Effective methods for bulk RNA-Seq deconvolution using scnRNA-Seq transcriptomes

RNA profiling technologies at single-cell resolutions, including single-cell and single-nuclei RNA sequencing (scRNA-Seq and snRNA-Seq, scnRNA-Seq for short), can help characterize the composition of tissues and reveal cells that influence key functions in both healthy and disease tissues. However, the use of these technologies is operationally challenging because of high costs and stringent sample-collection requirements. Computational deconvolution methods that infer the composition of bulk-profiled samples using scnRNA-Seq-characterized cell types can broaden scnRNA-Seq applications, but their effectiveness remains controversial. We produced the first systematic evaluation of deconvolution methods on datasets with either known or scnRNA-Seq-estimated compositions. Our analyses revealed biases that are common to scnRNA-Seq 10X Genomics assays and illustrated the importance of accurate and properly controlled data preprocessing and method selection and optimization. Moreover, our results suggested that concurrent RNA-Seq and scnRNA-Seq profiles can help improve the accuracy of both scnRNA-Seq preprocessing and the deconvolution methods that employ them. Indeed, our proposed method, Single-cell RNA Quantity Informed Deconvolution (SQUID), combined RNA-Seq transformation and dampened weighted least-squares deconvolution approaches to consistently outperform other methods in predicting the composition of cell mixtures and tissue samples. Furthermore, our analysis suggested that only SQUID could identify outcomes-predictive cancer cell subclones in pediatric acute myeloid leukemia and neuroblastoma datasets, suggesting that deconvolution accuracy improvements are vital to enabling its applications in the life sciences.