Prob a ble Anti-Glu ta mate De car box yl ase 65 (Gad65) An ti body-As so ci ated Cer e bel lar Ataxia. Clin i cal Case Re port and Lit er a ture Re view

ease pro gresses. Corticosteroids are one of the rec - om mended treat ment meth ods, which were ef fec tive in our case. Main te nance ther apy is es - sen tial to pre vent re lapse of the dis ease.


CLIN I CAL CASE PRE SEN TA TION
A 52-year-old woman was ad mit ted to the De part ment of Neu rol ogy in 2022 due to com plaints of in creased diz ziness, im paired co or di na tion, oc ca sional chok ing, and slurred speech.Med i cal his tory re vealed the pa tient's comorbidities: ar te rial hy per ten sion and de pres sion.The pa tient de nied al ler gies.The woman claimed to be tak ing sal buta mol in ha la tions be cause of a pre vi ous COVID-19 in fec tion, sertraline 100 mg once a day, agomelatine 50 mg once a day, clorazepate 5 mg twice a day, zolpidem 10 mg once in the eve ning, trazodone 50 mg once in the eve ning, and antihypertensive drugs.
The re sults of the anal y sis of pre vi ous med i cal data revealed that in 2018, the pa tient was ad mit ted to a lo cal hospi tal with sim i lar symp toms, such as nau sea, diz zi ness, and high blood pres sure af ter wak ing up.The ex am i na tion led to a ves tib u lar neuronitis di ag no sis.Af ter re ceiv ing treatment, the pa tient's con di tion im proved and she was discharged to con tinue treat ment at home.How ever, in 2019, the symp toms re curred, and the pa tient was re ferred to an otoneurologist, who con cluded that nys tag mus was pos sibly due to dam age to the cen tral sys tem.More over, brain mag netic res o nance im ag ing (MRI) was per formed but no sig nif i cant changes were found.
Later in 2019, the pa tient's con di tion wors ened, with weak ness and co or di na tion im pair ment of the right limbs and im paired walk ing (ataxic gait and re quir ing as sistance), and she was re ferred to the De part ment of Neu rology.Neu ro log i cal ex am i na tion re vealed bi lat eral hor izon tal third-de gree nys tag mus and ver ti cal nys tag mus.Ac cord ing to the Lov ett scale, the mus cle strength of the right limbs was 4 points and of the left limbs was 5 points.Re flexes in the right arm were slightly stron ger than in the left arm, and lower limb re flexes were stron ger sym met rically.There was a ten dency to the Babinski re flex on the right, weaker plan tar re flex bi lat er ally.Fin ger-to-nose and heel-to-shin tests were per formed, which showed ataxia of the right limbs.Move ments were slow, gait was ataxic, and ataxia was ob served more in the right limbs: while walk ing, the pa tient put the right leg higher.Also, a lower am pli tude of phys i o log i cal synkinesis was observed.
To ex clude var i ous causes of cer e bel lar ataxia, tests for vi ta min B12 (pos si ble de fi ciency), thy roid hor mones: thyroid-stim u lat ing hor mone (TSH) and tetraiodothyronine (T4) (sus pected hypothyroidism), as well as in fec tious disease mark ers (treponema pallidum hemagglutination assay (TPHA), hu man im mu no de fi ciency vi rus 1 and 2 (HIV1/HIV2) and p24 an ti gens, IgM and IgG against Borrelia burgdorferi, IgM and IgG against tick-borne enceph a li tis), were per formed.How ever, no changes were found: vi ta min B12, TSH, and T4 were within nor mal range and in fec tious mark ers were neg a tive, ex cept for IgG against tick-borne en ceph a li tis be cause of pre vi ous vac ci na tion.
