BBA - Re iews on Cance 1876 (2021) 188635
A ailable online 14 Oc obe 2021
0304-419X/© 2021 The Au ho s. Published by Else ie B.V. This is an open access a icle unde he CC BY-NC-ND license
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Re iew A icle
The mul i ace ed oles o gasde mins in cance biology and
oncologic he apies
Da id Sa i´
o
a
,
b
,
*
, Jeanne e Ma ínez-Val
c
, ´
Angela Molina-C espo
a
,
b
, Lau a S´
anchez
c
,
Gema Mo eno-Bueno
a
,
b
,
d
,
*
a
Biochemis y Depa men , UAM, & IIBm "Albe o Sols" CSIC-UAM, c/ A zobispo Mo cillo 4, 28029 Mad id, Spain.
b
Cen o de In es igaci´
on Biom´
edica en Red, ´
a ea de C´
ance (CIBERONC), c/ Melcho Fe n´
andez Almag o 3, 28029 Mad id, Spain.
c
Zoology, Gene ics and Physical An h opology Depa men , San iago de Compos ela Uni e si y, A da/ Al onso X O Sabio s/n, 27002 Lugo, Spain.
d
MD Ande son Cance Cen e Founda ion, c/ A u o So ia 270, 28033 Mad id, Spain.
ARTICLE INFO
Keywo ds:
Gasde min
Py op osis
cance cell dea h
cance ea men
In lamma ion
ABSTRACT
The in ol emen o he Gasde min (GSDM) p o ein amily in cance and o he pa hologies is one o he ho es
opics in biomedical esea ch. The e a e six GSDMs in humans (GSDMA, B, C, D, GSDME/DFNA5 and PJVK/
DFNB59) and, excep PJVK, hey can igge cell dea h mos ly by py op osis (a o m o ly ic and p o-
in lamma o y cell dea h) bu also o he mechanisms. The exac ole o GSDMs in cance is in ica e, since
depending on he biological con ex , hese p o eins ha e di e se cell-dea h dependen and independen unc-
ions, exhibi ei he p o- umo o an i- umo unc ions, and p omo e ei he sensi iza ion o esis ance o onco-
logic ea men s. In his e iew we p o ide a comp ehensi e o e iew on he mul i ace ed oles o he GSDMs in
cance , and we c i ically discuss he possibili ies o exploi ing GSDM unc ions as de e minan s o an i-cance
ea men and as no el he apeu ic a ge s, wi h special emphasis on inno a i e GSDM-di ec ed nano- he a-
pies. Finally, we discuss he issues o be esol ed be o e GSDM-media ed oncologic he apies became a eali y a
he clinical le el.
1. The gasde min gene amily
The gasde min amily (GSDM) comp ises six genes in humans (GSDMA,
B, C, D, GSDME/DFNA5, and PJVK/DFNB59) and en in mice (Gsdma1–3,
Gsdmc1–4, Gsdmd, Gsdme, and D nb59) [1–3]. GSDME (A.K.A DFNA5,
dea ness au osomal dominan 5) is he mos ancien GSDM gene, being
p esen in some in e eb a es like co als [2,4], while GSDMB appea s only
in speci ic mammalian species, being he only GSDM membe no p esen
in he mouse and a genomes [2]. GSDM usually clus e in speci ic ch o-
mosomal egions (17q21 con ains GSDMA and GSDMB; 8q24 GSDMC and
GSDMD; Table 1) indica ing ha gene duplica ions occu ed du ing
e eb a e e olu ion [2,4]. In ac , GSDMB likely o igina ed om a local
duplica ion and DNA-s and in e sion o GSDMA in some mammals [2,4].
The name Gasde min o igina es om he “Gas ic and de mal” exp ession
o GSDMA, he i s iden i ied GSDM gene [5]. GSDMs a e gene ally
exp essed in he diges i e ac , whe e hey show speci ic exp ession pa -
e ns: in he gas o-esophageal epi helium GSDMA is mos ly de ec ed in
di e en ia ed cells, GSDMB in he basal laye s, GSDMC in he sup abasal
and di e en ia ed egions and GSDMD in he di e en ia ing cells [1,3]. In
addi ion, GSDMs show speci ic exp ession pa e ns in mul iple issues/o -
gans. Fo example, GSDMA is exp essed in skin, lung, mamma y glands;
GSDMB in li e , lung, colon and immune cells; GSDMC in skin and spleen;
GSDMD in he in es ine and leukocy es; GSDME in ep oduc i e o gans and
ne ous sys em and PJVK in es es and he audi o y ne ous sys em, among
o he s [1–4,6,7].
2. GSDM a he c oss oads be ween py op osis and o he cell
dea h mechanisms
GSDMs a e cy oplasmic p o eins (a ound 50 KDa) wi h nine
Abb e ia ions: CT, C- e minal domain; DAMPs, damaged-associa ed molecula pa e ns; DCT, Deci abine; GZM, G anzyme; KO, Knock-ou ; LDH, Lac a e dehy-
d ogenase; LPS, lipopolysaccha ide; NLRP3, NOD-like ecep o p o ein 3; NC, Nanocapsules; NT, N- e minal domain; NP, Nanopa icles; PLK1, Polo like kinase 1;
ROS, eac i e oxygen species; SCC, Squamous Cell Ca cinoma; TGF-β, ans o ming g ow h ac o -β; TNF, Tumo nec osis ac o ; TSG, Tumo supp esso gene; WB,
Wes e n blo ; WT, Wild- ype.
* Co esponding au ho s a : Biochemis y Depa men UAM, & IIBm "Albe o Sols" CSIC-UAM, & CIBERONC
E-mail add esses: [email p o ec ed] (D. Sa i´
o), [email p o ec ed] (G. Mo eno-Bueno).
Con en s lis s a ailable a ScienceDi ec
BBA - Re iews on Cance
jou nal homepage: www.else ie .com/loca e/bbacan
h ps://doi.o g/10.1016/j.bbcan.2021.188635
Recei ed 30 June 2021; Recei ed in e ised o m 28 Sep embe 2021; Accep ed 10 Oc obe 2021
BBA - Re iews on Cance 1876 (2021) 188635
2
conse ed mo i s (con aining leucine- ich egions) o s ill unknown
unc ion, dis ibu ed h oughou he p o ein [3]. GSDMs, wi h he
excep ion o PJVK, sha e a mode a ely conse ed N- e minal (NT) and
C- e minal (CT) domains ha a e in e connec ed by a cen al hinge e-
gion, which is speci ic o each amily membe [2]. Fo many yea s,
GSDMs unc ions ha e been unclea un il he disco e y, in 2015, ha
GSDMD igge in lammasome-dependen ly ic cell dea h [8–10] by
o ming memb ane po es [11–15]. Fu he s udies e ealed ha o he
GSDMs exhibi a simila p o ein 3D s uc u e [15–18] and sha e he p o-
cell dea h unc ion ha is ac i a ed by a common mechanism
[15,19,20]. The common model shows ha cy osolic GSDMs ha e an
inac i e con o ma ion in which he NT po e- o ming domain is au o-
inhibi ed by binding o speci ic esidues in he hyd ophobic co e o
he CT [11–15,19,20]. This olded con o ma ion is main ained by he
linke in e domain egion, which is clea ed by speci ic p o eases ac i-
a ed a e pa icula s imuli (Fig. 1A). The eleased GSDM NT binds o
speci ic acid lipids (e.g., phosphoinosi ides and phospha idylse ine) and
inse s in o he cell memb ane in e nal laye , oligome izing and o ming
la ge ansmemb ane po es [11–15,19,20] (Fig. 1B). The ups eam
pa hways leading o GSDM NT elease a e speci ic o each GSDM
[7,19,20]. Rega ding GSDMD, mul iple pa hogen signals (e.g., LPS,
lagellin) o cell damage molecula pa e ns “DAMPs” (e.g., eac i e
oxygen species “ROS”, dsDNA) ac i a e he mul ip o ein complex
in lammasome, which hen p o okes he elease o GSDMD NT domain
by he p o-in lamma o y caspases1/4/5/11 [7,19,20]. GSDMD NT po es
acili a e he sec e ion o ma u e in lamma o y cy okines (IL1β, IL18)
and o he in acellula molecules (i.e., Lac a e dehyd ogenase; LDH),
while he in lux o wa e and ions migh be esponsible o cell swelling
[7,19,20]. La e , he ac i a ion and oligome iza ion o Ninju in-1
(NINJ1) p oduces he ex ensi e memb ane up u e and cell lysis
(Fig. 1B), which esul s in he elease o di e se in acellula con en ,
including he p o-in lamma o y DAMP HMGB1 [21]). This
in lammasome/GSDMD-media ed mechanism is e med py op osis
(“ ie y dea h”) and can be di ided in o “canonical” (caspase 1) o “non-
canonical” (caspase 4/5/11) depending on he s imuli and caspases
in ol ed [7,19,20]. Py op osis occu s mos ly in leukocy es and epi he-
lial cells and p o okes an in lamma o y eac ion o he su ounding cells
[7,19,20]. Py op osis is mechanis ically and biologically di e en o
o he p og ammed cell dea h ypes, such as apop osis, nec op osis o
e op osis, and dying cells exhibi dis inc mo phologic ea u es:
ex ensi e cell swelling and memb ane ballooning in absence o cell
de achmen , in ac nucleus wi h ch oma ic condensa ion, among o he s
[22]. Excep o PJVK, he NT o o he GSDMs can p oduce ly ic cell
dea h [15,19,20] wi h he elease o LDH and o he molecules. The e-
o e, py op osis was enamed as “Gasde min-media ed p og ammed cell
dea h” [23], bu accumula ing e idences p o ed ha GSDMs can
o ches a e o he cell dea h p ocesses and addi ional dea h-independen
unc ions.
