Phase II trial of the histone deacetylase inhibitor pivaloyloxymethyl butyrate (Pivanex, AN-9) in advanced non-small cell lung cancer

Lung Cancer (2004) 45, 381—386

Phase II trial of the histone deacetylase inhibitor pivaloyloxymethyl butyrate (Pivanex, AN-9) in advanced non-small cell lung cancer Tony Reid a , Frank Valone b , William Lipera c , David Irwin d , Warren Paroly e , Ron Natale f , Sunil Sreedharan b , Harold Keer b , Bert Lum g , Frank Scappaticci b , Anish Bhatnagar b,* a

VA Medical Center and Stanford University, 3801 Miranda Drive, MC111 ONC, Palo Alto, CA 94304, USA b Titan Pharmaceuticals Inc., 400 Oyster Point Boulevard, Suite 505, South San Francisco, CA 94080, USA c North Shore Hematology Oncology Associates, 235 N. Belle Mead Road, E. Setauket, NY 11733, USA d Alta Bates Cancer Center, 2001 Dwight Way, Berkeley, CA 94704, USA e Sidney Kimmel Cancer Center, 10835 Altman Row, San Diego, CA 92121, USA f The Cedars Sinai Comprehensive Cancer Center, 8700 Beverly Boulevard, RM AC-1237, Los Angeles, CA 90048, USA g Stanford University, 300 Pasteur Drive, Stanford, CA 94305, USA Received 15 October 2003 ; received in revised form 8 March 2004; accepted 15 March 2004

KEYWORDS Pivanex; Histone deacetylase inhibitor; Non-small cell lung cancer; AN-9

Summary This multicenter phase II trial evaluated the therapeutic activity and safety profile of pivaloyloxymethyl butyrate (Pivanex, AN-9) as a single agent in refractory non-small cell lung cancer (NSCLC). Pivanex (2.34 g/m2 per day) was administered as a 6-h continuous intravenous infusion, daily for 3 days, and repeated every 21 days until disease progression. Forty-seven patients were treated. More than 90% of patients had received both a platinum compound and a taxane and 32% had received three or more prior chemotherapy regimens. The most common toxicities were transient grade 1—2 fatigue (34%), nausea (17%), and dysgeusia (11%). Three patients had partial responses (6.4 and 95%; CI 1.4—18.7%) and 14 patients had stable disease for ≥12 weeks (30%). Median survival for all patients was 6.2 months with 1-year survival of 26%. For patients who received fewer than three prior chemotherapy regimens, median survival was 7.8 months and 1-year survival was 31%. Pivanex is well tolerated and appears to be active as a single agent in patients with advanced NSCLC refractory to previous chemotherapy. Based on its therapeutic activity and favorable safety profile, further studies of Pivanex in NSCLC, particularly in combination with current chemotherapeutic agents, are warranted. © 2004 Elsevier Ireland Ltd. All rights reserved.

1. Introduction * Corresponding

author. Tel.: +1-650-244-4990; fax: +1-650-244-4956. E-mail address: [email protected] (A. Bhatnagar).

Pivanex is a small molecule inhibitor of histone deacetylases (HDACs) that has anti-proliferative,

0169-5002/$ – see front matter © 2004 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.lungcan.2004.03.002

