Assays to study cancer cell responses to pharmacologic or genetic perturbations are typically restricted to using simple phenotypic readouts such as proliferation rate. Information-rich assays, such as gene-expression profiling, have generally not permitted efficient profiling of a given perturbation across multiple cellular contexts. Here, we develop MIX-Seq, a method for multiplexed transcriptional profiling of post-perturbation responses across a mixture of samples with single-cell resolution, using SNP-based computational demultiplexing of singlecell RNA-sequencing data. We show that MIX-Seq can be used to profile responses to chemical or genetic perturbations across pools of 100 or more cancer cell lines. We combine it with Cell Hashing to further multiplex additional experimental conditions, such as posttreatment time points or drug doses. Analyzing the high-content readout of scRNA-seq reveals both shared and context-specific transcriptional response components that can identify drug mechanism of action and enable prediction of long-term cell viability from shortterm transcriptional responses to treatment.

The recent approval of two PD-1 inhibitors for the treatment of non–small cell lung cancer (NSCLC) has rapidly led to the widespread use of these agents in oncology practices. Pneumonitis has been recognized as a potentially life-threatening adverse event among NSCLC patients treated with PD-1 inhibitors; however, the detailed clinical and radiographic manifestations of this entity remain to be described. We report two cases of anti–PD-1 pneumonitis in advanced NSCLC patients treated with nivolumab after its FDA approval. Both cases presented with ground-glass and reticular opacities and consolidations in a peripheral distribution on CT, demonstrating a radiographic pattern of cryptogenic organizing pneumonia (COP). Consolidations were extensive and rapidly developed within 8 weeks of therapy in both cases. Both patients were treated with corticosteroids with subsequent improvement of respiratory symptoms and radiographic findings. One patient experienced recurrent pneumonitis after completing corticosteroid taper, or a “pneumonitis flare”, in the absence of nivolumab retreatment, with subsequent improvement upon corticosteroid re-administration. With the increasing use of immune checkpoint inhibitors in a growing number of tumor types, awareness of the radiographic and clinical manifestations of PD-1 inhibitor–related pneumonitis will be critical for the prompt diagnosis and management of this potentially serious adverse event.

BackgroundTumor response characteristics using immune-related RECIST1.1 (irRECIST1.1) in advanced non-small-cell lung cancer (NSCLC) patients treated with nivolumab monotherapy in the clinical setting have not been previously described with a direct comparison with the assessments according to the conventional RECIST1.1.MethodsFifty-six advanced NSCLC patients treated with nivolumab monotherapy after its Food and Drug Administration (FDA) approval were retrospectively studied. Tumor burden was quantified on serial CT scans during therapy using irRECIST1.1, which uses unidimensional measurements and includes new lesion measurements in total tumor burden. Response assessments by irRECIST1.1 were compared with assessments by RECIST1.1. Responses of individual lesions in different organs were also compared.ResultsTumor burden change at best overall response ranged from −66.8 to +278.1% (median: +3.9%). Response rate was 14% (8/56; 8 partial responses, 0 complete responses) by irRECIST1.1 and by RECIST1.1. Time-to-progression (TTP) by irRECIST1.1 was longer than TTP by RECIST1.1 (median TTP: not reached vs. 1.9 months, respectively). No patients experienced pseudoprogression during the study. Among 128 target lesions, the lesion-based size change at best response differed significantly across different organs, with adrenal lesions and lymph nodes having greater size decrease, followed by lung, while liver and other miscellaneous lesions had lesser degree of size decrease (p = 0.002).ConclusionsImmune-related response evaluations using irRECIST1.1 in advanced NSCLC patients treated with nivolumab resulted in the identical response rate and longer TTP compared to RECIST1.1. No pseudoprogression cases were observed during the study. Adrenal lesions and lymph nodes were more responsive and liver lesions were less responsive to nivolumab.Electronic supplementary materialThe online version of this article (doi:10.1186/s40425-016-0193-2) contains supplementary material, which is available to authorized users.