Background Small cell lung cancer (SCLC) is an aggressive malignancy harboring a dismal 5-year survival rate of less than 10%. In 2019, the addition of an anti-PDL1 antibody to initial platinum-etoposide chemotherapy revolutionized the standard of care after decades of ineffective chemoradiation. Nevertheless, the observed increase in patient survival with immune checkpoint blockade (ICB) remains modest, highlighting the need for further improvements in immunotherapy for this disease. Recently, there has been a growing interest in exploring and targeting non-canonical cancer immune checkpoints. One promising target is hypersialylation, a glycosylation signature associated with poorer patient outcomes in various malignancies. Studies have shown that certain cancers can exploit sialylation as a mechanism of immune evasion through interactions with the SIGLEC family of inhibitory receptors on immune cells. In SCLC, hypersialylation and SIGLEC signaling remain entirely unstudied. We hypothesize that SCLC utilizes sialic acid (SA) for immune evasion and that targeting this axis could serve as a novel glycoimmune checkpoint therapy in treating SCLC (figure 1).

Methods Using CRISPR editing technology, we generated several mouse SCLC knockout (KO) cell lines in Gne and Cmas, two genes encoding enzymes involved in the endogenous SA biosynthesis pathway (figure 2). Validation was performed using western blot and lectin staining. In vitro, cells were stimulated with IFNy for 48 hours and accessed for MHC-I expression by flow cytometry. For in vivo experiments, cells were propagated as flank tumors into NSG and immunocompetent syngeneic C57BL/6 mice recipients, and kinetics were measured by caliper. Tumor immune infiltration was quantified by immunohistochemistry (IHC) at endpoint.

Results Loss of either gene significantly decreased surface sialylation in all KO lines. Desialylated cells exhibited higher MHC-I expression with IFNy stimulation as compared to controls, suggesting that loss of SA leads to increased antigen presentation and immunogenicity. These results are significant as recent clinical data revealed SCLC patients with lower MHC-I expression respond weaker to classical ICB therapy. In vivo, we observed slower tumor kinetics with the KO lines in the syngeneic flank models. However, kinetic differences were not seen in the NSG model, suggesting that the anti-tumoral effects of desialylation are immune dependent. On IHC, we detected increased infiltration of CD8+ T cells and macrophages in the KO tumors.

Conclusions Our data suggests that the loss of sialylation in SCLC promotes pro-immune and anti-tumoral effects. Ultimately, our work enhances the understanding of immune evasion mechanisms in SCLC and unveils a novel immunotherapeutic approach for treating SCLC.

Ethics Approval Animal experiments were performed at the Fred Hutchinson Cancer Research Center with approval from the Institutional Animal Care and Use Committee (IACUC).

• Download figure
• Open in new tab
• Download powerpoint

Abstract 495 Figure 1

Illustration of the sialic acid - SIGLEC immune axis and its similarities to other known canonical cancer immune checkpoints

• Download figure
• Open in new tab
• Download powerpoint

Abstract 495 Figure 2

Overall schematic of the experimental approaches employed to study sialylation using mouse models of small cell lung cancer