EXPAREL™, an investigational drug product, is a new formulation of a local anesthetic (numbing medicine) that is designed to be longer acting than the currently-available local anesthetics. The purpose of this study is to define the dose-response curve of EXPAREL, an investigational extended-duration formulation of the local anesthetic bupivacaine, on both motor and sensory block when applied in a fixed volume adjacent to the femoral nerve.

Putting local anesthetic next to a nerve is a common way of decreasing the pain that patients feel after surgery. For knee surgery, the local anesthetic is placed next to the femoral nerve in the middle of the crease where the leg meets the body when bending at the hip joint. However, the local anesthetic takes away not only sensation-and therefore pain-but also motor control, leaving muscles weaker. It would greatly improve patient safety if the investigators could administer a very long-acting local anesthetic that decreased postoperative pain, but affected muscle strength only minimally. The purpose of this study is to define the dose-response curve of EXPAREL (also termed SKY0402), an investigational extended-duration formulation of the local anesthetic bupivacaine, on both motor and sensory block when applied in a fixed volume adjacent to the femoral nerve.

This investigation will be a Phase 1, prospective, double-masked, human-subjects dose-response clinical trial.

Enrollment. Subjects will be volunteers of both sexes, age 18 and older. If a volunteer meets inclusion/exclusion criteria and desires study participation, written, informed consent will be obtained. Selection for inclusion will not be based on race or socioeconomic status. The study population of interest includes men and women of all races and socioeconomic status.

Subject preparation. Following written, informed consent, subjects will be admitted to the UCSD CTRI and have demographic/morphometric data recorded (e.g., age, weight, height), a medical history recorded, and a brief physical examination. Block placement itself may occur either in the UCSD CTRI or Hillcrest Outpatient Surgical Center Post Anesthesia Care Unit (PACU: this is where regional anesthetics are administered on a regular basis for surgical patients). If the blocks are placed in the PACU, subjects will remain there for two hours and then moved (accompanied by a physician) on their gurney to the UCSD CTRI where they will spend the remainder of the study period. Women of childbearing potential will have a urine pregnancy test. Prior to dosing, baseline quadriceps strength and sensory level measurements will also be obtained. An intravenous line will be placed in an upper extremity, followed by external monitors (pulse oximeter, blood pressure, and EKG), and oxygen by nasal cannula (1-6 L/min). Sedation will be provided by a combination of one or more of the following, titrated to effect: oral valium (10 mg), intravenous fentanyl (50 mcg), and/or intravenous midazolam (1 mg). Following sterile preparation with chlorhexidine gluconate and isopropyl alcohol, and once the topical antiseptic is dry, subjects will have bilateral, single-injection, ultrasound-guided femoral nerve blocks placed using standard UC San Diego techniques as previously published by the current P.I.

Treatment Group Assignment. The dominant side (left or right) will be randomized to one of two treatment groups: the higher or lower concentration of the local anesthetic SKY0402. The non-dominant contralateral side will receive the other possible treatment. The volume of each and every single-injection femoral nerve block will be 30 mL (standard for femoral nerve blocks is 30-40 mL). Since volume will remain constant, we will vary the dose of SKY0402 by varying concentration (volume x concentration = dose). Of note, SKY0402 may be mixed with normal saline to vary the concentration. Randomization will be based on computer-generated codes. Randomization will be in blocks of two.

Dose Determination. Initial doses of SKY0402 will begin at 0 mg (low: exclusively normal saline as the treatment) and 2 mg (high) for the first subject; and 1 mg (low) and 3 mg (high) for the second subject. Since the volume of each of the bilateral blocks will be 30 mL, the first subject's concentrations will be 0 mg / 30 mL (0%) and 2 mg / 30 mL (0.007%); while the second subject's concentrations will be 1 mg / 30 mL (0.003%) and 3 mg / 30 mL (0.010%). Unmasking of treatments will occur following data collection for each subject to allow determination of dosing for subsequent subjects. The subsequent doses will be between 0-160 mg (0.534%) per side, determined prior to randomization of each subsequent subject. The specific subsequent doses will increase, remain the same, or decrease, determined by the P.I. in consultation with the manufacturer of SKY0402. Since the dose-response for SKY0402 remains unknown for single-injection peripheral nerve blocks, a set "tier" or dose-increase plan prior to experience with each subject is impossible. However, the dose will never be increased by more than 20 mg for each side. In addition, doses will always remain within the range of 0-160 mg (0% - 0.534%) per side, and never exceed a total dose of 160 mg for both sides combined. Dose escalation will be stopped and only lower doses will be administered subsequently, if either of the following occurs after study drug administration: one subject experiences clinically significant motor block persisting more than 7 days or four consecutive subjects experience clinically significant motor block persisting more than 5 days.

