Human Epidermal Growth Factor Receptor 2 (HER2)-Negative Breast Cancer.

Background : PD 0332991 is a potent, oral, small molecule inhibitor of CDK 4/6 kinase. Preclinical studies identified that human luminal ER+ breast cancer cell lines were uniquely sensitive to G0/G1 arrest and growth inhibition by PD 0332991 and this appears to be related to an intact Rb pathway in this subtype (Finn, SABCS 2008, abstract 5064). These studies also identified the ability of PD 0332991 to act synergistically with anti-hormonal treatment in vitro and to restore sensitivity to hormonal treatment in models of acquired resistance. Based on these observations, a phase I/II randomized study of PD 0332991 in combination with letrozole versus letrozole alone as first-line treatment of metastatic ER+ breast cancer was launched. Methods: Post-menopausal women with ER+, HER2-negative breast cancer were eligible. Other eligibility criteria included Eastern Cooperative Oncology Group performance status of 0 or 1, no brain metastases, and adequate organ function. The phase I part of the study was conducted to assess the safety and tolerability of the combination including dose-limiting toxicities (DLTs). Enrollment in phase I is planned for up to 12 patients. The starting dose was PD 0332991 125 mg daily for 3 weeks followed by 1 week off (Schedule 3/1) and letrozole 2.5 mg daily. Cycle 1 consisted of PD 0332991 administered for 2 weeks alone followed by 1 week off. Subsequent cycles consisted of PD 0332991 on Schedule 3/1 and letrozole administered continuously. Pharmacokinetic samples were collected to assess the potential for a drug–drug interaction (DDI). Archival tumor tissue was required for biomarker evaluation. The tumor assessments were performed every 8 weeks. Results: Nine patients have currently been enrolled and treated; median duration of treatment was 3 months (range Conclusions: The combination of PD 0332991 and letrozole is well tolerated. Encouraging activity is seen with the combination and a randomized phase II part of the study will be initiated. Updated results on the phase I cohort will be presented at the meeting. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5069.

Abstract 906: Gas1 and Kif27 genes are strongly up-regulated biomarkers of Hedgehog inhibition (PF-04449913) on leukemia stem cells in Phase I Acute Myeloid Leukemia and Chronic Myeloid Leukemia treated patients

Hedgehog (Hh) pathway activation contributes to leukemia development and growth, and that targeted pathway inhibition is likely to offer an efficient therapeutic opportunity. PF-04449913, a Hh pathway inhibitor, is a new selective and potent inhibitor of leukemia self-renewal and is currently being evaluated in phase I clinical trials. In order to identify new potential clinical biomarkers for the PF-04449913, we studied CD34+ leukemia stem cell population (LSC) collected before and after 28 days treatment in a phase I dose escalation protocol (Clinical Trial Gov. NTC00953758) enrolling selected hematological malignancies. This experimental clinical trial enrolled Myelofibrosis (MF), MDS, blastic phases CML, chronic myelomonocytic leukemia (CMML) and AML patients (pts). We were able to collect and separate highly purified (98%) bone marrow CD34+ cells from 5 AML, 1 MF and 2 CML pts by immunomagnetic separation, and analysed them for gene expression profile using Affimetrix HG-U133 Plus 2.0 platform. 1197 genes were differentially expressed between CD34+ cells collected before and after 28 days of PF-04449913 dose finding oral therapy. Clustering of their expression profiles showed that mostly genes differentially expressed are mainly related to Hh signaling, this providing further evidences that PF-04449913 really therapeutically targets the Hh pathway. Regarding genes involved in Hh signaling pathway, Gas1 and Kif27 were strongly upregulated (fold change 1.0947 and 1.12757 respectively; p-value 0.01 and 0.02 respectively) in CD34+ LSC after 28 days exposure to PF-04449913 as compared to baseline, suggesting these two genes have potential as new biomarkers of activity. GAS-1 is a Sonic Hedgehog (Shh)-binding protein; it acts to sequester Shh and inhibit the Shh signalling pathway. Kif27 mainly acts as a negative regulator in the Hh signaling pathway, and inhibits the transcriptional activator activity of Gli1 by inhibiting its nuclear translocation. Other genes were differentially expressed after ‘ex- vivo’ treatment with PF-04449913 as compared to baseline: we observed a down regulation of Bcl2 (fold change -1.03004), ABCA2 (fold change -1.08966), LEF1 (fold change -1.28457), Gli1 (fold change -1.0775), Smo (fold change -1.07702), and an upregulation of Gli2 (fold change 1.08191). Conclusions: This data demonstrates that PF-04449913 specifically targets the Hh Pathway in CD34+ cells, suggesting that Hh inhibition may impair leukemia stem cell maintenance. In addition, we identify several new potential biomarkers (e.g. Gas1 and KIF27). Taken together, these data may be useful for pts selection strategies and subsequent eradication of the LSC. Acknowledgments: Work supported by Pfizer, European LeukemiaNet, FIRB 2006, AIRC, AIL, COFIN, University of Bologna and BolognAIL. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 906. doi:1538-7445.AM2012-906

