GAD is predominantly expressed in neurons, which might be linked to SPSSD, and insulin-secreting pancreatic β cells, which might be linked to type I diabetes (107). GAD regulates the decarboxylation of glutamate to gamma-aminobutyric acid (GABA), the main inhibitory neurotransmitter within the CNS, and is related to SPSSD (108, 109). There are two GAD isoforms—GAD65 and GAD67. GAD65 is expressed in the presynaptic end of nerve terminals in its inactive form and is converted to its active form at the post-natal stage to rapidly synthesize GABA for synaptic transmission (110). GAD65 is also responsible for packaging GABA after its synthesis (111). Additionally, GAD67 is expressed in the cell body and dendrites and is responsible for synthesizing basal levels of GABA (100).

GAD65 antibody (Ab) titers and epitope specificities are present in different diseases and different subtypes of SPSSD (112, 113). For example, the GAD65 Ab titer is 348 U/mL in type I diabetes, 6.0 × 105 U/mL in cerebellar ataxia (CA), 6.2 × 105 U/mL in LE, and 1.1 × 106 U/mL in SPS (112). GAD65 binding in the presence of rFab b78 is 99% in type I diabetes, 81% in CA, 88% in LE, and 77% in SPS (112). Moreover, positive GAD immunoreactivity is ≥1,800 U/mL in SPS rat brain sections as detected via immunohistochemistry or cell-based assays (7, 114). A high range of GAD65 Ab levels is associated with SPS, whereas a lower one is associated with type I diabetes (13). A possible mechanism underlying this distinction could be that the GAD Ab in type I diabetes primarily reacts with conformational epitopes, whereas GAD antibodies in SPS recognize linear epitopes (115–117). Furthermore, GAD Ab-positive type I diabetes or SPS, CA, and LE are associated with different HLA class II haplotypes (118–121).

GABAergic neurons are responsible for inhibitory signals in the CNS and express high levels of GAD65. They are mainly located in the hippocampus, cerebellum, basal ganglia, brainstem nuclei, and spinal gray matter (122, 123). GABA binds GABAA and GABAB receptors to mediate the hyperpolarization of post-synaptic neurons, comprising an inhibitory signal (124, 125). The GAD Ab inhibits GAD65 to block GABA synthesis, thereby reducing the uptake of newly synthesized GABA in synaptic vesicles and its synaptic release (111, 126–128).

Inhibiting GABA synthesis results in decreased GABAergic transmission, which is linked to neuronal hyperexcitability and is the core pathophysiological mechanism in SPS (129, 130). For example, the possible mechanism underlying SPS might be mediated by the inhibition of GABAergic neurons in the spinal cord, resulting in a state of motor neuron hyperexcitability, ultimately causing the simultaneous contraction of agonist and antagonist muscles (100, 131). GABAergic interneurons are located at different levels of the CNS, other than the spinal cord, leading to other subtypes of SPSSD, such as PERM (128), which has also been supported by animal studies (132, 133). However, this is not the case for LE and temporal lobe epilepsy, owing to insufficient data.

The major SPSSD subtype is classic SPS (11, 68, 69, 73, 77, 80, 92–98, 134, 135), followed by SPS with myasthenia gravis (21–25, 29, 31, 34–36, 86, 89, 90, 135, 136), PERM (91), SLS (57), progressive dizziness and unstable gait (87), and NMDAR-SPS (9), as shown in Table 5. Moreover, the major carcinoma associated with SPSSD subtypes is breast cancer (21, 39, 42, 47, 50, 51, 56, 57, 137), followed by thymoma (11, 80, 91–98, 104–106, 134, 135), lymphoma (77, 80), lung cancer (68, 69, 73), and other carcinomas (21–25, 29, 31, 34–36, 86, 89, 90, 135, 136).

One report showed that the median concentration of anti-GAD65 Ab, measured via ELISA, is 30-fold higher in the serum (74,700 IU/mL) than in the cerebrospinal fluid (CSF) (2,430 IU/mL). However, these data were from 34 patients with classical anti-GAD65-associated syndromes, including SPS, CA, chronic epilepsy, and LE, with overlapping syndromes in some of the cases (138). The serum/CSF ratio of anti-GAD65 Ab was reported to be approximately 20 in patients with SPS (138). Moreover, serum and CSF anti-GAD65 Ab titers decreased, with those of CSF decreasing more rapidly than serum titers after patients with SPS received immunotherapy (138).

