Status of alternative angiogenic pathways in glioblastoma resected under and after bevacizumab treatment.

Glioblastoma multiforme (GBM) acquires resistance to bevacizumab (Bev) treatment. Bev affects angiogenic factors other than vascular endothelial growth factor (VEGF), which are poorly understood. We investigated changes in angiogenic factors under and after Bev therapy, including angiopoietin-1 (ANGPT1), angiopoietin-2 (ANGPT2), placental growth factor (PLGF), fibroblast growth factor 2, and ephrin A2 (EphA2). Fifty-four GBM tissues, including 28 specimens from 14 cases as paired specimens from the same patient obtained in three settings: initial tumor resection (naïve Bev), tumors resected following Bev therapy (effective Bev), and recurrent tumors after Bev therapy (refractory Bev). Immunohistochemistry assessed their expressions in tumor vessels and its correlation with recurrent MRI patterns. PLGF expression was higher in the effective Bev group than in the naïve Bev group (p = 0.024) and remained high in the refractory Bev group. ANGPT2 and EphA2 expressions were higher in the refractory Bev group than in the naïve Bev group (p = 0.047 and 0.028, respectively). PLGF expression was higher in the refractory Bev group compared with the naïve Bev group for paired specimens (p = 0.036). PLGF was more abundant in T2 diffuse/circumscribe patterns (p = 0.046). This is the first study to evaluate angiogenic factors other than VEGF during effective and refractory Bev therapy in patient-derived specimens.

Integrative Transcriptomic and Single-Cell Protein Characterization of Colorectal Carcinoma Delineates Distinct Tumor Immune Microenvironments Associated with Overall Survival.

Colorectal carcinoma (CRC) is a heterogeneous group of tumors with varying therapeutic response and prognosis, and evidence suggests the tumor immune microenvironment (TIME) plays a pivotal role. Using advanced molecular and spatial biology technologies, we aimed to evaluate the TIME in patients with CRC to determine whether specific alterations in the immune composition correlated with prognosis. We identified primary and metastatic tumor samples from 31 consented patients, which were profiled with whole-exome sequencing and bulk RNA-seq. Immune cell deconvolution followed by gene set enrichment analysis and unsupervised clustering was performed. A subset of tumors underwent in situ analysis of the TIME spatial composition at single-cell resolution through Imaging Mass Mass Cytometry. Gene set enrichment analysis revealed two distinct groups of advanced CRC, one with an immune activated phenotype and the other with a suppressed immune microenvironment. The activated TIME phenotype contained increased Th1 cells, activated dendritic cells, tertiary lymphoid structures, and higher counts of CD8+ T cells whereas the inactive or suppressed TIME contained increased macrophages and a higher M2/M1 ratio. Our findings were further supported by RNA-seq data analysis from the TCGA CRC database, in which unsupervised clustering also identified two separate groups. The immunosuppressed CRC TIME had a lower overall survival probability (HR 1.66, p=0.007). This study supports the pertinent role of the CRC immune microenvironment in tumor progression and patient prognosis. We characterized the immune cell composition to better understand the complexity and vital role that immune activity states of the TIME play in determining patient outcome.

The evolution of metastatic upper tract urothelial carcinoma through genomic-transcriptomic and single-cell protein markers analysis.

The molecular characteristics of metastatic upper tract urothelial carcinoma (UTUC) are not well understood, and there is a lack of knowledge regarding the genomic and transcriptomic differences between primary and metastatic UTUC. To address these gaps, we integrate whole-exome sequencing, RNA sequencing, and Imaging Mass Cytometry using lanthanide metal-conjugated antibodies of 44 tumor samples from 28 patients with high-grade primary and metastatic UTUC. We perform a spatially-resolved single-cell analysis of cancer, immune, and stromal cells to understand the evolution of primary to metastatic UTUC. We discover that actionable genomic alterations are frequently discordant between primary and metastatic UTUC tumors in the same patient. In contrast, molecular subtype membership and immune depletion signature are stable across primary and matched metastatic UTUC. Molecular and immune subtypes are consistent between bulk RNA-sequencing and mass cytometry of protein markers from 340,798 single cells. Molecular subtypes at the single-cell level are highly conserved between primary and metastatic UTUC tumors within the same patient.