untitled h1
9.0 drug information 9.1 esk981

Other names for the drug: formerly known as CEP-11981  
Description: ESK981 drug product is an off-white to orange opaque, hard gelatin capsule containing 1 mg, 5 mg, 20 mg or 40 mg (free base equivalents) of ESK981 monotosylate. The drug product is packaged in a high-density polyethylene (HDPE) bottle with a child resistant closure and an inductionsealed inner seal. The formulation also contains lactose monohydrate.  
Classification - type of agent: Small molecular inhibitor of multiple tyrosine kinases such as vascular endothelial growth factor receptor 1 (VEGFR-1), VEGFR-2, and Tie-2, (IC50 of 3, 4, 22nM,
respectively)
Pharmacokinetics:
Note: CEP-11981 refers to the development name for ESK981. The pharmacokinetic, ADME, and toxicokinetic characteristics of CEP-11981 have been extensively studied in Sprague Dawley rats and cynomolgus monkeys after intravenous and oral doses and preliminarily in Balb/c nude mice after oral doses. In order to facilitate comparisons between studies and species, all doses are presented in this summary as milligrams of CEP-11981 free base per kilogram of body weight, regardless of the form administered, unless specifically noted otherwise. The intravenous dose in rats was administered in 3% dimethylsulfoxide, 30% Solutol®, and 67% phosphate-buffered saline at a volume of 1 mL/kg. In monkeys, the intravenous vehicle was 47.4% polyethylene glycol (PEG) 400, 31.1% water, 20.7% hydroxy-β-cyclodextrin (HPBCD), and 0.8% Pluronic® F-68 at a volume of 0.5 mL/kg. Pharmacokinetic parameters obtained after single intravenous
(bolus) administration of CEP-11981 are summarized in Table 3.  
In rats, the systemic clearance is low relative to the estimated total hepatic plasma flow; the volume of distribution is approximately that of total body water. There are apparent sex-related differences in the pharmacokinetics of CEP-11981 in rats, with lower clearance and longer half-life in female rats than in male rats. The volume of distribution is numerically less in female rats, but the significance of this difference is doubtful. In monkeys, the half-life is similar to that in rats, but the systemic clearance is substantially larger than that in rats, as is the volume of distribution. The difference in clearance is consistent with the higher rate of metabolism of CEP-11981 in incubations with monkey liver microsomes than with rat liver microsomes. The origin of the larger volume of distribution in monkeys is not known. Pharmacokinetic parameters obtained after single, low-dose oral administration of CEP-11981 are summarized in Table 4.  
| Table 4. Pharmacokinetics After Single Oral Administration of CEP-11981   |
|---------------------------------------------------------------------------|  
Dose administration in the mice was as a suspension of the free base in 0.6% methylcellulose:Tween 80 (99.5:0.5) at a volume of 25 mL/kg. In the rats and monkeys, the dose was administered as a suspension of the tosylate salt in Ora-Plus oral suspending vehicle at volumes of 2 and 5 mL/kg, respectively.  
Significant oral bioavailability was obtained in both rats and monkeys. The lack of an intravenous comparator dose in mice precluded calculation of the absolute bioavailability in that species, but the similarity in systemic exposure in mice relative to rats would suggest that oral bioavailability is comparable. The effect of fed versus fasted state on the pharmacokinetics of CEP-11981 was assessed in a preliminary study in rats. Cmax and AUC0-6 were minimally affected by the fed/fasted state of the animals, but the tmax was shifted from 2.7 to 5.3 hours post dose when the animals had been fed before dosing. At higher oral doses of CEP-11981, there was evidence of dose-dependent pharmacokinetics in all 3 species. In mice, single-dose systemic exposure increased in a slightly more than dose-proportional manner between 1 and 10 mg/kg, but did not show consistent further increase at 20 or 30 mg/kg. After 23 daily doses, the dose response and concentrations were similar to those after single doses at 1, 3, or 10 mg/kg, but were approximately 2 times higher in the 20- and 30-mg/kg groups on day 23 versus day 1 of dosing. Systemic exposure after multiple daily doses of 20 and 30 mg/kg was dose-related, although not dose-proportional, relative to that at 10 mg/kg. In male rats, Cmax was approximately dose proportional between 5 and 15 mg/kg, but was less than dose proportional between 15 and 40 mg/kg. The increase in AUC0-∞ was greater than dose-proportional between 5 and 15 mg/kg, but was approximately dose proportional between 15 and 40 mg/kg.  
In monkeys, increases in both Cmax and AUC0-∞ were greater than dose proportional between 3 and 13.5 mg/kg. Similar (i.e., greater-than-dose-proportional) increases in systemic exposure were also noted in a preliminary study that tested single oral doses as high as 100 mg/kg in monkeys.  
Due to changes in the form of CEP-11981 that was obtained from successive synthetic campaigns, a series of single-dose oral studies were conducted in rats and monkeys to which different forms of the compound and/or different dosage regimens were administered. The outcome of those studies indicated that milled CEP-11981 tosylate, administered as a suspension in aqueous methylcellulose, provides systemic exposures that are in the targeted ranges in both species, and it is that form with which the bulk of the nonclinical toxicity testing was done and which will be the form of the compound used in early clinical testing.  
The tissue distribution of [14C]-CEP-11981 was studied in male albino (Sprague Dawley) and pigmented
(Lister hooded) rats after single doses orally at 10 mg/kg and intravenously at 1 mg/kg, respectively, as the tosylate salt. Radioactivity was broadly distributed, with highest concentrations (exclusive of the gastrointestinal tract) in the liver, kidney, and adrenal gland.  
Minimal radioactivity was detected in barriered tissues, i.e., brain and testis, but there was apparent association with melanin-containing structures such as uveal tract and pigmented skin in the pigmented animals. The bulk of the radioactivity was removed from the tissues during the 168- and 24-hour test periods in the albino and pigmented animals, respectively.  
CEP-11981 is extensively metabolized in vitro by liver microsomes from male Swiss CD1 mice, male Sprague Dawley rats, male New Zealand rabbits, male beagle dogs, male cynomolgus monkeys, and humans. The relative order of loss of parent compound during 20 minutes of incubation was: monkeys
> humans > mice ≈ dogs > rats ≈ rabbits.  