Indeed, each GSDM is ac i a ed by speci ic p o eases in a cell
con ex -dependen way (Fig. 1A) ha could lead o di e en biological
consequences: GSDMD NT is eleased by in lamma o y caspases du ing
canonical and non-canonical py op osis [9,10], whe eas Ye sinia in ec-
ion [24,25] o RIPK3 signaling [26] p o okes caspase-8-media ed
GSDMD ac i a ion. Ca hepsin-G ac i a es GSDMD du ing NOD-like e-
cep o p o ein 3 (NLRP3)-s imula ed py op osis in mac ophages and
neu ophils [27], bu neu ophil elas ase (NE) clea age in neu ophils
could esul in py op osis [28] o NETosis [29]. O no e, GSDMD po es
can elease cy okines in leukocy es in absence o cell dea h [30], and
GSDMD-cell dea h can be inhibi ed by he Endosomal so ing complexes
equi ed o anspo (ESCRT-III) mechanism [31]. GSDME is ac i a ed
by apop o ic caspase-3, in esponse o mul iple s imuli [32,33], leading
o cell lysis conside ed by some au ho s as “nec osis seconda y o
apop osis” [32,34], while GSDME p ocessing ia kille cell-de i ed
G anzyme-B (GZMB) induces py op o ic and immunogenic cell dea h
in umo cells [35]. Su p isingly, e en caspase-8 (a e Ye sinia in ec-
ion) and caspases 4/11 can ini ia e signaling pa hways leading o
GSDME clea age and cell dea h [36]. Immunocy e- eleased G anzyme-A
(GZMA) clea age o GSDMB can p oduce ei he ly ic cell dea h in cance
cells [37] o selec i e killing o Shigella in acellula bac e ia bu no he
in ec ed cells [38]. Mo eo e , unp ocessed GSDMB enhances caspase-4/
GSDMD non-canonical py op osis [39]. Finally, bo h TNF-
α
+hypoxia
[40] and
α
-Ke oglu a a e [41] induce Caspase-8 clea age o GSDMC bu
in di e en esidues (Fig. 1A). GSDMC NT p o okes py op osis in cance
cells bu i could esul in ch onic umo nec osis [40]. The p o eases
ac i a ing human and mouse GSDMA and PJVK p o eins a e s ill un-
known. No ably, cell-dea h unc ion can be inhibi ed by p o ease p o-
cessing wi hin he GSDM NT domain, like GSDMD clea age by caspases-
3/7 [42] o en e o i us 3C p o ease [43]. Likewise, di e se caspases (1/
3/6/7) can clea e and inac i a e he GSDMB NT [16,39], con adic ing
he wo k by Panganiban and collabo a o s, which sugges ed ha
caspase-1 clea age o GSDMB linke induced py op osis du ing as hma
[44].
O no e, GSDM-media ed cell dea h is mo e complex han me ely
o ming cell memb ane po es, since some GSDM NT can also a ge
in acellula o ganelle (mi ochond ia, neu ophil g anules and possibly
he nucleus) [4,7] (Fig. 1C). In pa icula , ac i a ed GSDMs (GSDMA/
A3/D/E) igge mi ochond ial damage [45–48], and con e sely,
al e ed mi ochond ial unc ion induces GSDMD/E clea age [49]. In ac ,
ime lapse mic oscopy e eals ha du ing py op osis mi ochond ial
damage p ecedes cell lysis [45,46]. Mi ochond ia dys unc ion can occu
in di e se ways. A e caspase-3/GSDME clea age, GSDME po es pe -
meabilize he mi ochond ial memb ane leading o downs eam apop-
osome ac i a ion [49]. Thus, GSDME ac i a ion cause a posi i e
eedback loop enhancing mi ochond ial apop osis and/o py op osis. By
con as , cons i u i ely ac i e mu an GSDMA3 p o eins could cause
mi ochond ial damage by wo mechanisms: binding o he mi ochon-
d ial chape one T ap1, which p omo es oxida i e s ess and loss o he
mi ochond ia memb ane po en ial igge ing apop osis-independen cell
dea h [47], o h ough s imula ion o p o cell-dea h au ophagy [48].
Consis en wi h he GSDM mi ochond ial a ge ing, GSDM NTs exhibi
s ong binding a ini y o ca diolipin, a lipid en iched in he in e nal
mi ochond ial memb anes [15,50].
Taken oge he , hese e idences p o ed ha GSDMs coo dina e an
ex ensi e and complex c oss- alk be ween di e se cell dea h pa hways
[22,34], bu he p ecise unc ions o each GSDM in physiology and
disease has only s a ed o eme ge.
The molecula mechanisms egula ing in lammasome-dependen
and independen py op osis and he implica ion o GSDMs in physio-
logical p ocesses (e.g., esponse o in ec ious agen s) and pa hologies has
been ex ensi ely e ised elsewhe e [7,19,20]. As examples, GSDM p o-
cell dea h unc ions a e in ol ed in mul iple in lamma o y pa hologies
such as sepsis, au oimmune encephalomyeli is (GSDMD), as hma and
in lamma o y bowel diseases (GSDMB), among o he s. Addi ionally,
a e GSDM mu a ions (Fig. 1) p o oke pa hogenic pheno ypes in mice
(Gsdma3 mu a ions ha impede CT p o ein inhibi o y unc ion igge
skin in lamma ion and hai loss, among o he de ec s) and humans
(GSDME and PJVK unca ing mu a ions p oduce he edi a y dea ness
diso de s) [7,19,20].
In his e iew we p o ide a comp ehensi e o e iew on he mul i-
ace ed oles o he GSDM amily in cance , co e ing no only hei p o-
cell dea h ac i i ies in pa icula umo con ex s, bu also o he unc ions
ha e en ually could lead o umo p og ession. Mo eo e , we will ocus
on he implica ion o GSDMs in media ing cance esponse o ea men
and he ele ance o GSDMs as no el he apeu ic a ge s o clinical
oncology managemen , wi h special emphasis on no el GSDM-di ec ed
nano- he apies.
3. An i- umo and p o- umo e ec s o GSDMs
The p o-cell dea h ac i i y o GSDMs could lead o he idea ha hese
p o eins play mainly an an icance unc ion. Ye , GSDM genes a e o en
loca ed in genomic egions equen ly ampli ied in cance s (Table 1),
D. Sa i´
o e al.
BBA - Re iews on Cance 1876 (2021) 188635
3
Table 1
Exp ession and unc ional oles o GSDMs in human cance s.
