Annals of Oncology Advance Access originally published online on December 1, 2005
Annals of Oncology 2006 17(2):313-321; doi:10.1093/annonc/mdj067
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© 2005 European Society for Medical Oncology
A phase I, pharmacokinetic and biologic correlative study of oblimersen sodium (GenasenseTM, G3139) and irinotecan in patients with metastatic colorectal cancer
1 Institute for Drug Development, Cancer Therapy and Research Center, San Antonio, TX; 2 Department of Pharmacology, University of Texas Health Science Center, San Antonio, TX; 3 Brooke Army Medical Center, San Antonio, TX; 4 Department of Pathology, University of Texas Health Science Medical Center, San Antonio, TX, USA
* Correspondence to: Dr A. W. Tolcher, Institute for Drug Development, Cancer Therapy and Research Center, 7979 Wurzbach Road, Suite Z414, San Antonio, TX 78229, USA. Tel: +1-210-616-5914; Fax: +1-210-692-7502; E-mail: atolcher{at}idd.org
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Abstract |
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Purpose: To assess the feasibility and antitumor activity of oblimersen sodium, an antisense oligonucleotide directed to the Bcl-2 mRNA, combined with irinotecan in patients with advanced colorectal carcinoma, characterize the pharmacokinetic behavior of both oblimersen sodium and irinotecan, and examine Bcl-2 protein inhibition in peripheral blood mononuclear cells (PBMC).
Patients and methods: Patients were treated with escalating doses of oblimersen sodium administered by continuous intravenous infusion (CIVI) days 1–8, and irinotecan administered intravenously on day 6 once every 3 weeks.
Results: Twenty patients received a total of 84 courses at doses ranging from 3 to 7 mg/kg/day for oblimersen sodium and from 280 to 350 mg/m2 for irinotecan. Febrile neutropenia and diarrhea limited escalation of oblimersen sodium and irinotecan to 5 mg/kg/day and 350 mg/m2, respectively. Other toxicities included nausea, vomiting, fever and fatigue. Steady-state plasma concentrations were achieved within 48 h of beginning oblimersen sodium treatment and the agent was undetectable 24 h after the discontinuation of the infusion. Reduction in levels of Bcl-2 protein in PBMC was documented following treatment with oblimersen sodium. One patient experienced a partial response and 10 additional patients had stable disease lasting 2.5–10 months.
Conclusions: The combination is well tolerated at the recommended phase II oblimersen sodium dose of 7 mg/kg/day CIVI days 1–8 with irinotecan 280 mg/m2 intravenously on day 6 every 3 weeks.
Key words: antisense oligonucleotide, Bcl-2, irinotecan, oblimersen, phase I pharmacokinetics
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introduction |
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Apoptosis or programmed cellular death is a complex, multistep, precisely regulated process that ultimately depends on the orderly activation of a family of intracellular cysteine proteases called caspases [1
, 2]. Restoring apoptosis pathways to improve the effectiveness of chemotherapy, irradiation and hormone therapy represent a novel approach in cancer treatment. Bcl-2 represents an attractive target for therapeutic intervention considering its pivotal role in the regulation of caspase activation and apoptosis [3–7]. Specifically, Bcl-2 overexpression has been reported in numerous solid tumors including colorectal cancer (CRC) [8–12]. In the normal colonic epithelium, Bcl-2 is weakly expressed in the basal proliferative layer [8]. In contrast, Bcl-2 overexpression has been detected in 30–94% of clinicopathological colorectal carcinoma specimens [9–12]. Moreover, it has been suggested that Bcl-2 overexpression could be a negative prognostic factor for recurrence and death in Dukes' C resected CRC [13]. The Bcl-2 family of proteins could play a role not only in cancer pathogenesis, but also in resistance to anticancer treatment. Indeed, Bcl-2 protein expression aborts apoptosis induced by diverse signals including chemotherapy, glucocorticoids and radiation, and confers a multidrug resistant phenotype in several cell lines [14–16]. Bcl-2 gene transfer studies indicate induced resistance to drugs that are diverse in structure and mechanism of action and include irinotecan, etoposide, vincristine, cisplatin, cyclophosphamide, doxorubicin and nitrogen mustard compounds. Furthermore, antisense oligonucleotide therapy directed against Bcl-2 was shown to significantly enhance the chemosensitivity in several cancer cells lines compared with controls in vitro, further supporting the critical role of Bcl-2 in mediating chemotherapy resistance [17–19].