Au to im mune en ceph a li tis was then sus pected, and several tests were per formed to ex clude it.The brain MRI in T2W/Flair mode showed sin gle, non spe cific, hyperintense sig nals in the right fron tal and pa ri etal subcortical ar eas, pos si bly microangiopathic foci, with out atro phic changes in the cer e bel lum (Fig. 1 and Fig. 3A).Anal y sis of the cerebrospinal fluid (CSF) showed: 0.4 g/L of to tal pro tein con cen tra tion, 3.55 mmol/L of glu cose, 9×10 6 /L of white blood cells, no oligoclonal bands were found.An ti bod ies for paraneoplastic syn drome and autoantibodies against neuronal sur face an ti gens were in ves ti gated.Only se rum anti-GAD65 Ab were pos i tive (1+).Elec tro en ceph a log raphy (EEG) was per formed and showed no changes.On sus pi cion of stiff per son syn drome, electro neuro myography (ENMG) was per formed and re vealed no patho logi cal changes.Fur ther more, some di ag nos tic pro ce dures were taken to ex clude oncological pro cesses: thy roid ul trasound, ul tra sound of the up per ab do men, mammogram, fae cal oc cult blood test, and chest X-ray.No sig nif i cant devi a tions were ob served.Dur ing hos pi tal iza tion, the pa tient was treated with methylprednisolone 1 g once a day for 3 days.The pa tient's neu ro log i cal sta tus had not changed dur ing the time of hos pi tal iza tion, and the pa tient was discharged.
In 2022, the pa tient was again ad mit ted to the De partment of Neu rol ogy be cause of per sis tent symp toms.Ad dition ally, oc ca sional chok ing and slurred speech ap peared.Neu ro log i cal ex am i na tion showed slurred and chant ing speech, bi lat eral hor i zon tal and ver ti cal nys tag mus, a tendency to the Babinski re flex on the right, and a pos i tive Babinski re flex on the left.Fin ger-to-nose and heel-to-shin tests were per formed with dysmetria, this time, es pe cially on the left.The pa tient had an ataxic gait and needed as sistance walk ing.
Sev eral tests were re peated: for vi ta min B12, TSH, and T4 and the same in fec tious mark ers for neurosyphilis, Lyme dis ease, tick-borne en ceph a li tis, and HIV1/HIV2.All tests were neg a tive, and there were no new find ings.On sus pi cion of glu ten ataxia, anti-Gliadin (GAF-3X) IgA and IgG, an ti bod ies to tis sue transglutaminase (anti-tTG) IgA and IgG were searched, but the re sults were neg a tive.Wilson's dis ease was also con sid ered, but the level of ceruloplasmin was only 0.33 g/L (nor mal range 0.22-0.58g/L).MRI of the brain was re peated and showed, as be fore, non-spe cific, hyperintense sig nals, prob a bly of microangio pathic or i gin, in the ce re bral hemi spheres in the T2W/Flair mode.New find ings were also dis cov ered: atro phic changes in the up per parts of the cer e bel lum (Fig. 2 and Fig. 3B).
A con sul ta tion with a clin i cal phar ma col o gist was requested about the med i ca tions the pa tient was tak ing to eval u ate whether they might worsen the woman's con dition.The clin i cal phar ma col o gist stated that abruptly stopping sertraline or us ing trazodone and clorazepate could cause ataxia as a side ef fect.How ever, no in for ma tion on the abrupt dis con tinu a tion of sertraline was gath ered, and the symp toms ap peared be fore trazodone and clorazepate were ad min is tered.Blood tests and CSF anal y sis were performed again, show ing no sig nif i cant changes: to tal protein con cen tra tion 0.39 g/L, glu cose 3.77 mmol/L, oligoclonal bands not found.On the other hand, the cru cial point was the rise in se rum anti-GAD65 an ti bod ies from 2019 (+1) to 2022 (+2).More over, the se rum con cen tration of anti-GAD65 Ab de tected by en zyme immunoassay was 2000 kU/L (nor mal val ues: 0-5 kU/L).On re peated inves ti ga tion, where an ti bod ies and cer e bel lar atro phic changes were found, and in the ab sence of any other cause that could ex plain the changes, anti-GAD65 an ti body-as -so ci ated cer e bel lar ataxia was con sid ered the most likely cause.In this case, a de fin i tive di ag no sis could not be confirmed since this re quires the de tec tion of high titers of anti-GAD65 an ti bod ies, which were not pres ent in our case.Ev i dence of intrathecal syn the sis of anti-GAD65 Ab and pos i tive immunohistochemistry on brain tis sue could also help con firm the di ag no sis, how ever, this was not performed in this case.