Incubations with individual recombinant human CYP enzymes suggested that 3 enzymes, i.e., CYP1A2, CYP2C8, and CYP3A4, may play significant roles in the metabolic elimination of CEP-11981. Other CYP
enzymes that were also capable of metabolizing the compound in vitro are either minimally expressed in adult human liver (i.e., CYP1A1 and CYP3A7) or are polymorphic, being expressed in a minority of individuals (i.e., CYP3A5). The results of experiments assessing the effect of CYP-specific inhibitors on loss of CEP-11981 during incubation with human liver microsomes also identified CYP3A enzymes as playing a key role in metabolic elimination of the compound. After oral administration of [14C]-CEP-11981, radioactivity was predominantly recovered in the feces in both rats and monkeys, with less than 1% of the radiochemical dose recovered in urine of either species. Metabolic profiling was hence conducted in rat bile, in which parent [14C]-CEP-11981 was the most prominent radioactive component, especially in the female rat. The most prominent metabolite in both sexes, but especially in the male, was a monohydroxylated derivative. Overall, the reactions previously observed in vitro, i.e., ring hydroxylation, N-demethylation, and reduction (didehydrogenation) accounted for all of the CEP-11981-derived compounds detected in vivo. The potential for CEP-11981 to affect the metabolic elimination of co-administered drugs was tested in vitro through examination of its capacity for induction of CYP enzymes activities in primary human hepatocyte cultures and for inhibition of CYP activities in human hepatic microsomal preparations. The levels of mRNA for CYP1A1, CYP1A2, CYP2C9, and CYP3A4 were measured after incubation of CEP-11981 with hepatocytes for 30 hours at concentrations of 1 to 30 μM. No indication of induction was obtained, but suppression of CYP2C9 and CYP3A4 message was observed at the highest concentration. However, due to apparent cellular toxicity at that concentration, this result was likely an artifact.  
CEP-11981 did not inhibit the activities of CYP2B6, CYP2C9, CYP2C19, CYP2D6, or CYP4A9/11. Marginal inhibition of CYP1A2 was observed, but with an estimated Ki that is higher than exposures expected clinically. CYP3A4/5 activity was inhibited with a Ki of 2.6 and 2.2 μM for the 2 reactions examined, i.e., testosterone 6β-hydroxylation and midazolam 1′-hydroxylation, respectively. Much of the inhibition was reversible and competitive in nature, with a metabolism-dependent component, but at least some of the inhibition was irreversible for both reactions tested.  
In addition to the pharmacokinetic and ADME studies, bioanalytical and pharmacokinetic support has been provided to the toxicokinetics portions of nonclinical safety studies in rats and monkeys.  
The longest nonclinical safety study conducted to date in rats was a 4-week oral toxicity study at daily doses of 0 (vehicle only), 5, 10, 20 (lowered to 15 on day 8), and 40 mg/kg. CEP-11981 was administered as suspensions of its tosylate salt in Ora-Plus oral suspending vehicle. Key toxicokinetic parameters from the study are tabulated in Table 5.  
| Table 5. Toxicokinetics Results for the 4-Week Oral Toxicity Study in Rats   |                         |           |            |           |
|------------------------------------------------------------------------------|-------------------------|-----------|------------|-----------|
| Day 1/Day 28                                                                 |                         |           |            |           |
| 20 → 15 mg/kg                                                                |                         |           |            |           |
| Parameter a                                                                  | 5 mg/kg                 | 10 mg/kg  | 40 mg/kg   |           |
| Males (day 1/day 28)                                                         |                         |           |            |           |
| Czuz (Ug/mL)                                                                 | 0.94/1.03               | 1.70/2.20 | 3.45/NAV ® | 5.28/NAV  |
| 4/1                                                                          | 2/4                     | 2/NAV     | 4/NAV      |           |
| trear (hr)                                                                   |                         |           |            |           |
| Half-life (hr)                                                               | NC 6 /3.6               | 5.7/3.2   | 4.0/NAV    | 4.2/NAV   |
| AUC ~ or AUC ~                                                               | 5.34 (AUCo.)/9.47       | 17.7/27.1 | 42.4/NAV   | 73.6/NAV  |
| (IIg-hr/mL) a                                                                |                         |           |            |           |
| Females (day 1/day 28)                                                       | 1.17/1.73               | 2.69/2.72 | 5.19/NAV   | 8.21/NAV  |
| Cmx (Ug/mL)                                                                  |                         |           |            |           |
| tensy (hr)                                                                   | 4/2                     | 4/2       | 2/NAV      | 2/NAV     |
| Half-life (hr)                                                               | 2.9/3.2                 | 4.6/NAV   |            |           |
| 2.6/5.7                                                                      | 3.7/NAV                 |           |            |           |
| AUCo... or AUCo...                                                           | 12.5/15.8               | 28.5/37.2 | 70.6/      | 118.7/NAV |
| (ug·hr/mL) 9                                                                 |                         |           |            |           |
| a Parameters were generated from composite mean plasma concer                | ration-versus-time data |           |            |           |
| 6 NAV=not available (insufficient data to estimate parameter)                |                         |           |            |           |
| ' NC=not calculable                                                          |                         |           |            |           |  
4 AUCo... for day 1 and AUCo.. for day 28.  
Czux=maximum observed plasma drug concentration; teax=time to maximum observed plasma drug concentration; AUCow=area under the plasma drug concentration by time curve from time zero to infinity; AUC ox =area under the plasma drug concentration versus time curve from time zero to the time of the last measurable plasma drug concentration; AUCo-r=AUC from time zero through 1 dosing interval.  
As noted in the pharmacokinetic studies, the systemic exposures were higher in female rats than in male rats and, in groups having both single-dose and multiple-dose datasets, were higher on day 28 than on day 1. Values of C max and AUC were dose-related over the dose range tested, but increases in AUC
were more than dose-proportional over portions of the dose range, suggesting that 1 or more of the pharmacokinetic characteristics of CEP-11981 are dose-dependent.  
