Fludarabine Phosphate Injection (Mylan Pharmaceuticals) Product Monograph

SUMMARY PRODUCT INFORMATION

INDICATIONS AND CLINICAL USE

FLUDARABINE PHOSPHATE INJECTION is indicated for:

• Second line therapy in patients with chronic lymphocytic leukemia (CLL) and low-grade non-Hodgkin's lymphoma (Lg-NHL) who have failed other conventional therapies. Such patients should be treated only by physicians skilled in the use of chemotherapeutic agents.

Geriatrics (> 75 years of age):

Since there are limited data for the use of fludarabine phosphate in elderly persons (> 75 years), caution should be exercised with the administration of FLUDARABINE PHOSPHATE INJECTION in these patients. The total body clearance of the principal plasma metabolite 2F-ara-A shows a correlation with creatinine clearance, indicating the importance of the renal excretion pathway for the elimination of the compound. Patients with reduced kidney function demonstrated an increased total body exposure (AUC of 2F-ara-A). Limited clinical data are available in patients with impairment of renal function (creatinine clearance below 70 mL/min). Since renal impairment is frequently present in patients over the age of 70 years, creatinine clearance should be measured. If creatinine clearance is between 30 and 70 mL/min, the dose should be reduced by up to 50%, and close hematologic monitoring should be used to assess toxicity. Fludarabine phosphate treatment is contraindicated, if creatinine clearance is <30 mL/min. (See WARNINGS AND PRECAUTIONS and DOSAGE AND ADMINISTRATION).

Pediatrics:

The safety and effectiveness of fludarabine phosphate in children have not been established.

CONTRAINDICATIONS

• Patients who are hypersensitive to this drug or to any ingredient in the formulation or component of the container. For a complete listing, see the Dosage Forms, Composition and Packaging section of the product monograph.
• Renally impaired patients with creatinine clearance <30 mL/min.
• Patients with decompensated hemolytic anemia
• Pregnancy
• Lactation

• In a clinical investigation using fludarabine phosphate in combination with pentostatin

  (deoxycoformycin) for the treatment of refractory CLL, there was an unacceptably high incidence of fatal pulmonary toxicity. Therefore, the use of fludarabine phosphate in combination with pentostatin is contraindicated.

WARNINGS AND PRECAUTIONS

General

FLUDARABINE PHOSPHATE INJECTION is a potent antineoplastic agent with potentially

Serious Warnings and Precautions

FLUDARABINE PHOSPHATE INJECTION should be administered under the supervision of, or prescribed by, a qualified physician experienced in the use of antineoplastic therapy. Fludarabine phosphate can severely suppress bone marrow function. When used at high doses in dose-ranging studies in patients with acute leukemia, intravenous fludarabine phosphate was associated with severe irreversible neurologic effects, including blindness, coma, and death. This severe central nervous system toxicity occurred in 36% of patients treated intravenously with doses approximately four times greater (96 mg/m²/day for 5-7 days) than the recommended dose. In patients treated at doses in the range of the dose recommended for chronic lymphocytic leukemia (CLL) and low-grade non-Hodgkin's lymphoma (Lg-NHL), severe central nervous system toxicity occurred rarely (coma, seizures and agitation) or uncommonly (confusion). Patients should be closely observed for signs of neurologic side effects.

Instances of life-threatening and sometimes fatal autoimmune hemolytic anemia have been reported to occur during or after treatment with fludarabine phosphate. The causality of the development of this complication has not been identified. Patients undergoing treatment with FLUDARABINE PHOSPHATE INJECTION should be evaluated and closely monitored for signs of autoimmune hemolytic anemia (a decline in hemoglobin linked with hemolysis and a positive Coombs' test). Discontinuation of therapy with FLUDARABINE PHOSPHATE INJECTION is recommended in the event of hemolysis. The transfusion of irradiated blood and the administration of corticosteroids are the most common treatment measures for autoimmune hemolytic anemia.

In a clinical investigation using fludarabine phosphate in combination with pentostatin (deoxycoformycin) for the treatment of refractory CLL, there was an unacceptably high incidence of fatal pulmonary toxicity. Therefore, the use of fludarabine phosphate in combination with pentostatin is contraindicated.

significant toxic side effects. Patients undergoing therapy should be closely observed for signs of hematologic and nonhematologic toxicity. Periodic assessment of peripheral blood counts is recommended to detect the development of neutropenia, thrombocytopenia, anemia and leukopenia.

Vaccination with live vaccines should be avoided during and after treatment with FLUDARABINE PHOSPHATE INJECTION

Gastrointestinal

In clinical trials with oral fludarabine phosphate, nausea/vomiting and/or diarrhea were reported in approximately 38% of patients. In most cases, the severity was mild to moderate (WHO toxicity grading). Only a small percentage of patients, approximately 1 % with nausea/vomiting and 5% with diarrhea, required therapy. Patients with prolonged, clinically relevant, nausea/vomiting and diarrhea should be closely monitored to avoid dehydration.

Hematologic

In patients with an impaired state of health, FLUDARABINE PHOSPHATE INJECTION should be given with caution and after careful risk/benefit consideration. This applies especially to patients with severe impairment of bone marrow function (thrombocytopenia, anemia, and/or granulocytopenia), immunodeficiency or with a history of opportunistic infection. Prophylactic treatment should be considered in patients at increased risk of developing opportunistic infections (see ADVERSE REACTIONS).

Bone marrow suppression, notably thrombocytopenia, anemia, leukopenia and neutropenia, may occur with administration of fludarabine phosphate and requires careful hematologic monitoring. In a Phase I study in solid tumor patients, the median time to nadir counts was 13 days (range, 3-25 days) for granulocytes and 16 days (range, 2-32 days) for platelets. Most patients had hematologic impairment at baseline either as a result of disease or as a result of prior myelosuppressive therapy. Cumulative myelosuppression may be seen. While chemotherapy-induced myelosuppression is often reversible, administration of FLUDARABINE PHOSPHATE FOR INJECTION requires careful hematologic monitoring.

Several instances of trilineage bone marrow hypoplasia or aplasia resulting in pancytopenia, sometimes resulting in death, have been reported in adult patients. The duration of clinically significant cytopenia in the cases reported has ranged from approximately 2 months to approximately 1 year. These episodes have occurred in both previously treated and untreated patients.

Instances of life-threatening and sometimes fatal autoimmune phenomena (e.g. autoimmune hemolytic anemia, autoimmune thrombocytopenia, thrombocytopenic purpura, pemphigus, acquired hemophilia and Evans' syndrome) have been reported to occur during or after treatment with fludarabine phosphate in patients with or without a previous history of autoimmune processes or a positive Coombs' test and who may or may not be in remission from their disease. Steroids may or may not be effective in controlling these hemolytic episodes. One study was performed with 31 patients with hemolytic anemia related to the administration of fludarabine phosphate. Since the majority (90%) of these patients rechallenged with fludarabine phosphate developed a recurrence in the hemolytic process, rechallenge with FLUDARABINE PHOSPHATE INJECTION should be avoided. The mechanisms which predispose patients to the

development of this complication have not been identified. Patients undergoing treatment with fludarabine phosphate should be evaluated and closely monitored for signs of autoimmune hemolytic anemia (a decline in hemoglobin linked with hemolysis and a positive Coombs' test). Discontinuation of therapy with FLUDARABINE PHOSPHATE INJECTION is recommended in the event of hemolysis. The transfusion of irradiated blood and the administration of corticosteroids are the most common treatment measures for autoimmune hemolytic anemia.

Tumor lysis syndrome associated with fludarabine phosphate treatment has been reported in CLL patients with large tumor burdens. Since fludarabine phosphate can induce a response as early as the first week of treatment, precautions should be taken in those patients at risk of developing this complication.

