Ectopic NMDAR expression in cancer unmasks germline-encoded autoimmunity | Nature

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Subjects

- Cancer

- Cryoelectron microscopy

- Ion channels in the nervous system

- Neurotoxicity syndromes

Abstract

Autoimmunity and anti-cancer immunity lie on the same biological continuum1,2, but their link remains obscure. The paraneoplastic neurological syndrome ANRE (anti-NMDA receptor (NMDAR) encephalitis) is a paradigm for their connectivity3, given that intratumoural NMDAR expression is correlated with the generation of anti-NMDAR antibodies4,5. Here we verify ectopic expression of GluN1 and GluN2B NMDAR subunits in triple-negative breast cancer (TNBC)6 and model this using orthotopic TNBC tumours with inducible expression of GluN1–GluN2B NMDARs. We show that NMDAR expression is sufficient to induce the recruitment of B cells and their affinity maturation, consistent with an integrated adaptive immune response. Reconstruction of extended intratumoural B cell phylogenies and cryogenic electron microscopy structural analyses demonstrate that affinity-matured hypermutated and class-switched antibodies emerged from pre-existing germline-configuration lower-affinity anti-NMDAR antibodies. Distinct matured antibodies targeted specific epitopes and induced conformational rearrangements within the NMDAR amino-terminal domain, predictive of their functional effects, ranging from inhibition to potentiation. Passive transfer of an NMDAR-potentiating antibody caused autonomic dysregulation and lowered the seizure threshold in healthy female mice, recapitulating key diagnostic criteria of ANRE4,5. We further identify a correlation between intratumoural NMDAR expression and anti-NMDAR antibody titres in patients with TNBC. Taken together, our data establish a direct connection between intratumoural NMDAR expression, antibody maturation and the onset of autoimmunity. These findings suggest that germline-encoded anti-NMDAR antibodies contribute to immune surveillance but can also trigger autoimmune disease after maturation, revealing a mechanistic trade-off between cancer immunity and neurotoxicity.

Main

The adaptive immune system evolved to distinguish between self and non-self7. Nevertheless, immune responses against the self (that is, autoimmunity) occur, but their origin remains unclear8. In paraneoplastic syndromes, ectopic intratumoural expression of non-mutant self-proteins triggers antibody-mediated autoimmune disease9. These autoantigens typically have restricted expression in immunologically privileged sites, such as the central nervous system4,10 (onconeural antigens)11. Germline-configuration antibodies12 with low-affinity self-binding have been described13,14,15 but not traced to disease-causing high-affinity antibodies.

Paraneoplastic autoantibodies can modulate the host, for example, by altering neuronal function, and the tumour, for example, by restraining tumour growth and metastasis16. In fact, spontaneous tumour regression has even been observed after the onset of autoimmunity17. It is accepted that both antibody- and T-cell-mediated immunity synergize to drive immune control of tumours18. Therapies that activate T-cell-mediated immunity can precipitate considerable antibody-mediated neuronal autoimmunity19 that correlates with improved anti-tumour efficacy20.

These observations suggest ectopic onconeural antigen expression recruits germline-configuration B cells, which undergo affinity maturation in tertiary lymphoid structures21 to generate high-affinity IgG autoantibodies22 that cross the blood–brain barrier (BBB). However, whether onconeural autoantibodies are causative of autoimmunity remains unclear23. For example, ANRE is characterized by autoantibodies targeting the NMDAR and a stereotypical clinical syndrome3,4,5. NMDARs are voltage-dependent cation channels activated by glycine and glutamate24. They comprise two GluN1 subunits (encoded by GRIN1) and two GluN2 subunits (GRIN2A–GRIN2D) and/or GluN3 subunits (GRIN3A–GRIN3B)24,25 and mediate synaptic plasticity in the central nervous system26. ANRE is correlated with the expression of NMDAR in multiple tumour types, including ovarian, breast and pancreas5, and is associated with the production of high-affinity anti-NMDAR IgG antibodies15 from tertiary lymphoid structures22. The anti-NMDAR humoral response constrains tumour growth4,27 but is also associated with a debilitating neuropsychiatric syndrome characterized by psychosis, autonomic dysregulation (hyperthermia) and epileptic seizures3,4,5. Here using a mouse model of NMDAR-expressing cancer, we define how germline-encoded NMDAR-binding antibodies undergo affinity maturation to modulate receptor function, thereby constraining tumour growth and engendering neurological disease.

GluN1–GluN2B expression in TNBC

NMDAR expression has been detected in human breast cancer and is enriched in TNBC6. We reanalysed RNA-sequencing (RNA-seq) data from the METABRIC and TCGA breast cancer cohorts, and identified a subset of tumours that express both GRIN1 and GRIN2B (Extend