Oblimersen sodium (G3139, GenasenseTM; Genta Inc., San Diego, CA, USA) is a phosphorothioate antisense oligonucleotide (sequence: 5' TCT CCC AGC GTG CGC CAT 3') that hybridizes to the first six codons of the Bcl-2 open reading frame mRNA. The hybrid recruits endogenous RNAse H to cleave the Bcl-2 mRNA, thus blocking the production of the target protein Bcl-2. Experiments using oblimersen in vivo and in vitro in different tumor cell lines including follicular lymphoma, melanoma, breast, prostate, colon and ovarian carcinomas showed dose-dependent inhibition of Bcl-2 at mRNA and protein expression levels [20–23]. In the initial clinical studies exploring a continuous 14-day subcutaneous infusion of oblimersen, dose-limiting toxicities (DLTs) were fever, thrombocytopenia and fatigue. Antitumor activity, including a complete response, was seen in non-Hodgkin's lymphoma. In subsequent studies, oblimersen could be administered for 14 or 21 days by continuous intravenous infusion (CIVI) in patients with solid tumors with limited side-effects including fever, fatigue and lymphopenia [24–26]. Additionally, disease-directed trials exploring oblimersen alone or combined with chemotherapy have been performed in solid tumors such as melanoma, small cell lung, prostate, breast and colorectal carcinomas [27–30].
Irinotecan hydrochloride (CPT-11, CamptosarTM; Pharmacia, Michigan, USA) is a semisynthetic derivative of the topoisomerase I inhibitor alkaloid camptothecin [31]. Irinotecan is a prodrug that is rapidly metabolized in vivo to its more active metabolite SN-38. Although the liver is the major site of bioactivation, extrahepatic metabolism including intratumor bioactivation also occurs. Following the initial approval as second-line therapy for metastatic colorectal cancer after failure of 5-fluorouracil, irinotecan is currently widely used for the treatment of metastatic CRC [32, 33].
The role of Bcl-2 and its family members in the pathogenesis of CRC as well as in the resistance to cytotoxic drugs strongly supports the study of a combination of oblimersen and irinotecan for patients with metastatic CRC. The main objectives of this phase I, pharmacokinetic (PK) and biologic correlative study were: (i) to determine the maximum tolerated dose (MTD) of oblimersen sodium administered by CIVI days 1–8 when combined with irinotecan intravenously administered over 90 min on day 6, every 3 weeks; (ii) to characterize the toxicities of the combination; (iii) to characterize the PK behavior of oblimersen and irinotecan when co-administered; (iv) to document Bcl-2 protein expression in peripheral blood mononuclear cells (PBMC) before and after the treatment with oblimersen; and (v) to seek preliminary evidence of antitumor activity in patients with advanced CRC.
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patient selection
Patients with histologically or cytologically documented colon or rectal carcinoma (unresectable, metastatic or recurrent) who had failed prior chemotherapy were eligible for this study. Prior therapy with irinotecan was allowed. The eligibility criteria also included:
18 years of age; an Eastern Cooperative Oncology Group performance status 
2; estimated life expectancy of at least 12 weeks; no chemotherapy, investigational agents or wide-field radiation therapy within 4 weeks prior to the study; adequate organ function including adequate hematopoietic [hemoglobin 9 g/dl, absolute neutrophil count (ANC) 1500/ml, platelet count 100 000/ml], hepatic [bilirubin 1.5 mg/dl, aspartate transaminase and alanine transaminase 3x the institutional upper normal limit (5x if due to hepatic metastasis)] and renal (creatinine 1.5 mg/dl) functions; coagulation tests within normal limits; and evaluable or measurable disease. Patients were excluded if they had an active infection, a coexisting medical problem of sufficient severity to limit compliance with the study, active brain metastasis or were pregnant or breast feeding. All patients gave written informed consent before treatment according to federal and institutional guidelines.
dosage and drug administration
Patients received oblimersen as a CIVI for 7 days (days 1–8) and irinotecan administered intravenously over 90 min on day 6. Courses of treatment were repeated every 21 days following the start of administration of oblimersen. Oblimersen was supplied as a sterile solution by the National Cancer Institute (NCI) in 10 ml vials containing 300 mg of product. The appropriate dose of the drug was further diluted with 0.9% saline solution to a total volume of 100 ml for the infusion pump cassette. The final solution was infused continuously for 148 h. Irinotecan was commercially available and was supplied in 40 mg (2 ml) and 100 mg (5 ml) vials. Each milliliter of solution contains 20 mg of irinotecan trihydrate, 45 mg of sorbitol powder and 0.9 mg of lactic acid. Prior to infusion, irinotecan was diluted in 500 ml 5% dextrose solution or 0.9% sodium chloride. Irinotecan was infused over 90 min using a separate intravenous access.