382 differentiating, and pro-apoptotic effects [1,2]. Pivanex inhibits clonogenicity of fresh tumor cells from a wide range of human solid tumors including non-small cell lung carcinoma (NSCLC), colo-rectal cancer, breast cancer, and melanoma [3]. It is also cytotoxic to fresh human chronic lymphocytic leukemia (CLL) cells [4] and acute lymphocytic leukemia (ALL) cells [5]. Pivanex inhibits growth of Lewis lung, EBC-1, SK-Mes-1, Calu-6, PC9, and AS49 lung cancer cell lines in vitro [6—8]. In pre-clinical animal models, Pivanex decreases pulmonary metastases after intravenous injection of 3LLD122, Lewis lung carcinoma cells [6]. Pivanex also increases survival of mice injected with Lewis lung carcinoma [6] or monocytic leukemia MmA cells [8]. Inhibition of histone deacetylases by Pivanex results in hyperacetylation of histones leading to altered gene expression and cell differentiation or cell death [9—11]. Pivanex decreases expression of the anti-apoptotic protein Bcl-2 and increases expression of the pro-apoptotic protein BAX in fresh human CLL cells and this may account for Pivanex’s apoptotic effect on CLL cells [4]. Furthermore, Pivanex increases expression of p53, which may contribute to tumor cell apoptosis [7]. Inhibition of cell proliferation may also result from Pivanex-induced increases in active (hypophosphorylated) retinoblastoma protein and decreases in inactive (hyperphosphorylated) retinoblastoma protein [10]. Finally, Pivanex promotes differentiation of leukemia and solid tumor cell lines, perhaps through modulation of expression of oncogenes such as c-myc and c-jun [7,11]. Pivanex was developed as a pro-drug of the differentiating agent butyric acid [1]. Pre-clinical studies, however, demonstrated that Pivanex is substantially more potent than butyric acid for altering gene expression and inducing tumor cell differentiation or death [1,8,11]. More rapid and complete cell uptake may account for Pivanex’s enhanced efficacy [12]. In a phase I trial, doses of Pivanex up to 3.3 g/m2 per day were well tolerated when given on days 1—5 of a 21-day treatment cycle [13]. One patient with squamous cell lung cancer had an objective clinical response to Pivanex. Based on the extensive pre-clinical data that suggested Pivanex might be active in NSCLC, we undertook a phase II trial of Pivanex in patients with chemotherapy-resistant NSCLC. The goals of the trial were to assess anti-cancer activity of Pivanex as a single agent and to obtain additional safety information on potential toxicities.

T. Reid et al.

2. Patients and methods 2.1. Patient eligibility Patients with histologically or cytologically confirmed NSCLC that was locally recurrent or metastatic and had progressed after standard chemotherapy were eligible for the trial. Additional eligibility criteria were: (1) age ≥18; (2) ECOG performance status ≤2; (3) no chemotherapy, radiotherapy, or investigational therapy within 4 weeks prior to Pivanex treatment; (4) one or more index pulmonary lesions that were bidimensionally measurable by radiographs; (5) adequate hematological function (WBC >2500 ␮l−1 , absolute neutrophil count >1500 ␮l−1 , platelets ≥100,000 ␮l−1 , and hemoglobin ≥9 g/dl); (6) adequate hepatic function (total bilirubin level ≤1.5 times the upper normal limit, aspartate aminotransferase and alanine aminotransferase of ≤2.0 times the upper limit of normal); (7) normal serum creatinine; (8) serum amylase <1.5 times the upper normal limit; (9) serum triglycerides <400 mg/ml; (10) New York Heart Association Class I or II; and (11) no coexisting medical conditions that would interfere with treatment. Written informed consent was obtained from patients according to federal and institutional guidelines. The study was approved by the appropriate ethics committees at each institution.

2.2. Treatment Pivanex (Titan Pharmaceuticals Inc., South San Francisco) was infused at a dose of 2.34 g/m2 over 6 h by peripheral vein. Treatment was given on days 1—3 and repeated every 21 days until disease progression. For the infusions, Pivanex was mixed with 20% intralipid at a final concentration of 27 mg Pivanex/ml.

2.3. Evaluation of response and toxicity Bidimensionally measurable tumor masses were evaluated at baseline and at the end of every two cycles. Complete response (CR) was defined as a 100% disappearance of all tumor masses (measurable or not) and partial response (PR) was defined as 50—99% decrease in the sum of products of the longest perpendicular diameters (SPPD) of all measurable tumors. Stable disease (SD) was defined as <50% decrease or <25% increase in SPPD of all measurable tumors. Progressive disease (PD) was defined as ≥25% increase in SPPD or appearance of new lesions. Reported responses are best responses seen while on study based on the WHO criteria. Toxi-

Phase II trial of Pivanex, AN-9 in advanced non-small cell lung cancer cities were graded according to the National Cancer Institute Common Toxicity Criteria (version 1.0).