Two 4-week oral toxicity studies of CEP-11981 have been conducted in monkeys. In the first study (study DS-2006-020), daily doses of 0 (vehicle only), 5, 10, and 25 mg/kg were administered as suspensions of the free base in Ora-Plus oral suspending vehicle. Due to a subsequent change in the form of the compound to its tosylate salt, a second 4-week study (study DS-2006-042) was conducted testing daily doses of 0 (vehicle only) and 3 mg/kg. Key toxicokinetic parameters from the study are tabulated in Table 6.  
| Table 6. Toxicokinetics Results for the 4-Week Oral Toxicity Studies in Monkeys                                   |                                        |                                                                       |                     |                     |
|-------------------------------------------------------------------------------------------------------------------|----------------------------------------|-----------------------------------------------------------------------|---------------------|---------------------|
| Day 1/Day 28                                                                                                      |                                        |                                                                       |                     |                     |
| Parameter 2                                                                                                       | 5 mg/kg                                | 10 mg/kg                                                              | 25 mg/kg            | 3 mg/kg             |
| Males (day 1/day 28)                                                                                              |                                        |                                                                       |                     |                     |
| C EE                                                                                                              | 0.061±0.026/0.093±0.058                | 0.53±0.33/0.67±0.23                                                   | 0.70±0.27/0.11 c e  | 0.24±0.12/0.20±0.09 |
| (//g/mL)                                                                                                          |                                        |                                                                       |                     |                     |
| trens                                                                                                             | 4 (all 4)/3 (2-6)]                     | 4 (4-6)/2 (2-4)                                                       | 4 (2-6)/8 cre       | 4 (2-4)/2 (2-4)     |
| (hr)                                                                                                              |                                        |                                                                       |                     |                     |
| Half-life                                                                                                         | 1.4 (n=1)/1.9                          | 2.1/2.0                                                               | 2.3/NC 2            | 2.2/2.0             |
| (hr)                                                                                                              |                                        |                                                                       |                     |                     |
| AUCo= or AUC                                                                                                      | 0.24±0.13 (AUC يو)/                    | 3.65±2.31/3.13±1.58                                                   | 5.23=2.24/1.32 c    | 1.30±0.71/1.19±0.39 |
| 0.86±0.18                                                                                                         |                                        |                                                                       |                     |                     |
| (..g-hr/mL) 2                                                                                                     |                                        |                                                                       |                     |                     |
| Females (day 1/day 28)                                                                                            |                                        |                                                                       |                     |                     |
| CEE                                                                                                               | 0.12±0.09/0.06±0.04                    | 0.44±0.27/0.43±0.32     0.59±0.40/0.46±0.28     0.20±0.09'0.14 ± 0.05 |                     |                     |
| (/ig/mL)                                                                                                          |                                        |                                                                       |                     |                     |
| tees                                                                                                              | 2 (2-4)/3 (2-4)                        | 4 (4-6)/3 (2-4)                                                       | 5 (4-6)/2 (all 2)*  | 2 (2-4)/2 (2-4)     |
| (hr)                                                                                                              |                                        |                                                                       |                     |                     |
| Half-life                                                                                                         | 1.6/1.8                                | 1.6/1.6                                                               | 1.9/1.6 8           | 2.0/2.2             |
| (fir)                                                                                                             |                                        |                                                                       |                     |                     |
| AUCom or AUC                                                                                                      | 0.86±0.18/0.27±0.23                    | 3.13±1.58/2.5±1.99                                                    | 3.87±2.28/2.92±2.73 | 1.19±0.39/0.83±0.31 |
| (ug·br/mL)                                                                                                        |                                        |                                                                       |                     |                     |
| * Parameters were generated from composite                                                                        | psite mean plasma concentration-versus |                                                                       |                     |                     |
| b Values are mean#standard deviation, except tear, which is median (range), and half-life, which is harmonic mean |                                        |                                                                       |                     |                     |
| c n=1 due to mortality in the group.                                                                              |                                        |                                                                       |                     |                     |
| d NC = not calculable.                                                                                            |                                        |                                                                       |                     |                     |
| * Sampled on day 14 or 15 of dose administration.                                                                 |                                        |                                                                       |                     |                     |
| 1 AUC 0... for day 1 and AUC 0.. for day 28.                                                                      |                                        |                                                                       |                     |                     |  
C mx maximum observed plasma drug concentration; t mx -time to maximum observed plasma drug concentration; AUCo==area under the plasma drug concentration by time curve from time zero to infinity; AUCo==area under the plasma drug concentration versus time curve from time zero to the time of the last measurable plasma drug concentration; AUC r =AUC from time zero through 1 dosing interval.  
The monkeys did not show the sex-related differences in systemic exposure that were evident in the rats, but there was a suggestion, as in the pharmacokinetic studies, that some aspect(s) of the pharmacokinetics of the compound might be dose-dependent, especially at higher doses.  
Overall, the pharmacokinetic and ADME properties of CEP-11981 have been studied in vitro using blood and tissue fractions from mice, rats, rabbits, dogs, monkeys, and humans and in vivo in mice, rats, and monkeys. The estimated half-life in rats and monkeys after an intravenous dose is approximately 2 to 3 hours. Despite its low aqueous solubility, CEP-11981 is generally well absorbed in mice, rats, and monkeys at dose levels associated with pharmacologic responses in preclinical disease models. At higher doses, evidence of dose-dependent pharmacokinetics has been obtained in all 3 species. After an oral or intravenous dose, [14C]-CEP-11981-derived radioactivity is broadly distributed into tissues, generally at concentrations higher than those in blood. Only in barriered tissues, e.g., brain and testis, are concentrations lower than those in blood. The compound is extensively metabolized, but is also excreted in rats via the bile as the parent drug. Excretion is predominantly in the feces in both rats and monkeys.  
.  