Cance ype
(sugges ed unc ion*)
Exp ession in umo s (me hod) & e ec s in un ea ed cance cells. Re
Human GSDMA (GSDM, GSDM1) [17q21.1]
Mouse Gsdma1/2/3 [11D]
B eas , gas ic & o a ian
(an i- umo ) No exp ession was de ec ed in any o he 24 cance cell lines (No he n blo ), e en hough GSDMA gene was ampli ied in 4/4 (100%) b eas and 2/8 (25%) gas ic HER2 cell lines. [52]
Esophageal & Gas ic (an i-
umo ) Unde ec able exp ession in 58/60 (97%) umo s and 17/21 (81%) cell lines (RT-PCR). O e exp ession dec eases colony o ma ion in MKN28 cells. [1]
Gas ic
(an i- umo )
Unde ec able exp ession in 11/18 (61%) umo s and 8/11 (73%) cell lines (RT-PCR). GSDMA is es o ed by inhibi ion o p omo e hype me hyla ion in 6/8 (75%) lines. O e exp ession in cell lines
dec eases cell g ow h and induces cell dea h. [51]
O a ian
(p o- umo ) G ea e mean exp ession in umo s (n =379) compa ed o no mal issue (n =88) (mRNA DBs). Highe GSDMA cance exp ession associa es wi h poo su i al. [53]
Human GSDMB (GSDML, PRO2521) [17q21.1]
No p esen in mouse
Bladde
(p o- umo )
La ge mean exp ession in umo s compa ed o no mal issue (n =19) (mRNA DBs). GSDMB and USP24 p o ein exp ession a e posi i ely co ela ed in bladde cance s (n =80; IHC). GSDMB silencing in
T24 and 5637 cells dampens p oli e a ion, mig a ion and in asion and educes in i o umo g ow h o T24 xenog a s. GSDMB con ols glycolysis and in i o cance g ow h ia STAT3 signaling. [59]
B eas
(p o- umo )
G ea e mean exp ession in umo s compa ed o no mal issue (RT-PCR, n =18). Highe mRNA umo exp ession associa es wi h poo p ognosis (n =1628; mRNA DBs). I s o e -exp ession in MCF7 cells
induces cell mo ili y, in asion, gela in deg ada ion in i o & in i o umo g ow h and me as asis (mos ly iso o m 2). GSDMB silencing educes mig a ion and in asion o HCC1954 cells. [56]
HER2 B eas
(p o- umo )
O e exp essed (80/212; 66%) (IHC) and gene-ampli ied (FISH) (73/123; 59%) in HER2-posi i e umo s. Highe le els (n =212; IHC & n =2096; mRNA DBs) associa e wi h me as asis, poo p ognosis
and ea men elapse. [55]
HER2 B eas (p o- umo ) GSDMB exp ession p omo es mig a ion in HCC1954, SK-BR-3, and BT474 cells. Enhances lung me as asis in MDA-MB-231 cells. [61]
Ce ical
(p o- umo ) Mo e equen ly exp essed in umo s (16/21; 76%) han co esponding non-neoplas ic a eas (8/21; 38%) (IHC). I s silencing educes cell g ow h in Hela cells. [57]
Ce ical (unce ain) Equal exp ession equency in umo s (23/27; 85%) han co esponding non-neoplas ic a eas (24/27; 89%) (IHC). [37]
Colo ec al (unce ain) Simila exp ession equency in umo s (166/230; 72%) han co esponding non-neoplas ic a eas (154/230; 67%) (IHC). [37]
Esophageal (an i- umo ) Less equen exp ession in umo s (44/80; 55%) han co esponding non-neoplas ic a eas (72/80; 90%) (IHC). [37]
Esophageal & Gas ic
(p o- umo ) Exp essed in 47/60 (78%) umo s and 21/21 (100%) cell lines (RT-PCR). Co-ampli ied wi h HER2 in 2/8 (25%) gas ic cance s. I s o e exp ession in MKN28 cells does no a ec colony o ma ion. [1]
Gas ic
(an i- umo ) Less equen exp ession in umo s (34/75; 45%) han co esponding non-neoplas ic a eas (66/75; 88%) (IHC). [37]
Gas ic
(p o- umo ) Mo e equen ly exp essed in umo s (44/52; 85%) han no mal issues (29/82; 35%) (RT-PCR). S onge exp ession in umo cells compa ed o no mal cells (ISH, n =5). [58]
Gas ic, CRC & HCC
(unce ain) Simila mean exp ession be ween umo s (gas ic n =21; HCC n =15; colon n =9) han co esponding non-neoplas ic a eas (RT-PCR). [6]
HCC (unce ain) Simila exp ession equency in umo s (17/21; 81%) han co esponding non-neoplas ic a eas (21/21; 100%) (IHC). I s silencing does no a ec cell g ow h in HepG2 cells. [57]
Lung
(p o- umo ) Highe mean exp ession in lung adenoca cinomas han no mal issue (n =515; mRNA DBs). [54]
Mul iple cell line ypes GSDMB p o ein endogenous exp ession de ec ed only in 18/54 (33%) cell lines (WB). I s exp ession could be induced by IFN-Ɣ in 11 cell lines (WB). GZMA induces py op osis killing o GSDMB-exp essing
cell lines. [37]
OSCC
(p o- umo ) Highe mean exp ession in lymph-node me as a ic cance s s non-me as a ic (n =53; mic oa ays and RT-PCR) [60]
Panc ea ic (unce ain) Simila exp ession equency in umo s (41/77; 53%) han co esponding non-neoplas ic a eas (45/77; 45%) (IHC). [37]
Human GSDMC (MLZE) [8q24.21]
Mouse Gsdmc1–4 [15D1]
B eas
(p o- umo ) Highe le els associa e wi h wo se o e all su i al (n =626; IHC). [40]
Colo ec al
(p o- umo )
Highe mean exp ession in umo s s adjacen no mal issue (n =44; RT-PCR). GSDMC p o ein de ec ed only in umo s, no in no mal issue (n =44; IHC). GSDMC was silenced (DLD-1 and LoVo cells) o
o e -exp essed (SW480 and WiD cells). GSDMC up egula ion p omo ed in i o p oli e a ion, ancho age-independen g ow h and in i o umo igenesis. [68]
Esophageal & Gas ic (an i-
umo ) Exp essed in 34/60 (57%) o umo s and 16/21 (76%) cell lines (RT-PCR). I s o e exp ession educes sligh ly colony o ma ion in MKN28 cells. [1]
(con inued on nex page)
D. Sa i´
o e al.
BBA - Re iews on Cance 1876 (2021) 188635
4
Table 1 (con inued)
Cance ype
(sugges ed unc ion*)
Exp ession in umo s (me hod) & e ec s in un ea ed cance cells. Re
Lung
(p o- umo )
G ea e mean exp ession in lung adenoca cinomas s no mal issue (n =515; mRNA DBs). Highe GSDMC exp ession associa es wi h poo p ognosis, me as asis and adio- esis ance (mRNA DBs).
Hype me hyla ion associa es wi h gene silencing. [54]
Melanoma
(p o- umo ) Exp essed in 8/26 (30%) melanomas bu no in ne i (n =5). Mo e equen ly exp essed in me as a ic (6/11; 55%) han non-me as a ic (2/15; 13%) melanomas (IHC). [67]
Human GSDMD (GSDMDC1, DFNA5L, DF5L, FKSG10) [8q24.3]
Mouse Gsdmd [15D3-E1]
ESCC (unce ain) Highe mean exp ession in umo s han in no mal issues (n =148; mRNA DBs and n =30; IHC). [133]
Esophageal & Gas ic (an i-
umo ) Exp essed in 38/60 (63%) o umo s and 21/21 (100%) cell lines (RT-PCR). I s o e exp ession s ongly educes colony o ma ion in MKN28 cells. [1]
Gas ic
(an i- umo )
S ong exp ession mo e equen in no mal issue (41/61; 67%) han in umo s (27/102; 27%) (IHC). mRNA le els highe in no mal s ma ched umo issue (n =39; RT-PCR). GSDMD exp ession in
BGC823 cells educes in i o p oli e a ion and in i o cance g ow h. [69]
Lung
(p o- umo )
Highe exp ession (IHC sco e; n =168) in adenoca cinomas and SCCs han no mal issue. O e exp ession associa es wi h inc eased umo size, s age and lowe su i al in adenoca cinomas. GSDMD
silencing in PC9, H1703 and H1975 cells educes p oli e a ion, p omo es apop osis and a enua es in i o umo g ow h. [70]
Os eo-sa coma
(p o- umo )
Exp essed in 20/41 (49%) umo s and none o non-neoplas ic a eas (IHC). G ea e mean p o ein exp ession in umo s han ma ched no mal issues (n =61; WB). Highe le els associa e wi h me as asis,
esis ance o chemo he apy, and poo su i al. [134]
Sali a y ACC (p o- umo ) Mo e equen ly exp essed in ACC (33/33; 100%), han adenomas (23/29; 79%) & no mal issues (24/33; 73%) (IHC). S ong IHC exp ession only seen in ACC. O e -exp ession in ACC-LM and ACC-83
cells enhances in i o in asion. [71]
Human GSDME (DFNA5) [7p15.3]
Mouse D na5 [6B2.3]
B eas
(an i- umo )
Lowe mean exp ession in umo s s no mal samples (n =1142; mRNA DBs). Reduced GSDME in umo s does no associa e wi h su i al. G ea e GSDME le els in ER-nega i e and lobula ca cinomas.