2.4. Statistical analysis Statistical analyses were performed using SAS v.8e (SAS Institute Inc., Cary, North Carolina). Survival probabilities were computed using the Kaplan—Meier product limit estimates and plotted (survival plots) using the STAT/GRAPH procedures (PROC PLOT). Patient variables were of binomial nature (yes/no) and included histology (squamous and non-squamous), and number of prior chemotherapy regimens (<3 and ≥3). Analysis of the influence of patient variables on survival was performed using the Logrank test to compare survival distributions. Time to disease progression and overall survival endpoints were calculated as the time from the first Pivanex dose to the specified event. All enrolled patients were evaluated for the study’s endpoints.

383

Table 1 Frequent (≥5%) adverse events potentiallya related to Pivanex (n = 47) Event

Fatigue Nausea Dysgeusia Asthenia Cough Anorexia Dyspnea Injection site pain Injection site burning Vision blurred Headache Chest pain Pyrexia

n (%) Overall

Grade 3

Grade 4

16 8 5 5 5 4 4 4 3 3 3 3 3

1 (2%) 0 0 1 (2%) 0 0 0 0 0 0 0 1 (2%) 0

0 0 0 0 0 0 2 (4%) 0 0 0 0 1 (2%) 0

(34%) (17%) (11%) (11%) (11%) (9%) (9%) (9%) (6%) (6%) (6%) (6%) (6%)

a

Includes events considered unlikely to be related to Pivanex.

3. Results

ing two full courses of Pivanex therapy. The median number of Pivanex courses was two (mean: 3.6; range: 0—25).

3.1. Patient characteristics

3.2. Adverse events

Patients were enrolled at 10 separate clinical centers in this multicenter trial from 14 January 1999 to 16 July 2001. The median age of patients was 66 (range 43—82). The patients were a relatively poor prognostic group with only seven patients (15%) having a performance status of 0 and eight patients (17%) having a performance status of 2. Forty-three percent of patients had squamous cell tumors and the rest had a variety of non-squamous cell tumor histologies. A majority of patients (72%) had metastatic disease (stage IV) at disease presentation and the remainder presented with stage IIIa/IIIb disease that was refractory or relapsed at the time of protocol registration. The patients were heavily pre-treated with 15 patients (32%) having received ≥3 different chemotherapy regimens. Forty-five patients (96%) received a platinum compound (cisplatin and/or carboplatin) and 42 patients (89%) had received a taxane (paclitaxel or docetaxel). Prior chemotherapy agents and the number of patients who received them were carboplatin (40), cisplatin (12), paclitaxel (38), docetaxel (6), gemcitabine (18), navelbine (17), and other (23). The dose of Pivanex was not reduced for toxicity in any patient. Eight patients experienced rapid disease progression (n = 5) or an adverse event (n = 3) leading to treatment discontinuation before receiv-

Pivanex was generally well tolerated. Hematologic toxicity was minimal and no patient experienced grade 3 or 4 neutropenia. One patient experienced grade 3 thrombocytopenia (45,000 mm−3 ) and one patient experienced grade 4 anemia (hemoglobin = 5.4 g/dl). There were no episodes of hepatotoxicity or nephrotoxicity. Table 1 presents the most frequent (≥5%) clinical adverse events that were scored as Pivanex-related even if the investigator considered the relationship to be unlikely. Almost all treatment-related adverse events were mild/moderate in severity (grades 1—2), occurred during the Pivanex infusion, and were rapidly reversible. Two grade 3 adverse events were reported as possibly related to Pivanex: fatigue and hypokalemia. One episode of grade 4 hypersensitivity was reported as possibly related to Pivanex. This episode occurred at the onset of the patient’s second dose of Pivanex. The patient was treated with supportive care, with complete resolution (without hospitalization), and Pivanex was discontinued.