Dur ing her stay in the De part ment of Neu rol ogy, the pa tient be gan pulse ther apy with methylprednisolone 1 g Fig. 2. Brain MRI in T2W/Flair mode (ax ial plane) shows atro phic changes in the cer e bel lum.once a day for 5 days.The treat ment was con tin ued with prednisolone 60 mg per day for 7 days, re duc ing the dose to 50 mg per day and fi nally to 30 mg per day.Dur ing treatment, the pa tient's con di tion im proved: trem ors decreased, in di ca tors on co or di na tion tests im proved.However, chant ing speech and gait dis tur bance re mained.The woman eval u ated the im prove ment in her con di tion by 50%.Be fore dis charge, the pa tient was as sessed ac cord ing to the In ter na tional Co op er a tive Ataxia Rat ing Scale.The re sults were as fol lows: pos ture and gait score 27/34, kinetic score 25/52, dysarthria score 3/8, and oculomotor move ment score 2/6.The to tal ataxia score was 54/100.The pa tient was rec om mended to con tinue us ing prednisol one 30 mg per day for 2-3 months, re duc ing the dose to 15 mg per day for 2 months, and re ferred to re ha bili ta tion.

DIS CUS SION AND LIT ER A TURE RE VIEW Prev a lence and pathogenetic mech a nism of anti-GAD65 an ti body-as so ci ated cer e bel lar ataxia
Anti-glu ta mate de car box yl ase 65 an ti body-as so ci ated cere bel lar ataxia is a rare au to im mune neu ro log i cal dis or der.This con di tion most com monly oc curs in women in their 60s, es pe cially those with type 1 di a be tes mellitus, au to immune thy roid dis eases, or other au to im mune dis or ders.Fur ther more, about 70% of pa tients have serological ev idence of glu ten sen si tiv ity [2][3][4].How ever, in our case, the woman had no con firmed au to im mune dis or ders, but her age cor re sponded to the data in the lit er a ture.The pa tient was also tested for glu ten sen si tiv ity, but there was no serological ev i dence of this.
Glu ta mate de car box yl ase (GAD) is mainly found in two lo ca tions: in pan cre atic is let b cells and CNS in hib itory (gamma-aminobutyric acid (GABA)-nergic) neurons.It ex ists as two isoforms: GAD65 and GAD67.GAD67 reg u lates the basal lev els of GABA and is pres ent in the cy to plasm of in hib i tory neu rons.GAD65 is re sponsi ble for GABA syn the sis and exocytosis (trans port of GABA into syn ap tic ves i cles and con trol of syn ap tic release) [5,6].When anti-GAD65 an ti bod ies are pres ent in the or gan ism, they ap proach GAD65 and in ter fere with GABA exo cyto sis.As a re sult, GABA lev els de crease.Such dis tur bances can cause a def i cit in cer e bel lar tem poral co or di na tion, which can lead to cer e bel lar ataxia [3].In ad di tion, a de crease in GABA lev els af fects an in crease in the amount of glu ta mate.This leads to the ac ti va tion of microglia and ac ti va tion of the N-methyl-D-aspartate (NMDA) re cep tor, and then to the dis rup tion of NMDA recep tor-me di ated re sponses.This causes dys func tional signal trans mis sion and fi nally cell de gen er a tion.This is the main rea son why cer e bel lar at ro phy is no tice able in anti-GAD Ab-as so ci ated CA [5,6].Our clin i cal case also had vis i ble at ro phy, which took time to de velop (about three years).