Drug Interactions: Prohibited before enrollment and during administration of study treatment
Concomitant systemic treatments for CRPC (other than a GnRH agonist/antagonist) are prohibited including: CYP-17 inhibitors (e.g. ketoconazole, abiraterone), antiandrogens (e.g. bicalutamide, nilutamide), second generation antiandrogens (e.g. enzalutamide, ARN-509, Galeterone),
immunotherapy (e.g. sipuleucel-T, ipilimumab) and chemotherapy (e.g. docetaxel, cabazitaxel).  
.  
Storage and stability: ESK981 capsules should be stored refrigerated at 2°C to 8°C (36°F to 46°F).  
The stability of the drug substance and drug product continues to be monitored.
Preparation and Dispensing:
ESK981 is supplied in 28 count HDPE bottles.  
Supply and packaging ESK981 monotosylate is a fully synthetic drug substance containing a core indolocarbazolone ring system. ESK981 monotosylate drug substance is a yellow to orange powder that has a molecular weight of 649.77 g/mol. The drug substance contains 73.5% theoretical weight percent of the active moiety ESK981 (free base).  
ESK981 drug product is an off-white to orange opaque, hard gelatin capsule containing 40 mg (free base equivalents) of ESK981 monotosylate. The drug product is packaged in a high-density polyethylene (HDPE) bottle with a child resistant closure and an induction-sealed inner seal. The formulation also contains lactose monohydrate.  
Administration:
Treatment will be administered on an outpatient basis. Study drug will be affixed with a clinical label in accordance with regulatory requirements. This trial is open-label; therefore, the subject, the study team, the Sponsor and/or designee are not blinded. Treatment (name, strength or potency) is included in the label text; Subjects will be given a one-month (28 day) supply of study drug. Subjects will be instructed to take 4 capsules per, with or without food, day for 5 consecutive calendar days, then take a drug holiday for 2 consecutive days before repeating the 5 days on-2 days off cycle in sets of 4 weeks or 28 calendar days. Subjects will be asked to keep a pill diary noting the date they take their study drug. They will be asked to bring their pill diary to each study visit along with all used and unused study drug containers.  
Availability:
Provided by Esanik Therapeutics, Inc.  
Under no circumstance will the study medication *ESK981* be used other than as directed by the protocol.  
Return and Retention of Study Drug:
The investigator is responsible for keeping accurate records of the study drug received from the Sponsor or designee, the amount dispensed to and returned by the subjects and the amount remaining at the conclusion of the trial.  
A member of the study team will review the pill dairy during each visit study drug is returned and discuss compliance or other concerns the subject may have. The investigator or designee will provide appropriate documentation that must be completed for drug accountability and return, or local discard and destruction if appropriate.  
Where local discard and destruction is appropriate, the investigator is responsible for ensuring that a local discard/destruction procedure is documented.  
Drug Accountability:
The investigator, or a responsible party designated by the investigator, must maintain a careful record of the inventory and disposition of the investigational drug ESKI981. The drug accountability records will capture drug receipt, drug dispensing, drug return and final disposition.
untitled h1
9.2 nivolumab

Other names for the drug: Opdivo Description: Injection: 40 mg/4 mL and 100 mg/10 mL solution in a single-dose vial.  
Classification - type of agent: immunomodulatory; checkpoint inhibitor Mode of action:
Binding of the PD-1 ligands, PD-L1 and PD-L2, to the PD-1 receptor found on T cells, inhibits Tcell proliferation and cytokine production. Upregulation of PD-1 ligands occurs in some tumors and signaling through this pathway can contribute to inhibition of active T-cell immune surveillance of tumors. Nivolumab is a human immunoglobulin G4 (IgG4) monoclonal antibody that binds to the PD-1 receptor and blocks its interaction with PD-L1 and PD-L2, releasing PD-1 pathway-mediated inhibition of the immune response, including the anti-tumor immune response. In syngeneic mouse tumor models, blocking PD-1 activity resulted in decreased tumor growth.  
Pharmacokinetics:
Nivolumab pharmacokinetics (PK) was assessed using a population PK approach for both single-agent OPDIVO and OPDIVO with ipilimumab.  
OPDIVO as a single agent: The PK of single-agent nivolumab was studied in patients over a dose range of 0.1 to 20 mg/kg administered as a single dose or as multiple doses of OPDIVO every 2 or 3 weeks. Nivolumab clearance decreases over time, with a mean maximal reduction
(% coefficient of variation [CV%]) from baseline values of approximately 24.5% (47.6%) resulting in a geometric mean steady state clearance (CLss) (CV%) of 8.2 mL/h (53.9%); the decrease in CLss is not considered clinically relevant. The geometric mean volume of distribution at steady state (Vss) (CV%) is 6.8 L (27.3%), and geometric mean elimination half-life (t1/2) is 25 days (77.5%). Steady-state concentrations of nivolumab were reached by approximately 12 weeks when administered at 3 mg/kg every 2 weeks, and systemic accumulation was approximately 3.7-fold. The exposure to nivolumab increased dose proportionally over the dose range of 0.1 to 10 mg/kg administered every 2 weeks.  
Side effects:
OPDIVO as a single agent: fatigue, rash, musculoskeletal pain, pruritus, diarrhea, nausea, asthenia, cough, dyspnea, constipation, decreased appetite, back pain, arthralgia, upper respiratory tract infection, and pyrexia.  
Drug Interactions:
No formal pharmacokinetic drug-drug interaction studies have been conducted with OPDIVO.  
Storage and stability:
The product does not contain a preservative. After preparation, store the OPDIVO infusion either:
- at room temperature for no more than 8 hours from the time of preparation. This includes room temperature storage of the infusion in the IV container and time for administration of the infusion or - under refrigeration at 2°C to 8°C (36°F to 46°F) for no more than 24 hours from the time of
untitled h1
infusion preparation. do not freeze.

Preparation and Dispensing:
Visually inspect drug product solution for particulate matter and discoloration prior to administration. OPDIVO is a clear to opalescent, colorless to pale-yellow solution. Discard the vial if the solution is cloudy, discolored, or contains extraneous particulate matter other than a few translucent-to-white, proteinaceous particles. Do not shake the vial. - Withdraw the required volume of OPDIVO and transfer into an intravenous container. - Dilute OPDIVO with either 0.9% Sodium Chloride Injection, USP or 5% Dextrose Injection, USP to prepare an infusion with a final concentration ranging from 1 mg/mL to 10 mg/mL.  
