Nicotinic Acetylcholine Receptors of Muscle and Neuronal (α₇) Types Coexpressed in a Small Cell Lung Carcinoma

Abstract

SCC‐37 is a small cell lung carcinoma line that aberrantly expresses muscle‐type nicotinic acetylcholine receptors (nAChRs). It was established from a patient with a paraneoplastic autoimmune neuromuscular disorder, myasthenia gravis. When grown as a xenograft tumor, SCC‐37 cells express plasma membrane receptors that bind 125I‐labeled α‐bungarotoxin (125I‐α‐BTx), cosediment with 9S nAChR pentamers, and bind to a monoclonal antibody (MAb 35) specific for muscle‐type (α1 subunit) α‐BTx receptors. The agonist carbamylcholine (carbachol) stimulates influx of 22Na+ in SCC‐37 cells; this is inhibited by α‐BTx and by d‐tubocurarine. Long‐term cultured SCC‐37 cells have functional and ligand‐binding evidence for surface coexpression of both α1 and neuronal‐type (α7 subunit) α‐BTx receptors. A subclone of SCC‐37, designated SCC‐A9, expresses only the neuronal‐type (α7 subunit) α‐BTx receptors on its surface. Carbachol does not stimulate 22Na+ influx in SCC‐A9 cells, but cytisine initiates a sustained influx of Ca2+. Activation of this response is inhibited by α‐BTx and by the α7‐selective antagonist methyllycaconitine. Addition of Co2+ abrogates the sustained elevation of intracellular free Ca2+ concentration, implying that the cytisine‐stimulated influx of Ca2+ is sustained by secondary opening of voltage‐sensitive channels in the plasma membrane. Surface receptors for 125I‐α‐BTx are blocked by methyllycaconitine and d‐tubocurarine. Solubilized α‐BTx receptors from plasma membranes of SCC‐A9 cells cosediment with 10S neuronal nAChR pentamers and bind to an α7‐specific monoclonal antibody (MAb P27) but not to the muscle nAChR‐reactive MAb 35. However, MAb P27 and MAb 35 both bind to α‐BTx receptors solubilized from the cytoplasmic compartments of SCC‐A9 and the parental SCC‐37 line. Reverse transcription‐PCR analysis revealed RNA transcripts for α7 and α1 subunits in both SCC‐A9 and SCC‐37 cells. The nAChRs that are expressed in these novel human cell lines can regulate cation fluxes directly as well as indirectly by synergizing with the activity of voltage‐sensitive Ca2+ channels. These activities may influence the secretion of autocrine growth factors and the transcription of growth regulatory genes and thus be pertinent to the growth and metastasis of malignant neuroendocrine neoplasms.

Immunization with neuronal nicotinic acetylcholine receptor induces neurological autoimmune disease

“…There is a recognized link between ectopic expression of neuronal and muscle autoantigens in certain cancers, particularly SCLC (14,(28)(29)(30), and the finding in a patient's serum of corresponding marker antibodies, some of which have pathogenic potential (14,28,29,31). This relationship is best illustrated by the calcium channel antibodies that cause presynaptic failure of neuromuscular transmission in the Lambert-Eaton syndrome (20,28,29).…”

“…We prepared membranes from cell lines grown as xenografts in immunodeficient mice (14). After homogenization, we isolated postnuclear membrane fractions by differential centrifugation, extracted nAChR in 2% Triton X-100, and removed insoluble residue by ultracentrifugation.…”

Immunization with neuronal nicotinic acetylcholine receptor induces neurological autoimmune disease

“…Several neuronal nAChR subtypes, as well as muscle nAChR, have previously been found in SCLC cell lines. Functional studies in SCLC cell lines suggest that these ligandgated cation channels may act in synergy with voltagegated calcium channels (14) to influence transcription of growth-regulatory genes. They also regulate cell proliferation and secretion of autocrine growth factors (32)(33)(34)(35).…”

“…Under these conditions, [ 125 I]epibatidine binds negligibly to α1 and α7 nAChRs (15). Subtype antigenicity was confirmed by quantitative immunoprecipitation using the following nAChR-specific rat monoclonal IgGs: mAb 3 (muscle and α3 nAChR) (15), mAb P20 (muscle nAChR) (15), mAb P27 (α7 nAChR) (14), and mAb 299 (α4 nAChR; Covance Inc., Princeton, New Jersey, USA). Rat mAb 1, specific for Torpedo electric organ nAChR (15), was the negative control.…”

Autoimmune and Paraneoplastic Channelopathies

“…2 In some cases (especially with poorly differentiated small cell lung carcinoma, SCLC), the cancer cells express proteins that are usually restricted to the nervous system, including ligand-and voltage-gated ion channels. 3,4 In other situations (i.e., thymoma or lymphoma), the malignancy may interrupt the maintenance of tolerance to self antigens. Many different neuron-specific autoantibodies have been described in association with the paraneoplastic neurological disorders.…”