Phenotyping of tumor-associated macrophages in colorectal cancer: Impact on single cell invasion (tumor budding) and clinicopathological outcome

Abstract

Tumor-associated macrophages (TAM) play a controversial role in epithelial-mesenchymal transition (EMT) and prognosis of colorectal cancer (CRC). In particular, the microlocalization, polarization and prognostic impact of TAM in the immediate environment of invading CRC cells has not yet been established. To address this clinically relevant question, intraepithelial (iCD68) and stromal macrophages (sCD68), M1-macrophages (iNOS), M2-macrophages (CD163), cytokeratin-positive cancer cells (tumor buds) and expression of the anti-phagocytic marker CD47 were investigated in primary tumors of 205 well-characterized CRC patients. Cell-to-cell contacts between tumor buds and TAM were detected using high-resolution digital scans. The composition of the tumor microenvironment was analyzed with clinicopathological and molecular features. High CD68 counts predicted long term overall survival independent of microlocalization (iCD68 p=0.0016; sCD68 p=0.03), pT, pN, pM and post-operative therapy. CD68 infiltration correlated with significantly less tumor budding (iCD68 p=0.0066; sCD68 p=0.0091) and absence of lymph node metastasis (sCD68 p=0.0286). Cell-to-cell contact of sCD68 and invading cancer cells was frequent and ameliorated the detrimental prognostic effect of the tumor budding phenotype. Subgroup analysis identified long-term survival with CD47 loss and predominance of CD163 M2 macrophages ( = 0.0366). CD163 macrophages represented 40% of the total population, and positively correlated with total CD68 macrophage numbers (r[CD68/CD163] = 0.32; = 0.0001). Strong CD163 infiltration predicted lower tumor grade ( = 0.0026) and less lymph node metastasis ( = 0.0056). This study provides direct morphological evidence of an interaction between TAM and infiltrating cancer cells. The prognostic impact of TAM is modulated by phenotype, microlocalization and the expression of anti-phagocytic markers in CRC.

The Role of Tumor-Associated Macrophages in Colorectal Carcinoma Progression

“…Surprisingly, heavy CD163 + M2 macrophage infiltration in CRC tumors was found in tumors of a lower grade and in patients with fewer lymph node metastases. A survival benefit was seen in patients with high CD163 + M2 macrophages, but not with iNOS + macrophages [11]. In addition, in a study with 158 CRC patients with liver metastases suggested that high TAMs infiltration in primary tumors is correlated with a better clinical outcome [12].…”

Carcinoma and Sarcoma Microenvironment at a Glance: Where We Are

“…In fact, the modulation of their activity seems to be localized within the tumor mass. TAMs at the tumor edges induce apoptosis in cancer cells, while TAMs localizing at the invasive front showed a resistant phenotype to suppressive TME (64,65). Recent studies highlight that TAM plasticity, modulated by different microenvironmental factors, has a prognostic impact on CRC patients (64,65).…”

“…TAMs at the tumor edges induce apoptosis in cancer cells, while TAMs localizing at the invasive front showed a resistant phenotype to suppressive TME (64,65). Recent studies highlight that TAM plasticity, modulated by different microenvironmental factors, has a prognostic impact on CRC patients (64,65). An integrative study, proposed by Mlecnik et al showed that among the heterogeneous group of CRC patients with a mismatch repair-deficient and microsatellite instability-high (MSI-H) phenotype, two groups can be distinguished: the most represented group with high T-cell activity and improved prognosis, and a minor group with reduced T-cell activity and poor prognosis (66).…”

Improving tumour budding evaluation in colon cancer by extending the assessment area in colectomy specimens

“…Although the use of H&E for TB counting is feasible in most cases, TB identification may be difficult in tumours with inflammatory infiltrate and/or striking fibrosis with reactive stromal cells. For these reasons, some authors favour the use of cytokeratin (CK) immunostaining to improve the visualisation of tumour buds and reproducibility of the assessment . Moreover, counting in such a limited area may result in overclassification of cases with focal TB activity that may not have the same aggressive potential as cases with diffuse and intense budding .…”

“…For these reasons, some authors favour the use of cytokeratin (CK) immunostaining to improve the visualisation of tumour buds and reproducibility of the assessment. [13][14][15] Moreover, counting in such a limited area may result in overclassification of cases with focal TB activity that may not have the same aggressive potential as cases with diffuse and intense budding. [16][17][18] In this study, we aimed to improve the accuracy and reproducibility of the recommended TB evaluation method by CK immunostaining, increasing the number of counted fields, and assessing cell proliferation, the apoptotic index and the emperipoletic index in tumour buds.…”