Highe p omo e hype me hyla ion in umo s. GSDME gene body me hyla ion associa es wi h educed o e all su i al. [79]
B eas
(an i- umo )
Reduced mean exp ession in cance s no mal samples (n =2509; mRNA DBs). Highe umo GSDME does no associa e wi h su i al bu co ela e wi h immune- ela ed genes. No associa ion be ween
p omo e me hyla ion and gene exp ession. [96]
B eas
(an i- umo )
Dec eased mean exp ession in cance s s no mal samples (n =10; RT-PCR). Highe me hyla ion in umo s. GSDME exp ession can be ac i a ed wi h deme hyla ing agen s. GSDME silencing inc eases
colony o ma ion, p oli e a ion and in asion o MDA-MB-231 cells. [75]
B eas and colon
(an i- umo )
Lowe mean exp ession in b eas (n =1097) and colon (n =286) cance s s no mal samples (mRNA DBs). GSDME supp esses in i o umo g ow h and p omo es an i- umo immuni y o mu ine cells
(EMT6, CT26, 4 T1 & B16–10) in immunocompe en mice. [35]
Colo ec al (an i- umo ) Lowe exp ession in cance s s no mal issue (n =5) (RT-PCR). Highe hype me hyla ion in umo s (65%) han no mal issue (3%). GSDME in HCT116 cells educes cell g ow h and colony o ma ion. [73]
Colo ec al (unce ain) Using Gsdme KO mice, no clea e ec s we e seen in wo expe imen al models o in es inal cance : he chemical induc ion by azoxyme hane “AOM” o c ossing wi h he Apc1638N/+s ain. [82]
ESCC
(an i- umo ) G ea e exp ession (IHC sco e) in cance s compa ed o no mal issue (IHC). Inc eased GSDME in SCCs (n =104; IHC) associa es wi h enhanced he apy esponse and be e p ognosis. [116]
Gas ic
(an i- umo )
F equen p omo e hype me hyla ion in umo s (53%). GSDME exp ession up egula ed by deme hyla ing agen s in gas ic cell lines. GSDME o e -exp ession educes colony o ma ion and augmen s
apop osis in NUGC3 cells. [74]
GBM (unce ain) Highe mean exp ession in umo s han in no mal b ain (mRNA DBs), bu GSDME le els do no associa e wi h pa ien s’ su i al. [89]
Lung
(unce ain) Exp essed in 59% o cance s, simila le els han no mal issues (IHC). Exp essed in 95% o cell lines (WB). GSDME does no a ec umo g ow h o human NCI-H3122 and HCC827 xenog a s. [78]
Mul iple
cell lines
(an i- umo )
P o ein de ec ed in 30/57 (53%) cell lines, s ong exp ession in 11/57 (19%) (WB). Gsdme KO mice do no exhibi de elopmen al de ec s. [33]
Melanoma (an i- umo ) GSDME silencing in B16-O a cells inc eases in i o umo g ow h. [49]
Human PJVK (Pej akin, DFNB59) [2q31.2]
Mouse D nb59 [2C3]
O a ian
(an i- umo ) Lowe mean exp ession in umo s (n =379) compa ed o no mal issue (mRNA DBs). Lowe PJVK cance exp ession associa ed wi h poo su i al. [53]
Table ows a e o de ed alphabe ically by GSDM gene name (aliases indica ed in pa en heses and gene locus in squa e b acke s) and hen by umo ypes. * The o e all e ec (p o en o in e ed) on umo biology
(excluding e ec s on cance ea men ) acco ding o s udy esul s. Abb e ia ions: ACC: Adenoid Cys ic Ca cinoma; CRC: Colo ec al; ESCC: Esophageal Squamous Cell Ca cinoma; GBM: Glioblas oma Mul i o me; IHC:
Immunohis ochemis y; ISH: in si u hyb idiza ion; mRNA DBs: Da a Bases o mRNA exp ession (mic oa ays o RNAseq); SCC: Squamous Cell Ca cinoma; WB: Wes e n Blo .
D. Sa i´
o e al.
BBA - Re iews on Cance 1876 (2021) 188635
5
such as he 17q12–21 egion (GSDMA and GSDMB), ha con ains he
HER2/ERBB2 oncogene, o he 8q24 egion (GSDMC and GSDMD), close
o MYC oncogene. Mo eo e , GSDMs a e ei he up egula ed o down-
egula ed (mRNA and/o p o ein) in a wide ange o cance ypes
compa ed o hei espec i e no mal issues (Table 1). In ac , accumu-
la ing e idences indica e ha GSDMs play complex oles in cance
biology, possibly ha ing ei he p o- umo o an i- umo unc ions
depending on he cellula con ex (summa ized in Table 1). The
in ol emen o GSDMs in oncologic ea men esponse will be p e-
sen ed in sec ion 4.
GSDMA was ini ially desc ibed as equen ly los in gas o-
esophageal cance s [1,51] and b eas cance cells [52], e en in HER2-
posi i e cell lines wi h GSDMA gene co-ampli ica ion [52]. GSDMA
exp ession could be es o ed by me hyla ion inhibi o s [51]. In gas ic
cance cells, GSDMA up egula ion, media ed by TGF-β ( ans o ming
g ow h ac o -β) and he ansc ip ion ac o LMO1 (LIM domain only
1), p omo ed an apop o ic signal leading o educed cell g ow h [51].
These da a poin o a po en ial umo -supp esso gene (TSG) ole o
GSDMA. Con a ily, in exp ession da abases GSDMA mRNA was up e-
gula ed in o a ian ca cinomas e sus no mal issue, and highe GSDMA
le els associa ed wi h wo se su i al [53]. Likewise, GSDMA mRNA is
up egula ed in some lung cance da ase s [54] and b eas umo s
[55,56], bu he ele ance o hese obse a ions is unce ain. In ac ,
despi e being he i s GSDM gene iden i ied [5], he mechanism o
clea age/ac i a ion and he p ecise unc ional e ec s o GSDMA in
cance emains unclea .
Unlike o he GSDMs, GSDMB does no consis en ly educe cell
g ow h o gas ic cance cells o o he cell models [1,56,57]. Mo eo e ,
con a y o i s neighbo gene GSDMA, GSDMB is equen ly exp essed
(mRNA and/o p o ein) in se e al human umo ypes and cance cell
lines, including gas ic, hepa ic, and b eas , among o he s (Table 1).
