Immune checkpoint inhibitors (ICIs) are widely used for various malignancies. However, their safety and efficacy in patients with a kidney transplant have not been defined. To delineate this, we conducted a multicenter retrospective study of 69 patients with a kidney transplant receiving ICIs between January 2010 and May 2020. For safety, we assessed the incidence, timing, and risk factors of acute graft rejection. For efficacy, objective response rate and

The heme-containing Cytochrome P450s (CYPs) are a major enzymatic determinant of drug clearance and drug-drug interactions. The CYP3A4 isoform is inhibited by antifungal imidazoles or triazoles, which form low spin heme iron complexes via formation of a nitrogen-ferric iron coordinate bond. However, CYP3A4 also slowly oxidizes the antifungal itraconazole (ITZ) at a site that is ∼ 25 Å from the triazole nitrogens, suggesting that large antifungal azoles can adopt multiple orientations within the CYP3A4 active site. Here, we report a surface plasmon resonance (SPR) analysis with kinetic resolution of two binding modes of ITZ, and the related drug ketoconazole (KTZ). SPR reveals a very slow off-rate for one binding orientation. Multiphasic binding kinetics are observed and one of the two binding components resolved by curve-fitting exhibits 'equilibrium overshoot'. Pre-loading of CYP3A4 with the heme ligand imidazole abolishes this component of the antifungal azole binding trajectories, and it eliminates the conspicuously slow off-rate. The fractional populations of CYP3A4 complexes corresponding to different drug orientations can be manipulated by altering the duration of the pulse of drug exposure. UV-vis difference absorbance titrations yield low spin spectra and K D values that are consistent with the high affinity complex resolved by SPR. These results demonstrate that ITZ and KTZ bind in multiple orientations, including a catalytically productive mode and a slowly-dissociating inhibitory mode. Most importantly, they provide the first example of an SPR-based method for the kinetic characterization of drug binding to any human CYP, including mechanistic insight not available from other methods. Keywords protein-ligand interactions; tight binding inhibitors; drug metabolism; drug interactionsThe hepatic and intestinal heme-containing Cytochrome P450s (CYPs) 1 oxidize most drugs, and thus play a critical role in their metabolism and disposition [1][2][3]. Among the various human CYP isoforms, CYP3A4 contributes most to drug metabolism. CYP3A4 exhibits complex allosteric kinetics, which may result from drug-drug interactions on a single CYP molecule through multiple drug binding, as suggested by steady state kinetic behavior [4][5][6], large active sites observed in crystal structures [7][8][9][10] and spectroscopic studies [11]. Complex kinetics † This work was supported by NIH grants GM32165 (WMA, NI, KLK) and GM07750 (JTP). * Corresponding author: Tel: (206) FAX: (206) [12][13][14][15], and equilibrium optical titrations that rely on the inhibitor-or substrate-dependent changes in ferric spin state equilibrium [16][17][18]. The complexity of CYP kinetics severely impedes prediction of drug clearance and drug-drug interactions [12][13][14][15]19]. Clearly, new methods are required to elucidate the molecular basis of CYP allosterism and complex inhibitor or substrate binding.The antifungal azoles, including ketoconazole (KTZ) and itraconazole (ITZ), historically have been considered to be potent inh...

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657 Background: Ligation of IL-2 receptor or blockade of PD-1 receptor may change lymphocytes to induce regression of cancers with diverse histology or site of origin. Single agent objective response rates of 14-25% have been reported for IL-2 therapy of metastatic clear cell RCC (ccRCC). A response rate of 33% was observed in pembrolizumab treated ccRCC patients. Nivolumab treated ccRCC patients were observed to have early intratumoral migration of lymphocytes. A case report of IL-2 induced major regression right after no change on nivolumab therapy suggested that combining the two means of lymphocyte stimulation could be effective. Other trials combining IL-2 receptor agonists (NKTR-214) and PD-1 blockade have also reported regression of ccRCC. Distinctive attributes of high dose IL-2 therapy are the required inpatient stay and the durability of the complete responses. Methods: This single-institution, single arm study addresses the safety and feasibility of the combination of IL-2 and pembrolizumab in the treatment of metastatic ccRCC. Subjects are treated on four nine-week blocks, as follows: Pembrolizumab is given on weeks 1, 4, and 7 of each block. Patients are admitted for 5 doses of high dose IL-2 (given over ~ 33 hours/3 days) on weeks 2, 3, 5, and 6 of blocks 2 and 3. Safety is monitored by a Pocock boundary of .05 likelihood of 0.15 dose limiting toxicity rate. The Simon 2-stage alternative hypothesis for the sample size was a 45+% major response rate vs null hypothesis < 20%, at alpha = 0.10, 90% power. Results: No accrual stop for safety was triggered. Thirteen of the first 18 patients responded, substantially exceeding the requirement of 8+/24 combination-treated patients to reject the null. Seven patients responded after receiving pembrolizumab alone, six after starting combination therapy in block 2. Accrual is completed; at 27. Kaplan-Meier analysis projects ORR of 69%, with ORR 90%-lower confidence bound of 55%. Conclusions: The combination of high dose IL-2 and pembrolizumab is feasible, with a high response rate, justifying further exploration of this dual immune treatment of metastatic ccRCC. Clinical trial information: NCT02964078.

