Although third-generation Epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKI) is standard of care for patients with EGFR-mutant Non-small cell lung cancer (NSCLC), little is known about the predictors of response or resistance. Here, we integrated single-cell RNA (scRNA) sequencing, bulk RNA sequencing, multiplexed immunofluorescence and flow cytometry data from pretreatment and post-resistant tumor samples of EGFR-mutant NSCLC patients received third-generation EGFR-TKIs. We show that resistant samples had a markedly enriched CXCR1+ neutrophils infiltration (P < 0.01) than pretreatment samples, which were distinguished from other subtypes of neutrophils and displayed immunosupressive characteristics. Spatial analysis showed that increased CXCR1+ neutrophils predominantly infiltrated into the tumor core in resistant samples and the average distance of neutrophils to tumor cells markedly reduced from 33 to 19 μm. Deep analysis of scRNA and bulk RNA sequencing data revealed the increased interactions between CXCR1+ neutrophils and tumor cells and activated TNF-α/NF-κB signaling pathway in tumor cells of resistant samples. In vitro and in vivo experiments validated that CXCR1+ neutrophils resulted in resistance to third-generation EGFR-TKI via activating TNF-α/NF-κB signaling pathway in tumor cells. Importantly, patients with low pretreatment CXCR1+ neutrophil infiltration abundance had a dramatically longer progression-free survival (11.8 vs. 7.5 months; P = 0.019) and overall survival (33.0 vs. 23.5 months; P = 0.029) than those with high infiltration abundance. Collectively, these findings suggest that CXCR1+ neutrophils infiltration was associated with the efficacy of third-generation EGFR-TKI in patients with EGFR-mutant NSCLC.
© 2024. The Author(s).

Background IL4, IL5, IL13, and IL17-producing CD4 T helper 2 (Th2)-cells and IL17-producing CD4 T helper 17 (Th17)-cells contribute to chronic eosinophilic and neutrophilic airway inflammation in asthma and allergic airway inflammation. Chemokines and their receptors are upregulated in Th2/Th17-mediated inflammation. However, the ability of CXCR1 and CXCR2 modulate Th2 and Th17-cell-mediated allergic lung inflammation has not been reported. Methods Mice sensitized and challenged with cat dander extract (CDE) mount a vigorous Th2-Th17-mediated allergic lung inflammation. Allosteric inhibitor of CXCR1 and CXCR2, ladarixin was orally administered in this model. The ability of ladarixin to modulate allergen-challenge induced recruitment of CXCR1 and CXCR2-expressing Th2 and Th17-cells and allergic lung inflammation were examined. Results Allergen challenge in sensitized mice increased mRNA expression levels of Il4, Il5, Il13, Il6, Il1β, Tgfβ1, Il17, Il23, Gata3, and Rorc , and induced allergic lung inflammation characterized by recruitment of CXCR1- and CXCR2-expressing Th2-cells, Th17-cells, neutrophils, and eosinophils. Allosteric inhibition of CXCR1 and CXCR2 vigorously blocked each of these pro-inflammatory effects of allergen challenge. CXCL chemokines induced a CXCR1 and CXCR2-dependent proliferation of IL4, IL5, IL13, and IL17 expressing T-cells. Conclusion Allosteric inhibition of CXCR1 and CXCR2 abrogates blocks recruitment of CXCR1- and CXCR2-expressing Th2-cells, Th17-cells, neutrophils, and eosinophils in this mouse model of allergic lung inflammation. We suggest that the ability of allosteric inhibition of CXCR1 and CXCR2 to abrogate Th2 and Th17-mediated allergic inflammation should be investigated in humans.

h4>Background: /h4> Although targeted therapies usually trigger great initial responses in patients, the efficacy is transient due to tumor metastasis. The formation of pre-metastatic niche was proposed as the main cause for metastasis, of which the blocking way may be a potential method for inhibiting metastasis. Sijunzi Tang (SJZ), as a complementary drug for targeted therapy, can reduce the recurrence and metastasis of tumors and prolong the survival time of patients. However, how SJZ regulates the formation of pre-metastatic niche to improve the efficacy of targeted therapy remains unclear. h4>Methods: /h4>: Here, we investigated the anti-tumor activity and immunological mechanism of SJZ plus gefitinib based on pre-metastatic niche in Lewis lung carcinoma (LLC) incubated spontaneous metastatic mouse model, using histopathology and immunological methods. h4>Results: /h4>: The results showed that SJZ can improve the effect of gefitinib by inhibiting tumor cell growth, promoting tumor cell apoptosis and preventing metastasis in the lung. Besides, SJZ plus gefitinib could inhibit tumor cell aggregation and the expression of characteristic proteins of Mmp2 and Mmp9 in the lung areas of mice. We also confirmed that SJZ could downregulate the expression levels of c-kit and VEGFR2 on DCs, c-kit on neutrophils, c-kit, VEGFR2 on B lymphocyte in the blood, and c-kit and CXCR1 on monocytes in the lung; Gefitinib could decrease the expression levels of c-kit and VEGFR2 on DCs in the blood and c-kit and CXCR1 on monocytes in the lung, but increase the amount of c-kit + monocytes in the blood. SJZ plus gefitinib decrease the proportion of c-kit + and CXCR1 + monocytes in the lung. SJZ could regulate pro-metastatic inflammatory responses represented by down-regulating the expression of IL-23，RANTES, GRO-α against that of gefitinib. Moreover, there were significant rises in the expression level of IL-12p70 and IL-15，while declines in the expression level of IL-1β、IL-18、GRO-α in co-treatment group. h4>Conclusions: /h4>: This work identified the immune cells and cytokines in pre-metastatic niche associated with lung cancer affected by gefitinib and SJZ, and further revealed the immunological mechanism of SJZ improving the efficacy of gefitinib.