3.3. Tumor response and patient survival For the overall population, three patients experienced partial responses (6.4 and 95%; CI 1.4—18.8%) (two confirmed, one unconfirmed) and 14 patients

384

T. Reid et al.

Table 2 Efficacy of Pivanex in advanced NSCLC All patients (n = 47)

Patients who received <3 prior therapies (n = 32)

Best tumor response Partial response Stable disease (≥12 weeks)

3 (6.4%) 14 (30%)

3 (9.4%) 12 (40%)

Survival Progression-free survival, median Overall, median 1-year survival

1.5 months (3—634 days) 6.2 months (6—772 days) 26%

2.5 months (6—634 days) 7.8 months (6 to >772 days) 31%

(30%) experienced stable disease for ≥12 weeks (Table 2). The three responding patients received fewer than three prior chemotherapy regimens. Disease stabilization ranged from 12 to >38 weeks with one long-term responding patient completing 25 cycles of Pivanex before stopping therapy while still in remission. The protocol did not collect quality of life data prospectively. For all patients median progression free survival was 1.5 months and median overall survival was 6.2 months (Fig. 1). Twenty-six percent of patients were alive at 1 year. Median progression-free survival was significantly greater for the 32 patients who were treated with fewer than three prior chemotherapy regiments (2.5 months) compared to patients who received three or more prior chemotherapy regimens (1.4 months) (P = 0.007). As can be seen in Fig. 1, patients who received fewer than three prior chemotherapy regimens

Fig. 1

also had significantly improved overall survival when compared to patients who received three or more prior regimens (median survival = 7.8 and 2.5 months, respectively; P = 0.003).

4. Discussion The results of this study show that Pivanex is well tolerated in patients with advanced non-small cell lung cancer and suggest that Pivanex has therapeutic activity in this setting. Overall, most adverse events related to Pivanex were mild to moderate in severity and transient in duration. The most common adverse events were fatigue, nausea, and dysgeusia. Unlike conventional chemotherapeutic agents, Pivanex has minimal hematological, renal, and hepatic toxicity. Pivanex’s safety profile compares favorably with other HDAC inhibitors. In a

Kaplan—Meier plot of overall survival according to the number of prior chemotherapy regimens.

Phase II trial of Pivanex, AN-9 in advanced non-small cell lung cancer phase I trial, dose-limiting toxicities for depsipeptide were fatigue, nausea and vomiting, thrombocytopenia, and cardiac arrhythmia [14]. Major toxicities with suberoylanilide hydroxamic acid (SAHA) include fatigue, diarrhea, anorexia, dehydration, and myelosuppression [15]. More than 25% of patients receiving MS-275 experience grade 1—2 fatigue, headache, nausea and vomiting, and dose-limiting toxicities include elevated AST, pleural effusion, and epigastric pain [16]. Phenylbutyrate has a safety profile comparable to Pivanex with the most frequent adverse events being reversible fatigue and somnolence [17]. This study suggests that Pivanex is active for treatment of advanced NSCLC. Three patients (6.4%) experienced partial remissions with one response lasting >38 weeks. An additional 14 patients (30%) experienced disease stabilization for 12 or more weeks. Pivanex induces tumor cell differentiation and this effect could result in prolonged survival without tumor regression. In this heavily pre-treated group of NSCLC patients, median survival was 6.2 months with 1-year survival of 26%. For patients who received Pivanex as second or third line therapy, median survival was 7.8 months and 31% of patients were alive at 1 year. Although patient selection may contribute to Pivanex’s apparent favorable effect on survival, the trial was a multicenter, community-based study making such selection bias less likely. These results are comparable to those observed for advanced NSCLC patients treated with docetaxel [18 (95% CI 3.1—13.1%), 19 (95% CI not reported)] or gefitinib [20 (95% CI 11.5—27.3%), 21 (95% CI not reported)]. For patients who had received one or two prior chemotherapy regimens, docetaxel achieves response rates of 5.5 and 6.7%. Median survival with docetaxel treatment is approximately 6 months with 1-year survival of approximately 35%. Patients treated with gefitinib experience higher response rates of 18.4—19%. However, median survival is 7.6 and 8.0 months with 1-year survival of 29 and 35%. Similarly, median survival and 1-year survival rates for patients who received more than two prior chemotherapy regimens are similar after treatment with Pivanex, docetaxel, and gefitinib [18—21]. However, the current study is not a randomized controlled study and the primary endpoint was not survival. Pivanex-treatment causes fewer adverse events than docetaxel and gefitinib, suggesting that Pivanex has a favorable risk-benefit profile. Additional studies that formally assess quality of life after Pivanex treatment are planned to confirm this suggestion. Pivanex’s favorable toxicity profile suggests that full doses of this agent could be combined safely

385

with standard cytotoxic agents. Pre-clinical studies of Pivanex combined with standard cytotoxic agents show synergistic interactions for combinations of Pivanex with docetaxel, gemcitabine, doxorubicin, and daunorubicin [6—8,22], as well as cisplatin, carboplatin, irinotecan, and topotecan (unpublished observations). Synergy appears to be schedule-dependent as maximal synergy with docetaxel requires pre-treatment with Pivanex [22] whereas pre-treatment is not required for synergy with doxorubicin [6]. These observations suggest that more than one molecular mechanism underlies the synergistic interactions of Pivanex and cytotoxic agents.