The MTD was defined as the highest dose at which no more than one of six patients experience DLT during course 1. A minimum of three patients were to be treated per dose level. The occurrence of DLT in one of three patients at any dose level required the enrollment of three additional patients at that dose level. After determination of the MTD, a total of six patients were to be treated at this dose level to further ascertain the toxicities. Toxicities were evaluated according to the NCI Common Toxicity Criteria, version 2.0. DLT was defined as any of the following occurring during course 1: grade 4 neutropenia lasting >5 days or associated with fever 100.5°F or infection; grade 4 thrombocytopenia; any grade 3–4 non-hematological toxicity except alopecia; grade 3–4 vomiting in patients who have received optimal prophylaxis and treatment; grade 3–4 diarrhea in patients who have received an optimal anti-diarrheal regimen; treatment delay >2 weeks due to unresolved toxicity in patients who experienced at least grade 3 thrombocytopenia, grade 4 neutropenia or any grade 3 non-hematological toxicity (except for alopecia and suboptimally treated nausea, vomiting or diarrhea). Oblimersen doses were not modified downward; only the dose of irinotecan was modified based on toxicities. No intrapatient dose-escalation was permitted.
All patients received antiemetics prior to and after each irinotecan infusion. Filgrastim use was permitted for prolonged neutropenia (neutrophils <500/ml for at least 5 days), neutropenic fever, diarrhea associated with grade 4 neutropenia or documented infection during neutropenia. Filgrastim was discontinued at least 24 h prior to the start of the next irinotecan dose. Patients receiving recombinant erythropoietin prior to study entry were allowed to continue this therapy during the study. Atropine and loperamide were permitted for cholinergic symptoms and early/late onset diarrhea, respectively.
pretreatment and follow-up studies
A complete history and physical examination that included performance status, concurrent medications and routine laboratory evaluations were performed pretreatment and weekly while on study. Routine laboratory evaluations included complete blood cell counts (CBC) with differential white blood cell, serum electrolytes, renal and liver function panels, prothrombin time and urinalysis. In addition, a CBC was assessed every other day if ANC was <750/ml or platelets <25 000/ml. Radiological studies for tumor assessment were conducted pretreatment and after every other course. Patients were able to continue on study in the absence of progressive disease, which was defined as a 25% increase in the sum of diameters of all measurable lesions or appearance of any new lesion. A complete response was scored if there was disappearance of all disease on two measurements at least 4 weeks apart, and a partial response required at least 50% reduction in the sum of the product of perpendicular diameters of all measurable lesions compared with baseline at least 4 weeks apart. Stable disease was defined as not meeting criteria for response or progression. Patients were followed 30 days after the last dose of study drug to detect delayed toxicity and every 30 days thereafter until toxicity resolved.
plasma pharmacokinetic sampling and assay
Blood samples were obtained for assessment of oblimersen plasma concentration during the first course prior to drug infusion and 2, 48, 120 and 168 h following the initiation of the infusion, and then 1, 2, 4, 6, 24 and 48 h after the discontinuation of the infusion.
Plasma samples containing oblimersen were prepared by adding 20 ml of buffer (25 mM Tris, 1 mM EDTA, pH 7) to 100 µl of sample plus 280 ml of IPEGAL [1% (weight/volume) IPEGAL CA-630 (Sigma, St Louis, MO, USA)]; 0.9% (weight/volume) sodium chloride/phosphate-buffered saline (PBS) (Dulbecco's PBS) solution, vortexed (30 s) and centrifuged for 7 min at 
10 000 r.p.m. in a microcentrifuge. Percentage recovery of oblimersen from plasma was 89%. One hundred and eighty microliters of supernatant was injected into a temperature controlled (30°C) DNA Pac PA-100 (13 x 2 x 10 mm) alkyl quaternary amine column (Dionex, Houston, TX). An equilibrating solution (100 mM Na perchlorate, 17.5 mM Tris, pH 7, 0.7 mM EDTA, 30% formamide) was pumped at 1 ml/min across the column. At 1.5 min, the flow rate (1.5 ml/min) and the concentration of Na perchlorate (800 mM) were increased. UV detection was monitored at 267 nm. Calibration curves were linear (r2 >0.99) from 0.2 to 20 µg/ml. The mean back-calculated concentrations for the calibration standards were all within 15% deviation from the theoretical concentration.