None heless, compa ed o no mal issues, GSDMB le els a e ei he
up egula ed [1,54–59], down egula ed [37] o unchanged [6,37,57]
depending on he umo ype and he me hods used o measu e i s
exp ession (Table 1). In e ms o mRNA, GSDMB o e -exp ession asso-
cia es wi h ad anced/me as a ic disease in o al SCC [60] and gas ic
umo s [58], sho e disease ee and me as asis ee su i al in b eas
cance [56], and o e all su i al in only 3 o 33 cance ypes es ed by
Zhou and colleagues [37]. Howe e , since GSDMB mRNA is de ec ed in
a ious no mal cell ypes [7] he ue GSDMB exp ession in umo s
should be assessed by immunohis ochemis y. Using a alida ed GSDMB
an ibody and a speci ic FISH p obe, He gue a-Redondo and colleagues
[55] demons a ed ha GSDMB is o e -exp essed/gene-ampli ied in
>60% o HER2 b eas cance s. Simila ly, in gas o-esophageal cance s
GSDMB and HER2 co-ampli ica ion is a common inding [1]. Impo -
an ly, in HER2 b eas ca cinomas, GSDMB p o ein up egula ion asso-
cia ed signi ican ly wi h a ious ad e se clinical pa ame e s: disease
p og ession, elapse and esponse a e neoadju an he apy, as well as
dis an me as ases and lymph node posi i i y in he adju an se ing,
independen ly o ho monal ecep o s s a us [55]. Mo eo e , GSDMB
exp ession media es mul iple p o- umo unc ions in b eas neoplasias:
inc eases in i o cell mig a ion, possibly by modula ing he Rac GTPa-
ses, in asion and gela in deg ada ion [56], s imula es in i o me as a ic
dissemina ion [56,61], and educes sensi i i y o an i-HER2 he apy.
Ac ually, all hese p o- umo ac i i ies could be dec eased wi h a no el
an i-GSDMB nano he apy [61], hus demons a ing ha GSDMB plays a
key ole in he biology and clinical beha io o HER2 b eas ca cinomas.
Simila ly, in bladde cance cells GSDMB p omo es p oli e a ion,
Fig. 1. GSDM po e- o ming unc ions. A: Regula ion o GSDM unc ion by clea age o mu a ion. Schema ic ep esen a ion o GSDM egions (NT, N- e minal
domain; CT, C- e minal; and linke in e domain) showing he known p o eases and espec i e clea age si es (in pa en heses, he clea age si es o mu ine p o eins a e
indica ed in g ey le e s). The clea age e en s inducing po e- o ming ac i i y a e shown in ed lines, and inhibi o y p ocessing in black. Gain-o – unc ion mu a ions
in he CT o GSDMA3 and GSDME a e indica ed by as e isks. B: Main s eps in he o ma ion o GSDM memb ane po es. The e a e di e ences in he s uc u e and size
o GSDM po es. GSDMA3 o m po es con aining a ound 27 monome s and wi h an in e nal po e diame e o 16–18 nm; GSDMD, 33 monome s and up o 21 nm
in e nal diame e . C: Summa y o mechanisms o GSDM ac i a ion and biological e ec s. Casp, Caspase; Ca G Ca hepsin G; GZM, G anzyme; NE, Neu ophil Elas ase.
(Fo in e p e a ion o he e e ences o colou in his igu e legend, he eade is e e ed o he web e sion o his a icle.)
D. Sa i´
o e al.

BBA - Re iews on Cance 1876 (2021) 188635
6
mig a ion and in asion [59]. These e idences and he p e ious obse -
a ions in gas ic ca cinomas [1,58] indica e ha GSDMB could ac like
an oncogene. Howe e , he in i o e ec o GSDMB on umo igenesis
and p og ession migh depend on he cance models, since i p omo es
umo g ow h in MCF7 b eas [56] and T24 bladde cance xenog a s
[59] bu no o he human [61] and mu ine [37] cance models (Table 1).
A his poin , i is impo an o highligh he exis ence o mul iple
GSDMB ansc ip ional a ian s [6,57] and a leas ou dis inc ans-
la ed p o ein iso o ms (ENSEMBL:ENSG00000073605). Though, o
unknown easons he 411 aa p o ein sequence -Q8TAX9-, which was
pa ially c ys alized [16], and is ega ded as “canonical” in all da a-
bases, does no ma ch any o he ansc ip ional a ian s. The ou
ansla ed iso o ms, which di e on he al e na i e usage o exons 6–7,
can play di e en unc ions in cance [56] and in lamma o y diseases
[44,62]. The iso o m di e en ial exp ession was epo ed in no mal and
cance issues [6,57,58,63–65] and could be egula ed by speci ic
genomic elemen s [58,64,65] and SNPs [44,63,66]. Howe e , he e is
sca ce in o ma ion on he p ecise unc ions o each ansla ed a ian in
no mal and pa hological con ex s. In his ega d, He gue a-Redondo
e al. epo ed ha in MCF7 b eas cance cells bo h GSDMB iso o m 1
(GSDMB-1; lacks exon 6) and 2 (GSDMB-2; he sho es iso o m, lacking
exons 6–7) enhanced mig a ion and in asion in i o, while only
GSDMB2 p omo ed in i o umo g ow h and me as a ic dissemina ion
in immunode icien mice [56].
Opposing i s p o- umo unc ions, GSDMB can, unde speci ic s im-
uli, exhibi py op o ic ac i i y in cance cells [37,61], and hus an
an i umo e ec . Speci ically, in a con ex o an ac i a ed an i umo
immune esponse, he lymphocy e-de i ed G anzyme A (GZMA) can
clea e GSDMB wi hin cance cells (Fig. 1A,C), ac i a ing caspase-
independen py op osis and p omo ing umo clea ance [37]. Con-
as ing wi h o he s udies (Table 1), Zhou and cols [37] epo ed a
lowe GSDMB immunohis ochemical s aining in gas o-esophageal u-
mo s compa ed o no mal issues, bu no o he cance ypes (Table 1),
sugges ing ha GSDMB could ac as umo supp esso in hese neo-
plasias. Un o una ely, his s udy did no e alua e he associa ion o
GSDMB s aining wi h clinicopa hological a iables o disease su i al,
hus he clinical ele ance o hese obse a ions emains unclea .
GSDMC was ini ially iden i ied as a ma ke o melanoma p og es-
sion, being pa icula ly o e -exp essed in me as a ic melanoma cells
[67]. Fu he s udies epo ed opposing esul s ega ding i s exp ession
and unc ional e ec s in umo s. On one hand, Saeki and cols ound
GSDMC exp ession in 50% o gas o-esophageal umo s and cell lines
bu was mos ly los in di use- ype gas ic ca cinomas. In gas ic cance
cells, GSDMC exp ession educed cell p oli e a ion, sugges ing a
possible umo supp esso unc ion [1]. On he o he hand, GSDMC
up egula ion in lung [54] and b eas [40] umo s associa es wi h
ad e se clinical pa ame e s (Table 1), and inc eases p oli e a ion along
wi h in i o umo g ow h in colo ec al cance cells [68], sugges ing a
p o- umo ole. Ac i a ion o GSDMC clea age by d ugs o me aboli es
(
α
-KG) [41] induces py op osis in cance cells (Fig. 1; Table 2), bu can
also p o oke ch onic nec osis [40]. In he la e case, unde hypoxia
nuclea PD-L1 ac i a es GSDMC ansc ip ion in b eas cance cells. In
pa allel, ac i a ed mac ophages sec e e TNF-
α
esul ing in caspase-8
ac i a ion wi hin cance cells. The coo dina ed up egula ion o
GSDMC and caspase-8 induces GSMDC clea age, causes py op osis and
subsequen nec osis in b eas cance cells [40]. Pa adoxically, ch onic
umo nec osis uels cance p og ession, and he e o e pa ien s wi h
GSDMC-o e exp essing b eas cance s show poo su i al [40]. This
s udy challenges he idea ha GSDM-media ed cance cell dea h can
ha e bene icial e ec s on cance p og ession.
Whe eas GSDMD ole in leukocy e py op osis is well known, i s
implica ion in cance is deba able. Depending on he s udy, bo h
GSDMD up egula ion and down egula ion (mRNA and/o p o ein) ha e
been desc ibed in umo s, whe e GSDMD o e exp ession associa es wi h
ei he good o un a o able p ognosis, as well as an i umo o p o- umo
e ec s (Table 1). Fo ins ance, GSDMD exp ession diminishes
p oli e a ion in gas ic cance cells, ei he ia cell dea h induc ion [1] o
by egula ing cell cycle- ela ed p o eins [69], bu i inc eases lung
cance g ow h in i o [70] and in asion o sali a y gland adenoca ci-
nomas [71].
Among he GSDMs, GSDME exhibi s he mos consis en an i- umo
unc ions in he li e a u e and is commonly conside ed as a po en ial
TSG o se e al easons. Fi s , GSDME exp ession in cance models
(un ea ed cells) inhibi s cell g ow h and/o p o okes cell dea h
[72,73]. Second, di e se an i-oncologic ea men s induce GSDME
clea age and enhance cance dea h (Table 2; discussed in sec ion 4).