For unsuccessful local anesthetic deposition (defined as local anesthetic that could not be deposited immediately adjacent to the femoral nerve as viewed by real-time ultrasound) or subject withdrawal from the study, the subjects' data will not be included in the analysis and the subject dropped from the study (subjects will always receive compensation for at least one night in the CTRI--$400-even if they are discharged the day of block placement due to failed deposition). Remaining subjects will remain within the CTRI and within their bed until discharged home. Discharge will occur when quadriceps strength has returned to at least 80% of its baseline value. We consider a difference of less than 20 percentage points to be clinically relevant because a 10% side-to-side strength difference is common, yet functionally unnoticeable in healthy individuals.4,5 Discharge will occur-at the very earliest-24 hours following initial block placement. Therefore, the amount of time subjects will spend in the CTRI will depend upon the duration of local anesthetic action, which will be variable among subjects and is currently unpredictable without a dose-response curve for peripheral nerve blocks using SKY0402. We anticipate the duration of study participation for each subject to be approximately 72 hours. Subjects may withdraw from participation at any time, and will receive compensation for the time they have participated up until withdrawal (subjects who withdraw the day of block placement will receive $400). However, if the research coordinator who is responsible for distributing the compensation is not available at the time of study withdrawal, then the subject will need to return to the CTRI the following day (or following Monday if withdrawal occurs over the weekend) for compensation.

Outcome Measurements. We have selected measures that have established reliability and validity. Staff blinded to treatment group assignment will perform all measures and assessments. For all measurements, the dominant side will always be tested first, followed by the contralateral side. Measurements will be performed prior to local anesthetic administration initiation ("baseline"; Hour 0); as well as specific post-block time points:

Hour Prior to Hour 0 0 0:10 0:20 0:30 0:40 0:50

1 1:15 1:30 1:45 2 2:30 3 3:30 4 5 6 7 8 9 10 24 (Day #2) 27-36 48 (Day #3) 51-60 72 (Day #4) 75-84 96 (Day #5) 99-108 120 (Day #6)

Primary Outcome Measurement

Quadriceps femoris muscle strength: Evaluated using a portable isometric force dynamometer to measure the maximum voluntary isometric contraction (MVIC) in a seated position with the knee flexed at 90º. This variable will be presented and analyzed as post / pre x 100. For all measurements, subjects will be asked to take 2 sec to come to maximum effort contracting the target muscle(s), maintain this effort for 5 sec, and then relax.

Secondary Outcome Measurement

Sensory level: Evaluated using transcutaneous electrical stimulation (TES) in the same manner as described throughout the anesthesia literature (this is a "gold standard" for regional anesthesia studies). After clipping any hair, EKG pads will be positioned 2 cm medial to the proximal patella and quadriceps tendon and attached to a nerve stimulator. The current will be increased from 0 mA until the subject identifies slight discomfort, at which time the current is recorded as the TES value and the nerve stimulator turned off. The secondary end point will be the post-administration maximum current (absolute values in mA).

Safety Assessments Safety assessments will include monitoring of AEs, SAEs, and vital signs (heart rate, respiratory rate, and blood pressure).

Inclusion Criteria:

• greater than or equal to 18 years old
• able and willing to have bilateral femoral nerve blocks placed and repeated motor/sensory testing for 24-120 hours (1-5 days), requiring 1-5 overnight stay(s) in the UCSD CTRI to allow dissipation of local anesthetic infusion effects to near-baseline values
• have the ability to adequately communicate with all study personnel
• willing and capable of providing written informed consent

Exclusion Criteria:

• daily analgesic use for over one week within the past 6 months
• opioid use within the previous 4 weeks
• any neuro-muscular deficit of either femoral nerves and/or quadriceps muscles
• body mass index > 30 kg/m2
• current pregnancy
• incarceration
• any coagulation disorder
• uncontrolled anxiety, schizophrenia, or other psychiatric disorder that, in the opinion of the investigators, may interfere with study assessments or adherence
• any previous allergic reaction to fentanyl, midazolam, or an amide local anesthetic (bupivacaine is of the amide local anesthetic class)
• any previous participation in a SKY0402/EXPAREL study
• nursing mothers
• suspected or known drug or alcohol abuse within the previous year; and/or
• planning on becoming pregnant in the one month following study participation.

Investigational Drug Service prepared all study medication, with each dose provided to the caregivers in a 30-mL syringe with IV line extension tubing, both wrapped in opaque tape to retain masking since the relative study drug concentration could be inferred by the opacity of the injectate. One syringe was labeled "Dominant" and the other labeled "Other." In this manner, all investigators, nursing staff, and subjects remained masked to the specific concentration/dose during all outcome measurements.