For assessment of irinotecan concentrations, PK samples were obtained at the end of infusion, and 2, 4, 6, 8, 24 and 48 h after the end of infusion. Blood samples were collected from the contralateral arm to the irinotecan and oblimersen infusion line. Plasma samples were analyzed for concentrations of irinotecan and SN-38 by a previously described HPLC method [34].
pK analyses
Oblimersen, irinotecan and SN-38 plasma concentrations were analyzed by standard non-compartmental methods. Irinotecan peak concentrations (Cmax) were determined by inspection of each individual's plasma concentration–time curve. Oblimersen steady-state plasma concentrations (Css) were determined by averaging the plasma concentrations at the 48, 120 and 192 h time points. Elimination rate constants were estimated by linear regression of the last two (irinotecan and SN-38) or last three (oblimersen sodium) data points on the terminal log linear portion of the concentration–time curve. Terminal half-life (T1/2) was calculated by dividing 0.693 by the elimination rate constant. The AUC for irinotecan/SN-38 was calculated using the linear trapezoidal rule up to the last measurable data point (AUC0–
), then extrapolated to infinity (AUC). The systemic clearance (CL) for irinotecan was determined by dividing the dose (in mg free base/m2) by the AUC. The clearance for oblimersen was calculated using the formula: CL = drug infusion rate/Css.
pharmacodynamic evaluation
Quantitation of Bcl-2 in PBMC.
Bcl-2 expression and downregulation in PBMC were analyzed pre-study, and days 6, 8 and 10. These end points were utilized to define the relationship between oblimersen administration and Bcl-2 downregulation. Peripheral blood specimens were collected from patients in EDTA tubes and mononuclear cell separation was performed with Histopaque 1077 (Sigma). Briefly, the PBMC were washed with RPMI 1640 containing 5% fetal calf serum, the cell pellet was resuspended in 5 ml PBS on ice and immediately frozen to –70°C until analyzed. PBMC pellets were lysed in Nonidet P-40 lysis buffer (20 mM HEPES pH 7.5, 5 mM MgCl2, 1% Nonidet P-40), and protease inhibitor cocktail (Sigma). The protein content was determined (Bradford reagent; Bio-Rad, Hercules, CA, USA) and 10–20 mg of total protein was loaded per lane and fractionated in 12% SDS–polyacrylamide gels. The gels were transferred to PVDF membranes and incubated in blocking buffer (PBS, 0.1% Tween 20, 1% milk) for 1 h at room temperature. Bcl-2 protein detection was performed using a monoclonal anti-Bcl-2 antibody (DAKO, Carpinteria, CA, USA) at 1:200 dilution, then incubated at room temperature with goat anti-mouse horseradish peroxidase-conjugated secondary antibody (Cell Signaling, Berkeley, CA, USA) at a dilution of 1:20 000. Visualization of Bcl-2 bands was performed using enhanced chemoluminescence. The blots were incubated in 20 ml of detection buffer (1.25 mM luminol, 200 mM paracoumaric acid, 2.9 mM H2O2 and 100 mM Tris, pH 8.5) for 5 min followed by exposure to Kodak Biomax Light film. ß-actin levels in the samples were quantified to normalize Bcl-2 expression. The change in Bcl-2 normalized protein expression was determined using the following formula: 100% x [1 – (Bcl-2 at day 6/Bcl-2 at day 1)].
immunohistochemical methods
Paraffin-embedded tumor blocks were obtained from the patients original diagnostic material for the detection of Bcl-2, Bcl-XL and Bax by immunohistochemistry. Standard immunohistochemical techniques were applied for detection of Bcl-XL, Bcl-2 and Bax [35]. Antigen retrieval using heat was utilized as pretreatment for all three antibodies. Antibody to Bcl-XL (monoclonal, clone 7D9) was obtained from Neomarkers (Freemont, CA, USA) and used at 1:1000 dilution. Anti-Bcl-2 (monoclonal, clone 124) and Bax (polyclonal) antibodies were both obtained from DAKO and used at 1:25 and 1:12 000 dilutions, respectively. The biotin–streptavidin system was used, with the addition of the DAKO CSA system for amplification of Bax. Cytoplasmic and membranous staining was considered positive. A semiquantitative scale was used to evaluate the degree of staining, with intensity graded between 0 and 3 and the proportion of cells stained graded as between 0 and 5 [34].