Thi d, GSDME is usually exp essed in no mal issues bu is equen ly
down egula ed in di e se cance ypes, being his al e a ion some imes
associa ed wi h inc eased malignancy o poo p ognosis (Table 1).
Fou h, p omo e hype me hyla ion (a common mechanism o silencing
TSGs) o GSDME is commonly obse ed in cance s [72–77]. Besides,
GSDME loss-o - unc ion mu a ions occu in a small p opo ion o umo s,
as an al e na i e mechanism o me hyla ion o inac i a ing GSDME-
media ed cance cell dea h [35]. Finally, he umo supp esso ac i i y
o GSDME is media ed by i s clea age by kille -cell GZMB, esul ing in
caspase-independen py op osis o cance cells and enhancemen o an i-
umo immuni y [35].
Despi e hese e idences, and he consis en e ec o GSDME on
umo sensi iza ion o cance he apies (Table 2), i is s ill ques ionable
i GSDME pe se gene ally ac s as a bona ide TSG, and whe he he
e alua ion o umo GSDME p o ein le els ha e a eliable p ognos ic
u ili y (Table 1). Fo ins ance, GSDME p o ein up egula ion was
obse ed in he majo i y in lung umo s and cell lines, and GSDME
clea age (py op o ically ac i e) was de ec ed e en in un ea ed speci-
mens [78]. Besides, GSDME p omo e hype me hyla ion does no uni-
e sally co ela e wi h gene silencing in di e se cance da ase s
(Table 1) [77]. In e es ingly, he gene body o GSDME is equen ly
hypome hyla ed in cance compa ed o no mal issues, and in ac , he
pa icula pa e ns o me hyla ion in he GSDME gene p omo e and
body egions, a he han GSDME exp ession, can be used as a powe ul
bioma ke o pan-cance de ec ion [79–81]. Apa om human sam-
ples, s udies wi h in i o animal models epo ed seemingly con adic-
o y esul s ega ding GSDME ole in umo de elopmen and
p og ession. In xenog a cance models, GSDME does no a ec umo
g ow h o human lung cance cells [78], bu educes umo g ow h o
mu ine melanoma [49], b eas , and colon cell lines [35]. Mo eo e ,
compa ing Gsdme Knock-ou (KO) mice wi h wild ype (WT) animals in
wo expe imen al models o in es inal cance , no clea e ec s on ca ci-
nogenesis, umo di e en ia ion and p og ession we e e idenced,
hough an inc eased umo al in lamma ion was obse ed in WT mice
[82]. By con as , Gsdme KO mice exhibi educed umo igenesis in
in lamma ion–media ed coli is-associa ed colo ec al cance models
[83], sugges ing ha GSDME-media ed py op osis and in lamma ion
play a ole in cance ini ia ion. None heless, hese con lic ing da a could
be pa ly explained conside ing new indings demons a ing ha
GSDME umo supp essi e e ec depends mos ly on he subsequen
umo in lamma ion and immunogenic ac i a ion (see sec ion 6).
Finally, hi he o, he implica ion o PVJK is la gely unknown.
Recen ly, using o a ian ca cinoma exp ession da abases, PJVK was
ound down egula ed in umo s compa ed wi h no mal samples, and
lowe PJVK mRNA le els in cance s associa ed wi h poo su i al a es
[53].
4. GSDMs modula e umo esponse o an i-cance ea men s
E ading apop osis is a cance hallma k ha can p o ide esis ance o
an i-cance he apies. In his scena io, ac i a ing py op osis o o he cell
dea h mechanisms could lead o umo eg ession. GSDM-media ed cell
dea h, which can p oceed in caspase-dependen and independen ways
(Fig. 1), occu s in esponse o mul iple cell-damaging s imuli and an i-
cance ea men s (chemo he apy, a ge ed d ugs and immuno-
he apy) (Table 2). These da a would b ing o he idea ha GSDM-
D. Sa i´
o e al.
BBA - Re iews on Cance 1876 (2021) 188635
7
Table 2
E ec o GSDMs on an i-cance he apy esponse.
Tumo s /cell lines T ea men E ec on cance cells Re
GSDMB
HER2 B eas umo s, PDXs &
cell lines T as uzumab Highe le els associa e wi h poo ea men esponse (adju an o neoadju an con ex s) in human umo s. T as uzumab esis ance associa es wi h GSDMB up egula ion
in human PDXs. GSDMB exp ession p omo es d ug su i al in SK-BR-3 and HCC1954 cells. [55]
HER2 B eas cell lines T as uzumab GSDMB up egula ion inc eases as uzumab su i al in HCC1954, SK-BR-3, and BT474 cells. This e ec can be pa ially blocked wi h an i-GSDMB nano he apy. [61]
Colon & Melanoma cell lines An i-PD1 Exogenous GSDMB o e -exp ession in mu ine CT26 and B16-F10 cell line xenog a s sensi izes o an i-PD1 ea men in i o. [37]
GSDMC
B eas cell line Mul iple chemo d ugs Mos d ugs up egula e GSDMC bu only he an ibio ic- ype induce caspase-8/GSDMC py op osis in MDA-MB-231 cells. [40]
Mul iple cell lines DM-
α
-KG Induces ROS and ac i a ion o DR6/caspase-8 axis leading o GSDMC py op osis in mul iple human and mouse cance cell lines in i o. DM-
α
-KG educes in i o umo
g ow h (Hela and B16 xenog a s) and me as asis (B16) and his e ec depends on he p esence o GSDMC and DR6. [41]
GSDMD
AML p ima y cul u es and
lines
Val-bo oP o (DPP8/9
inhibi o )
Ac i a es CARD8/caspase-1/GSDMD-media ed py op osis and e icien ly kills AML p ima y cells and 12/17 AML cell lines in i o. Hal s umo p og ession o one PDX
and MV4;11 cell line xenog a s in i o. [86]
NPC
cell lines Taxol Caspase-1 clea es GSDMD and ac i a es py op osis in i o in HNE-2, 5-8F cell lines. GSDMD silencing does no a ec umo g ow h in i o in un ea ed 5-8F xenog a s.
GSDMD sensi izes 5-8F xenog a s o axol ea men . [135]
Esophageal SCC cell lines Me o min Me o min up egula es miR-497, which in u n down egula es PELP1. PELP1 educ ion inc eases GSDMD py op osis in KYSE510 and KYSE140 cells. [133]
O a ian cell lines Alpha-NETA Induces caspase-4 and GSDMD up egula ion and cell dea h in Ho8910PM cells. GSDMD silencing pa ly inc eases cell su i al o he compound. [136]
GSDME
(TN) B eas cance cell lines Te a‑a senic hexoxide Induces mi ochond ial ROS-media ed caspase-3/GSDME clea age and py op osis in mouse EO771, 4T1 and human Hs578T, MDA-MB-231 cance cells bu no in non-
umo igenic cell lines (MCF10A, NMuMG). GSDME silencing educes d ug-induced cell dea h. [125]
(TN) B eas cell lines Ce uximab miR-155-5p an agomi up egula es GSDME and swi ches apop osis o py op osis a e ce uximab ea men in EGF-o e exp essing MDA-MB-231 and MDA-MB-468
cells. [112]
Colon
cell lines Lobapla in A e lobapla in, caspase-3 clea es GSDME and p omo es py op osis ia ROS/JNK/bax mi ochond ial signaling pa hway in HT-29 and HCT116 cells. GSDME silencing
swi ched om py op osis o apop osis bu did no a ec in i o umo g ow h upon lobapla in ea men . [90]
Colon
cell lines TNF +CHX o na i oclax These d ugs induce BAK/BAX/Caspase-3 ac i a ion and GSDME clea age and py op osis in HCT116 cells. GSDME silencing educes cell dea h o ea men . [121]
Colon
cell lines Ionizing adia ion (IR) IR induces GSDME-py op osis ia he LncRNA NEAT1 and miR488 in HCT116 cells. [113]
Gas ic
cell lines 5-FU S imula es Caspase-3 and GSDME clea age and py op osis in SGC-7901 and MKN-45 cells. GSDME knock ou swi ches py op osis o apop osis. [137]
GBM
cell lines
Galangin (na u al
la onoid)
Simul aneously induces au ophagy plus caspase-3/GSDME clea age and py op osis in U87MG and U251 cells. GSDME silencing swi ches py op osis o apop osis.