Transfusion-associated graft-versus-host disease (reaction by the transfused immunocompetent lymphocytes to the host) has been observed rarely after transfusion of non- irradiated blood in patients treated with fludarabine phosphate. Fatal outcome as a consequence of this disease has been reported with a high frequency. Therefore, to minimize the risk of transfusion-associated graft-versus-host disease, patients who require blood transfusion and who are undergoing, or who have received treatment with fludarabine phosphate should receive irradiated blood only.

Disease progression and transformation (e.g. Richter’s Syndrome) have been commonly reported in CLL patients.

Hepatic/Biliary/Pancreatic

No data are available concerning the use of fludarabine phosphate in patients with hepatic impairment. In this group of patients, FLUDARABINE PHOSPHATE INJECTION should be used with caution and administered if the perceived benefit outweighs any potential risk.

Neurologic

When high doses of fludarabine phosphate were administered in dose-ranging studies in acute leukemia patients, a syndrome with delayed onset, characterized by blindness, coma, and death was identified. Symptoms appeared from 21 to 60 days post dosing (however, in post marketing experience, cases of neurotoxicity have been reported to occur both earlier and later than seen in clinical trials). Demyelination, especially of the occipital cortex of the brain was noted. The majority of these cases occurred in patients treated intravenously with doses approximately four times greater (96 mg/m²/day for 5-7 days) than the recommended dose. Thirteen of 36 patients (36.1%) who received fludarabine phosphate at high doses (≥ 96 mg/m²/day for 5 to 7 days per course) developed severe neurotoxicity, while only one of 443 patients (0.2%) who received the drug at low doses (≤ 40 mg/m²/day for 5 days per course) developed the toxicity. In patients treated at doses in the range of the dose recommended for CLL and Lg-NHL, severe central nervous system toxicity occurred rarely (coma, seizures and agitation) or uncommonly (confusion).

The effect of chronic administration of fludarabine phosphate on the central nervous system is unknown. In some studies, however, patients tolerated the recommended dose, for relatively long treatment periods (up to 26 courses of therapy). Periodic neurological assessments are recommended.

Renal

The total body clearance of the principal plasma metabolite 2F-ara-A shows a correlation with creatinine clearance, indicating the importance of the renal excretion pathway for the elimination of the compound. Patients with reduced renal function demonstrated an increased total body exposure (AUC of 2F-ara-A). Limited clinical data are available in patients with impairment of renal function (creatinine clearance below 70 mL/min). Therefore, if renal impairment is clinically suspected, or the patient is over the age of 70 years, creatinine clearance should be measured. If creatinine clearance is between 30 and 70 mL/min, the dose should be reduced by up to 50% and close hematological monitoring should be used to assess toxicity. FLUDARABINE PHOSPHATE INJECTION treatment is contraindicated, if creatinine clearance is < 30 mL/min. (See DOSAGE AND ADMINISTRATION).

Sexual Function/Reproduction

Preclinical toxicology studies in mice, rats and dogs have demonstrated dose-related adverse effects on the male reproductive system. Observations consisted of a decrease in mean testicular weights in dogs and degeneration and necrosis of spermatogenic epithelium of the testes in mice, rats and dogs. The possible adverse effects on fertility in males and females in humans have not been adequately evaluated. Therefore, it is recommended that females of child-bearing potential and males take contraceptive measures during FLUDARABINE PHOSPHATE INJECTION therapy, and for at least 6 months after the cessation of FLUDARABINE PHOSPHATE INJECTION therapy.

Skin

Reversible worsening or flare-ups of pre-existing skin cancer lesions has been reported to occur in some patients during or after intravenous fludarabine phosphate therapy.

Special Populations

Pregnant Women: Fludarabine phosphate has been shown to be teratogenic in rats and in rabbits. A study in rats demonstrated a transfer of fludarabine phosphate and/or metabolites across the placental barrier.

One case of fludarabine phosphate use during early pregnancy leading to skeletal and cardiac malformation in the newborn has been reported.

FLUDARABINE PHOSPHATE INJECTION should not be used during pregnancy.

Women of child-bearing potential should be advised to avoid becoming pregnant and to inform the treating physician immediately should this occur.

Nursing Women: It is not known whether fludarabine phosphate is excreted in human milk. However, there is evidence from animal data that fludarabine phosphate and/or metabolites transfer from maternal blood to milk. Therefore, breast-feeding should be discontinued during FLUDARABINE PHOSPHATE INJECTION therapy.

Pediatrics: The safety and effectiveness of fludarabine phosphate in children have not been established.

Geriatrics (> 75 years of age): Since there are limited data for the use of fludarabine phosphate in elderly persons (> 75 years), caution should be exercised with the administration of FLUDARABINE PHOSPHATE INJECTION in these patients. The total body clearance of the principal plasma metabolite 2F-ara-A shows a correlation with creatinine clearance, indicating the importance of the renal excretion pathway for the elimination of the compound. Patients with reduced kidney function demonstrated an increased total body exposure (AUC of 2F-ara-A). Limited clinical data are available in patients with impairment of renal function (creatinine clearance below 70 mL/min). Since renal impairment is frequently present in patients over the age of 70 years, creatinine clearance should be measured. If creatinine clearance is between 30 and 70 mL/min, the dose should be reduced by up to 50%, and close hematologic monitoring should be used to assess toxicity. Fludarabine phosphate treatment is contraindicated, if creatinine clearance is <30 mL/min. (See DOSAGE AND ADMINISTRATION).

Monitoring and Laboratory Tests

During treatment, the patient's hematologic (particularly neutrophils and platelets) and serum chemistry profiles should be monitored regularly.

Effects on Ability to Drive and Operate Machines

The effect of treatment with fludarabine phosphate on the patient’s ability to drive or operate machinery has not been evaluated.

ADVERSE REACTIONS

Adverse Drug Reaction Overview

The most common adverse events occurring with fludarabine phosphate use include myelosuppression (anemia, leukopenia, neutropenia and thrombocytopenia), leading to decreased resistance to infection, including pneumonia, cough, fever, fatigue, weakness, nausea, vomiting and diarrhea. Other commonly reported events include chills, edema, malaise, peripheral neuropathy, visual disturbances, anorexia, mucositis, stomatitis and skin rash. Serious opportunistic infections have occurred in patients treated with fludarabine phosphate. Fatalities as a consequence of serious adverse events have been reported.

The most frequently reported adverse events and those reactions which are more clearly related to the drug are listed below according to body system regardless of their seriousness. Their frequency (very common ≥1/10, common ≥1/100 to < 1/10, uncommon ≥1/1000 to <1/100) is based on clinical trial data regardless of the causal relationship with fludarabine phosphate. The rare events (≥1/10000 to <1/1000) were mainly identified from post-marketing experience.

Body as a Whole

Infection, fever, fatigue and weakness have been reported very commonly in patients receiving fludarabine phosphate. Malaise and chills have been commonly reported.

Hematopoietic and Lymphatic System

Hematologic events (neutropenia, thrombocytopenia and anemia) have been reported in the majority of patients treated with fludarabine phosphate. Myelosuppression may be severe and cumulative. The prolonged effect of fludarabine phosphate on the decrease in the number of T-lymphocytes may lead to increased risk of opportunistic infections, including those due to latent

viral reactivation, eg. herpes zoster virus, Epstein-Barr-Virus (EBV) and progressive multifocal leukoencephalopathy. Evolution of EBV-infection/reactivation into EBV-associated lymphoproliferative disorder has been observed in immunocompromised patients. Life-threatening and sometimes fatal autoimmune hemolytic anemia has been reported to occur in patients receiving fludarabine phosphate. The majority of patients rechallenged with fludarabine phosphate developed a recurrence in the hemolytic process. (See WARNINGS AND PRECAUTIONS section for information on autoimmune hemolytic anemia associated with fludarabine phosphate).