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general
Twenty patients, whose pertinent demographics are listed in Table 1, received a total of 84 courses of the combination of oblimersen and irinotecan. Oblimersen doses ranging from 3 to 7 mg/kg/day combined with irinotecan 280–350 mg/m2 were evaluated. The dose escalation scheme is presented in Table 2. Patients enrolled at the first dose level received 3 mg/kg/day of oblimersen for the first course and 5 mg/kg/day for the subsequent courses. The median number of courses per patient was four (range one to 13). Dose reductions due to toxicity occurred in seven patients. One patient received the fifth course of treatment without oblimersen.
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Patients at the first and second dose level had only mild to moderate treatment-related side-effects. At the third dose level (oblimersen 5 mg/kg/day combined with irinotecan 350 mg/m2) two of six patients treated experienced DLTs (febrile neutropenia and diarrhea). Since the side-effects were consistent with the toxicity profile of irinotecan, the next dose level explored a reduced dose of irinotecan at 280 mg/m2 combined with an increased dose of oblimersen at 7 mg/kg/day. Only one out of the seven patients enrolled in this cohort experienced DLT (febrile neutropenia).
hematological toxicity
The distributions of grades of neutropenia and lymphopenia as functions of dose are listed in Table 3. Neutropenia was the principal side-effect, with five out of 13 (38%) patients experiencing grade 3 or 4 toxicity at dose levels 3 and 4. The median time to ANC nadir in course 1 was day 15 (range 5–22). Recovery of ANC to >1500/ml commonly occurred by the planned start date of the next oblimersen infusion in all patients except one. Moderate to severe lymphopenia (grade 2–4) attributable to oblimersen was observed in 12 out of 20 (60%) patients during the first course of treatment. Severe thrombocytopenia was uncommon, with only one patient experiencing a grade 4 (in course 6) at the fourth dose level and two patients experiencing grade 3 at dose levels 1 and 3. Drug-related anemia was generally mild (grade 1–2), with only one patient (5%) experiencing grade 3 anemia on day 13 of course 1 at the fourth dose level. Four (20%) patients required red blood cell transfusion.
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non-hematological toxicity
The most common grade 1 or 2 non-hematological toxicities were fatigue, nausea, vomiting, diarrhea, mucositis, fever and arthralgia. The distribution of these toxicities as a function of dose level is listed in Table 4. As a direct toxicity attributable to oblimersen, mild and moderate (grade 1–2) pyrexia was a common finding, occurring in six out of 20 (30%) patients during the first course, and in 22 of 84 (26%) total courses. The fever commenced 24–48 h after the initiation of the oblimersen infusion and was commonly associated with chills. The duration of the pyrexia generally did not exceed the end of the oblimersen infusion. No infectious etiology could be related to the fever, and acetaminophen or non-steroidal anti-inflammatory agents were successful for treating and preventing this symptom. Fatigue was another side-effect observed in the majority (16 out of 20; 80%) of patients, but was not severe enough to represent DLT for any patient. However, at the higher dose-level, three patients experienced grade 3 fatigue in courses beyond course 1. Diarrhea occurred in 13 out of 20 (65%) patients during the first course, and two patients experienced dose-limiting grade 3 diarrhea despite optimal treatment. These two patients also experienced grade 4 neutropenia. These toxicities were considered mainly related to irinotecan. Nausea and vomiting were common but never dose limiting, and were also considered mostly related to the irinotecan. Treatment with 5-hydroxy-tryptamine 3 receptor antagonists generally resulted in successful management or prevention of emesis. Other mild to moderate side-effects, included weakness, insomnia and increase in transaminases, occurred in <15% of patients. These effects were noted across the entire dosing range.