Amphiphysins are members of the Bin-Amphiphysin-Rvsp (BAR) family of proteins, which includes the mammalian bridging-integrators (Bin1 and Bin2), amphiphysins, and yeast Rvs161p and Rvs167p (139). Some members of the amphiphysin superfamily have conserved BAR domains, mainly in the N-terminus, and an SH3 domain in the C-terminus (139). Amphiphysin I is expressed in chicken and mammalian brains (140) and is associated with SPS and breast cancer (49, 55). Two members of amphiphysin II are also expressed in the brain. Amphiphysin II, also known as BIN1 (MYC box-dependent interacting protein-1 or bridging integrator-1) or SH3P9, is associated with cancer progression, several myopathies, heart failure, and late-onset Alzheimer's disease (141). Amphiphysin IIa shares a brain-specific domain with amphiphysin I (142, 143), and amphiphysin IIb has a skeletal muscle-specific domain with a tumor suppressor that interacts with the c-Myc oncoprotein (142, 144). In several cancers, such as breast, colon, prostate, and lung cancers, as well as hepatocarcinoma and neuroblastoma, the expression of amphiphysin II is reduced or altered (145–148). In addition, the ablation of amphiphysin II is linked to a poor cancer prognosis and increased metastasis (145, 148–151). Amphiphysin II can also inhibit Myc-dependent transformation and tumorigenesis (145, 148–151).

The major SPSSD subtype is classic SPS (28, 37, 38, 43–45, 47–50, 52–56, 135), followed by SLS (58, 65) and PERM (62, 72). Moreover, the major carcinoma associated with SPSSD is breast cancer (28, 37, 38, 43–45, 47–50, 52–56, 58, 135), followed by thymoma (66, 95, 99) and lung cancer (66, 72) (Table 6).

As an inhibitory neurotransmitter, glycine, as well as its receptor (GlyR), is critical for CNS development (152). Glycine is synthesized via serine hydroxymethyl transferase or a glycine synthase (glycine cleavage, GCS) enzyme, located between carbon dioxide, ammonium ion, N5, N10-methylene tetrahydrofolate, NADH, and a proton, producing glycine, tetrahydrofolate, and NAD+ (153), as confirmed from a rat study (154). Furthermore, the biological function of glycine requires specific transporters such as GlyT1 (glial cells) and GlyT2 (neurons) (155, 156). GlyT1 also regulates glutamatergic neurotransmission through NMDA receptors, affecting brain function and diseases (157).

There are four α subunits and one β unit in GlyR, and these are expressed in the spinal cord and retina, respectively (158–160). Microglia secrete glycine, enhance NMDA receptor-mediated responses (161), and express GlyR to induce membrane depolarization, increasing intracellular calcium and proliferation (162). In addition, glial cells modulate synaptic development by participating in the induction of the action potential conduction in white matter via GlyRs (163). Importantly, glycine has also been linked to rapid cancer cell proliferation due to glycine metabolism (164). For example, α1 and α3 GlyR subunits were found to be expressed in human brain tumor biopsies, and the lack of α1 GlyR protein expression resulted in inhibition of the self-renewal capacity and tumorigenicity of GL261 glioma cells (165). GlyR knockdown can increase P53 tumor suppressor protein expression (166, 167).

Benzodiazepines are the first-line treatment for patients with SPS. These drugs enhance the neurotransmitter effect of GABA at its receptor. Furthermore, benzodiazepines are widely used for their sedative, muscle-relaxant, and anticonvulsant effects (21). Long-term benzodiazepine therapy has been shown to benefit patients with classic or partial SPS and reduce GAD-65-positive Ab-mediated stiffness and spasm symptoms; however, this improvement might also be due to other adjunct medications.

The major drug for SPSSD treatment is diazepam, which results in a good response in most patients at high doses of up to 60 mg daily (169). However, owing to concerns about withdrawal from long-term use and high doses of diazepam therapy, tizanidine has emerged as a good candidate for alternative therapy. As an NMDAR, tizanidine is an α2 inhibitor that inhibits glutamate release and prevents glutamatergic hyperactivity, thereby resolving convulsions in patients with SMS. Nevertheless, the dose of tizanidine should be individualized (21, 169).