- Mix diluted solution by gentle inversion. Do not shake. - Discard partially used vials or empty vials of OPDIVO.  
Administration:
Administer the infusion over 30-60 minutes through an intravenous line containing a sterile, nonpyrogenic, low protein binding in-line filter (pore size of 0.2 micrometer to 1.2 micrometer). Do not coadminister other drugs through the same intravenous line. Flush the intravenous line at end of infusion.  
Availability: Commercially available Return and Retention of Study Drug:
- Discard partially used vials or empty vials of OPDIVO.  
Drug Accountability:
The investigator, or a responsible party designated by the investigator, must maintain a careful record of the inventory and disposition of the drug Nivolumab. The drug accountability records will capture drug receipt, drug dispensing, drug return and final disposition.
untitled h1
10.0 special studies

10.1 **Sample Collection Guidelines**
Refer to Section 6.0 for tumor tissue and blood collection time points and Lab Manual for collection and processing details.  
10.2 **Specimen Banking**
Patient blood and tissue samples collected for this study will be retained at the University of Michigan. Specimens will be stored indefinitely or until they are used up. If future use is denied or withdrawn by the patient, best efforts will be made to stop any additional studies and to destroy the specimens.  
Specimen being stored long-term for potential use or not outlined in the protocol will be stored and used in compliance with the University of Michigan policy University Policy Governing Tissue Sample Collection, Ownership, Usage, and Disposition within all UMMS Research Repositories -
https://research.medicine.umich.edu/office-research/biorepository/governance-policies.  
11.0 **STATISTICAL CONSIDERATIONS**
The primary objective of the study is to determine the efficacy of ESK981 in combination with nivolumab therapy in patients with metastatic renal cell carcinoma who may have progressed despite treatment with one VEGF-TKI. Response rate, defined as the sum of complete + partial response (CR+PR), as measured by RECIST v1.1 in Cohort B (combined therapy with ESK981 and Nivolumab) will be the primary statistical endpoint.  
11.1 **Study Design/Study Endpoints**
A two-stage Mini-max Simon design will be used to minimize the number of patients to be treated with the combination therapy (Cohort B). We assume that this combination will not be of further interest if the true response rate were less than 0.25 and assume that a true response rate of at least 0.45 would be of clinical interest. This phase II study design permits early termination of patient entry due to futility of efficacy after the first 17 objective response evaluable patients have been accrued to Cohort B if there are 4 or less responses. If the study passes the first phase, then a second stage of 19 more objective response evaluable patients will be accrued for a total of 36 objective response evaluable patients. If 14 or more patients out of 36 have a response, then the treatment combination will be deemed successful. This trial design has 80% power to detect a response rate of 45% compared to a null rate of 25% assuming a 5% type I error. The probability of early termination with this design is 0.57 if the true response rate is 25%.  
A single stage design will be used to determine efficacy of monotherapy treatment (Cohort A) with ESK981 efficacy in patients with metastatic renal cell carcinoma will be assessed as the RECIST v1.1 response rate. Assuming a response rate of clinical interest with monotherapy is 25% and null response proportion of 5% with a 1-sided type I error=0.102 there is 80% power with 11 patients to detect a response rate of 25%. Additionally, a futility analysis assuming a response rate of 25%
would have 5% probability of 0 responses out of 11 patients. If 3 or more patients have a response with monotherapy ESK981, then the monotherapy treatment will be deemed of interest in a future study. Secondary endpoints of the trial include assessment of toxicity in all patients who receive any study treatment. We will evaluate overall survival, progression free survival, duration of therapy, duration of response and QoL.  
11.2 **Sample Size and Accrual**
There are 2 cohorts for accrual with Cohort A being accrued first. Cohort B will follow and will have 2 stages of accrual. Cohort A will accrue 11 patients to monotherapy treatment. Then Cohort B will begin accrual of 17 patients to the first stage and if the interim analysis permits, the second stage will accrue an additional 19 patients. Our final sample size if all stages accrue will include 47 total objective response evaluable patients (11+17+19). We expect to accrue 1-2 patients per month for a total accrual period of 24 - 30 months.  
11.3 **Early Stopping Due to Toxicity**
Toxicity will be assessed in each cohort. If the DLT proportion is >30% during cycle 1 in Cohort A then dosing as is planned for the combination cohort will require modification before Cohort B can begin. Accrual of Cohort B will not begin until toxicity assessment of Cohort A is complete. Cohort B will assess DLT proportions in the first 6 patients. If 2 or more DLTs occur in the first 6 patients of Cohort B, then dosing in the combination treatment in Cohort B will be decreased one dose level. If a decreased dose level is opened, assessment of another 6 patients will be assess for DLTs; if 2 or more DLTs occur then the trial will be halted. Accrual will be held during toxicity evaluation.  
11.4 **Data Analyses Plans**
The primary analysis of efficacy will report the number of patients with an objective response and the associated proportion with the corresponding 95% exact binomial confidence interval separately in each cohort. In Cohort B, if stage 2 is initiated then the efficacy analysis methods will be adjusted for the interim check as described by Koyama and Chen [36]. Toxicity will be described by grade, attribution, and body system using counts and proportions. Toxicity will be described separately for each cohort.  
Overall survival, progression free survival, duration of therapy, and duration of response will be displayed using a Kaplan-Meier figure and the associated median (if reached), one-year and two-year estimates with 95% confidence intervals using Greenwood's formula will be reported if there is censoring. If there is no censoring in the endpoint, then the standard mean (or median) and 95% confidence interval (or 5th and 95th percentiles) will be reported. Each endpoint will be reported separately by cohort.  
Health-related quality of life assessment changes from baseline using EuroQoL-5D will be described using descriptive statistics.  