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pharmacokinetics
Eighteen patients had plasma sampling performed for oblimersen Css and for irinotecan PK parameters. Three patients on the first dose level had plasma samples in course 1 (at 3 mg/kg of oblimersen) and also in course 2 (at 5 mg/kg of oblimersen). All datasets were analyzed by standard non-compartmental methods. The mean oblimersen Css increased as the dose increased from 3 to 7 mg/kg/day, although there was a substantial overlap of individual Css values between 5 and 7 mg/kg/day dose levels. Oblimersen reached Css within 48 h of the infusion initiation and the drug was undetectable 24 h after the discontinuation of the infusion. The representative oblimersen plasma concentration–time curve is depicted in Figure 1. The mean clearance of oblimersen was 69 (±18), 68 (±22) and 97 (±34) ml/h/kg at 3, 5 and 7 mg/kg/day, respectively. The oblimersen PK characteristics are presented in Table 5. The mean values for PK parameters of irinotecan at each dose level are listed in Table 6. Interestingly, with the same dose of irinotecan (280 mg/m2) and increasing doses of oblimersen sodium (3, 5 and 7 mg/kg/day), the Cmax and AUC of SN-38 decreased. However, this trend is not seen for the Cmax and AUC of irinotecan.
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Since neutropenia was the predominant hematological toxicity observed with the combination, the relationship between irinotecan Cmax, AUC and oblimersen Css and the percentage decrement of neutrophils was explored. No relationship (either in linear or non-linear models) could be determined. Similarly, no relationship was detected between the exposure of oblimersen and/or irinotecan and the magnitude of lymphocytes decrement.
pharmacodynamics
Bcl-2 expression in normal PBMC.
For the determination of Bcl-2 protein expression, normal PBMCs were collected on day 1 (pretreatment), day 6 (pre-irinotecan), day 8 (post-infusion) and day 10 (post-infusion). Bcl-2 protein was markedly reduced (>90%) in four of four paired evaluable samples. A representative blot is depicted in Figure 2. No evaluable blot samples are available for day 10 to track Bcl-2 protein recovery following discontinuation of oblimersen on day 8. No dose–response relationship between time of oblimersen administration and the Bcl-2 downregulation was possible based upon the small number of patients with paired evaluable samples.
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Bcl-2, Bax, and Bcl-XL expression in tumor samples.
The results of immunohistochemistry staining for Bcl-2, Bax and Bcl-XL are summarized in Table 7. The majority of tumors demonstrated weak Bcl-2 expression intensity (0–1 for 10 out of 14 patients tested), although the proportion of the positive cells was greater than 50% in 12 out of 14 patients. The pattern for Bcl-XL testing was similar, with most tumors showing weak staining but in a high proportion of the cells. In contrast, most tumors expressed Bax with both high intensity and in a high proportion of cells. No correlation was found between Bcl-2, Bax and Bcl-XL expression and the antitumor efficacy of the combination of oblimersen and irinotecan. In this trial such assessments are precluded by the heterogeneous group of tumors.
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antitumor activity
One of the 20 evaluable patients had a partial response for 10 months. This patient had a history of colon cancer with liver metastasis and received therapy with 5-fluorouracil/leucovorin before being enrolled in the current study. One patient having previous exposure to irinotecan had persistent stable disease for 10 months. Nine additional patients had stable disease for 2–8 months (median 3 months), four (44%) of them had received irinotecan previously.
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The critical role of Bcl-2 in the regulation of apoptosis provides a strong rationale for using Bcl-2-directed antisense oligonucleotide in order to increase apoptosis and enhance the effectiveness of several cytotoxic agents [17
, 18]. The combination of oblimersen with irinotecan in patients with advanced CRC is an attractive approach supported by the frequent overexpression of Bcl-2 in CRC, the demonstration of in vitro synergy between oblimersen and chemotherapy, and their non-overlapping toxicities. Oblimersen was administered on a 7-day CIVI that overlapped on day 6 with the irinotecan infusion [36, 37]. The rationale for this schedule of administration was based on the premise that a prolonged oblimersen infusion would exert maximal downregulation of Bcl-2 and favor Bax homodimerization and apoptosis before and during exposure to the chemotherapy [25, 26].
