TAK1 expression is associated with increased PD-L1 and decreased cancer-specific survival in microsatellite-stable colorectal cancer.

BACKGROUND: Transforming growth factor ß-activated protein kinase-1 (TAK1) plays an important role in MAPK and NFκB pathways and has been associated with colorectal cancer. The aim of this study was to determine how cytoplasmic and juxtanuclear punctate staining of TAK1 relates to immune checkpoint expression and cancer specific survival in colorectal cancer. METHODS: Protein expression was assessed by immunohistochemistry on tissue microarrays from primary curative colorectal cancer resected specimens. Expression levels of cytoplasmic TAK1 by QuPath digital quantification and punctate TAK1 staining was scored using a manual point scoring technique and correlated with clinicopathological features, immune checkpoint expression and cancer-specific survival. Bulk RNA sequencing was performed in specimens to determine mutational profiles and differentially expressed genes. RESULTS: A cohort of 875 patients who had undergone colorectal cancer resection were assessed for TAK1 expression. Higher levels of cytoplasmic TAK1 expression correlated with elevated PD1 and PD-L1 expression (p < 0.010). High punctate TAK1 expression was more commonly identified in poorly differentiated colorectal cancers (p = 0.036), had dysregulated mutational and transcriptional profiles with decreased insulin-like growth factor 2(IGF2) expression (p < 0.010), and independently predicted poor cancer-specific survival (HR 2.690, 95% CI 1.419-5.100, p = 0.002). The association of punctate TAK1 expression and recurrence remained after subgroup analysis for microsatellite-stable colorectal cancer (p = 0.028). DISCUSSION: Punctate TAK1 expression is associated with worse cancer specific survival. TAK1 signalling may be an important pathway to investigate underlying mechanisms for recurrence in microsatellite-stable colorectal cancer.

Spatial transcriptomic analysis of tumour with high and low CAIX expression in TNBC tissue samples using GeoMx™ RNA assay.

PURPOSE: Prognostic significance and gene signatures associated with carbonic anhydrase IX (CAIX) was investigated in triple negative breast cancer (TNBC) patients. METHODS: Immunohistochemistry (IHC) for CAIX was performed in tissue microarrays (TMAs) of 136 TNBC patients. In a subset of 52 patients Digital Spatial Profiler (DSP) was performed in tumour (pan-cytokeratin+) and stroma (pan-cytokeratin-). Differentially expressed genes (DEGs) with P<0.05 and and log2 fold change (FC)>(±0.25 and ±0.3, for tumour and stromal compartment, respectively) were identified. Four genes were validated at the protein level. RESULT: Cytoplasmic CAIX expression was independently associated with poor recurrence free survival in TNBC patients [hazard ratio (HR)=6.59, 95% confidence interval (CI): 1.47-29.58, P=0.014]. DEG analysis identified 4 up-regulated genes (CD68, HIF1A, pan-melanocyte, and VSIR) in the tumour region and 9 down-regulated genes in the stromal region (CD86, CD3E, MS4A1, BCL2, CCL5, NKG7, PTPRC, CD27, and FAS) when low versus high CAIX expression was explored. Employing IHC, high CD68 and HIF-1α was associated with poorer prognosis and high BCL2 and CD3 was associated with good prognosis. CONCLUSIONS: DSP technology identified DEGs in TNBC. Selected genes validated by IHC showed involvement of CD3 and BCL2 expression within stroma and HIF-1α, and CD68 expression within tumour. However, further functional analysis is warranted.

Relationship between immune checkpoint proteins, tumour microenvironment characteristics, and prognosis in primary operable colorectal cancer.

The tumour microenvironment is an important factor for colorectal cancer prognosis, affecting the patient's immune response. Immune checkpoints, which regulate the immune functions of lymphocytes, may provide prognostic power. This study aimed to investigate the prognostic value of the immune checkpoints TIM-3, LAG-3 and PD-1 in patients with stage I-III colorectal cancer. Immunohistochemistry was employed to detect TIM-3, LAG-3, PD-1 and PD-L1 in 773 patients with stage I-III colorectal cancer. Immune checkpoint protein expression was assessed in tumour cells using the weighted histoscore, and in immune cells within the stroma using point counting. Scores were analysed for associations with survival and clinical factors. High tumoural LAG-3 (hazard ratio [HR] 1.45 95% confidence interval [CI] 1.00-2.09, p = 0.049) and PD-1 (HR 1.34 95% CI 1.00-1.78, p = 0.047) associated with poor survival, whereas high TIM-3 (HR 0.60 95% CI 0.42-0.84, p = 0.003), LAG-3 (HR 0.58 95% CI 0.40-0.87, p = 0.006) and PD-1 (HR 0.65 95% CI 0.49-0.86, p = 0.002) on immune cells within the stroma associated with improved survival, while PD-L1 in the tumour (p = 0.487) or the immune cells within the stroma (p = 0.298) was not associated with survival. Furthermore, immune cell LAG-3 was independently associated with survival (p = 0.017). Checkpoint expression scores on stromal immune cells were combined into a Combined Immune Checkpoint Stromal Score (CICSS), where CICSS 3 denoted all high, CICSS 2 denoted any two high, and CICSS 1 denoted other combinations. CICSS 3 was associated with improved patient survival (HR 0.57 95% CI 0.42-0.78, p = 0.001). The results suggest that individual and combined high expression of TIM-3, LAG-3, and PD-1 on stromal immune cells are associated with better colorectal cancer prognosis, suggesting there is added value to investigating multiple immune checkpoints simultaneously.