Figure 3: Clonally diverse B cells co-cluster with paired T cell clones with an activated phenotype.

A) Correlations of V-gene usage between those detected using TRUST4 analysis of scRNA-seq vs. those detected in Xenium in situ for TCRs. “Patient-matched” refers to correlations between Xenium samples and scRNA-seq samples originating from the same patient at the same clinical time point where “patient-mismatched” refers to correlations between different patients or different clinical timepoints.

B) Correlations of V-gene usage between scRNA-seq vs. Xenium for IG-genes.

C) Correlation of T cell “pseudo-clone” sizes with paired B cell “pseudo-clone” sizes as a function of PD-1 inhibitor exposure of the originating tumor tissue and degree of spatial co-clustering between those T and B “pseudo-clones”.

D) Representative images of tumor and lymph node samples from patient su001 highlighting a pair of co-clustered “pseudo-clones” and a pair of unclustered “pseudo-clones”. Green lines represent T/B cell clones within 20μm of each other.

E) Fraction of TCRα “pseudo-clone” pairs present in the draining lymph node samples that are also present in the respective tumor sample of that patient. All lymph node samples were collected post-PD-1 inhibitor treatment. Patient su005 had no remaining archived pre-PD-1 inhibitor tumor to be analyzed.

F) Fraction of TCRβ pairs present in both draining lymph nodes and tumors.

G) Normalized gene expression by patient and tissue compartment for T cell clones that are present in both lymph nodes and tumor (shared) vs. those present in a single compartment (exclusive).

H) Schematic diagram of a proposed mechanism of antigen cross-presentation by B cells to CD8+ T cells leading to a cycle of tumor clearance and generation of additional tumor neo-antigens.