Inhibi ion o au ophagy plus Galangin inc eases cell dea h in i o in U87MG xenog a s. [89]
Esophageal SCC
cell lines
Cispla in
and
BI2536
The Plk1 inhibi o BI2536 combined wi h cispla in p o okes caspase-3/GSDME clea age and py op osis in nine SCC cell lines in i o. The d ug combina ion se e ely
educes in i o umo g ow h o KYSE150 umo xenog a s. [116]
Head & Neck cell lines T ip olide T ip olide ac i a es BAD/BAX-caspase 3-GSDME py op osis by ep essing mi ochond ial associa ed hexokinase-II and p o oking ROS in HK1 and FaDu cells. Combining
T ip olide wi h e as in (SLC7A11 inhibi o ) hal s in i o umo g ow h in HK1 xenog a s. [88]
Li e
cell lines Mil i one Induces ROS and mi ochond ial damage leading o caspase-3/GSDME clea age and py op osis in HepG2 and Hepa1–6 cells in i o and cell dea h in i o using Hepa1–6
umo xenog a s. GSDME silencing swi ches py op osis o apop osis. [126]
Lung
cell lines
and p ima y human umo s
T ame inib, E lo inib
o
Ce i inib
GSDME sensi izes o T ame inib, e lo inib and ce i inib a ge ed he apies in i o and in i o using mul iple cell lines. GSDME knock ou swi ches py op osis o apop osis
bu does no a ec umo g ow h when un ea ed. GSDME py op osis was obse ed in pa ien s ea ed p ima y umo s. [78]
Lung
cell line
Cispla in
o
pacli axel
Cispla in p o okes a s onge ac i a ion o caspase-3/GSDME clea age and py op osis han pacli axel in A549 cells. GSDME knockdown inhibi s cispla in- bu no
pacli axel-induced pyop osis in i o. [138]
Lung & neu oblas oma
cell lines Dasa inib Dasa inib up egula es GSDMD and GSDME p o ein le els and p o okes caspase-3/GSDME clea age and py op osis in A549 and SH-SY5Y cells in i o. [139]
Melanoma
Cell lines Doxo ubicin S imula es au ophagy plus caspase-3/GSDME clea age and py op osis in SK-MEL-5, SK-MEL-28, and A-375 cells. Inhibi ion o eEF-2K dec eases au ophagy and
up egula es sensi i i y o doxo ubicin and py op osis in i o. [140]
Melanoma
cell line
I on
plus
ROS ac i a o s
I on up egula es ROS which in u n ac i a es Tom20/bax/caspase-3 and GSDME py op osis in A375 cells. GSDME silencing bu no GSDMD p e en s I on/CCCP
py op osis in i o. I on supplemen a ion plus sul asalazine inhibi s xenog a umo g ow h and me as asis h ough GSDME-py op osis induc ion. [124]
Melanoma cell lines BRAF and MEK inhibi o s BRAFi+MEKi ac i a e caspase-3 / GSDME clea age and py op osis in i o and in i o in di e se melanoma models. GSDME-py op osis s imula es immune an i- umo
esponse. [85]
Mul iple cell line ypes Mul iple chemo
d ugs
Chemo he apy d ugs p o oke caspase-3/GSDME clea age and py op osis in SH-SY5Y, Mewo, Hela, NCI-H522 and EMT6 cance cells bu also in no mal ke a inocy es,
placen al epi helial cells and smoo h muscle cells. Gsdme KO mice display educed in i o oxici y o no mal issues o Cispla in, 5-FU o bleomycin ea men . [33]
AML: Acu e myeloid leukemia; DM-
α
-KG: Dime hyl-
α
-Ke oglu a a e; GBM: Glioblas oma; NPC: nasopha yngeal ca cinoma; SCC: Squamous Cell Ca cinoma; TN: T iple Nega i e B eas Cance .
D. Sa i´
o e al.
BBA - Re iews on Cance 1876 (2021) 188635
8
exp essing cance cells may be gene ally mo e sensi i e o oncologic
he apeu ic challenge, bu he eali y is much mo e complex.
Fi s , he p ecise e ec o GSDMs on cell dea h depends on he bal-
ance o di e en s imuli, signaling pa hways, molecula al e a ions, and
cellula con ex s. In ac , he co-ac i a ion o speci ic signaling pa hways
migh be equi ed o GSDM py op osis. Fo ins ance, in HepG2 cells
GSDME enhances e oposide cell dea h only in he p esence o WT p53
[84] and GSDMC is up egula ed by di e se chemo he apy d ugs in MDA-
MB-231 cells, bu only he an ibio ic- ype (doxo ubicin/epi ubicin)
induce caspase-8/GSDMC-dependen py op osis [40]. Second, he
GSDMs media e an ex ensi e c oss alk among di e se cell dea h mech-
anisms, and hese could coope a e o compe e in a biological con ex -
dependen way. Fo example, in lammasome- igge ed caspases (1/4/
5/11) ac i a e GSDMD py op osis bu he apop o ic caspases 3/7 in-
hibi s py op osis in monocy es [42]. Thi d, umo cells could exhibi
in insic o acqui ed esis ance o py op osis. While loss o unc ion
mu a ions in human GSDM genes ha e been epo ed only o GSDME so
a [35], al e a ions in ups eam signaling pa hways o caspases could
also occu . Acco dingly, GSDME-posi i e BRAF
V600E/K
melanomas
esis an o BRAF+MEK inhibi o s exhibi weak induc ion o he p o-
apop o ic p o eins BIM-EL and BMF, and hus educed caspase-3 ac i -
i y [85]. Simila ly, 5/17 AML lines we e in insically esis an o Val-
bo oP o agen due o he lack o p o-caspase-1, which was equi ed
o GSDMD-media ed py op osis [86].
Apa om hese conside a ions, he gene al pic u e shows ha he
GSDMs a e mainly in ol ed in swi ching apop osis o ly ic and in lam-
ma o y cell dea h p ocesses (py op osis and nec osis, bu no nec op osis
o e op osis) [87] a e cance ea men (Table 2). GSDME is he main
de e minan o his apop osis- o-py op osis swi ch in mul iple in i o
and in i o models ea ed no only wi h a my iad o an i-cance agen s
(Table 2) bu also wi h speci ic na u al an i umo p oduc s, such as he
epoxide ip olide [88] o he la onol galangin [89]. In clinical speci-
mens, GSDME-py op osis was also e idenced (measu ing se um LDH
concen a ions) in lung cance pa ien s a e chemo- o EGFR inhibi o -
based ea men s [78]. None heless, in many s udies i is unclea i his
apop osis-py op osis swi ch ansla es in o a bigge cance killing. Fo
example, in lobapla in- ea ed colo ec al cance cells GSDME silencing
did no a ec umo esponse (g ow h a e) in i o and in i o [90], and
TRAIL ea men esul ed in simila Hela cell dea h quan i ies, i e-
spec i e o GSDME le els [35].
Like GSDME, GSDMD ac s as a he apy sensi ize o media es
apop osis-py op osis swi ch in pa icula cance cell ypes and oncologic
ea men s (Table 2), hus, hese GSDMs could be impo an modula o s
o cance he apy esponse. Un o una ely, many o he s udies lis ed in
Table 2 we e no pe o med in clinical human specimens, so he impac
o GSDM-media ed py op osis on cance pa ien su i al is s ill o be
e i ied.
Con a y o o he GSDMs, GSDMB up egula ion has been associa ed
wi h he apy esis ance. Speci ically, in HER2 b eas ca cinomas GSDMB
ende s cance cells mo e esis an o he an i-HER2 agen s as uzumab
and lapa inib, bu no o axol chemo he apy [55,61], likely in a
py op osis independen mechanism. Fo his eason, GSDMB signi i-
can ly associa es wi h wo se b eas cance pa ien s’ p ognosis in bo h
adju an and neoadju an an i-HER2 ea men s [55].
Summa izing, he inal e ec o GSDMs on umo ea men esponse
depends on mul iple ac o s, including he co-ac i a ion o speci ic
signaling pa hways, he d ug used, cellula con ex and, as discussed
la e , he e ec on he umo mic oen i onmen .