This study demonstrates that the combination of oblimersen and irinotecan is feasible in pretreated patients with advanced colorectal cancer. The recommended dose for future phase II studies is oblimersen 7 mg/kg/day CIVI for 7 days and irinotecan 280 mg/m2 intravenously over 90 min on day 6. The predominant hematological toxicity observed with this combination was neutropenia, which reached grade 3–4 in 50% of patients; however, only one out of seven patients treated at the recommended dose experienced hematological DLT. Febrile neutropenia was observed in only three of 84 courses (3.5%). Attempts to deliver 350 mg/m2 of irinotecan resulted in unacceptable toxicity (prolonged neutropenia associated with diarrhea in two of six patients), suggesting that the neutropenia was mostly related to the irinotecan. Similar to results from other studies, lymphopenia was common and the predominant hematologic toxicity ascribed to oblimersen alone [36]. Non-hematological toxicities were mild to moderate and included fatigue, nausea and vomiting, mucositis, and increase in transaminases. Additionally pyrexia was observed in a majority of patients and considered to be oblimersen-related. Sixty-five per cent of patients reported mild to moderate diarrhea during the first cycle that generally readily resolved with loperamide treatment.
The PK analysis suggested that Css for oblimersen increased with the increase in dose from 3 to 7 mg/kg/day, although a substantial overlap of individual Css values between 5 and 7 mg/kg/day was observed. Steady-state oblimersen concentrations (>5mg/ml) exceeding those demonstrated to have biological activity in preclinical studies (1–2 mg/ml) were reached within 48 h of the initiation of oblimersen, were maintained during the 7 days infusion and decreased rapidly to undetectable levels within 48 h after discontinuation of the infusion. The pharmacokinetic parameters for irinotecan were comparable with the results published from previous studies and a relationship was not found between the irinotecan PK parameters reflecting drug exposure and the decrements in ANC, despite the observation that neutropenia was mainly related to the irinotecan dose. Furthermore, oblimersen dose did not appear to influence the magnitude of neutropenia in this patient population and no correlation was found between lymphopenia and either oblimersen dose, Css or estimate of exposure. The decrease of the Cmax and AUC of SN-38 with the increase of the oblimersen dose when combined with the same dose of irinotecan could suggest a reduction of the conversion of irinotecan to SN-38 induced by oblimersen. This hypothesis should, however, be validated by further studies with the combination.
One irinotecan-naïve patient experienced a prolonged partial response and 10 patients (50%) had stable disease for 2–8 months. The high rate of stable disease is of particular interest, as 44% of these patients had previously received irinotecan. These results suggest that the addition of oblimersen may enhance the cytotoxicity in the presence of external apoptotic signals provided by the chemotherapeutic agent, although not reversing the resistant phenotype. However, these results are not robust enough to suggest favorable effects of oblimersen sodium. Similar results from other studies combining oblimersen with cytotoxic drugs have also been published [28, 30, 36].
The reduction in expression of Bcl-2 protein in normal PBMCs following oblimersen therapy demonstrated in previous studies and in our trial confirms the mechanism of action for oblimersen [26, 36]. Other recent studies, however, failed to demonstrate a significant change of Bcl-2 expression in PBMC after treatment with oblimersen [28, 30]. Furthermore, it is not clear whether Bcl-2 suppression in the peripheral blood is reflective of a similar pattern in the tumor cells mainly related to the differences in the Bcl-2 expression. Therefore, efforts should be undertaken to further identify and evaluate biomarkers that may be predictive for responsiveness to oblimersen or define optimal dose versus Bcl-2 inhibitory effect. Selecting the optimal patient population susceptible to achieve a tumor response to Bcl-2 inhibition may represent an important step for the successful development of oblimersen.
In conclusion, the combination of oblimersen and irinotecan administered on this schedule is safe and moderately effective in this population of previously treated CRC patients. Based on these results, oblimersen at 7 mg/kg/day days 1–8 with irinotecan 280 mg/m2/day on day 6 once every 3 weeks is recommended for phase II studies. Further testing in selected populations and in disease oriented randomized studies could be appropriate to evaluate the real impact of adding oblimersen or other proapoptotic drugs to classic cytotoxic drugs.
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Acknowledgements |
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This work was supported by the NIH Grant UO1-CA69853 and Cancer Center Council Grant P30-CA54174. Presented in part at the 37th Annual Meeting of the American Society of Clinical Oncology 2001, Abstract 297.
Received for publication September 2, 2005. Accepted for publication October 3, 2005.
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