In rare cases, the occurrence of myelodysplastic syndrome (MDS) has been described in patients treated with fludarabine phosphate. The majority of these patients also received prior, concomitant or subsequent treatment with alkylating agents or irradiation. Monotherapy with fludarabine phosphate had not been associated with an increased risk of development of MDS.

Nervous System

Following administration of fludarabine phosphate at doses of 20-30 mg/m²/day in 133 patients with CLL, reported events included weakness, visual disturbances, loss of hearing, numbness, agitation, confusion, seizure and coma. Peripheral neuropathy has been commonly observed. Confusion is uncommon. Coma, seizures and agitation occur rarely. There was one case of wrist drop. (See WARNINGS AND PRECAUTIONS section for information on neurotoxicity associated with high doses of fludarabine phosphate).

Special Senses

Visual disturbances are commonly reported events in patients treated with fludarabine phosphate. In rare cases, optic neuritis, optic neuropathy and blindness have occurred.

Respiratory System

Pneumonia has been commonly reported. Pneumonia, a frequent manifestation of infection in CLL patients occurred in 16% and 22% of those treated with fludarabine phosphate in the MDACC and SWOG studies, respectively. Pulmonary hypersensitivity reactions to fludarabine phosphate (pulmonary infiltrates, pneumonitis, fibrosis) characterized by dyspnea, and cough are uncommon.

Digestive System

Gastrointestinal disturbances such as nausea and vomiting, anorexia, diarrhea, mucositis and stomatitis are commonly reported. Gastrointestinal bleeding, mainly related to thrombocytopenia, has been reported in patients treated with fludarabine phosphate.

Skin and Appendages

Skin rashes have been commonly reported in patients treated with fludarabine phosphate. In rare cases, a Stevens-Johnson syndrome or toxic epidermal necrolysis (Lyell's disease) may develop.

Cardiovascular System

One patient developed a pericardial effusion possibly related to treatment with fludarabine phosphate. Rare instances of heart failure and arrhythmia have been reported in patients treated with fludarabine phosphate.

Urogenital System

Rare cases of hemorrhagic cystitis have been reported in patients treated with fludarabine phosphate.

Metabolic an Nutritional Disorders

Tumor lysis syndrome has been reported in CLL patients treated with fludarabine phosphate. This complication may include hyperuricaemia, hyperphosphatemia, hypocalcemia, metabolic acidosis, hyperkalemia, hematuria, urate crystalluria and renal failure. The onset of this syndrome may be heralded by flank pain and hematuria. Edema has been commonly reported. Changes in hepatic and pancreatic enzymes levels are uncommon.

The spectrum of adverse reactions reported in patients (n=3000) receiving fludarabine phosphate in studies of lymphomas, other leukemias and solid tumors is consistent with the above data.

DRUG INTERACTIONS

Drug-Drug Interactions

The therapeutic efficacy of fludarabine phosphate may be reduced by dipyridamole and other inhibitors of adenosine uptake.

DOSAGE AND ADMINISTRATION

Dosing Considerations

Incompatibilities

The formulation for intravenous use must not be mixed with other drugs.

Recommended Dose and Dosage Adjustment

The usual starting dose of fludarabine phosphate is 25 mg/m² administered intravenously over a period of approximately 30 minutes, daily for five days every 28 days. Dosage may be decreased based on evidence of hematologic or nonhematologic toxicity.

Note that in patients with decreased renal function (creatinine clearance between 30 and 70 mL/min) the dose should be reduced by up to 50%. Fludarabine phosphate treatment is contraindicated, if creatinine clearance is <30 mL/min. (See WARNINGS AND PRECAUTIONS).

The duration of treatment depends on the treatment success and the tolerability of the drug. FLUDARABINE PHOSPHATE INJECTION should be administered up to the achievement of a maximal response (complete or partial remission, usually 6 cycles) and then the drug should be

Serious Drug Interactions In a clinical investigation using fludarabine phosphate in combination with pentostatin (deoxycoformycin) for the treatment of refractory CLL, there was an unacceptably high incidence of fatal pulmonary toxicity. Therefore, the use of fludarabine phosphate in combination with pentostatin is contraindicated.

discontinued.

Administration

Studies in animals have shown that even in cases of misplaced injections, no relevant local irritation was observed after paravenous, intraarterial, and intramuscular administration of an aqueous solution containing 7.5 mg fludarabine phosphate/mL.

It is strongly recommended that fludarabine phosphate should be only administered intravenously. No cases have been reported in which paravenously administered fludarabine phosphate led to severe local adverse reactions. However, unintentional paravenous administration should be avoided.

FLUDARABINE PHOSPHATE INJECTION comes prepared for parenteral use. Each mL of the solution contains 25 mg of fludarabine phosphate, 25 mg of mannitol and 3.30 mg of sodium hydroxide. The pH range of the final solution is 6.0-7.1.

The product must be further diluted for intravenous infusion administration in PVC bags to a concentration of 1 mg/mL in 5% Dextrose Injection USP, or in 0.9% Sodium Chloride Injection USP.

Fludarabine Phosphate injection (25 mg/mL) is NOT for direct injection. Fludarabine Phosphate injection MUST be diluted with recommended intravenous infusion solutions.

Missed Dose

In the event that a dose is missed the opinion of an oncologist should be sought.

OVERDOSAGE

Higher than recommended doses of fludarabine phosphate have been associated with an irreversible central nervous system toxicity characterized by delayed blindness, coma and death. High doses are also associated with bone marrow suppression manifested by thrombocytopenia and neutropenia. There is no known specific antidote for fludarabine phosphate overdosage. Treatment consists of drug discontinuation and supportive therapy.

ACTION AND CLINICAL PHARMACOLOGY

Mechanism of Action

FLUDARABINE PHOSPHATE INJECTION is a fluorinated analog of adenine that is relatively resistant to deamination by adenosine deaminase.

Fludarabine phosphate (2F-ara-AMP) is a water-soluble prodrug, which is rapidly dephosphorylated to 2-fluoro-ara-A (2F-ara-A) and then phosphorylated intracellularly by deoxycytidine kinase to the active triphosphate 2-fluoro-ara-ATP (2F-ara-ATP). The antitumor activity of this metabolite is the result of inhibition of DNA synthesis via inhibition of ribonucleotide reductase, DNA polymerase α, δ and g, DNA primase and DNA ligase. Furthermore, partial inhibition of RNA polymerase II and consequent reduction in protein synthesis occur. While some aspects of the mechanism of action of 2F-ara-ATP are as yet unclear, it is believed that effects on DNA, RNA and protein synthesis all contribute to the

inhibition of cell growth, with inhibition of DNA synthesis being the dominant factor. In addition, in vitro studies have shown that exposure of CLL lymphocytes to 2F-ara-A triggers extensive DNA fragmentation and apoptosis.

Two open-label studies of fludarabine phosphate have been conducted in patients with CLL refractory to at least one prior standard alkylating agent-containing regimen. Overall objective response rates were 32% in one study, and 48% in the other, with median time to response at 21 and 7 weeks respectively.

Pharmacokinetics

Cellular pharmacokinetics of fludarabine triphosphate: Maximum 2F-ara-ATP levels in leukemic lymphocytes of CLL patients were observed at a median of 4 hours and exhibited considerable variation with a median peak concentration of approximately 20 μM. 2F-ara-ATP levels in leukemic cells were always considerably higher than maximum 2F-ara-A levels in the plasma, indicating an accumulation at the target sites. In vitro incubation of leukemic lymphocytes showed a linear relationship between extracellular 2F-ara-A exposure (product of 2F-ara-A concentration and duration of incubation) and intracellular 2F-ara-A enrichment. Two independent investigations respectively reported median half-life values of 15 and 23 hours for the elimination of 2F-ara-ATP from target cells.

