Immune Response to Immunotherapy in Lung Cancer: Study of Sputum and Blood Samples

Lung cancer is the leading cause of cancer death in France and worldwide. In 2022, it affected nearly 2.48 million people and caused 1.8 million deaths. Despite therapeutic advances, it remains a major public health challenge due to often late diagnosis and a high mortality rate. For the past decade, the discovery of immune checkpoint inhibitors has revolutionized treatments, particularly with the advent of therapeutic antibodies (Abs) targeting PD-1 and PD-L1 proteins, such as nivolumab, pembrolizumab, and atezolizumab. These immune checkpoint inhibitors (ICIs) have improved the management of certain patients by reactivating T lymphocytes. Initially used as second-line therapy for metastatic non-small cell lung cancer (NSCLC), ICIs have gradually been integrated into first-line treatment for less advanced stages. They are currently indicated as monotherapy or in combination with chemotherapy in several contexts: metastatic disease, postoperative adjuvant therapy, or consolidation therapy after chemoradiotherapy. However, only a small percentage of patients actually benefit from these therapies, and their side effects can be severe.

Current research aims to improve the response to immune checkpoint inhibitors (ICIs) by combining them with chemotherapy, radiotherapy, or other immunotherapies targeting new checkpoints (anti-TROP2, anti-TIGIT). However, reliable clinical criteria for identifying responding patients are still lacking. PD-L1 expression appears to be a potential marker, but its efficacy remains variable across studies. Numerous other biomarkers have been explored, such as mutational burden, tumor infiltration by T lymphocytes, and certain blood parameters, but none has yet provided reliable prediction. The lung tumor microenvironment plays a key role in progression and treatment resistance. Composed of tumor, stromal, and immune cells, it promotes immunosuppression and limits the effectiveness of treatments by activating resistance mechanisms. It therefore represents a promising therapeutic target for improving the response to immune checkpoint inhibitors (ICIs).

Beyond the local pulmonary response, the blood compartment represents a valuable source of information for studying the systemic response to immunotherapy. Indeed, peripheral blood reflects, at least in part, the patient's overall immune status and allows for the dynamic monitoring of changes induced by ICIs during treatment. Several studies have shown that certain circulating cell populations can be correlated with clinical response or the occurrence of immuno-induced toxicities.

For example, an early increase in activated CD8+ T lymphocytes (HLA-DR+, CD38+) in the blood has been associated with a better response to treatment in patients with melanoma or non-small cell lung cancer (NSCLC). Similarly, the readily available neutrophil-to-lymphocyte ratio (NLR) has been identified as an adverse prognostic biomarker for overall survival and progression-free survival in patients undergoing immunotherapy.

The correlation between phenotypic changes in immune cells in blood and sputum could thus reveal immune signatures predictive of response or resistance.

Lung cancer is the leading cause of cancer death in France and worldwide. In 2022, it affected nearly 2.48 million people and caused 1.8 million deaths. Despite therapeutic advances, it remains a major public health challenge due to often late diagnosis and a high mortality rate. For the past decade, the discovery of immune checkpoint inhibitors has revolutionized treatments, particularly with the advent of therapeutic antibodies (Abs) targeting PD-1 and PD-L1 proteins, such as nivolumab, pembrolizumab, and atezolizumab. These immune checkpoint inhibitors (ICIs) have improved the management of certain patients by reactivating T lymphocytes. Initially used as second-line therapy for metastatic non-small cell lung cancer (NSCLC), ICIs have gradually been integrated into first-line treatment for less advanced stages. They are currently indicated as monotherapy or in combination with chemotherapy in several contexts: metastatic disease, postoperative adjuvant therapy, or consolidation therapy after chemoradiotherapy. However, only a small percentage of patients actually benefit from these therapies, and their side effects can be severe.

Current research aims to improve the response to immune checkpoint inhibitors (ICIs) by combining them with chemotherapy, radiotherapy, or other immunotherapies targeting new checkpoints (anti-TROP2, anti-TIGIT). However, reliable clinical criteria for identifying responding patients are still lacking. PD-L1 expression appears to be a potential marker, but its efficacy remains variable across studies. Numerous other biomarkers have been explored, such as mutational burden, tumor infiltration by T lymphocytes, and certain blood parameters, but none has yet provided reliable prediction. The lung tumor microenvironment plays a key role in progression and treatment resistance. Composed of tumor, stromal, and immune cells, it promotes immunosuppression and limits the effectiveness of treatments by activating resistance mechanisms. It therefore represents a promising therapeutic target for improving the response to immune checkpoint inhibitors (ICIs).

Beyond the local pulmonary response, the blood compartment represents a valuable source of information for studying the systemic response to immunotherapy. Indeed, peripheral blood reflects, at least in part, the patient's overall immune status and allows for the dynamic monitoring of changes induced by ICIs during treatment. Several studies have shown that certain circulating cell populations can be correlated with clinical response or the occurrence of immuno-induced toxicities.

For example, an early increase in activated CD8+ T lymphocytes (HLA-DR+, CD38+) in the blood has been associated with a better response to treatment in patients with melanoma or non-small cell lung cancer (NSCLC). Similarly, the readily available neutrophil-to-lymphocyte ratio (NLR) has been identified as an adverse prognostic biomarker for overall survival and progression-free survival in patients undergoing immunotherapy.

The correlation between phenotypic changes in immune cells in blood and sputum could thus reveal immune signatures predictive of response or resistance.

The RICEPS study, conducted at the University Hospital of Tours and within INSERM U1100 from 2021 to 2023, analyzed the blood and lung immune response in 19 patients receiving pembrolizumab, collecting blood and sputum samples. It demonstrated the presence of specific non-circulating lymphocytes, resident memory T cells (RMCs), as well as key cells involved in the immune response to immune checkpoint inhibitors (ICIs), CD8+ T cells, in patient sputum, with longitudinal follow-up. Preliminary results from this study showed that, in blood, independently of treatment, patient, and time, CD8+ T cells, monocytes, B cells, and several cytokines (GCSF, interleukin-4 (IL-4), IL-6, IL-7, IL-12, IL-17, IL-33) influence treatment response. In sputum, TRM cells and two cytokines (IL-7 and CCL2) appear to contribute to treatment response (unpublished results).

Based on these results, the RICEPS-2 study aims to include a larger number of patients treated with immunotherapy, regardless of the type of NSCLC. It will analyze different cell populations in sputum and blood to better understand the involvement of the lung mucosa and the systemic immune response in the response to ICIs. It will serve as a validation cohort for the first RICEPS cohort. Longitudinal monitoring of these populations at several treatment time points (M0, M3, M6) will allow us to...