1148 A natural antibody response associated with improved survival of SCLC patients can be actively induced in a SCLC mouse model and leads to significantly improved survival

<h3>Background</h3> Small cell lung cancer (SCLC) has a 5-year survival of 8%, killing over 20,000 Americans annually. New therapies are urgently needed. ~15% of SCLC patients exhibit a naturally occurring ‘anti-Hu’ antibody response against ELAVL4 and show improved response to therapy and significantly better survival.^{1 2} Rare SCLC patients develop a severe and sometimes lethal autoimmune response against ELAVL4 and can show complete SCLC regression. We determined that the antibody initiating antigenic epitope consists of ELAVL4 that is isoaspartylated in its unstructured N-terminal region (aa 1-38).³ Isoaspartylation is an immunogenic type of protein damage that is normally repaired but appears to be abnormally present in ELAVL4 in SCLC.³ ELAVL4 constitutes a cancer-specific neo-antigen that is expressed on the outside of SCLC cells.⁴ In rare cases this antibody response can lead to autoimmunity through epitope spreading. <h3>Methods</h3> Here we tested whether actively inducing an immune response against recombinant isoaspartylated ELAVL4 (isoAsp-ELAVL4) in a mouse SCLC model improves survival. We used a <i>Trp53</i>^fl/fl; <i>Rb1</i>^fl/fl inducible SCLC mouse model⁵ to examine: 1) Whether immunization with isoAsp-ELAVL4 prior to SCLC induction is protective, and 2) Whether immunization with isoAsp-ELAVL4 following completion of cisplatin+etoposide therapy increases survival. Mice were immunized with a recombinant N-terminal fragment of ELAVL4 (isoAsp-ELAVL4, amino acids 1-117) generated in <i>Clear Coli</i> and incubated under isoaspartyl-inducing conditions (‘aged’ at 37°C for 7 days), then emulsified in incomplete Freund’s adjuvant (IFA). The negative control was phosphate-buffered saline in IFA; recombinant ELAVL4 naturally undergoes isoaspartylation even after short periods under physiological conditions, preventing its use as a negative control. The trajectories of mice were closely monitored with a detailed body metric score and serum was collected from bi-weekly blood draws to monitor the anti-ELAVL4 antibody response. <h3>Results</h3> ELISA data showed that all isoAsp-ELAVL4-immunized animals became immune responsive against isosp-ELAVL4, and just as in human SCLC patients, a small fraction of control-immunized animals spontaneously developed anti-ELAVL4 reactivity. Immunization alone before SCLC induction in the absence of chemotherapy did not improve survival (figure 1A). In contrast, immunization following cisplatin+etoposide therapy significantly increased survival (p = 0.00088) (figure 1B). <h3>Conclusions</h3> An immune response against isoAAsp-ELAVL4 can be actively induced, and when given after chemotherapy can significantly improve SCLC survival in a mouse model. We are leveraging these observations for the development of a new SCLC therapy. <h3>Acknowledgements</h3> This work is supported by a Wright Foundation Transformative Cancer Grant Award (IAO), the Norris Comprehensive Cancer Center Core Grant (NIH/NCI P30CA014089), the USC Provost’s Undergraduate Research Award (HL and SE); the Bravo Medical Magnet High School STAR/EHA program (JV), and the Walter A. Richter Cancer Research Chair (WMK). Diego is a Grad+ trainee of CaRE2, the Cancer Research Education and Engagement Health Equity Center, which is supported by NIH grants U54CA233396, U54CA233444, and U54233465. The figure was generated using Biorender. <h3>References</h3> Graus F, Dalmou J, Reñé R, <i>et al</i>. Anti-Hu antibodies in patients with small-cell lung cancer: association with complete response to therapy and improved survival. <i>JCO.</i> 1997;<b>15</b>(8):2866–2872. Gozzard P, Chapman C, Vincent A, Lang B, Maddison P. Novel humoral prognostic markers in small-cell lung carcinoma: a prospective study. Zhang L, ed. <i>PLoS ONE</i>. 2015;<b>10</b>(11). Pulido MA, DerHartunian MK, Qin Z, <i>et al</i>. Isoaspartylation appears to trigger small cell lung cancer-associated autoimmunity against neuronal protein ELAVL4. <i>Journal of Neuroimmunology</i>. 2016;<b>299</b>:70–78. Tora M, Graus F, de Bolos C. Cell surface expression of paraneoplastic encephalomyelitis/sensory neuronopathy- associated Hu antigens in small-cell lung cancers and neuroblastornas. <i>American Academy of Neurology</i>. <b>1997</b>:734–741. Meuwissen R, Linn SC, Linnoila RI, Zevenhoven J, Mooi WJ, Berns A. Induction of small cell lung cancer by somatic inactivation of both Trp53 and Rb1 in a conditional mouse model. <i>Cancer Cell</i>. 2003;<b>4</b>(3):181–9. <h3>Ethics Approval</h3> This study was approved by the University of Southern California’s Institutional Animal Care and Use Committee, protocol number 20105.