Clin i cal man i fes ta tion and di ag nos tics of anti-GAD65 an ti body-as so ci ated cer e bel lar ataxia
Most pa tients have ataxia of gait and pos ture.Al though 60 to 70% of pa tients have nys tag mus, dysarthria, dysphagia, and limb ataxia.Usu ally, limb ataxia pro ceeds asym met rically [2][3][4] (this was ob served in our clin i cal case as well).Many ex hibit mild ataxia and can walk un aided, but some may have se vere or mod er ate ataxia and re quire a walk ing aid or a wheel chair [4].Some pa tients may ex pe ri ence lum bar pain, mus cle ri gid ity, and spasms, which can lead to fre quent falls.Some pa tients may have drug-re sis tant fo cal ep i lepsy [2,7].In our clin i cal case, the pa tient ex pe rienced the symp toms men tioned in the lit er a ture, ex cept for lum bar pain, mus cle ri gid ity, spasms, and she did not suffer from drug-re sis tant fo cal ep i lepsy.
The on set of symp toms is sub acute or chronic, progress ing over months or years [1].It is cru cial to know the pa tient's com plete med i cal his tory and per form a body assess ment along with a neu ro log i cal ex am i na tion.It would be in for ma tive if pa tients with ataxia were as sessed us ing the In ter na tional Co op er a tive Ataxia Rat ing Scale.In our case, the pa tient was as sessed on this scale only at the time of dis charge in 2022, but it would be im por tant to com pare the re sults be tween 2019 and 2022 in or der to mon i tor the pro gres sion of the dis ease over the years.
Blood tests to as sess elec tro lytes and the anal y sis of CSF are also nec es sary.Oc ca sion ally, the oligoclonal bands may be pres ent in the CSF [3].How ever, in our case, CSF was per formed twice (in 2019 and 2022), but no oligo clonal bands were found.Se rum and CSF lev els of anti-GAD65 an ti bod ies are cru cial for dif fer en tial di ag nosis since anti-GAD-Ab are found not only in pa tients with cer e bel lar ataxia but also in pa tients with other neu ro log ical dis eases.Most of ten, the symp toms over lap with limbic en ceph a li tis, stiff per son syn drome, pro gres sive encephalo myelitis with ri gid ity and myoclonus, and re frac tory ep i lepsy, which makes ac cu rate clin i cal di ag no sis dif fi cult [5,8].This is the rea son our pa tient un der went var i ous diag nos tic pro ce dures, such as brain MRI, anal y sis of CSF, search for an ti bod ies against neuronal sur face an ti gens and against intracellular an ti gens, EEG, and ENMG to ex clude other neu ro log i cal dis eases.How ever, only pa tients with anti-GAD65 cer e bel lar ataxia have high titers of these an tibod ies, usu ally more than 10,000 U/mL (or 10-100 times higher) [2,5,6,9].Fur ther more, cer e bel lar vermian or hemi spheric at ro phy may be pres ent on brain MRI, but the de gree of at ro phy is usu ally milder com pared to the se verity of ataxia [3,4].In our case, el e vated se rum anti-GAD65 an ti body titers (2000 kU/L (nor mal val ues: 0-5 kU/L)), an in crease in se rum an ti bod ies from +1 in 2019 to +2 in 2022, the dy nam ics of brain MRI from 2019 to 2022, when vis ible cer e bel lar at ro phy ap peared, and the ex clu sion of other causes of cer e bel lar ataxia (vi ta min B12 de fi ciency, hypothyroidism, glu ten ataxia, neurosyphilis, Lyme dis ease, tick-borne en ceph a li tis, and other in fec tious dis eases) led to the di ag no sis of prob a ble anti-GAD65 an ti body-as so ciated cer e bel lar ataxia.How ever, we can not con firm a de -fin i tive di ag no sis, as in this case, not only were symp toms of cer e bel lar dys func tion pres ent (dysphagia and patholog i cal re flexes were ob served in the pa tient), morever, the anti-GAD65 titers were also not as high as in di cated in the lit er a ture, and the changes ob served on brain MRI were not in flam ma tory, but atro phic.Graus F et al. rec om mend that syn dromes such as CA, which can be caused by var i ous other mech a nisms and eti ol o gies, should be con firmed when there are high lev els of anti-GAD65 an ti bod ies in the CSF and these an ti bod ies are syn the sized intrathecally.Con di tions with out intrathecal anti-GAD65 Ab syn the sis should be con sid ered as 'prob a bly au to im mune' [10].Posi tive immunohistochemistry on the brain tis sue may be also help ful to con firm the diagnosis [11].
