Role of Combined Anti-PD-1 and Anti-CTLA-4 Therapy in Melanoma Immune Response

Background

Immune checkpoint inhibitors have made significant progress in clinical oncology, especially anti-PD-1 (anti-programmed death protein 1) and anti-CTLA-4 (anti-cytotoxic T-lymphocyte-associated antigen 4) therapies, which have been proven to induce long-term remission effects in various cancers, including melanoma. However, the combination therapy of anti-PD-1 and anti-CTLA-4 (combined therapy) is more effective than single agents. Previous studies have shown that this combination therapy improves the five-year survival rate of melanoma patients to 52%, compared to 44% and 26% for anti-PD-1 or anti-CTLA-4 used alone, respectively. Nevertheless, the specific mechanisms of these therapies in humans, especially the dynamics of the immune response induced by combination therapy, have yet to be fully elucidated.

Study Summary

This paper was completed by a research team from the Perelman School of Medicine at the University of Pennsylvania, Immunai, and the Grossman School of Medicine at New York University, published in the 2024 issue of Cancer Cell (Wang et al., 2024). The study conducted a longitudinal analysis of 36 stage IV melanoma patients using single-cell RNA sequencing and T-cell receptor (TCR) sequencing techniques to investigate how anti-CTLA-4 and anti-PD-1 combination therapy elicits T-cell clonal responses within patients, particularly the changes in exhausted CD8+ T cells (Tex) at different time points. Additionally, the research team developed a new algorithm named Cyclone to analyze the dynamic trajectories of clones.

Research Methods

The study collected peripheral blood samples from patients before and during different stages of treatment to analyze the clonal changes of immune cells. The important steps of the study are as follows:

1. Sample Collection: Samples from 36 stage IV melanoma patients, including those receiving anti-PD-1, anti-CTLA-4, and combination treatments.
2. Experimental Procedure: Utilized single-cell RNA and TCR sequencing methods to trace immune responses at 0, 3, 6, and 9 weeks post-treatment.
3. Data Analysis: The research team developed the Cyclone algorithm, treating each clone as a time series, to identify clonal expansion and contraction trajectories under different treatment regimens.
4. Clone Grouping: Using the Cyclone algorithm, the research team defined six clonal response patterns, each exhibiting specific expansion and contraction time points post-treatment (e.g., 3rd week, 6th week, and 9th week).

Main Findings

The study revealed the significant impact of combination therapy on immune cells, analyzed as follows:

Temporal Dynamics of Clonal Fluctuations

The study demonstrated that anti-PD-1 and anti-CTLA-4 combination therapy induced larger-scale clonal responses at 6 weeks and 9 weeks, respectively, showing a stronger immune response compared to single agents. This combination therapy induced specific melanoma CD8+ T cell and exhausted Tex clonal expansion. Anti-CTLA-4 alone primarily triggered early T-cell proliferation and expansion with relatively smaller changes. Moreover, different treatments caused immune clonal fluctuations at different time points, exhibiting clonal expansion and contraction phenomena post-treatment (e.g., 3rd, 6th, and 9th weeks).

Characteristics of Tex Subgroups

The study found that the proportion of Tex in peripheral blood was low but differentiated into several subtypes such as progenitor (Progex), intermediate or NK-like cells, and terminally exhausted cells (Terminal Tex), each displaying different expansion and differentiation characteristics post-treatment. Anti-CTLA-4 effectively promoted Progex subgroup expansion, while anti-PD-1 facilitated the transition of Progex into more differentiated terminal Tex. During combination therapy, these therapies worked synergistically, causing Tex responses to peak at 3 and 6 weeks. The study also noted that Tex in peripheral blood predominantly exhibited characteristics of Progex and intermediate/NK-like Tex compared to those in tumor tissues.

Melanoma-Specific Immune Response

The study used specific antigen tetramer staining to detect melanoma-specific CD8+ T cells, showing significant expansion of melanoma-specific CD8+ T cells at 3 and 6 weeks of combination therapy, while virus-specific (e.g., EB virus and influenza virus) CD8+ T cells expanded significantly after 9 weeks. This phenomenon suggests that the early immune response may be related to melanoma-specific antigens.

Immune Checkpoint Inhibition and Tex Reactivation

The study found that anti-CTLA-4 alone could activate the expansion of Progex, while anti-PD-1 mainly acted on the reactivation of Tex, pushing it towards terminal Tex differentiation over time. The combination therapy achieved significant early Progex proliferation, followed later by the enhanced differentiation into more effective Tex driven by anti-PD-1.

Significance of the Study

This research is the first to reveal the Tex clonal fluctuations induced by anti-PD-1 and anti-CTLA-4 combination therapy in melanoma patients, presenting the dynamic changes of immune cell clones through the new Cyclone algorithm. The main values of the study are highlighted as follows:

1. Revealing the Immune Mechanism of Combination Therapy: The study elucidated the temporal dynamics of Tex responses induced by anti-PD-1 and anti-CTLA-4 combination therapy in humans, providing foundational data for more effective combined immune strategies in clinical settings.
2. Predicting Treatment Timing and Effectiveness: Understanding the clonal dynamics and time points of Tex provides guidance for optimizing drug administration and treatment planning, aiding in the improvement of clinical efficacy of immunotherapy.
3. Improving Immunotherapy: By clarifying the functional effects of different immune checkpoint inhibitors, the study aids in developing more refined combination therapies, enhancing cancer treatment effects, and reducing adverse reactions.

Study Highlights

1. Development of New Algorithm Cyclone: This algorithm identifies clonal expansion and contraction patterns under different treatment regimens through time series analysis, providing new methods for clinical research in cancer immunotherapy.
2. Revealing Tex Clonal Fluctuation Characteristics: The first observation of Tex clonal fluctuations in human samples, with significant early Tex expansion induced by combination therapy.
3. Expanded Understanding of Immunotherapy: The study emphasizes the advantages of combined use of anti-CTLA-4 and anti-PD-1 in immunotherapy, offering important basis for the optimization of future anticancer immunotherapies.

Study Limitations and Future Prospects

Despite encouraging results, the study also has some limitations. The sample size is relatively small, and some patients could not provide adequate samples for longitudinal analysis. Furthermore, the lack of paired analysis of pre- and post-treatment tumor tissue samples restricts a comprehensive understanding of Tex clonal dynamics within tumors. Future research can further validate these findings through larger-scale clinical trials and paired analysis with tumor samples.

The study by Wang et al. (2024) made important progress in understanding the efficacy of immune checkpoint inhibition and clonal response mechanisms in melanoma, providing innovative ideas and technologies for future cancer immunotherapies.