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Background Immune checkpoint inhibitors (ICIs) have shown modest antitumor activity in unselected advanced sarcomas. Histology driven approach to patient selection is the current standard for off-label anti-programmed cell death 1 (PD1) immunotherapy use. Methods We retrospectively reviewed the clinical characteristics and outcomes of patients with advanced sarcoma who were treated with off label anti-PD1 immunotherapy at our center. Results A total of 84 patients with 25 histological subtypes were included. Nineteen patients (23%) had a cutaneous primary tumor site. Eighteen patients (21%) were classified as having clinical benefit, including 1 patient with complete response, 14 with partial response, and 3 with stable disease lasting over 6 months with previously progressive disease. Cutaneous primary site location was associated with higher clinical benefit rate (58% vs. 11%, p < 0.001), longer median PFS (8.6 vs. 2.5 months, p = 0.003) and OS (19.0 vs. 9.2 months, p = 0.011), compared to non-cutaneous primary. Patients with histological subtypes that pembrolizumab is indicated per current National Comprehensive Cancer Network guidelines had modestly higher rate of clinical benefit versus other histologies, however, the difference was statistically insignificant (29% vs. 15%, p = 0.182) and no statistically significant difference in PFS or OS was observed between these groups. Immune-related adverse events were more frequently seen among patients with clinical benefit (72% vs. 35%, p = 0.007). Conclusions Anti-PD1-based immunotherapy is highly efficacious in advanced sarcomas of cutaneous primary site. Cutaneous primary site location is a stronger predictor of ICI response than histologic subtype and should be accounted for in treatment guidelines and clinical trial design.

Background Soft tissue sarcomas (STS) are rare and heterogeneous tumors making chemotherapy use controversial. Our goal was to identify a subset of patients with primary STS that benefit with the addition of chemotherapy. Methods A retrospective chart review included intermediate to high-grade localized primary STS of the extremity/trunk, and tumor size > 5 cm. The effect of chemotherapy was evaluated for local control (LC), distant control (DC), progression free survival (PFS), and overall survival (OS). Results In this cohort (n = 273), patients were treated with surgery (98%), radiation (81%), and chemotherapy (24.5%). With a median follow-up of 51 months, the entire cohort’s 5-year LC, DC, PFS, and OS are 79.1%, 59.9%, 43.8%, and 68.7%, respectively. The addition of chemotherapy did not provide a DC benefit (p = 0.238) for the entire cohort. High-grade disease (n = 210) experienced a 5-year benefit in DC (68% vs. 54.4%, p = 0.04), which was more pronounced with MAI (Mesna, Adriamycin, Ifosfamide) based regimens (74.2%, p = 0.016), and a 5-year PFS (50.8% vs 45%, p = 0.025) and OS benefit (76.2% vs 70%, p = 0.067) vs. no chemotherapy. On multivariate analysis of the high-grade subset, chemotherapy independently predicted for a DC benefit (HR 0.48 95% CI 0.26–89, p = 0.019). The benefit of chemotherapy was more pronounced with MAI, showing a significant benefit in DC (HR 0.333 95% CI 0.145–0.767, p = 0.01) and PFS (HR 0.52 95% CI 0.28–0.99, p = 0.047). Conclusion In patients with localized STS > 5 cm, the high-grade subset had a distant control benefit with the addition of chemotherapy, leading to improved progression free survival. This is more pronounced with the use of MAI and should be considered in patients eligible for this regimen.