12.0 **DATA MANAGEMENT**
All information will be recorded locally and entered into Case Report Forms (CRFs) on the web-based electronic data capture (EDC) system of the University of Michigan. Online access will be provided to each site by the Coordinating Center. CRFs will be reviewed and source verified by the MSC during annual monitoring visits and prior to and between visits. Discrepant, unusual and incomplete data will be queried by the MSC. The investigator or study coordinator will be responsible for providing resolutions to the data queries, as appropriate. The investigator must ensure that all data queries are dealt with promptly.  
The data submission schedule is as follows:
At the time of registration o Subject entry into the EDC
Subject Status Demographics During study participation o All data should be entered online within 10 business days of data acquisition. *[Information on* dose limiting toxicity events must be entered within one business day.] Information on Serious Adverse Events must be entered within the reporting timeframe specified in Section 8 of the protocol.  
All study information should be recorded in an appropriate source document (e.g. clinic chart).  
13.0 **DATA AND SAFETY MONITORING**
This study will be monitored in accordance with the NCI approved University of Michigan Rogel Cancer Center Data and Safety Monitoring Plan, with oversight by the Rogel Cancer Center Data and Safety Monitoring Committee (DSMC)..  
The Sponsor-Investigator (S-I)/Study Principal Investigator will provide ongoing monitoring of data and patient safety in this trial and conduct regular data review with participating sites.The Sponsor-Investigator
(S-I)/Study Principal Investigator and/or the Project Manager/Delegate will review data and patient safety issues with participating sites per a defined **quarterly** meeting cadence. Depending on the protocol activity, the meeting cadence may be more frequent. This data review meeting may be achieved via a teleconference or another similar mechanism to discuss matters related to:
Enrollment rate relative to expectations, characteristics of participants Safety of study participants (Serious Adverse Event & Adverse Event reporting) Adherence to protocol (protocol deviations) Completeness, validity and integrity of study data
untitled h1
retention of study participants

Participating sites are required to ensure all pertinent data for the review period are available in the database at the time of the discussion.  
Participating sites unable to participate in the data review meeting are required to provide written confirmation that their site has reviewed the relevant data and patient safety issues for the review period and their site's data are in alignment with the data reported in the database. Written confirmation is to be provided to the Project Manager/Delegate within the timeline requested to retain compliance with monitoring timelines.  
Documentation of the teleconference or alternate mechanism utilized to review items above is to be retained in the Trial Master File.  
The Project Manager/Delegate is responsible for collating the data from all participating sites and completing the Protocol Specific Data and Safety Monitoring Report (DSMR) form to document the data review meeting discussion.  
The DSMR will be signed by the Sponsor-Investigator (S-I)/Study Principal Investigator or designated CoInvestigator and submitted to the DSMC on a **quarterly** basis for independent review.  
14.0 **QUALITY ASSURANCE AND AUDITS**
The DSMC can request a 'for cause' quality assurance audit of the trial if the committee identifies a need for a more rigorous evaluation of study-related issues. A regulatory authority (e.g. FDA) may also wish to conduct an inspection of the study, during its conduct or even after its completion. If an inspection has been requested by a regulatory authority, the site investigator must immediately inform the Coordinating Center that such a request has been made.
untitled h1
15.0 clinical monitoring procedures

Clinical studies coordinated by University of Michigan Rogel Cancer Center must be conducted in accordance with the ethical principles that are consistent with Good Clinical Practices (GCP) and in compliance with other applicable regulatory requirements.  
This study will be monitored by a representative of the Coordinating Center of the University of Michigan Rogel Cancer Center. Monitoring visits will be made during the conduct of the study and at study close-out. Prior to subject recruitment, a participating site will undergo site initiation meeting to be conducted by the Coordinating Center. This will be done as an actual site visit; teleconference, videoconference, or web-based meeting after the site has been given access to the study database and assembled a study reference binder.  
The site's principal investigator and his study staff should make every effort in attending the site initiation meeting. Study–related questions or issues identified during the site initiation meeting will be followed-up by the appropriate University of Michigan Rogel Cancer Center personnel until they have been answered and resolved.  
Monitoring of this study will include both 'Centralized Monitoring', the review of source documents at the Coordinating Center and 'On-site Monitoring', an actual site visit. The first 'Centralized' visit should occur after the first subject enrolled completes first treatment cycle. The study site will send the de-identified source documents to the Coordinating Center for monitoring. 'Centralized' monitoring may be requested by the Coordinating Center if an amendment requires changes to the protocol procedures. The site will send in pertinent de-identified source documents, as defined by the Coordinating Center for monitoring.  
The first annual 'On-site' monitoring visit should occur after the first five study participants are enrolled or twelve months after a study opens, whichever occurs first. The annual visit may be conducted as a 'Centralized' visit if less than three subjects have enrolled at the study site. The type of visit is at the discretion of the Coordinating Center. At a minimum, a routine monitoring visit will be done at least once a year, or once during the course of the study if the study duration is less than 12 months. The purpose of these visits is to verify:
Adherence to the protocol Completeness and accuracy of study data and samples collected Proper storage, dispensing and inventory of study medication Compliance with regulations During a monitoring visit to a site, access to relevant hospital and clinical records must be given by the site investigator to the Coordinating Center representative conducting the monitoring visit to verify consistency of data collected on the CRFs with the original source data. While most patient cases will be selected from patients accrued since the previous monitoring visit, any patient case has the potential for review. At least one or more unannounced cases will be reviewed, if the total accruals warrant selection of unannounced cases. The Coordinating Center expects the relevant investigational staff to be available to facilitate the conduct of the visit, that source documents are available at the time of the visit, and that a suitable environment will be provided for review of study-related documents. Any issues identified during these visits will be communicated to the site and are expected to be resolved by the site in a timely manner. For review of study-related documents at the Coordinating Center, the site will be required to ship or fax documents to be reviewed.  
Participating site will also undergo a site close-out upon completion, termination or cancellation of a study to ensure fulfillment of study obligations during the conduct of the study, and that the site Investigator is aware of his/her ongoing responsibilities. In general, a site close-out is conducted during a site visit; however, site close-out can occur without a site visit.  
16.0 **REFERENCES**
1. Yang, Z.J., et al., *The role of autophagy in cancer: therapeutic implications.* Mol Cancer Ther, 2011. 10(9): p. 1533-41.  