Baclofen is an agonist of GABA type B receptors that inhibits reflexive muscle contraction by blocking the release of excitatory neurotransmitters through voltage-gated calcium channels (170). It is also a second-line therapy for patients with SPS. However, to date, the use of oral baclofen therapies is still being debated. In one report, high doses (however, the dose is unknown) of oral baclofen therapy were found to result in serious side effects, such as sedation and respiratory depression (171). However, oral baclofen had good effects on SPS patients without serious side effects. For example, oral baclofen therapy (5 mg, three times per day) plus clonazepam resulted in improvements in a 69-year-old man with SPS and amphiphysin antibodies (172). Symptomatic treatment initiated with oral clonazepam and baclofen (5 mg Bid), followed by intravenous immunoglobulin (IVIG) resulted in improvements in a 60-year-old man with SPS associated with critical illness polyneuropathy (173). Baclofen (30 mg/day) combined with oral diazepam and steroids resulted in improvements in a 55-year-old GAD-Ab-positive female patient with SLS and breast carcinoma (57). For childhood-onset SMS, three SMS patients had good clinical responses with oral baclofen (dose range, 60–80 mg) combined with diazepam, IVIG, plasma exchange or dantrolene, and botulinum toxin (174).

Alternatively, intrathecal therapy is an effective route for baclofen treatment (2). The chronic infusion of intrathecal baclofen can improve SPS patient outcomes, including the pain Numeric Rating Scale, Spasm Frequency Scale, and lower extremity Modified Ashworth Scale (171). Intrathecal baclofen (100 μg) followed by a rehabilitation program resulted in substantial clinical and functional improvements in a 59-year-old female SPS patient, who had no therapeutic response with oral benzodiazepines and botulinum toxin injections (175). In addition, intrathecal baclofen (started from 50 μg/d up to 100 μg/d) improved motor functions in a 48-year-old male GAD-negative SPS patient (176). Baclofen can be used to effectively treat SPS because it is a direct agonist of GABA-B receptors and does not require endogenous GABA to induce presynaptic inhibition (176).

Levetiracetam binds to synaptic vesicle glycoprotein 2A (SV2A), resulting in the release of the neurotransmitter stored within the vesicle, rapidly inhibiting firing neurons and potassium and N-type calcium channels (177, 178). In a previous study, three patients with high anti-GAD65 Ab levels did not respond satisfactorily to IVIG and diazepam treatment with or without plasmapheresis (179). These patients were treated with 500 mg oral levetiracetam twice daily, which improved axial rigidity and the disappearance of paroxysmal respiratory arrest within 3 days of therapy initiation, with markedly reduced leg stiffness and ameliorated walking difficulties (179). However, to date, there is no evidence of the effects of long-term levetiracetam therapy. The possible mechanism by which levetiracetam achieves its effects could be by stabilizing and strengthening GABAA and decreasing hyperexcitability in spinal cord neurons (179).

Structurally, pregabalin is classified as a GABA analog or gabapentinoid (180). In a previous study, a female patient with SMS who did not respond to diazepam treatment, owing to excessive sedation, was successfully treated with a 3-month pregabalin regimen (181). The possible mechanism underlying the effects of pregabalin might be the inhibition of calcium influx and subsequent release of excitatory neurotransmitters, including glutamate and norepinephrine, resulting in compensation for the imbalance between inhibitory and excitatory intracortical circuits (181).

The mechanism of action of propofol in the CNS is unclear. Propofol might enhance the function of GABA receptors, evoking the chloride current in central neurons at clinically relevant concentrations, ultimately activating the GABA receptor–chloride ionophore complex (182). Notably, a low dose of propofol improves symptoms in patients with SPS who do not respond to high-dose benzodiazepines, baclofen, corticosteroids, levetiracetam, IVIG, or IV ethanol. Furthermore, propofol is effective for patients with SMS that is refractory to therapy (183). Unfortunately, long-term propofol therapy has unsatisfactory effects in patients with SPS (184).