5. Conclusion The data from this phase II trial show that Pivanex is well tolerated in patients with advanced NSCLC and is indicative of anti-cancer activity. Moreover, the response rate and patient survival characteristics are similar to other agents recently approved in this disease setting. Given Pivanex’s excellent safety profile and pre-clinical data showing synergy with cytotoxic agents used for treatment of NSCLC, further clinical development of Pivanex will focus on combination chemotherapy regimens. A phase I trial of the combination of Pivanex and docetaxel indicate that the combination is well tolerated [23]. We recently opened a randomized phase IIb trial comparing docetaxel monotherapy with the combination of Pivanex and docetaxel as second line therapy in patients with NSCLC.

Acknowledgements We would like to thank David Magnuson for critical review of this manuscript and Kim Lang for administrative assistance. This study was supported by Titan Pharmaceuticals Inc.

References [1] Rephaeli A, Rabizadeh E, Aviram A, Shaklai M, Ruse M, Nudelman A. Derivatives of butyric acid as potential anti-neoplastic agents. Int J Cancer 1991;49:66—72. [2] Nudelman A, Ruse M, Aviram A, Rabizadeh E, Shaklai M, Zimrah Y, et al. Novel anticancer prodrugs of butyric acid 2. J Med Chem 1992;35:687—94. [3] Siu LL, Von Hoff DD, Rephaeli A, Izbicka E, Cerna C, Gomez L, et al. Activity of pivaloyloxymethyl butyrate, a novel anticancer agent, on primary human tumor colony-forming units. Invest New Drugs 1998;16:113—9. [4] Rabizadeh E, Bairey O, Aviram A, Ben Dror I, Shaklai M, Zimra Y. Doxorubicin and a butyric acid derivative ef-

386

[5]

[6]

[7]

[8]

[9]

[10]

[11]

[12]

[13]

[14]

T. Reid et al. fectively reduce levels of Bcl-2 protein in the cells of chronic lymphocytic leukemia patient. Eur J Haematol 2001;66:263—71. Batova A, Shao LE, Diccianni MB, Yu AL, Tanaka T, Rephaeli A, et al. The histone deacetylase inhibitor AN-9 has selective toxicity to acute leukemia and drug-resistant primary leukemia and cancer cell lines. Blood 2002;100: 3319—24. Niitsu N, Kasukabe T, Yokoyama A, Okabe-Kado J, Yamamoto-Yamaguchi Y, Umeda M, et al. Anticancer derivative of butyric acid (pivalyloxymethyl butyrate) specifically potentiates the cytotoxicity of doxorubicin and daunorubicin through the suppression of microsomal glycosidic activity. Mol Pharmacol 2000;58:27—36. Reid T, Lee Y, Eng L, Gadol N, Sreedharan S, Keer H, et al. Pivanex represses oncogene expression and sensitizes tumor cells to chemotherapeutic agents. Proc Am Assoc Cancer Res 2002;43:944a. Kasukabe T, Rephaeli A, Honma Y. An anti-cancer derivative of butyric acid (pivalyloxmethyl butyrate) and daunorubicin cooperatively prolong survival of mice inoculated with monocytic leukaemia cells. Br J Cancer 1997;75:850— 4. Aviram A, Zimrah Y, Shaklai M, Nudelman A, Rephaeli A. Comparison between the effect of butyric acid and its prodrug pivaloyloxymethylbutyrate on histones hyperacetylation in an HL-60 leukemic cell line. Int J Cancer 1994;56:906—9. Sreedharan SP, Wirtz UF, Carrizale G. Pivanex, a short-chain fatty acid histone deacetylase inhibitor, induces changes in key regulatory protein expression in human non-small cell lung carcinoma cells [abstract]. AACR-NCI-EORTC International Conference–—Molecular Targets and Cancer Therapeutics; 2003. Rabizadeh E, Shaklai M, Nudelman A, Rephaeli A. Rapid alteration of c-myc and c-jun expression in leukemic cells induced to differentiate by a butyric acid prodrug. FEBS Lett 1993;328:225—9. Zimra Y, Nudelman A, Zhuk R, Rabizadeh E, Shaklai M, Aviram A, et al. Uptake of pivaloyloxymethyl butyrate into leukemic cells and its intracellular esterase-catalyzed hydrolysis. J Cancer Res Clin Oncol 2000;126:693—8. Patnaik A, Rowinsky EK, Villalona MA, Hammond LA, Britten CD, Siu LL, et al. A phase I study of pivaloyloxymethyl butyrate, a prodrug of the differentiating agent butyric acid, in patients with advanced solid malignancies. Clin Cancer Res 2002;8:2142—8. Sandor V, Bakke S, Robey RW, Kang MH, Blagosklonny MV, Bender J, et al. Phase I trial of the histone deacetylase inhibitor, depsipeptide (FR901228, NSC 630176), in patients

