Table 5.1. Agent Administration and Timing of Assessments Cohort INCB024360 Dosing MELITAC 12.1 Dosing Blood draws Tumor biopsies including FNA A Pts.enrolled >6 wks after failing anti- PD-1/PDL-1 300 mg po bid continuous for 98 days (3 months) Intradermal/subcutaneous Days 21, 28, 35, 56, 77, 98 (Weeks 3, 4, 5, 8, 11, and 14) Pretreatment day 0 and on days 21, 42, 98, and 112 Days 0, 21, 42, and (optional day 112) and (optional when coming off study including for progression) B Pts.enrolled 2-6 wks after failing anti- PD-1/PDL-1 100 mg po bid continuous for 98 days (3 months) Intradermal/subcutaneous Days 21, 28, 35, 56, 77, 98 (Weeks 3, 4, 5, 8, 11, and 14) Pretreatment day 0 and on days 21, 42, 98, and 112 Days 0, 21, 42, and (optional day 112) and (optional when coming off study including for progression) Abbreviations: bid, twice per day; FNA, fine-needle aspiration; po, orally; pr e-tx, pretreatment; SC, subcutaneous Enrollment will be staggered so that only 3 patients will initially be treated at the 300-mg bid dose in order to observe their response to the INCB024360-MELITAC 12.1 combination. After the first patient is st arted on INCB024360, treatment will proceed for 2 weeks before the second patient is st arted on INCB024360 monotherapy. Before the second patient starts the combination of INCB024360 and MELITAC 12, the first patient will have already received the second vacci nation of MELITAC 12.1 in combination with INCB024360 and observed for any toxicity. The third patient will start INCB024360 monotherapy 2 weeks after patient number 2 has received the combination of INCB024360 and MELITAC 12.1 and been obser ved for toxicities. After the first 3 patients are treated at the 300-mg bid dos e of INCB024360 in combination with the MELITAC 12.1 and found to tolerate the co mbination well, the dose of INCB024360 will be 300 mg BID for subsequent patients. Those patients treated with anti-PD- 1/PDL1antibody who have failed therapy and enrolled in the st udy 6 weeks or longer after progression of disease will be enroll ed in Cohort A and receive 300 mg BID of INCB024360. Patients who fail anti-PD-1/PDL-1 th erapy and enroll within 2-6 weeks of progression of disease will receiv e100 mg BID in Cohort B. All other patients enrolled in the study will receive INCB024360 300 mg BID. A total of 12 patients will be enrolled in the study. Assignment to Cohort A or B is for dose assignment only. NCI Protocol #: CITN-04 Version Date: 27 Jan 2016 30 5.1.1 CTEP IND Agent(s ): N/A 5.1.2 Other Agent(s): INCB024360 Combining INCB024360 with the multipep tide vaccine MELITAC 12.1 is a novel approach. Although no adverse findings in toxicology or safety pharmacology assessments have been described herein, a nd the most common grade 1 or 2 AEs at the recommended Phase-II dose (600 mg) include fatigue, nausea and vomiting, constipation, and anorexia; to decrease the possibility of una nticipated toxicities with the combination, the dose of INCB024360 in the first 3 patients tr eated in the study will be at the dose of 300 mg bid. Weak cross-reactivity of INCB024360 to the human vasopressin 1a receptor, dopamine receptor, and carbonic anhydrase II may lead to changes in blood pressure or blood electrolytes. In 28-day toxicology studies, C max values have exceeded the IC 50 for the IDO1 enzyme (72 nM) by up to 37-fold and the IC 50 for the vasopressin 1a receptor by up to 4-fold (13-fold in single- dose studies) in the absence of any toxicity, so the risk of unintended pharmacological activity is expected to be low. 5.1.3 Other Agent (s): MELITAC 12.1 MELITAC 12.1 vaccine comprises 100 μg each of the 12 melanoma peptides plus 190 μ g tetanus toxoid peptide (Pep tide-tet). Peptides are synt hesized under good manufacturing practice (GMP) conditions by PolyPeptide Gr oup (San Diego, CA; formerly Multiple Peptide Systems), reconstituted, mixed, sterile -filtered, and vialed under GMP conditions by Merck (Germany). The 12–melanoma peptide mixture and tetanus peptide preparation are made as separate sterile aqueous solutions to be administered in an emulsion with Montanide ISA-51 VG adjuvant (1 mL), us ing the 2-syringe method after emulsion stability and quality is verified. MELITAC 12.1 stimulated immune responses with an excellent safety profile and w ith presumptive evidence of efficacy in the adjuvant setting. At this writing, the 12-MP has been administ ered in combination with Peptide-tet and GM-CSF in Montanide ISA-51 VG adjuvant to mo re than 200 participants enrolled in 3 studies run through the University of