Multi-modal single-cell and whole-genome sequencing of minute, frozen specimens to propel clinical applications

Abstract

Single-cell genomics are enabling technologies, but their broad clinical application remains challenging. We report an easily adaptable approach for single-cell transcriptome and T cell receptor (TCR)-sequencing, and matched whole-genome sequencing from tiny, frozen clinical specimens. We achieve similar quality and biological outputs while reducing artifactual signals compared to data from matched fresh tissue samples. Profiling sequentially collected melanoma samples from the KEYNOTE-001 trial, we resolve cellular, genomic, and clonotype dynamics that encapsulate molecular patterns of tumor evolution during anti-PD-1 therapy. To demonstrate applicability to banked biospecimens of rare diseases, we generate a large uveal melanoma liver metastasis single-cell and matched WGS atlas, which revealed niche-specific impairment of clonal T cell expansion. This study provides a foundational framework for propelling single-cell genomics to the clinical arena.

Starfysh reveals heterogeneous spatial dynamics in the breast tumor microenvironment

“…These features enabled the extraction of new biological insights from a limited cohort of breast cancer patients. In a recent study, we applied Starfysh to disentangle the spatial dynamics of activated and exhausted T cell subsets in Slide-Seq V2 77 data from anti-PD-1 treated melanoma tumors 78 , showing its applicability to other ST technologies and cancer systems. In future work, the incorporation of archetypal analysis in the probabilistic framework and extensions to multi-omics integration with proteomics or chromatin accessibility will improve our ability to achieve comprehensive characterization of spatial heterogeneity.…”

The CD58:CD2 axis is co-regulated with PD-L1 via CMTM6 and governs anti-tumor immunity

“…We demonstrate that these mechanisms are orthogonal to clinically validated mechanisms of resistance to immunotherapies, such as impaired antigen presentation 4,5 or IFN-ɣ-JAK/STAT signaling 3,33 . In addition to melanoma 7 , we also show that these mechanisms may play a role in kidney cancer patients treated with immune checkpoint blockade 21 . Furthermore, CD58 loss has previously been implicated in immune escape by hematological cancers 34-37 and has recently been identified as a mechanism of resistance to CD19-targeting CAR-T cells in patients with diffuse large B cell lymphoma (DLBCL) 20 .…”

“…Prior reports indicated that CD58 downregulation or loss is associated with cancer immune evasion and resistance to ICB in melanoma or CAR-T cell therapy in lymphoma [6][7][8]20 ; however, the underlying mechanisms are poorly understood. Analysis of The Cancer Genome Atlas (TCGA) indicates that baseline CD58 expression is variable in cutaneous melanoma and virtually absent in uveal melanoma, a rare subtype of melanoma that arises in the eye and has an extremely low response rate to immunotherapies (Fig.…”

“…Using single-cell RNA-seq in melanoma patients, we recently found that downregulation of CD58 (also known as lymphocyte adhesion factor 3, LFA-3) is associated with resistance to ICB and inferred T cell exclusion [6][7][8] ; however, the underlying mechanisms are poorly understood. CD58 is physiologically expressed on antigen presenting cells, namely macrophages, where it ligates with its T cell receptor CD2 to deliver "Signal 2" following T cell receptor (TCR) and MHC-presented antigen engagement 9,10 .…”