2. Nelson, E.C., C.P. Evans, and P.N. Lara, Jr., Renal cell carcinoma: current status and emerging therapies. Cancer Treat Rev, 2007. 33(3): p. 299-313.  
3. Coppin, C., et al., *Targeted therapy for advanced renal cell carcinoma.* Cochrane Database Syst Rev, 2008(2): p. CD006017.  
4. Rini, B.I. and E.J. Small, *Biology and clinical development of vascular endothelial growth factortargeted therapy in renal cell carcinoma.* Journal of Clinical Oncology, 2005. 23(5): p. 1028-1043.  
5. Joosten, S.C., et al., *Resistance to sunitinib in renal cell carcinoma: From molecular mechanisms to* predictive markers and future perspectives. Biochim Biophys Acta, 2015. **1855**(1): p. 1-16.  
6. Dvorak, H.F., et al., Vascular permeability factor/vascular endothelial growth factor, microvascular hyperpermeability, and angiogenesis. Am J Pathol, 1995. 146(5): p. 1029-39.  
7. Benjamin, L.E., et al., *Selective ablation of immature blood vessels in established human tumors* follows vascular endothelial growth factor withdrawal. J Clin Invest, 1999. 103(2): p. 159-65.  
8. Watanabe, Y., et al., Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF)
delays and induces escape from senescence in human dermal microvascular endothelial cells.  
Oncogene, 1997. 14(17): p. 2025-32.  
9. Rini, B.I. and E.J. Small, *Biology and clinical development of vascular endothelial growth factortargeted therapy in renal cell carcinoma.* J Clin Oncol, 2005. 23(5): p. 1028-43.  
10. Hanahan, D. and R.A. Weinberg, *The hallmarks of cancer.* Cell, 2000. 100(1): p. 57-70.  
11. Ferrara, N., *The role of vascular endothelial growth factor in pathological angiogenesis.* Breast Cancer Res Treat, 1995. 36(2): p. 127-37.  
12. Rini, B.I., et al., Axitinib with or without dose titration for first-line metastatic renal-cell carcinoma: a randomised double-blind phase 2 trial. Lancet Oncology, 2013. 14(12): p. 12331242.  
13. Sternberg, C.N., et al., Pazopanib in Locally Advanced or Metastatic Renal Cell Carcinoma:
Results of a Randomized Phase III Trial. Journal of Clinical Oncology, 2010. 28(6): p. 1061-1068.  
14. Motzer, R.J., et al., Overall Survival and Updated Results for Sunitinib Compared With Interferon Alfa in Patients With Metastatic Renal Cell Carcinoma. Journal of Clinical Oncology, 2009. 27(22): p. 3584-3590.  
15. Schor-Bardach, R., et al., Does Arterial Spin-labeling MR Imaging-measured Tumor Perfusion Correlate with Renal Cell Cancer Response to Anti-angiogenic Therapy *in a Mouse Model? (vol* 251, pg 731, 2009). Radiology, 2009. 253(3): p. 900-900.  
16. Schnurch, H. and W. Risau, *Expression of Tie-2, a Member of a Novel Family of Receptor Tyrosine* Kinases, in the Endothelial-Cell Lineage. Development, 1993. 119(3): p. 957-968.  
17. Maisonpierre, P.C., et al., *Angiopoietin-2, a natural antagonist for Tie2 that disrupts in vivo* angiogenesis. Science, 1997. 277(5322): p. 55-60.  
18. Suri, C., et al., Requisite role of angiopoietin-1, a ligand for the TIE2 receptor, during embryonic angiogenesis. Cell, 1996. 87(7): p. 1171-80.  
19. Davis, S., et al., *Isolation of angiopoietin-1, a ligand for the TIE2 receptor, by secretion-trap* expression cloning. Cell, 1996. 87(7): p. 1161-9.  
20. Valenzuela, D.M., et al., *Angiopoietins 3 and 4: diverging gene counterparts in mice and humans.*
Proc Natl Acad Sci U S A, 1999. 96(5): p. 1904-9.  
21. Hu, B. and S.Y. Cheng, *Angiopoietin-2: development of inhibitors for cancer therapy.* Curr Oncol Rep, 2009. 11(2): p. 111-6.  
22. Casanovas, O., et al., Drug resistance by evasion of antiangiogenic targeting of VEGF signaling in late-stage pancreatic islet tumors. Cancer Cell, 2005. 8(4): p. 299-309.  
23. Bullock, A.J., et al., Plasma angiopoietin-2 (ANG2) as an angiogenic biomarker in renal cell carcinoma (RCC). Journal of Clinical Oncology, 2010. 28(15).  
24. Oliner, J., et al., Suppression of angiogenesis and tumor growth by selective inhibition of angiopoietin-2. Cancer Cell, 2004. 6(5): p. 507-16.  
25. Leow, C.C., et al., MEDI3617, a human anti-angiopoietin 2 monoclonal antibody, inhibits angiogenesis and tumor growth in human tumor xenograft models. Int J Oncol, 2012. 40(5): p.  
1321-30.  
26. Imanishi, Y., et al., *Angiopoietin-2 stimulates breast cancer metastasis through the alpha(5)beta(1)*
integrin-mediated pathway. Cancer Res, 2007. 67(9): p. 4254-63.  
27. Wang, X.E., et al., *The Role of Angiopoietins as Potential Therapeutic Targets in Renal Cell* Carcinoma. Translational Oncology, 2014. 7(2): p. 188-195.  
28. Atkins, M.B., et al., *Trebananib (AMG 386) in Combination With Sunitinib in Patients With* Metastatic Renal Cell Cancer: An Open-Label, Multicenter, Phase II Study. Journal of Clinical Oncology, 2015. 33(30): p. 3431-+.  
29. Motzer, R.J., et al., *Pazopanib versus sunitinib in metastatic renal-cell carcinoma.* N Engl J Med, 2013. 369(8): p. 722-31.  