Virg inia Human Immune Therapy Center (HITC) in collaboration with other centers (Mel39, Mel43, E1602) [ Slingluff 2007 ; Slingluff 2009; Slingluff 2010 ] and 12-MP has been administ ered in Montanide ISA-51 VG adjuvant, with or without Pe ptide-tet in about 300 part icipants (Mel43, Mel44, E1602) [Slingluff 2010 ; Slingluff 2011 ; Slingluff 2009 ]. No data are available regarding the distribution and excretion of th e vaccine preparation. In vitr o studies have indicated the half-life of synthetic peptides in the presence of serum peptidases is short (approximately 22 seconds) [ Brinckerhoff 1999 ]. However, in vivo data describing the metabolism of the vaccine are not available. In the first 51 participants vaccinated with the 12-MP and Peptide-tet with GM-CSF in Montanide ISA-51 VG adjuvant, the majority of toxicities described have been of minimal severity. The most frequent AEs for participants receiving the 12-MP in NCI Protocol #: CITN-04 Version Date: 27 Jan 2016 31 combination with Peptide-tet include maximu m grade 1 fatigue (49%), maximum grade 1 chills (49%), maximum grade 1 headache (4 2%), and maximum grade 2 injection site reactions (69%). Other common AEs include maximum grade 1 sweating (27%), maximum grade 1 hyperglycemia (33%), maximum grade 1 myalgia (24%), and maximum grade 3 injection site reaction (27%). At this writing, at least 1 case of vitiligo and no cases of visual change s related to the vaccine after vaccination with the 12-MP and Peptide-tet administered with GM-CSF in Montanide ISA-51 VG adjuvant have been reported. 5.1.4 Other Modalities or Procedures: N/A 5.1.5 Investigational Imaging Agent Administration: N/A 5.2 Definition of Dose-Limiting Toxicity: Dose-limiting toxicities (DLTs) include all Grade 3 AEs regardless of investigator attribution or relatedness except for those listed below. Based on investigator determination the follo wing Grade 3 toxicities can be deemed non- DLT:  adverse event with a clear alternative explana tion (e.g. due to disease progression)  transient toxicities ( ≤72 hours)  injection site reactions that are expe cted and do not require any surgical intervention or repair  abnormal laboratory values without associ ated clinically significant signs or symptoms There are two exceptions to the above. Grade 1 or greater ocular toxicities will be considered a DLT. Grade 2 LFT elev ations will be considered a DLT. In the event of a DLT, it will be determined by the safety team whether both the INCB024360 AND the MELITAC 12.1 will be perm anently discontinued. In cases of vaccine injection site reacti on or other vaccine related adverse events the INCB024360 may be continued if the vaccine related a dverse event continues to show improvement. Management and dose modifications associated with the above adverse events are further outlined in Section 6 . 5.3 General Concomitant Medication and Supportive Care Guidelines 5.3.1 INCB024360 INCB024360 is neither a substrate nor an in hibitor or inducer of cytochrome P450 enzymes. Analysis of human plasma sample s indicated that INCB024360 is metabolized by 2 major pathways: O-glucuronidation and re duction. The identifica tion of the 2 major pathways and the enzymes responsible for thos e pathways has enable d the refinement of the inclusion/exclusion criteria around c oncomitant medications. However for safety purposes, the case report form must capture th e concurrent use of all other drugs, over- NCI Protocol #: CITN-04 Version Date: 27 Jan 2016 32 the-counter medications, or alternative therapies. Hypothetically, inhibition of IDO could cause an increase in serotonin levels that could precipitate a cluster of AEs termed serot onin syndrome. This rare syndrome has been associated with some MAO inhibitors and w ith combinations of serotonergic drugs. Symptom onset is usually rapid, often occurr ing within minutes, and encompasses a wide range of clinical findings: o Mild symptoms may only consist of increa sed heart rate, shivering, sweating, dilated pupils, myoclonus, and over responsive reflexes. o Moderate symptoms include additional abnormalities such as hyperactive bowel sounds, high blood pressure, and hyperther mia. A temperature as high as 40°C (104°F) is common in moderate intoxication. Mental status changes include hypervigilance and agitation. o Severe symptoms include severe increases in heart rate and blood pressure that may lead to shock. In life-thr eatening cases, temperature may rise to above 41.1°C (106.0°F). Other abnormalities include meta bolic acidosis, rhabdomyolysis, seizures, renal