# Association of fibronectin 1 deregulation with tyrosine kinase inhibitor resistance in chronic myeloid leukemia

## TL;DR

This study shows that changes in fibronectin 1 (FN1) levels are linked to resistance to tyrosine kinase inhibitors in chronic myeloid leukemia patients.

## Contribution

The study identifies FN1 as a novel factor in tyrosine kinase inhibitor resistance and suggests it could be a new therapeutic target or biomarker.

## Key findings

- FN1 levels are significantly reduced in CML cell lines resistant to BCR::ABL1 inhibitors.
- Restoring FN1 expression in resistant cells re-sensitizes them to tyrosine kinase inhibitors.
- FN1 deregulation was also observed in peripheral blood cells from CML patients.

## Abstract

Therapy of chronic myeloid leukemia (CML) with tyrosine kinase inhibitors (TKIs) targeting the BCR::ABL1 kinase has become a paradigm for precision oncology. Despite the tremendous success of this strategy, with an overall long-term survival rate of 83%, approximately 25% of CML patients experience therapy failure within 5 years of treatment. TKI resistance is multifaceted, involving mutations in BCR::ABL1, but also BCR::ABL1-independent mechanisms. Among them, deregulation of cell adhesion and motility of CML cells has been observed in TKI-resistance. The extracellular matrix protein fibronectin 1 (FN1) has been shown to be deregulated in solid tumors promoting proliferation and metastasis. However, the role of FN1 in hematopoietic neoplasms remains to be fully elucidated. The aim of our study was to gain deeper insights into the role of FN1.

FN1 mRNA and protein levels were analyzed using qPCR and immunoblotting. Transfection was performed using nucleofection or stable transfection, followed by analyses of cell number, proliferation and viability. Cell adhesion was assessed using Matrigel-coated surfaces, and FN1 localization was analyzed using immunofluorescence.

FN1 levels were significantly downregulated in CML cell lines resistant against BCR::ABL1 inhibitors in vitro. SiRNA-mediated FN1 knockdown reduced the cell’s susceptibility to all generations of TKIs employed in treatment of CML, including asciminib. In contrast, the restoration of FN1 expression in TKI-resistant cells re-sensitized the cells to TKI treatment. This effect was also observed in K-562 cells that intrinsically harbor the BCR::ABL1 mutation p. E255K (−35.2%, p < 0.001), as well as in K-562 and Ba/F3 cells after stable transfection of the BCR::ABL1 wild-type or the p. T315I gatekeeper mutation. Clinically, deregulation of FN1 was also observed in peripheral blood cells derived from CML patients.

Our data indicate that FN1 may serve as a potential therapeutic target to address TKI resistance or as a suitable biomarker for the treatment.

Diagram explaining chronic myeloid leukemia (CML) progression and resistance to BCR::ABL1 tyrosine kinase inhibitors (TKIs), affecting 25% of patients. CML cells transition from TKI-sensitive to TKI-resistant. FN1 knockdown in sensitive cells reduces TKI sensitivity, while FN1 restoration in resistant cells improves sensitivity. The extracellular matrix's role is highlighted, with FN1's involvement. TKI non-responders are depicted.