No clear correlation was found between 2F-ara-A pharmacokinetics and treatment efficacy in cancer patients; however, the occurrence of neutropenia and hematocrit changes indicated that the cytotoxicity of fludarabine phosphate depresses hematopoiesis in a dose-dependent manner.

Plasma and urinary pharmacokinetics of fludarabine (2F-ara-A): Phase I studies in humans have demonstrated that fludarabine phosphate is rapidly converted to the active metabolite, 2F-ara-A, within minutes after intravenous infusion. Consequently, clinical pharmacology studies have focused on 2F-ara-A pharmacokinetics. After single doses of 25 mg 2F-ara-AMP/m² to cancer patients infused over 30 minutes, 2F-ara-A reached mean maximum concentrations in the plasma of 3.5 - 3.7 μM at the end of infusion. Corresponding 2F-ara-A levels after the fifth dose showed a moderate accumulation with mean maximum levels of 4.4 - 4.8 μM at the end of infusion. During a 5-day treatment cycle, 2F-ara-A plasma trough levels increased by a factor of about 2. Accumulation of 2F-ara-A over several treatment cycles does not occur. Post maximum levels decayed in three disposition phases with an initial half-life of approximately 5 minutes, an intermediate half-life of 1-2 hours and a terminal half-life of approximately 20 hours.

An interstudy comparison of 2F-ara-A pharmacokinetics resulted in a mean total plasma clearance (CL) of 79 mL/min/m² (2.2 mL/min/kg) and a mean volume of distribution (V_ss) of 83 L/m² (2.4 L/kg). The data showed a high interindividual variability. After i.v. and peroral administration of fludarabine phosphate plasma levels of 2F-ara-A and areas under the plasma level time curves increased linearly with the dose, whereas half-lives, plasma clearance and volumes of distribution remained constant independent of the dose indicating a dose-linear behaviour.

After oral fludarabine phosphate doses, maximum 2F-ara-A plasma levels reached approximately 20-30% of corresponding i.v. levels at the end of infusion and occurred 1-2 hours after dosing. The mean systemic 2F-ara-A availability was in the range of 50-65% following

single and repeated doses and was similar after ingestion of a solution or an immediate release tablet formulation. After oral doses of 2F-ara-AMP with concomitant food intake a slight increase (<10%) of systemic availability (AUC), a slight decrease in maximum plasma levels (C_max) of 2F-ara-A and a delayed time to occurrence of C_max was observed; terminal half-lives were unaffected.

The mean steady-state volume of distribution (Vd_ss) of 2F-ara-A in one study was 96 L/m² suggesting a significant degree of tissue binding. Another study, in which Vd_ss, for patients was determined to be 44 L/m², supports the suggestion of tissue binding.

Based upon compartmental analysis of pharmacokinetic data, the rate-limiting step for excretion of 2F- ara-A from the body appears to be release from tissue binding sites. Total body clearance of 2F-ara-A has been shown to be inversely correlated with serum creatinine, suggesting renal elimination of the compound.

Special Populations and Conditions

Renal Insufficiency: A pharmacokinetic study in patients with and without renal impairment revealed that, in patients with normal renal function, 40 to 60% of the administered i.v. dose was excreted in the urine. Mass balance studies in laboratory animals with ³H-2F-ara-AMP showed a complete recovery of radio-labelled substances in the urine. Another metabolite, 2F-ara-hypoxanthine, which represents the major metabolite in the dog, was observed in humans only to a minor extent. Patients with impaired renal function exhibited a reduced total body clearance, indicating the need for a reduced dose. Total body clearance of 2F-ara-A has been shown to be inversely correlated with serum creatinine, suggesting renal elimination of the compound. This was confirmed in a study of the pharmacokinetics of 2F-ara-A following administration of 2F-ara-AMP to cancer patients with normal renal function or varying degrees of renal impairment. The total body clearance of the principal metabolite 2F-ara-A shows a correlation with creatinine clearance, indicating the importance of the renal excretion pathway for the elimination of the compound. Renal clearance represented on average 40% of the total body clearance. In vitro investigations with human plasma proteins revealed no pronounced tendency of 2F-ara-A protein binding.

STORAGE AND STABILITY

Store FLUDARABINE PHOSPHATE INJECTION under refrigeration between 2°C and 8°C. Do not freeze. Discard unused portion.

FLUDARABINE PHOSPHATE INJECTION should be used immediately upon dilution with the recommended infusion solutions and not stored.

FLUDARABINE PHOSPHATE INJECTION contains no antimicrobial preservative and thus care must be taken to ensure the sterility of prepared solutions.

Parenteral drug products should be inspected visually for particulate matter and discoloration prior to administration.

SPECIAL HANDLING INSTRUCTIONS

FLUDARABINE PHOSPHATE INJECTION should not be handled by pregnant staff. Proper

handling and disposal procedures should be observed, with consideration given to the guidelines used for cytotoxic drugs. Any spillage or waste material may be disposed of by incineration.

Caution should be exercised in the preparation of FLUDARABINE PHOSPHATE INJECTION solution. The use of latex gloves and safety glasses is recommended to avoid exposure in case of breakage of the vial or other accidental spillage. If the solution comes into contact with the skin or mucous membranes, the area should be washed thoroughly with soap and water. In the event of contact with the eyes, rinse them thoroughly with copious amounts of water. Exposure by inhalation should be avoided.

DOSAGE FORMS, COMPOSITION AND PACKAGING

Medicinal ingredients: Each vial contains 50 mg of fludarabine phosphate.

Non-medicinal ingredients: Each vial contains 50 mg mannitol and 6.60 mg sodium hydroxide.

pH: 6.0-7.1

Availability :

FLUDARABINE PHOSPHATE INJECTION is supplied as 2 mL per vial of 50 mg fludarabine phosphate, 50 mg of mannitol and 6.60 mg of sodium hydroxide.

FLUDARABINE PHOSPHATE INJECTION is a single use vial.

FLUDARABINE PHOSPHATE INJECTION is supplied in a single vial carton.

FLUDARABINE PHOSPHATE INJECTION uses a latex free stopper.

PART II: SCIENTIFIC INFORMATION

PHARMACEUTICAL INFORMATION

Drug Substance

Common name: Fludarabine Phosphate

Chemical name: 9H Purin-6-amine, 2-fluoro-9-(5-O-phosphone-β-D-arabinofuranosyl)

Molecular formula and molecular mass: C₁₀H₁₃FN₅O₇P 365.21

Structural formula:

Physicochemical properties: Fludarabine phosphate is a white to almost white, crystalline powder. It has pKa values of 3.2 ± 0.1 and 5.8 ± 0.1 and pH value of 2.0 (9mg/mL in water).

Fludarabine phosphate is freely soluble in

dimethylsulphoxide and in dimethylacetamide; sparingly soluble in water; slightly soluble in methanol; insoluble in acetone and in dichloromethane.

CLINICAL TRIALS

Two single-arm open-label studies of fludarabine phosphate have been conducted in patients with CLL refractory to at least one prior standard alkylating-agent-containing regimen. In a study conducted by M.D. Anderson Cancer Center (MDACC), 48 patients were treated with a dose of 22-40 mg/m² daily for 5 days every 28 days. Another study conducted by the Southwest Oncology Group (SWOG) involved 31 patients treated with a dose of 15-25 mg/m² for 5 days every 28 days. The overall objective response rates were 48% and 32% in the MDACC and SWOG studies, respectively. The complete response rate in both studies was 13%; the partial response rate was 35% in the MDACC study and 19% in the SWOG study. These response rates were obtained using standardized response criteria developed by the National Cancer Institute CLL Working Group and achieved in heavily pre-treated patients. The ability of fludarabine

phosphate to induce a significant rate of response in refractory patients suggests minimal cross-resistance with commonly used anti-CLL agents.