failure, and disseminated intravascular coagulation; these effects usually arise as a consequence of hyperthermia. At this writing, no laboratory te st exists for serotonin syndrom e; therefore diagnosis is by symptom observation. Management is based primarily on stopping the usage of the precipitating drugs, the administration of sero tonin antagonists such as cyproheptadine, and supportive care including the control of agitation, autonomic instability, and hyperthermia. Additionally, those who ingest la rge doses serotonergic agents may benefit from gastrointestinal decontamination with act ivated charcoal if it can be administered within an hour of overdose. The intensity of therapy depends on the severity of symptoms. If the symptoms are mild, treatm ent may only consist of discontinuation of the offending medication or medications , offering supportive measures, giving benzodiazepines for myoclonus, and waiting for the symptoms to resolve. Moderate cases should have all thermal and cardiorespirato ry abnormalities corrected and can benefit from serotonin antagonists. As mentioned a bove, the serotonin anta gonist cyproheptadine is the recommended initial therapy. Additional pharmacological treatment for severe case includes administering atypical antipsychotic drugs with se rotonin antagonist activity (e.g., olanzapine). Critically ill patients shoul d receive the above th erapies as well as sedation and/or neuromuscular paralysis. Patients who have autonomic instability such as low blood pressure require treatment with direct-acting sympathomimetics, such as epinephrine, norepinephrine, or phenylephrin e. Conversely, hypertension or tachycardia can be treated with short-acting antihypertensi ve drugs such as nitroprusside or esmolol; longer-acting drugs such as propranolol s hould be avoided as they may lead to hypotension and shock. As mentioned above, weak cr oss-reactivity of INCB024360 to the human vasopressin 1a receptor, dopamine receptor, and carbonic anhydrase II may lead to changes in blood pressure or blood electrolytes. In 28-day toxicology studies, C max values have exceeded the IC 50 for the IDO1 enzyme (72 nM) by up to 37-fold and the IC 50 for the vasopressin 1a receptor by up to 4-fold (13-fold in single- dose studies) in the absence of any toxicity, so the risk of unintended pharmacological activity is expected to be low. NCI Protocol #: CITN-04 Version Date: 27 Jan 2016 33 Studies imply that IDO1 may be a negative inhibition pathway to control autoimmune diseases associated with inflammation. Recent studies with the anti-CTLA-4 antibody ipilimumab have shown dysregul ation of mucosal immunity and gastrointestinal toxicity, which manifests as diarrhea. For these reasons, subjects w ith a history of autoimmune disease should be excluded fr om participation in this study, and study participants should be closely monitored for signs or symptoms of developing autoimmune or inflammatory disease. Analysis of human plasma samples indicated that INCB024360 is metabolized by 2 major pathways: O-glucuronidation and reduc tion. The O-glucuronide (M9) was formed by UGT1A9 and the reduced metabolite (M11) a ppears to be catalyzed by intestinal microflora. Metabolite M11 is also further metabolized to M12, a product formed by oxidative cleavage catalyzed by CYPs. The id entification of the 2 major pathways and the enzymes responsible for those pathwa ys has enabled the refinement of the inclusion/exclusion criteria around concomitant medications. It should be noted that all three major metabolites (M9, M11 & M12) are inactive against IDO1 enzyme. Use of any UGT1A9 inhibitor including: acitretin, amitriptyline, androsterone, cyclosporine, dasatinib, diclofenac, diflunisal, efavirenz, erlotinib, estradiol (17-beta), flutamide, geftinib, gemfibrozil, glycyrrhetin ic acid glycyrrhizin, imatinib, imipramine, ketoconazole, linoleic acid, mefenamic acid, mycophenolic acid, niflumic acid, nilotinib, phenobartital, phenylbutazone, phenytoin, and probenecid propofol, qui nidine, ritonavir, Sorafenib, sulfinpyrazone, valproic acid, and verapamil should be avoided from screening through follow-up period. Low-dose Coumadin (1 mg) is acceptable; however, doses that increase INR are not permitted. If an alternative to Coumadin-bas ed anticoagulants cannot be used, the INR should be monitored weekly after initiation of therapy and upon discontinuation of INCB024360, until INR normalization. 