CD271 activation prevents low to high-risk progression of cutaneous squamous cell carcinoma and improves therapy outcomes

Abstract

Background Cutaneous squamous cell carcinoma (cSCC) is the second most prevalent form of skin cancer, showing a rapid increasing incidence worldwide. Although most cSCC can be cured by surgery, a sizeable number of cases are diagnosed at advanced stages, with local invasion and distant metastatic lesions. In the skin, neurotrophins (NTs) and their receptors (CD271 and Trk) form a complex network regulating epidermal homeostasis. Recently, several works suggested a significant implication of NT receptors in cancer. However, CD271 functions in epithelial tumors are controversial and its precise role in cSCC is still to be defined. Methods Spheroids from cSCC patients with low-risk (In situ or Well-Differentiated cSCC) or high-risk tumors (Moderately/Poorly Differentiated cSCC), were established to explore histological features, proliferation, invasion abilities, and molecular pathways modulated in response to CD271 overexpression or activation in vitro. The effect of CD271 activities on the response to therapeutics was also investigated. The impact on the metastatic process and inflammation was explored in vivo and in vitro, by using zebrafish xenograft and 2D/3D models. Results Our data proved that CD271 is upregulated in Well-Differentiated tumors as compared to the more aggressive Moderately/Poorly Differentiated cSCC, both in vivo and in vitro. We demonstrated that CD271 activities reduce proliferation and malignancy marker expression in patient-derived cSCC spheroids at each tumor grade, by increasing neoplastic cell differentiation. CD271 overexpression significantly increases cSCC spheroid mass density, while it reduces their weight and diameter, and promotes a major fold-enrichment in differentiation and keratinization genes. Moreover, both CD271 overexpression and activation decrease cSCC cell invasiveness in vitro. A significant inhibition of the metastatic process by CD271 was observed in a newly established zebrafish cSCC model. We found that the recruitment of leucocytes by CD271-overexpressing cells directly correlates with tumor killing and this finding was further highlighted by monocyte infiltration in a THP-1-SCC13 3D model. Finally, CD271 activity synergizes with Trk receptor inhibition, by reducing spheroid viability, and significantly improves the outcome of photodynamic therapy (PTD) or chemotherapy in spheroids and zebrafish. Conclusion Our study provides evidence that CD271 could prevent the switch between low to high-risk cSCC tumors. Because CD271 contributes to maintaining active differentiative paths and favors the response to therapies, it might be a promising target for future pharmaceutical development.

Ex vivo therapeutic screening of metastatic cSCC: A review of methodological considerations for clinical implementation

“…Such cell lines have also been translated to in vivo models of both mice and zebrafish to model the metastatic process. Furthermore, freshly dissociated primary cSCC tumours have been cultured short‐term as ex vivo spheroids using the liquid overlay method to evaluate both drug response and CD271 overexpression 19 . Whilst primary cSCC models exist, the relatively poorer prognosis of metastatic disease beckons closer investigation.…”

The Role of the Neurotrophin Network in Skin Squamous Cell Cancer and the Novel Use of the Zebrafish System

“…Recently, a stage-specific localization and an initial functional evaluation of NT receptors in cSCC have been elucidated ( Dallaglio et al, 2014 ; Palazzo et al, 2019 ; Quadri et al, 2023 ). A summary of the most important work reporting the expression and function of NT and their receptors in cSCC is displayed in Table 2 .…”

“…A critical analysis of the expression of the NT receptors during the transition from actinic keratosis to cSCC development and progression has been given by Quadri et al (2023) . Their findings proved that CD271 is upregulated in well-differentiated cSCC compared with that in the more aggressive moderately/poorly differentiated cSCC, whereas the expression of TRKA directly correlates with the level of malignancy.…”

“…However, given the difficulty found in generating neoplastic transformation in zebrafish KCs, most of the cSCC research takes advantage of the xenografting procedure, which allows to obtain information on cell growth and invasive capacity within a week ( Martins et al, 2016 ; Paulson et al, 1990 ; Quadri et al, 2023 , 2022b ). Moreover, the use of the fluorescent transgenic reporter zebrafish strains significantly increases the power of the information obtainable related to the interaction between cancer cells and host, such as angiogenesis and immune response, which remain the major hallmarks of aggressive tumors ( Codolo et al, 2022 ; Pan et al, 2022 ; Quadri et al, 2022a ; Roth et al, 2021 ; Wang et al, 2024 ; Zhao et al, 2024 ).…”

“…Recently, Quadri et al (2023) reported for the first time the use of zebrafish to elucidate the role of the common NT receptor CD271 in cSCC. Their work revealed the role of CD271 in inhibiting the aggressiveness of cSCC in vivo.…”