Abstract 4619: PF-04449913 reverts multi drug resistance (MDR) by a strong down-regulation of ABCA2 and BCL2 on leukemia stem cells in phase I acute myeloid leukemia and chronic myeloid leukemia treated patients

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL PF-04449913 is a selective and potent inhibitor of the Hh pathway and leukemia self-renewal and is currently being evaluated in Phase I clinical trials. We studied leukemia stem cell population (CD34+ subpopulation) collected before and after 28 days treatment in a phase I dose escalation protocol (Clinical Trial Gov. NTC00953758) enrolling selected hematological malignancies including MF, MDS, CML, CMML and AML. We collected and separated highly purified (98%) bone marrow hematopoietic progenitor cells (CD34+ populations) in 5 AML, 1 MF and 2 CML patients, by immunomagnetic separation, and analyzed them for gene expression profile (GEP) using Affimetrix HG-U133 Plus 2.0 platform. We have observed that 1197 genes were differentially expressed between CD34+ cells collected before and after 28 days of PF-04449913 dose finding oral therapy. We demonstrated a down regulation of Bcl2 (fold change –1.03004; p value= 0.01), ABCA2 (fold change –1.08966; p value=0.03), Bcl2l13 (fold change –1,04259; p-value=0,027642), Bcl2l2 (fold change –1,17214; p-value=0,000768), Casp4 (fold change –1,06551; p-value=0,032428), Casp7 (fold change –1,01569; p-value=0,006688), Casp10 (fold-change –1,3076; p-value=0,050431), ABCF1 (fold change –1,04999; p-value=0,07213). On the contrary, ABCB1 (fold change 1,46592) and ABCG2 (fold change –1,16103) are respectively up and down regulated, with a not statistically significant p-value. Bcl2 (B-cell lymphoma 2), Bcl2l2 (Bcl2-like protein 2) and Bcl2l13 (Bcl2-like 13) are the founding members of the Bcl-2 family of apoptosis regulator proteins. Recent studies showed that Hh signals upregulate Bcl2 to promote cellular survival. Casp 4,7,10 (Caspases) are a family of cysteine proteases that play essential roles in apoptosis, necrosis, and inflammation. ABCA2 (ATP-binding cassette sub-family A member 2), ABCF1 (ATP-binding cassette sub-family F member 1), ABCB1 (ATP-binding cassette sub-family B member 1, MDR1), ABCG2 (ATP-binding cassette sub-family G member 2) belong to the superfamily of adenosine triphosphate-binding cassette (ABC) transporters. One mechanism of MDR is the increased expression of ABC drug transporters, resulting in low intracellular drug concentrations. We evaluated Gli1 and Smo expression by GEP, comparing data before and after 28 days of treatment with PF-04449913 and we observed a down regulation of both genes (fold changes –1.0775 and –1.07702 respectively). PF-04449913 is able to revert MRD mechanisms of LSC by a strong down regulation of genes (Bcl-2, Bcl-2l13, Bcl-2l2, ABCA2, and ABCF1), which are critical for chemoresistance in acute and chronic leukemia patients. Acknowledgments. Pfizer, European LeukemiaNet, FIRB 2006, AIRC, AIL, COFIN, University of Bologna and BolognAIL. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4619. doi:1538-7445.AM2012-4619