The median time to response in the MDACC and SWOG studies was 7 weeks (range of 1 to 68 weeks) and 21 weeks (range of 1 to 53 weeks), respectively. The median duration of disease control was 91 weeks (MDACC) and 65 weeks (SWOG). The median survival of all refractory CLL patients treated with fludarabine phosphate was 43 weeks and 52 weeks in the MDACC and SWOG studies, respectively. Normalized lymphocyte count, one measure of disease regression, occurred at a median of 2 weeks (complete responders), 2 weeks (partial responders) and 22 weeks (non-responders).

Rai stage improved to Stage II or better in 7 of 12 MDACC responders (58%) and in 5 of 7 SWOG responders (71%) who were Stage III or IV at baseline. In the combined studies, mean hemoglobin concentration improved from 9.0 g/dL at baseline to 11.8 g/dL at the time of response in a subgroup of anemic patients. Similarly, average platelet count improved from 63,500/mm³ to 103,300/mm³ at the time of response in a subgroup of patients who were thrombocytopenic at baseline.

DETAILED PHARMACOLOGY

Mechanism of Action

The biological activity of 2F-ara-A was assessed in a number of models. 2F-ara-A has been shown to inhibit DNA synthesis in cultured mouse leukemia L1210 cells and in an in vivo mouse L1210 leukemia model. Total RNA synthesis in vitro was not inhibited by treatment with 2F-ara-A; however, protein synthesis was reduced substantially. It has been shown that 2F-ara-A is not deaminated by adenosine deaminase, contributing to the stability of the compound.

The activity, metabolism and toxicity of 2F-ara-A in the human lymphoblastoid T-cell line (CCRF-CEM) were compared with 9-β-D-arabinofuranosyl-adenine (ara-A). Inhibition of cell growth was equivalent for these two agents, provided that ara-A was protected from deamination. Similar studies conducted with CCRF-CEM showed that ara-A and 2F-ara-A exerted early killing effects preferentially during the S-phase of cell proliferation. Both compounds were converted to the triphosphate form, which accumulated intracellularly and inhibited DNA synthesis. This nucleoside metabolite, 2F-ara-ATP, was also shown to inhibit DNA polymerase α and to a lesser extent ribonucleotide reductase in mouse leukemia cells (L1210), human epithelial cells (HEp-2), and HeLa cells.

In the systems tested, 2F-ara-ATP is the active metabolite which acts by inhibiting DNA polymerase α and ribonucleotide reductase thus preventing DNA synthesis. In addition, in vitro studies have shown that exposure of CLL lymphocytes to 2F-ara-A triggers extensive DNA fragmentation and apoptosis.

Antitumor Activity

The effects of schedule and route of administration on the antitumor activity of fludarabine phosphate were examined using an in vivo mouse leukemia model (implanted L1210 leukemia cells). The drug was active following intraperitoneal administration on all treatment schedules. Antitumor activity increased almost three-fold when the number of drug treatments was

increased. In addition, the administration of several doses in one day was more effective than administration of one larger dose.

A single administration (900 mg/kg) on day 1 produced an increased life span (ILS) of 42% while administration of a smaller dose (250 mg/kg) 3 times a day on day 1 (total dose 750 mg/kg) gave a 98% ILS. This pattern of increased activity with administration of several doses in a day was also observed with the intermittent treatment schedule. A single administration on each of 3 days (total dose 2010 mg/kg) produced an ILS of 122% while administration of a smaller dose 3 times a day over 3 days (total dose 1125 mg/kg) produced the greatest activity, a 525% ILS with 6 long-term survivors (50 day) among the tumor-bearing mice.

With the administration of the drug 3 times a day on day 1, negative animal weight differences (body weight change over 5 days for test animals minus that for controls) of more than 4 grams at the highest dose evaluated suggests some acute drug toxicity. Based on equivalent total doses, administration of 3 smaller doses per day at 3-hour intervals was much more effective than a single administration for each day of treatment using the in vivo mouse leukemia model.

A single oral administration of fludarabine phosphate on day 1 was not effective against the L1210 leukemia. However, when given as 5 daily oral doses, the highest non-toxic dose of the drug, defined as the dose which results in at least 7 or 8 50-day survivors among the normal mice (800 mg/kg daily on days 1-5), was effective in a maximal ILS of 50%.

When the drug was administered i.v., it was more effective with daily administration for 5 days than it was with a single injection on day 1. Daily treatment for 5 days at a non-toxic dose level increased the life span of tumor-bearing mice by 71% and a higher, more toxic, treatment for 5 days produced an ILS of 95%; in contrast, in single i.v. treatment on day 1 produced a maximum ILS of 28%.

The intraperitoneally (i.p.) implanted L1210 leukemia was less sensitive to fludarabine phosphate when the drug was given either intravenously (i.v.) or orally than when compared to i.p. administration. A maximal ILS value of 122% was produced following i.p. administration of 266 mg/kg on days 1-5. This same dose given by i.v. administration on days 1-5 produced an ILS value of 95%. In contrast, a dose of 1600 mg/kg given orally on days 1-5 produced only a 75% ILS. However, with both i.p. and i.v. administration, the dose that produced the maximum ILS value was toxic to the non-tumored animals.

Fludarabine phosphate also demonstrated activity against the intraperitoneal implanted P388 leukemia. In two different experiments, the drug increased the life span of mice bearing the P388 leukemia by 115% and 53% following i.p. administration of 200 and 100 mg/kg injections, respectively, on days 1-9.

Cytotoxicity of Fludarabine Phosphate

Fludarabine phosphate has demonstrated significant antitumor activity against intraperitoneally (i.p.) implanted murine L1210 leukemia and the human LX-1 lung tumor xenograft. The drug has shown moderate activity against the murine subcutaneously (s.c.) implanted CD8F_l mammary epithelioma and the i.p. implanted P388 lymphocytic leukemia. Fludarabine phosphate was not active against the i.p. implanted B16 melanoma, the s.c. implanted colon tumor, or the intravenously (i.v.) implanted Lewis lung epithelioma, nor was it effective against the human CX-1 colon or MX-1 mammary xenografts in the subrenal assay.

Effects on Bone Marrow Survival and Tumor Cell Sensitivity

Fludarabine phosphate was tested in an in vitro human bone marrow cell survival assay and tumor cell sensitivity assay. The sensitivity of normal human granulocyte-macrophage colony-forming units in culture (GM-CFUC) showed a simple negative exponential curve characterized by a logarithmic decrease in survival as a function of drug concentration. Fludarabine phosphate exhibited an LD₆₃ of 0.51μg/mL for normal human granulocyte-macrophage colony-forming units in culture (GM-CFUC). In the tumor sensitivity assay, fludarabine phosphate demonstrated an LD₄₀ and LD₇₈ of 0.26 and 0.77 μg/mL, respectively.

Blood and bone marrow samples obtained from patients with relapsed leukemia and lymphoma after treatment with a single dose of 20-125 mg/m² fludarabine phosphate revealed that the area under the concentration-time curves for 2F-ara-A and 2F-ara-ATP were increased in proportion to the product dose. There was a high correlation between 2F-ara-ATP level in circulating leukemic cells and those in bone marrow cells aspirated at the same time. DNA synthetic capacity of leukemic cells was inversely related to the associated 2F-ara-ATP concentration. 2F-ara-ATP concentrations were three times higher in bone marrow cells from patients with lymphomatous bone marrow involvement than from those without evidence of marrow disease.

