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Ex Vivo Profiling of PD-1 Blockade Using Organotypic Tumor Spheroids.

ABSTRACT: Ex vivo systems that incorporate features of the tumor microenvironment and model the dynamic response to immune checkpoint blockade (ICB) may facilitate efforts in precision immuno-oncology and the development of effective combination therapies. Here, we demonstrate the ability to interrogate ex vivo response to ICB using murine- and patient-derived organotypic tumor spheroids (MDOTS/PDOTS). MDOTS/PDOTS isolated from mouse and human tumors retain autologous lymphoid and myeloid cell populations and respond to ICB in short-term three-dimensional microfluidic culture. Response and resistance to ICB was recapitulated using MDOTS derived from established immunocompetent mouse tumor models. MDOTS profiling demonstrated that TBK1/IKK? inhibition enhanced response to PD-1 blockade, which effectively predicted tumor response in vivo Systematic profiling of secreted cytokines in PDOTS captured key features associated with response and resistance to PD-1 blockade. Thus, MDOTS/PDOTS profiling represents a novel platform to evaluate ICB using established murine models as well as clinically relevant patient specimens.Significance: Resistance to PD-1 blockade remains a challenge for many patients, and biomarkers to guide treatment are lacking. Here, we demonstrate feasibility of ex vivo profiling of PD-1 blockade to interrogate the tumor immune microenvironment, develop therapeutic combinations, and facilitate precision immuno-oncology efforts. Cancer Discov; 8(2); 196-215. ©2017 AACR.See related commentary by Balko and Sosman, p. 143See related article by Deng et al., p. 216This article is highlighted in the In This Issue feature, p. 127.

SUBMITTER: Jenkins RW

PROVIDER: S-EPMC5809290 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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Ex Vivo Profiling of PD-1 Blockade Using Organotypic Tumor Spheroids.

Jenkins Russell W RW Aref Amir R AR Lizotte Patrick H PH Ivanova Elena E Stinson Susanna S Zhou Chensheng W CW Bowden Michaela M Deng Jiehui J Liu Hongye H Miao Diana D He Meng Xiao MX Walker William W Zhang Gao G Tian Tian T Cheng Chaoran C Wei Zhi Z Palakurthi Sangeetha S Bittinger Mark M Vitzthum Hans H Kim Jong Wook JW Merlino Ashley A Quinn Max M Venkataramani Chandrasekar C Kaplan Joshua A JA Portell Andrew A Gokhale Prafulla C PC Phillips Bart B Smart Alicia A Rotem Asaf A Jones Robert E RE Keogh Lauren L Anguiano Maria M Stapleton Lance L Jia Zhiheng Z Barzily-Rokni Michal M Cañadas Israel I Thai Tran C TC Hammond Marc R MR Vlahos Raven R Wang Eric S ES Zhang Hua H Li Shuai S Hanna Glenn J GJ Huang Wei W Hoang Mai P MP Piris Adriano A Eliane Jean-Pierre JP Stemmer-Rachamimov Anat O AO Cameron Lisa L Su Mei-Ju MJ Shah Parin P Izar Benjamin B Thakuria Manisha M LeBoeuf Nicole R NR Rabinowits Guilherme G Gunda Viswanath V Parangi Sareh S Cleary James M JM Miller Brian C BC Kitajima Shunsuke S Thummalapalli Rohit R Miao Benchun B Barbie Thanh U TU Sivathanu Vivek V Wong Joshua J Richards William G WG Bueno Raphael R Yoon Charles H CH Miret Juan J Herlyn Meenhard M Garraway Levi A LA Van Allen Eliezer M EM Freeman Gordon J GJ Kirschmeier Paul T PT Lorch Jochen H JH Ott Patrick A PA Hodi F Stephen FS Flaherty Keith T KT Kamm Roger D RD Boland Genevieve M GM Wong Kwok-Kin KK Dornan David D Paweletz Cloud Peter CP Barbie David A DA

Cancer discovery 20171103 2

Ex vivo systems that incorporate features of the tumor microenvironment and model the dynamic response to immune checkpoint blockade (ICB) may facilitate efforts in precision immuno-oncology and the development of effective combination therapies. Here, we demonstrate the ability to interrogate ex vivo response to ICB using murine- and patient-derived organotypic tumor spheroids (MDOTS/PDOTS). MDOTS/PDOTS isolated from mouse and human tumors retain autologous lymphoid and myeloid ce ...[more]