[15]

[16]

[17]

[18]

[19]

[20]

[21]

[22]

[23]

with refractory neoplasms. Clin Cancer Res 2002;8:718— 28. Heany M, O’Connor OA, Richon V. Clinical experience with the histone deacetylase (HDAC) inhibitor suberoylanilide hydroxamic acid (SAHA) in heavily pretreated patients with hematological malignancies. Proc Am Soc Clin Oncol 2003;22:200A (abstract 2321). Ryan QC, Headlee D, Sparreboom A, Figg W, Zhai J, Trepel J, et al. A phase I trial of an oral histone deacetylase inhibitor, MS-275, in advanced solid tumor and lymphoma patients. Proc Am Soc Clin Oncol 2003;22:200A (abstract 802). Gore SD, Weng LJ, Zhai S, Figg WD, Donehower RC, Dover GJ, et al. Impact of the putative differentiating agent sodium phenylbutyrate on myelodysplastic syndromes and acute myeloid leukemia. Clin Cancer Res 2001;7(8):2330— 9. Fossella FV, DeVore R, Kerr RN, Crawford J, Natale RR, Dunphy F, et al. Randomized phase III trial of docetaxel versus vinorelbine or ifosfamide in patients with advanced non-small-cell lung cancer previously treated with platinum-containing chemotherapy regimens. The TAX 320 non-small cell lung cancer study group. J Clin Oncol 2000;18(12):2354—62. Shepherd FA, Dancey J, Ramlau R, Mattson K, Gralla R, O’Rourke M, et al. Prospective randomized trial of docetaxel versus best supportive care in patients with non-small-cell lung cancer previously treated with platinum-based chemotherapy. J Clin Oncol 2000;18(10):2095—103. Fukuoka M, Yano S, Giaccone G, Tamura T, Nakagawa K, Douillard JY, et al. Multi-institutional randomized phase II trial of gefitinib for previously treated patients with advanced non-small-cell lung cancer. J Clin Oncol 2003;21(12):2237—46. Cappuzzo F, Gregorc V, Rossi E. Gefitinib in pretreated non-small-cell lung cancer (NSCLC): analysis of efficacy and correlation with HER2 and epidermal growth factor receptor expression in locally advanced or metastatic NSCLC. J Clin Oncol 2003;21(14):2658—63. Sreedharan S, Revilla M, Estlack L, Wirtz U, Valone F, Piazza G. Pivanex, a histone deacetylase inhibitor, is synergistic with chemotherapy in inhibiting growth of human non-small cell lung cancer lines. Proc Am Assoc Cancer Res 2003;44:853. Reid T, Cosgriff TM, Yen K, Scislowski P, Valone F, Bhatnagar A. Dose escalation study of Pivanex (a histone deacetylase inhibitor) in combination with docetaxel for advanced non-small cell lung cancer. Lung Cancer 2003;41(2):S182 (abstract P-358).