30. Hudkins, R.L., et al., *Synthesis and biological profile of the pan-vascular endothelial growth factor* receptor/tyrosine kinase with immunoglobulin and epidermal growth factor-like homology domains 2 (VEGF-R/TIE-2) inhibitor 11-(2-methylpropyl)-12,13-dihydro-2-methyl-8-(pyrimidin-2-ylamino)- 4H-indazolo[5, 4-a]pyrrolo[3,4-c]carbazol-4-one (CEP-11981): a novel oncology therapeutic agent. J Med Chem, 2012. 55(2): p. 903-13.  
31. Pili, R., et al., An open-label study to determine the maximum tolerated dose of the multitargeted tyrosine kinase inhibitor CEP-11981 in patients with advanced cancer. Invest New Drugs, 2014.  
32(6): p. 1258-68.  
32. Qu, X., et al., *Promotion of tumorigenesis by heterozygous disruption of the beclin 1 autophagy* gene. J Clin Invest, 2003. 112(12): p. 1809-20.  
33. Saltz, L.B., et al., *Phase II trial of sunitinib in patients with metastatic colorectal cancer after* failure of standard therapy. J Clin Oncol, 2007. 25(30): p. 4793-9.  
34. Motzer, R.J., et al., *Nivolumab versus Everolimus in Advanced Renal-Cell Carcinoma.* N Engl J
Med, 2015. 373(19): p. 1803-13.  
35. Wallin, J.J., et al., Atezolizumab in combination with bevacizumab enhances antigen-specific T-cell migration in metastatic renal cell carcinoma. Nat Commun, 2016. 7: p. 12624.  
36. Koyama, T. and H. Chen, *Proper inference from Simon's two-stage designs.* Stat Med, 2008.  
27(16): p. 3145-54.
untitled h1
appendices appendix 1 karnofsky performance

| 100   | Normal, no complaints, no evidence of disease                                                              |
|-------|------------------------------------------------------------------------------------------------------------|
| 90    | Able to carry on normal activity, minor symptoms of disease                                                |
| 80    | Normal activity with effort, some signs of symptoms of disease                                             |
| 70    | Cares for self (consistent with age), unable to carry on normal activity or do active  work/school/play    |
| 60    | Requires occasional assistance (beyond age-appropriate care), but is able to care for  most of their needs |
| 50    | Requires considerable assistance and frequent medical care                                                 |
| 40    | Disabled, requires special care and assistance                                                             |
| 30    | Severely disabled, hospitalization is indicated although death is not imminent                             |
| 20    | Hospitalization is necessary, very sick, active support treatment is necessary                             |
| 10    | Moribund, fatal processes progressing rapidly                                                              |  
APPENDIX 2 MEDICATIONS WITH THE POTENTIAL FOR DRUG-DRUG INTERACTIONS
POTENT INHIBITORS OF THE CYP1A2, 2C8, OR 3A4 INCLUDE, BUT ARE NOT LIMITED TO, THE
FOLLOWING:
CYP1A2 FLUVOXAMINE CIPROFLOXACIN MEXILETINE PROPAFENONE
ZILEUTON
CYP2C8 GEMFIBROZIL
CYP3A4 KETOCONAZOLE
INDINAVIR RITONAVIR
ITRACONAZOLE CLARITHROMYCIN TELITHROMYCIN APREPITANT
DILTIAZEM
ERYTHROMYCIN FLUCONAZOLE GRAPEFRUIT JUICE
VERAPAMI
THIS LIST IS NOT COMPREHENSIVE AS NEW INFORMATION IS CONTINUALLY BEING IDENTIFIED.  
POTENT INDUCERS OF THE CYP1A2, 2C8, OR 3A4 INCLUDE, BUT ARE NOT LIMITED TO, THE FOLLOWING: CYP1A2 OMEPRAZOLE
SMOKING
CYP2C8 RIFAMPICIN
CYP3A4 RIFAMPICIN CARBAMAZEPINE
GRISEOFULVIN
THIS LIST IS NOT COMPREHENSIVE AS NEW INFORMATION IS CONTINUALLY BEING IDENTIFIED.  
SUBSTRATES OF CYP3A4/5 WITH A NARROW THERAPEUTIC RANGE INCLUDE, BUT ARE NOT LIMITED
TO:
ALFENTANIL ASTEMIZOLE CISAPRIDE CYCLOSPORINE
DIERGOTAMINE
ERGOTAMINE FENTANYL PIMOZIDE
QUINDINE
SIROLIMUS TACROLIMUS
TERFENADINE
THIS LIST IS NOT COMPREHENSIVE AS NEW INFORMATION IS CONTINUALLY BEING IDENTIFIED.
untitled h1
appendix 3 study management during

Due to ongoing government and clinical changes necessary to effectively manage the COVID-19 pandemic, the following changes to protocol-required items were made to minimize or eliminate immediate hazards or to protect the life and well-being of research participants (e.g., to limit exposure to COVID-19).
untitled h1
a. covid-19 testing

COVID-19 is not currently being added to the protocol as part of the screening requirements, but may be done as part of the clinical assessment, as needed during the course of the pandemic.  
COVID-19 tests/results will be recorded in the subject's source documents but will only be added as an Adverse/ Serious Adverse Event in the eCRF should the test yield a COVID-19 positive result.
untitled h1
b. study visit schedule

Virtual clinic visits will be allowed per clinician/subject discretion.  
Vitals are allowed to be measured and recorded by patient/family. If patient has items such as blood pressure cuff, thermometer, weight scale, etc. at home, clinical staff may request him/her to use such apparatuses and record values in source documents (e.g. heart rate, respiratory rate, temperature, weight, etc.) - noting that home equipment was used for the reading.
untitled h1
c. laboratory assessments

All institutional requirements regarding in person contact with subjects and facility closures (including laboratories) will be followed. Blood samples will not be collected or processed when labs are closed due to COVID-19 restrictions.
untitled h1
d. study medications

Adjustments for alternate drug administration have been permitted by the Supporter (Esanik Therapeutics) and Sponsor-Investigator, to allow for study medication to be shipped directly to patient's home, via certified temp-controlled container for overnight delivery, with tracking number and delivery signature required.