5.3.2 MELITAC 12.1 In several studies, the vaccine has been well tolerated. In the most recent study, in which the adjuvant was Montanide ISA-51 VG alone , Mel44 (n=170), 8% of patients reported unexpected grade 3 treatment-related AEs, pl us 10% of patients with grade 3 injection- site ulceration and 7% with grade 3 injection- site induration; the in jection-site reactions were expected and not dose-limiting. One gr ade 4 toxicity was reported (hypoglycemia, <1%). Other common toxicities were evid ent for approximately 24 hours included fatigue, dyspnea, headache, and myalgia. Mild AEs of rigors/chills , anorexia, nausea, sweating, fever, and flushing were commonly experienced [ Slingluff 2011 ]. 5.4 Duration of Therapy Patients with surgically incurable stage III or IV melanoma and tissue amenable to serial biopsy will receive the sequential thera py with INCB024360 alone for 3 weeks followed NCI Protocol #: CITN-04 Version Date: 27 Jan 2016 34 by continued INCB024360 plus 6 MELITAC 12.1 vaccines administered intradermally or subcutaneously on days 21, 28, 35, 56, 77, and 98 (weeks 3, 4, 5, 8, 11, and 14). Serial biopsies will be obtained up to 14 days before starting INCB024360, on day 21 (before the first vaccine), and on day 42 (a week after the third vaccine ). When feasible a biopsy will be obtained at week 16 (2 week s after the sixth vacci nation). If possible, tumors will be biopsied from patients coming off study, including for progression of disease. In the absence of treatment delays due to AE(s), treatment may continue for up to 3 additional cycles of INCB024360 (a total of 4 cycles) after consulta tion with the trial Principal Investigator, CTEP, and the CITN, or until one of the following criteria applies:  Disease progression by Response Evaluation Cr iteria In Solid Tumors (RECIST) or Immune-Related Response Criteria (irRC)  Intercurrent illness that prevents further administration of treatment  Unacceptable AE(s)  Patient decides to withdraw from the study  General or specific changes in the pa tient’s condition rendering the patient unacceptable for further treatment in the judgment of the investigator If additional cycles of INCB024360 are d eemed appropriate for the patient, the INCB024360 will be given without the admini stration of the MELITAC 12.1 vaccines. Only the end of treatment biopsy will be obta ined if additional cycles of INCB024360 are approved for each patient. In patients who are approved to receive additional cycles of INCB024360, a reduced volume of blood will be drawn for specific assays. (Study Calendar, footnote ***) A delay of up to 60 days between treatment cycles will be allowed. In the event of unexpected se rious AEs related to either component of the combination, treatment of the affected participant with INCB024360 and/or MELITAC 12.1 will be discontinued after analysis by the DSMC, Incyte, and CITN. 5.5 Duration of Follow-up Patients will be followed for 1 year after re moval from study or until death, whichever occurs first. Patients removed from study for unacceptable AE(s) will be followed until resolution or stabilization of the AE. Unaccep table toxicity is defined as any of the following: 1) grade 3 or 4 adverse event requiri ng the patient to be removed from study treatment, 2) DLT’s of grade 1 treatment related ocular toxi city, grade 2 alle rgic reactions, 3) >2 cm ulceration at the vaccinatio n site or a vaccination adverse event requiring antibiotic treatment, debrid ement or narcotic management, NCI Protocol #: CITN-04 Version Date: 27 Jan 2016 35 NOTE: Chronic inflammatory reactions at the vaccine sites are expected to occur in all participants who receive Montanid e ISA-51. Induration may persist for days to months, but is not expected to requir e additional therapy. Sterile abscesses may occur in some participants. These are not a basis for discontinuation of the vaccines. However, if the inflammatory reactions develop significant ulceration of the skin (> 2 cm), then the patient would be taken off the study, however, this would not be considered a dose-limiting toxicity. 4) any unexpected grade 3 or greater non- hematologic or hematologic toxicity. 5.6 Criteria for Removal from Study Patients will be removed from study wh en any of the criteria listed in Section 5.4 applies. The reason for study removal and the date the patient was removed must be documented in the case report form. The patient should be followed per protocol requirements. AEs will be described and coded based upon the National Cancer Institute Common Terminology Criteria for Adverse Events v4.0 (NCI CTCAE v4.0). Safety will be evaluated by assessing and chronicling treatment-related AEs.