Rituximab binds to the CD20 antigen on mature B cells, leading to B cell lysis, while sparing precursor B cells. Rituximab improves SPS and other neurological autoimmune disorders, such as Devic's disease, myasthenia gravis, autoimmune neuropathies, and inflammatory myopathies (185). SPS is associated with elevated titers of anti-GAD65 Abs and glycine receptor α-subunits in patients (186). Four reports have demonstrated the benefits of rituximab for patients with SPS (186–189). Although rituximab improved the clinical conditions of patients, the decrease in the anti-GAD titer was inconsistent in different reports. Some reports demonstrated that after rituximab treatment, the anti-GAD titer was rapidly (17 days, from positive to undetectable) or slowly (1 year, from 1,000 to 400 U/mL) reduced (187). However, another case report showed that the anti-GAD Ab titer remained elevated, even during treatment with rituximab (188).

Tacrolimus inhibits the calcium calcineurin pathway and exerts its immunosuppressive effect by reducing the proliferation of activated T cells (190). Furthermore, tacrolimus decreases IL-2 levels and impairs T-helper cell functions, finally reducing the activation of B cells to produce antibodies. It also suppresses the function of anti-GAD Abs, thereby blocking GABAergic neurotransmission and interfering with GABA synthesis (191). Tacrolimus directly blocks calcineurin in the GABAergic inhibitory system. Nonetheless, the neuroprotective effect of tacrolimus therapy on SPS demonstrated based on the reduced density of neurons with somal areas and improved pathological conditions, remains debatable (192); evidence that macrolide antibiotics inhibit the function of immunophilins and provide neuroprotective and neuroregenerative effects contradicts this assertion (184). Tacrolimus combined with IVIG or prednisone treatment greatly improved symptoms and reduced Ab titers in two patients who showed no response to other medicines (192). After 4 weeks of treatment with tacrolimus, serum anti-GAD Ab titers in patients with SPS were decreased, with an increase in motor ability, and the patients became completely self-dependent (191).

After immune globulin therapy, 11 patients with SPS showed improvements in their movement disorder and decreased serum anti-GAD65 Ab titers (169). As we previously described, serum and CSF anti-GAD65 Ab titers were found to decrease, with those of CSF decreasing more rapidly than those of serum after patients with SPS received immunotherapy (138).

A 26-year-old woman with anti-NMDAR encephalitis and SPS with an ovarian teratoma was successfully treated via laparoscopic removal of the ovarian tumor. She received immune-suppressant medications (methylprednisolone followed by a combination with baclofen) preoperatively and postoperatively, and her symptoms were gradually resolved (9).

A 53-year-old male patient had anti-amphiphysin-positive SPS and breast cancer, as previously mentioned. After undergoing surgery to excise the cancer, he received adjuvant chemotherapy with cyclophosphamide, methotrexate, and 5-fluorouracil, followed by post-mastectomy radiation and adjuvant endocrine therapy with tamoxifen. After 1 year of surgery, the stiffness in his upper extremity, but not his lower extremities, greatly improved (42). However, for a 30-year-old female patient with anti-amphiphysin, GAD Ab-negative SPS, and breast cancer symptoms were not alleviated following surgery (41).

A 56-year-old woman with anti-amphiphysin-positive SPS associated with small-cell lung cancer received treatment with benzodiazepines and corticosteroids, followed by cancer therapy with cisplatin/etoposide and radiotherapy. Following treatment, she exhibited signs of improved stiffness and was able to walk independently for short distances (72).

The main reported methods for passive transfer animal models using rats or mice are single cerebellar or paraspinal injections (132) and intrathecal (210, 211) or intraperitoneal injections (208, 211). Unfortunately, these animal models do not effectively mimic the clinical symptoms of SPS. However, some symptoms, such as paraspinal electrophysiological evidence of continuous motor activity (132), increased anxiety-like behavior (212), worsened rotarod results, and deficits in postural control (211), were partially matched.

Active immunization using the human GAD65 protein has been effectively performed in type I diabetes studies; however, it has failed for neurologic diseases, including SPS, despite the high titers of GAD Abs (213) developing in these studies. This suggests that the GAD65 protein is also important for the pathogenesis of SPS; however, it is regulated by other autoantigens that contribute to the pathogenesis of SPS.