A dose response relationship between fludarabine phosphate concentration and inhibition of DNA synthesis in leukemia cells and bone marrow cells in culture was obtained.

Bone marrow progenitor cells from a normal subject and 10 patients with solid tumors, whose bone marrow was free of metastases, were treated with fludarabine phosphate and other cytotoxic drugs, using a bilayer soft agar culture. The in vitro effect of the drugs on bone marrow progenitor cells was not as toxic as expected relative to the myelosuppressive potency observed in vivo. In the case of fludarabine phosphate, it has been postulated that these findings might be related to incomplete in vitro phosphorylation to the triphosphate, 2F-ara-ATP.

Lymphocytotoxicity in Humans

Fludarabine phosphate was assessed for its lymphocytotoxicity in 11 patients receiving the investigational drug for treatment of nonhematologic cancers refractory to standard treatment. Fludarabine phosphate was administered by intravenous infusion at doses ranging from 18 mg/m²/day to 40 mg/m²/day, with each dose given on a 5-day dosing regimen.

Lymphocyte subsets were determined prior to treatment and on day 5 of treatment, 4 hours after the infusion. Observations indicated that lymphocytopenia developed rapidly but was reversible. Total T- lymphocyte counts fell during all treatment regimens, with a 90% decrease in mean absolute T-cell count. All major T-lymphocyte subsets were affected. B-lymphocyte counts decreased by 50% on average. Recoveries of total mononuclear cells, total T-cells and non-T,

non-B cells were reduced substantially by fludarabine phosphate treatment. B-cell recovery was not affected.

These results indicate that T-cells are more sensitive than B-cells to the cytotoxic effects of fludarabine phosphate.

Modulation of T-Cell Function by Fludarabine Phosphate

The effects of fludarabine phosphate on the growth and function of bone marrow and peripheral blood mononuclear cells (PBMC) from cancer patients were evaluated. Drug toxicity was dependent on time of incubation and concentration of fludarabine phosphate tested. After a 3- hour incubation of PBMC with 1 μg/mL of fludarabine phosphate, there was no effect on cell number whereas, after 48 hours, the cell count was 59% of control, untreated cells. In contrast, a 3-hour or 48-hour incubation of PBMC with 100 μg/mL of fludarabine phosphate reduced cell number to 65.7% or 63% of control, respectively.

Lymphocyte subpopulations of normal PBMCs were evaluated after treatment in vitro with fludarabine phosphate for 72 hours. A dose-dependent decrease in total T-cell number was noted. Incubation with 1 μg/mL of fludarabine phosphate reduced T-cells by 16.7%; 100 μg/mL reduced T-cells by 42%. The subset of T-cells predominantly affected was T-helper cells, reduced by 53.5% after incubation with 100μg/mL of fludarabine phosphate. B-cells, monocytes, and natural killer cells were not reduced, but rather increased relative to control. Fludarabine phosphate also inhibited the response of PBMC to mitogens in a dose-and time-dependent manner.

In Vitro Testing of Fludarabine Phosphate in Glioma Cell Cultures

Fludarabine phosphate was tested for growth inhibitory effects on human glioma cells isolated from patient specimens. Cells were treated with 1-10 μM of fludarabine phosphate beginning 4 days after cells were plated. After 3 more days of incubation, cell number was determined. Inhibition of cell growth was dose-dependent and approximately equal to inhibition seen after treatment with the same concentrations of 5-fluorouracil. Dose-dependent growth inhibition was also observed when interferon-beta (1-1000 IU/mL) was incubated with glioma cell cultures. Although the combination of fludarabine phosphate and 5-fluorouracil or interferon-beta produced additive inhibitory effects, no synergistic effects were observed

Pharmacokinetics (Animals)

Fludarabine phosphate and its metabolites have been studied in mice, dogs, miniature pigs, and monkeys to elucidate their pharmacokinetic, distribution, and excretion profiles.

In the mouse, dog, and monkey, the pharmacokinetics of fludarabine phosphate and its major metabolite, 2F-ara-A, generally exhibited bi-compartmental characteristics after intravenous administration, with rapid clearance and relatively large volumes of distribution.

The pharmacokinetic parameters of fludarabine phosphate and its metabolites are presented in Table 1 and Table 2, located on the following pages.

Tissue Distribution, Metabolism and Excretion in Animals

Tissue distribution and excretion studies were conducted with fludarabine phosphate in mice, dogs, and monkeys at doses between 30 and 500 mg/m².

Fludarabine phosphate is metabolized to 2F-ara-A and, to a lesser extent, 2F-ara-HX in the mouse and monkey, while in the dog, 2F-ara-A and 2F-ara-HX are both major metabolites. The majority of the administered compound is metabolized and then eliminated in the urine within 24 hours after dose administration.

The metabolism, distribution and excretion information is presented in Table 3 located on the following pages.

Table 1 PHARMACOKINETIC PARAMETERS OF FLUDARABINE PHOSPHATE AND 2F-ARA-A

HPLC: High performance liquid chromatography

TLC: thin layer chromatography

Table 1 (Continued) PHARMACOKINETIC PARAMETERS OF FLUDARABINE PHOSPHATE AND 2F-ARA-A

HPLC: High performance liquid chromatography

TLC: thin layer chromatography

Table 2 PHARMACOKINETIC PARAMETERS OF FLUDARABINE PHOSPHATE AND METABOLITES

Cmax: maximal concentration I.P.: intraperitoneal

Table 3 METABOLISM, DISTRIBUTION AND EXCRETION OF FLUDARABINE PHOSPHATE

Table 3 (Continued) METABOLISM, DISTRIBUTION AND EXCRETION OF FLUDARABINE PHOSPHATE

Table 3 (Continued) METABOLISM, DISTRIBUTION AND EXCRETION OF FLUDARABINE PHOSPHATE

Table 3 (Continued) METABOLISM, DISTRIBUTION AND EXCRETION OF FLUDARABINE PHOSPHATE

Pharmacokinetics (Humans)

The pharmacokinetics of fludarabine phosphate given intravenously have been determined in adult patients undergoing Phase I clinical trials at the University of Texas Health Science Center at San Antonio (UT), the University of Texas System Cancer Center at the M.D. Anderson Cancer Center (MDACC) and at Ohio State University (OSU). In addition, the pharmacokinetics of intraperitoneal fludarabine phosphate was also determined at UT and the pharmacokinetics of intravenous fludarabine phosphate in pediatric patients with leukemias and solid tumors were determined at the Children's Hospital of Los Angeles, the National Cancer Institute (NCI) and the Mayo Clinic.

Preliminary nonclinical and Phase I human studies demonstrated that fludarabine phosphate is rapidly converted to 2F-ara-A within minutes after intravenous infusion, and then phosphorylated intracellularly by deoxycytidine kinase to the active triphosphate, 2F-ara-ATP. Consequently, clinical pharmacology studies have focused on 2F-ara-A pharmacokinetics.

Described on the following pages are three principal pharmacokinetic studies that characterize the pharmacokinetic parameters of 2F-ara-A. Despite the differences in dosage and dosing schedules between these various studies discussed on the following pages, several consistent results were obtained. For the infusion studies, a mean terminal half-life of 9.2 hours was found in the population of patients studied at UT, and a median terminal half-life of approximately 8 hours was observed in the patients studied at MDACC. These values compare favorably to the 10.16-hour mean terminal half-life reported by the OSU investigators following large intravenous bolus injections. The terminal half-life of 2F-ara-A does not appear to be dose-dependent, as the doses used in these studies ranged from 18 to 260 mg/m².