PMID: 29101162

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Project description:Blockade of PD-1/PD-L1 interactions is proving an exciting, durable therapeutic modality in a range of cancers whereby T cells are released from checkpoint inhibition to revive their inherent anti-tumour activity. Here we have studied various ways to model ex vivo T cell function in order to compare the impact of the clinically utilised anti-PD-1 antibody, pembrolizumab (Keytruda) on the activation of human T cells: focussing on the release of pro-inflammatory IFNγ and anti-inflammatory IL-10 to assess functionality. Firstly, we investigated the actions of pembrolizumab in an acute model of T-cell activation with either immature or mature allogeneic dendritic cells (DCs); pembrolizumab enhanced IFNγ and IL-10 release from purified CD4+ T-cells in the majority of donors with a bias towards pro-inflammatory cytokine release. Next, we modelled the impact of pembrolizumab in settings of more chronic T-cell activation. In a 7-day antigen-specific response to EBV peptides, the presence of pembrolizumab resulted in a relatively modest increase in both IFNγ and IL-10 release. Where pembrolizumab was assessed against long-term stimulated CD4+ cells that had up-regulated the exhaustion markers TIM-3 and PD-1, there was a highly effective enhancement of the otherwise exhausted response to allogeneic DCs with respect to IFNγ production. By contrast, the restoration of IL-10 production was considerably more limited. Finally, to assess a direct clinical relevance we investigated the consequence of PD-1/PD-L1 blockade in the disease setting of dissociated cells from lung and colon carcinomas responding to allogeneic DCs: here, pembrolizumab once more enhanced IFNγ production from the majority of tumour preparations whereas, again, the increase in IL-10 release was modest at best. In conclusion, we have shown that the contribution of PD-1-revealed by using a canonical blocking antibody to interrupt its interaction with PD-L1-to the production of an exemplar pro- and anti-inflammatory cytokine, respectively, depends in magnitude and ratio on the particular stimulation setting and activation status of the target T cell. We have identified a number of in vitro assays with response profiles that mimic features of dissociated cell populations from primary tumours thereby indicating these represent disease-relevant functional assays for the screening of immune checkpoint inhibitors in current and future development. Such in vitro assays may also support patient stratification of those likely to respond to immuno-oncology therapies in the wider population.

Project description:BACKGROUND:Epilepsy is a prevalent neurological disorder worldwide. It is characterized by an enduring predisposition to generate seizures and its development is accompanied by alterations in many cellular processes. Organotypic slice cultures represent a multicellular environment with the potential to assess biological mechanisms, and they are used as a starting point for refining molecules for in vivo studies. Here, we investigated organotypic slice cultures as a model of epilepsy. METHODS:We assessed, by electrophysiological recordings, the spontaneous activity of organotypic slices maintained under different culture protocols. Moreover, we evaluated, through molecular-based approaches, neurogenesis, neuronal death, gliosis, expression of proinflammatory cytokines, and activation of NLRP3 inflammasome (nucleotide-binding, leucine-rich repeat, pyrin domain) as biomarkers of neuroinflammation. RESULTS:We demonstrated that organotypic slices, maintained under a serum deprivation culture protocol, develop epileptic-like activity. Furthermore, throughout a comparative study with slices that do not depict any epileptiform activity, slices with epileptiform activity were found to display significant differences in terms of inflammation-related features, such as (1) increased neuronal death, with higher incidence in CA1 pyramidal neurons of the hippocampus; (2) activation of astrocytes and microglia, assessed through western blot and immunohistochemistry; (3) upregulation of proinflammatory cytokines, specifically interleukin-1? (IL-1?), interleukin-6, and tumor necrosis factor ?, revealed by qPCR; and (4) enhanced expression of NLRP3, assessed by western blot, together with increased NLRP3 activation, showed by IL-1? quantification. CONCLUSIONS:Thus, organotypic slice cultures gradually deprived of serum mimic the epileptic-like activity, as well as the inflammatory events associated with in vivo epilepsy. This system can be considered a new tool to explore the interplay between neuroinflammation and epilepsy and to screen potential drug candidates, within the inflammatory cascades, to reduce/halt epileptogenesis.

Dataset Information

Ex Vivo Profiling of PD-1 Blockade Using Organotypic Tumor Spheroids.

Publications

<i>Ex Vivo</i> Profiling of PD-1 Blockade Using Organotypic Tumor Spheroids.

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