Innate Immune Remodeling Drives Therapy Resistance via Macrophage-NK Cell Crosstalk.

The dynamic evolution of the immune tumor microenvironment (TME) during targeted therapy is a critical yet poorly understood determinant of treatment response and resistance. While most studies compare immune states before and after treatment, temporal immune changes during therapy remain largely uncharacterized, limiting development of effective combination strategies. Here, we investigated immune dynamics throughout targeted therapy using mouse melanoma models that recapitulate human therapeutic responses. Single-cell RNA sequencing (scRNA-seq) identified a previously unrecognized inflection point where the inflamed TME during tumor regression, characterized by robust NK cell infiltration, transitions to an immune-excluded state upon onset of drug-tolerant residual disease. We uncovered a unique macrophage subset (F4/80hiCCL5MHCIICD63) that orchestrates NK cell recruitment through CCR2/5 signaling during regression. Depletion of these macrophages using LysM-cre;iDTR mice significantly reduced NK cell infiltration. Specifically during residual disease, pharmacological inhibition of Ptpn22, a phosphatase that negatively regulates immune activation, reprogrammed macrophages, restored NK cell recruitment and enhanced therapeutic efficacy. Extending these findings to human cancer, longitudinal scRNA-seq analysis of melanoma and lung cancer patient samples revealed dynamic NK cell infiltration during targeted therapy, establishing a direct link between innate immune remodeling and treatment outcome. Unlike prior prognostic studies assessing immune states at single time points, our results provide mechanistic evidence of a temporal relationship between NK cell infiltration and therapeutic efficacy. Together, these findings position immune evolution as a driver of acquired resistance and identify macrophage-NK cell crosstalk as a therapeutically actionable axis to overcome immune exclusion and improve targeted therapy across multiple cancer types. One Sentence SummaryCancer therapy resistance emerges from a dynamic evolution of the tumor microenvironment, characterized by macrophage-driven NK cell infiltration during initial tumor regression, followed by exclusion of NK cells during residual disease, highlighting macrophage-NK cell interactions as a promising therapeutic target to improve clinical outcomes.