The discrepancies between the studies regarding the biphasic or triphasic elimination patterns appear to be due to differences in sampling schedules and duration of intravenous administration. In addition, sampling duration has an impact upon the calculated value of the terminal half-life (t_½γ). The majority of pharmacokinetic studies use a blood sampling duration of 24 to 30 hours, which gives a calculated terminal half-life (t_½γ) of 8 - 10 hours. However, when the sampling duration is increased to 72 hours, the additional time points give a calculated t_½γ of up to 31 hours. Because the plasma concentration of 2F-ara-A declines more than 50-fold from the peak concentration before this long elimination phase, the consequences of the relatively low 2F-ara- A concentration remaining in the plasma after 24 hours (<0.1 pmol) remain uncertain, as far as drug scheduling is concerned.

In addition, both the UT and OSU investigators found a positive correlation between area under concentration-time curves and degree of neutropenia reinforcing the assertion that toxicity (myelosuppression) is dose related.

Phase I-II Study of Fludarabine in Hematologic Malignancies (Study No. T83-1275) conducted at University of Texas, San Antonio

Methods

The pharmacokinetic parameters of the principal metabolite of fludarabine phosphate, 2F-ara-A, were determined in seven adult patients (6 male; 1 female) who received fludarabine phosphate at doses of 18 or 25 mg/m²/day as a thirty-minute intravenous infusion daily for five consecutive days. Blood and urine samples were analyzed by HPLC for concentrations of 2F-ara-A.

The plasma concentration-time data, which were determined by HPLC, were analyzed by non-linear least squares regression analysis (NONLIN) using a zero order infusion input with first order elimination from the central compartment. Both a two and a three compartment model were tested and the data fitted to the two compartment open model.

Pharmacokinetic Parameters

Peak plasma concentrations of 2F-ara-A ranged from 0.199 to 0.876 μg/mL and appeared to be related to the dose and rate of infusion. Mean plasma concentrations of 2 fluoro-ara-A on days 1 and 5 in patients receiving 18 mg/m²/day were 0.39 and 0.51 μg/mL, respectively. Mean plasma concentrations of 2F-ara-A on days 1 and 5 in patients receiving 25 mg/m²/day were 0.57 and 0.54 μg/mL, respectively. There was no drug accumulation during the five-day treatment period.

The pharmacokinetic parameters derived from this study are presented in Table 4.

Table 4 2F-ARA-A KINETIC PARAMETERS

^aPatient omitted from calculation of mean and SD

^bDay 5 levels drawn on day 4

^cDay 5 levels not studied

^dHarmonic mean half-life

The mean central compartment volume of distribution (Vd) was 37.8 L/m² with a mean steady-state volume of distribution (Vdss) of 96.2 L/m². The mean tissue clearance was 20.1 L/h/m² and the mean plasma clearance was 9.1 L/h/m². Plasma concentrations declined bi-exponentially with a harmonic mean initial half-life (t_½α) of 0.6 hours and a harmonic mean terminal half-life (t_½β) of 9.2 hours. As presented in Table 5, approximately 24% of the parent compound,

fludarabine phosphate, was excreted in the urine as 2F-ara-A during the five-day treatment period.

Table 5 URINARY EXCRETION OF 2F-ARA-A

Correlation of Pharmacokinetic Parameters with Clinical Parameters

As presented in Table 6, a correlation was observed between decreasing absolute granulocyte count and the area under the concentration-time curve (AUC). The Spearman rank correlation coefficient between absolute granulocyte count and AUC was -0.94 which was statistically significant (p<0.02). The Spearman rank correlation coefficient was also calculated between absolute granulocyte count and total plasma clearance (TPC). Here the correlation coefficient was 0.94 which was also statistically significant (p<0.02). The correlation coefficient between creatinine clearance and TPC was 0.828 (0.05<p<0.1). No correlation was observed between TPC and any of the liver function measurements.

Table 6 COMPARISON OF AUC WITH ABSOLUTE GRANULOCYTE NADIR AND

CREATININE CLEARANCE

^aDays 0 - 5

^bAbsolute granulocyte count

Summary and Conclusions

Intravenous doses of 18 and 25 mg/m²/day for 5 days exhibited bi-exponential decay with a mean initial half-life (t_½α) of 0.6 hours and a mean terminal half-life (t_½β) of 9.2 hours. The mean

plasma clearance was 9.1 L/h/m² and the mean tissue clearance was 20.1 L/h/m². The mean Vd_ss was 96.2 L/m², which is approximately twice body weight, suggesting that tissue binding of the drug occurs. In addition, there was a significant inverse correlation between AUC and absolute granulocyte count (r=-0.94, p<0.02) suggesting that myelosuppression is dose related. Phase I-II Study of Fludarabine in Hematologic Malignancies (Study No. T83-1275) conducted at the M.D. Anderson Cancer Center

Methods

The pharmacokinetic parameters of the fludarabine phosphate metabolite, 2F-ara-A, were determined in 19 adult patients (12 male; 7 female) who received the drug as a 30-minute intravenous infusion daily for five consecutive days. Ten of the patients were diagnosed as having lymphoma and nine as having leukemia. In this study, five patients received doses of 20 mg/m²/day, five patients received doses of 25 mg/m²/day, one patient received 30 mg/m²/day, four patients received 50 mg/m²/day, two patients received 100 mg/m²/day, and an additional two patients received 125 mg/m²/day. Pharmacokinetic profiles were generally determined after the first dose of fludarabine phosphate. Plasma concentrations of 2F-ara-A and intracellular concentrations of 2F-ara-ATP were determined by HPLC. Intracellular concentrations were determined for mononuclear cells obtained from blood and bone marrow samples. The incorporation of 2F-ara-ATP into nucleic acids was determined using HPLC and liquid scintillation counting methods.

Pharmacokinetic Parameters

Plasma concentrations of fludarabine phosphate were undetectable at the times when the first samples were obtained. Of the patients receiving 20 or 25 mg/m²/day, only two had detectable peak 2F-ara-A concentrations (1.4 and 2.2 μM) and, in this group of patients, 2F-ara-A levels were completely undetectable three hours after the completion of infusion of fludarabine phosphate.

At fludarabine phosphate dose levels of 50-125 mg/m²/day, the disappearance of 2F-ara-A was biphasic and independent of dose with a median initial half-life (t_½α) of 1.41 hours and a median terminal half life (t_½β) of approximately 8 hours. Plasma pharmacokinetic parameters for patients with relapsed leukemia (N=8, Patients #5-12) are presented in Table 7.

Table 7 PHARMACOLOGICAL CHARACTERISTICS FOR 2F-ARA-A IN THE

PLASMA OF PATIENTS WITH RELAPSED LEUKEMIA

^a Initial rate of elimination

^bTerminal rate of elimination

^cArea under the concentration-time curve calculated to 24 h

^dAs the 2-h sample was the earliest obtained, this value is based on extrapolation of the line to 30 minutes

^eThe median value excluding patients 5 and 6 whose elevated creatinine levels may signal impaired renal function and thus a longer t_½β

A wide range of variation of pharmacokinetic parameters of 2F-ara-ATP in circulating leukemic cells was observed; however, when the median peak 2F-ara-ATP concentrations of 24 hour AUC values were compared at each dosage increment (20 or 25 mg/m², 50 mg/m², and 100 or 125 mg/m²), a clear dose-dependence emerged (Table 8). Cellular elimination was not dose-dependent, with a half-life of approximately 15 hours at all dose levels. There was a strong correlation between the 2F-ara-ATP levels in leukemic cells obtained from peripheral blood and those found in bone marrow (r=0.84, p=0.01) suggesting that there were no pharmacological barriers in the bone marrow. Those patients with bone marrow involvement had the highest 2F-ara- ATP levels. In addition, intracellular 2F-ara-ATP levels in circulating leukemic cells at 12-14 hours after fludarabine phosphate infusion were inversely related to the DNA synthetic capacity of the cells relative to pretreatment. DNA synthesis remained maximally inhibited (>80%) until cellular concentrations of 2F-ara-ATP fell below 90 μM.