An ex am ple to take into ac count is a ret ro spec tive study (con ducted in 2020) of 56 pa tients with pos i tive anti-GAD65 an ti bod ies who were tested and di vided into two groups: low con cen tra tion (n=20) of anti-GAD65 Ab and high con cen tra tion (n=36), where the cut off value was 10,000 IU/mL in se rum by au to mated quan ti ta tive ELISA.It was found, that high con cen tra tions of an ti bod ies were as so ci ated with clas si cal anti-GAD65 syn dromes (94%), such as cer e bel lar ataxia, chronic ep i lepsy, stiff-per son syn drome, and other.How ever, in 12 pa tients with low con cen tra tions of these an ti bod ies, the clin i cal pic ture was pres ent due to other causes, such as limbic en ceph a li tis with anti-GABABR and pan cre atic adenocarcinoma, var ious sub acute and chronic polyradiculoneuropathies, and other non-im mune me di ated dis eases such as mul ti ple system at ro phy and etc.These dis eases were not re lated to the de tec tion of anti-GAD65 Ab.The other 8 pa tients with low anti-GAD65 Ab had non spe cific ataxia and gait dis or der or chronic ep i lepsy, but the as so ci a tion with anti-GAD65 anti bod ies in these pa tients was not de ter mined [12].This study con firms that low titers of anti-GAD65 are not always as so ci ated with au to im mune neu ro log i cal dis or ders such as CA, which may also be pos si ble in our case.

Man age ment of anti-GAD65 an ti body-as so ci ated cer e bel lar ataxia
First-line immunotherapy is used un til re mis sion.Immuno therapy be gins with high doses of in tra ve nous cortico steroids, in tra ve nous im mu no glob u lin (IVIG), plasma pheresis, or rituximab.De pend ing on the sit u a tion, monotherapy or a com bi na tion of treat ment meth ods can be cho sen [6,9,13].Some clin i cal cases have re ported good out comes with IVIG [14] or corticosteroids [15,16] as first-line immunotherapy.Com bi na tions can be var i ous.A good ex am ple of com bi na tion ther apy is IVIG and rituximab, which have shown great re sults [17].The response to immunotherapy de pends on the clin i cal course.In the sub acute type, the re sponse to com bi na tion ther apy is higher than in the chronic type [13].Main te nance therapy is usu ally needed.It con sists of oral corticosteroids, IVIG, azathioprine, or mycophenolate mofetil [2,16].Good out comes with the use of in tra ve nous and oral cortico steroids were also ob served in our case.Al though dur ing the first hos pi tal iza tion in the De part ment of Neurol ogy, no re sponse to in tra ve nous immunotherapy was ob served, af ter the last hos pi tal iza tion and immunotherapy the pa tient's con di tion im proved: trem ors de creased and in di ca tors of co or di na tion tests im proved.At the be ginning of the last hos pi tal iza tion, the treat ment with methylpredni solone pulse ther apy was started and dis con tin ued af ter 5 days, leav ing the pa tient on oral prednisolone mainte nance ther apy for sev eral months.
Anti-GAD titers should be mon i tored af ter each immuno therapy treat ment to fol low the clin i cal course.Long-term fol low-up re sults show that immunotherapy im proved CA in pa tients with the sub acute type.A study in volv ing 25 pa tients with anti-GAD65 Ab-as so ci ated CA showed that af ter immunotherapy, 35% of pa tients ex pe rienced an im prove ment of at least 1 point on the mod i fied Rankin Score (mRS).Un for tu nately, the CA pro gressed in pa tients, who had chronic type of CA and who did not receive immunotherapy [2].