Table 8 PHARMACOLOGICAL CHARACTERISTICS OF 2F-ARA-ATP IN

CIRCULATING LEUKEMIC CELLS

^aElimination half-life

^bArea under the concentration-time curves calculated to 24 h

^cChronic lymphocytic leukemia

^dDiffuse, well-differentiated lymphoma

^eDiffuse, large cell lymphoma

^fNodular mixed cell lymphoma

^gAcute myelomonocytic leukemia

^hAcute myeloblastic leukemia

ⁱAcute lymphoblastic leukemia

^jChronic myelogenous leukemia in blast crisis

Summary and Conclusions

Intravenous doses of 20-125 mg/m²/day exhibited bi-exponential decay in plasma with a median initial half-life (t_½α) of 1.41 hours and a median terminal half-life (t_½β) of approximately 8 hours for 2F-ara-A. The median intracellular half-life for 2F-ara-ATP was approximately 15 hours. The terminal half-lives of both 2F-ara-A and 2F-ara-ATP were not dependent on the dose of fludarabine phosphate. In addition, there was a high correlation between 2F-ara-ATP levels in circulating leukemic cells and bone marrow cells aspirated at the same time. DNA synthetic capacity of leukemic cells was inversely related to intracellular 2F-ara-ATP levels. Finally, 2F-ara-ATP levels were approximately three times higher in bone marrow cells from patients with bone marrow involvement than from those patients without evidence of bone marrow disease, suggesting that tumor cells may have a greater capacity to accumulate and retain nucleoside analogue triphosphates than do normal cells.

Phase I - Pharmacokinetic Study of Fludarabine (NSC-312887) (Study No. W83-328) conducted at Ohio State University

Methods

Twenty-six patients participated in this study, in which fludarabine phosphate was administered as a rapid intravenous (I.V.) infusion of 2-5 minutes duration. Seven patients received fludarabine phosphate at a dose of 260 mg/m², one patient received a dose of 160 mg/m², eight patients received a dose of 120 mg/m², four patients received 100 mg/m², and an additional six patients received 80 mg/m². Plasma concentrations of fludarabine phosphate could not be detected five minutes after the discontinuation of the infusion. Plasma concentrations of 2F-ara- A, the principal metabolite of fludarabine phosphate, were determined by HPLC over a time period of 0-30 hours post dosing. The plasma concentration-time data were analyzed by the NONLIN computer program and fitted to a three-compartment open model with first-order elimination from the central (blood) compartment, using the equations for rapid intravenous infusion.

Pharmacokinetic Parameters

Harmonic mean half-lives, mean residence time and total body clearance of 2F-ara-A for each of the dose levels are shown in Table 9. This metabolite exhibited a very short initial half-life (mean t_½α) of 5.42 minutes, followed by an intermediate half-life (mean t_½β) of 1.38 hours and terminal half-life (mean t_½γ) of 10.16 hours. In the 26 patients, the terminal half-lives ranged

from 4.92 to 19.7 hours. The harmonic mean residence time (Vd_ss/C1T) was 10.4 hours, and total body clearance (C1T) ranged from 26.5 to 120.4 mL/min/m² with a mean of 68.98 mL/min/m².

Table 9 2F-ARA-A HARMONIC MEAN HALF-LIVES, MEAN RESIDENCE TIME,

AND TOTAL BODY CLEARANCE IN PATIENTS

C.V.: coefficient of variation

Table 10 2F-ARA-A MEAN VOLUME PHARMACOKINETIC PARAMETERS

The mean volume parameters for each dosage level are shown in Table 10. The central compartment volume of distribution was approximately 20% of body weight (V₁ = 7.30 L/m²). The steady-state volume of distribution indicated significant binding of the drug to tissue components (Vd_ss=44.22 L/m²). The smallest of the microscopic rate constants was k₃₁, indicating release of drug from the deep tissue compartment to be the rate-determining step in the elimination of 2F-ara-A from the body. Table 11 lists the microscopic rate constants for the first nine patients studied.

Table 11 2F-ARA-A MICROSCOPIC RATE CONSTANTS (N= 9)

Correlation of Pharmacokinetic Parameters with Clinical Parameters

Upon completion of the pharmacokinetic studies, a multivariate correlation analysis was undertaken of all pharmacokinetic parameters with the following clinical parameters: bilirubin, serum creatinine, creatinine clearance, BUN, SGOT, SGPT, LDH, alkaline phosphatase, hemoglobin, hematocrit, baseline WBC, baseline platelets, WBC nadir, platelet nadir, WBC toxicity grade, platelet toxicity grade, nausea and vomiting grade, age and sex. Pearson correlation coefficients were substantiated by Spearman correlations. Despite the small number of patients, total body clearance correlated well with creatinine clearance and serum creatinine indicating that renal excretion is important for the elimination of the drug from the body. The volume parameters, particularly Vd_ss and Vd_γ, correlated with creatinine clearance and serum creatinine (p ≤0.011). A positive correlation of Cl_T, with hemoglobin and hematocrit was observed (p≤0.035) and may be due to the metabolism of 2F-ara-A in the RBC. In addition, apparent correlations of Vd_γ with WBC toxicity (p=0.025) and γ with hematocrit (p=0.035) were observed. Tables 12 and 13 list the correlation coefficients and p values for the above correlations.

Table 12 CORRELATION OF 2F-ARA-A PHARMACOKINETIC PARAMETERS

WITH CREATININE CLEARANCE AND SERUM CREATININE

^aPearson correlation coefficients which were substantiated by Spearman correlations.

Table 13 CORRELATION OF 2F-ARA-A PHARMACOKINETIC PARAMETERS

WITH OTHER CLINICAL PARAMETERS

^aPearson correlation coefficients which were substantiated by Spearman correlations.

A rank ordering of the areas under the plasma concentration-time curve (AUC) for the first nine patients enrolled in the study showed good agreement with the corresponding severity of neutropenia developed by each patient (Table 14). Thus, the capacity of the compound to depress hematopoiesis appears to be dos-related.

Table 14 AREAS UNDER THE PLASMA CONCENTRATION-TIME CURVE AND

NEUTROPENIA GRADE

TOXICOLOGY

Toxicology information from acute toxicity (Table 15 and Table 16), long term toxicity (Table 17), mutagenicity (Table 18), and reproductive studies (Table 19) is presented in the following pages.

Table 15 ACUTE TOXICITY STUDIES – MOUSE

Table 15 (Continued) ACUTE TOXICITY STUDIES – MOUSE

^a=½LD₁₀ ^b=LD₁₀ ^c=LD₅₀

Table 16 ACUTE TOXICITY STUDIES - RAT AND DOG

MELD = Mouse Equivalent Lethal Dose ^a=1/10 MELD₁₀

^b=MELD₁₀

^c=2 x MELD₁₀

^d=3 x MELD₁₀

^e=4 x MELD₁₀

Table 17 SUBCHRONIC STUDIES INTRAVENOUS 13-WEEK TOXICITY STUDIES IN RATS AND DOGS

Table 18 MUTAGENICITY STUDIES

Table 18 (Continued) MUTAGENICITY STUDIES

Table 18 (Continued) MUTAGENICITY STUDIES

Table 19 REPRODUCTIVE STUDIES INTRAVENOUS DEVELOPMENTAL TOXICITY STUDIES OF FLUDARABINE PHOSPHATE

Table 19 (Continued) REPRODUCTIVE STUDIES INTRAVENOUS DEVELOPMENTAL TOXICITY STUDIES OF FLUDARABINE PHOSPHATE

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