5. Ta ge ed ac i a ion o GSDM cy o oxici y in umo cells as
no el he apeu ic op ions
Di e se oncologic ea men s induce GSDM-media ed cell dea h,
hus sugges ing ha a ge ed app oaches speci ically aimed a ac i a ing
o GSDM cy o oxici y in umo s can be exploi ed as no el he apeu ic
op ions (Fig. 2). To his end, p omising he apeu ic e ec s has been
ecen ly ob ained using wo gene al ypes o app oaches: a) Ta ge
ac i a ion o he in insic GSDM p o-cell dea h unc ion in cance cells
h ough nanomedicines; b) Deli e ing GSDM cy o oxic pep ides/
exp ession cons uc s in o umo s.
Wi hin he i s ype o app oaches, h ee s udies a ge ed GSDME-
py op osis wi h nanoca ie s ha combined agen s o induce GSDME
ansc ip ion (such as he DNA me hyla ion inhibi o deci abine, DCT)
wi h an i-cance d ugs (Fig. 2A). Fan and colleagues [91] used umo -
a ge ing nanoliposomes loaded wi h cispla in (LipoDDP) and DCT o
igge GSDME-py op osis in 4T1 b eas xenog a s. The in a enous
adminis a ion o LipoDDP-DCT up egula ed GSDME exp ession (by
inhibi ing i s gene p omo e hype me hyla ion), induced caspase-3/
GSDME clea age and he subsequen py op osis led o a educed
umo g ow h and me as a ic sp ead in i o. Simila ly, Zhao and col-
labo a o s [92] designed biomime ic nanopa icles (BNP), by using
b eas cance memb anes on o a poly-lac ic-co-glycolic acid (PLGA)
co e, ha we e loaded wi h he pho osensi ize indocyanine g een (ICG)
and DCT. In 4T1 umo xenog a s he “BNP” nanomedicine was ac i-
a ed by low-dose pho o-ac i a ion (local hype he mia) and induced
GSDME-dependen py op osis by he combina ion o wo e ec s: On one
hand, ICG-media ed punc u e o cance cell memb anes p omp ed a
sha p inc ease in cy oplasmic Ca
2+
, p o oking cy och ome c elease and
caspase-3 ac i a ion. On he o he hand, DCT elease up- egula ed
GSDME ansc ip ion. Impo an ly, his “ac i a able” he apeu ic agen
educed signi ican ly he size o p ima y umo and me as asis by ac i-
a ing a s ong sys emic an i umo immune esponse. Finally, Hu e al.
[93] designed a nano-d ug deli e y sys em based on mPEG-PLGA-PLL
copolyme ha was loaded wi h a senic ioxide (As
2
O
3
) as he apeu-
ic agen o hepa ocellula ca cinoma (HCC) ea men . In a umo
adminis a ion o hese nanopa icles (As
2
O
3
-NPs) esul ed in caspase-
3/GSDME clea age and py op osis o Huh7 xenog a ed umo cells.
Mo eo e , As
2
O
3
educed he exp ession o DNA me hyl ans e ases
(Dnm 1, 3a and 3b) which modula e he ansc ip ional le els o GSDMD
o GSDME in a cell line-dependen way. In e ms o p e-clinical e icacy,
compa ed o ee As
2
O
3
, As
2
O
3
-NPs g ea ly inhibi ed umo g ow h and
showed no ob ious sys emic oxici y.
Ins ead o using d ugs/compounds a ec ing GSDM exp ession and/
o ac i a ion, Molina-C espo and collabo a o s [61] c ea ed he i s
nano he apy a ge ing di ec ly a GSDM p o ein wi h a he apeu ic
an ibody (Fig. 2B). Speci ically, he nanomedicine ( e med AbGB-NC)
combined a he apeu ic an i-GSDMB monoclonal an ibody (AbGB)
wi h biocompa ible nanocapsules (NCs) unc ionalized wi h hyalu onic
acid (HA). Since GSDMB up egula ion p omo es mul iple p o- umo
e ec s in HER2 b eas ca cinomas [55,56], he au ho s nex alida ed
he he apeu ic e ec o his nanomedicine in di e en HER2/GSDMB+
b eas cance models. The esul s demons a ed ha he in acellula
eleased AbGB, bu no an i ele an an ibody, was able o impac
signi ican ly on umo beha io in di e se ways: a) dec easing in i o
cell mig a ion; b) inc easing he sensi i i y o as uzumab an i-HER2
he apy; c) educing umo g ow h in i o o o ho opic b eas cance
xenog a s; and d) limi ing lung me as asis coloniza ion in i o, wi h no
ob ious unwan ed cy o oxic e ec s. A he mechanis ic le el, in i o
es s indica ed ha he binding o he AbGB o GSDMB (wi hin he CT
egion and close o he hinge egion) could a ec he p o ein con igu-
a ion hus enhancing he in e ac ion o GSDMB p o ein wi h sul a ides.
This in e ac ion would subsequen ly dec ease cell mig a ion, and
impo an ly may elease he in insic p o-cell dea h ac i i y o GSDMB.
In ag eemen wi h his, he AbGB-NCs, bu no he con ol ea men s,
inc eased he cell dea h a e (TUNEL assay, no speci ic es o py op-
osis) o cance cells in i o, and speci ically o GSDMB-o e exp essing
umo s (Fig. 2B). O e all, his s udy p o ed ha GSDMB is a no el
he apeu ic a ge o agg essi e HER2/GSDMB+b eas cance s, and
indica ed, o he i s ime, ha GSDMB cy o oxic e ec and o he
unc ions in cance can be modula ed di ec ly wi h in acellula
an ibodies.
Whe eas he abo e nano he apies ely on he endogenous exp ession
D. Sa i´
o e al.
BBA - Re iews on Cance 1876 (2021) 188635
9
o he GSDMs by he umo cells, o he he apeu ic app oaches ha e
been designed o ex insically deli e ing GSDM cy o oxic cons uc s
in o cance cells. As a po en ial ea men o schwannoma umo s
Ahmed and collabo a o s [94] gene a ed an adeno-associa ed se o ype-
1 i us (AAV1)-based ec o ha , unde he con ol o a Schwann cell-
speci ic p omo e (P0), exp esses he py op o ic NT egion o GSDMD
( esidues 1–276). The in a umo al injec ion in o in a-scia ic ne e o
he AAV1-P0-GSDMD-NT in human and syngeneic mouse cells xenog a
schwannoma models educed umo bu den by inc easing cell dea h and
educing cell p oli e a ion. Mo eo e , his ea men alle ia ed umo -
associa ed pain while causing no e iden neu ologic oxici y.
Recen ly, Wang e al. [95] de ised a e y cle e app oach by which
he elease o he py op o ic GSDMA3 p o ein occu s speci ically wi hin
umo cells (Fig. 2C). Speci ically, he au ho s designed a complex nano-
bioo hogonal sys em based on wo in e ac ing componen s. As i s
componen , he mouse GSDMA3 clea ed p o ein (NT plus CT domain)
was conjuga ed ia a ie hylsilyl (TES) e he linke o gold nanopa icles
(NP) o gene a e he NP-GSDMA3, a biocompa ible nanosys em ha
accumula ed mos ly in o umo issue o 4T1 cells xenog a s. As second
componen , he au ho s used he umo -imaging p obe phenylalanine
i uo obo a e (Phe-BF
3
), which was aken up speci ically cance cells.
Phe-BF3 desilya ed he silyl e he bond o NP-GSDMA3 esul ing in he
in acellula elease o a py op o ic ac i e GSDMA3 p o ein and he
killing o 4T1 umo cells in i o. A e h ee ounds o injec ion wi h
Phe-BF3 plus NP-GSDMA3 he umo bu den educed eno mously, while
he adminis a ion o Phe-BF
3
o NP-GSDMA3 alone, o a loss-o -
unc ion GSDMA3-NP did no ha e any e ec . No ob ious cy o oxic
e ec s we e obse ed in o he o gans in mice ea ed Phe-BF
3
plus NP-
GSDMA3 [95].
Rema kably, some o hese GSDM- a ge ed nano he apies achie ed
umo eg ession h ough he ac i a ion o a po en an icance in lam-
ma o y eac ion (Fig. 2) (discussed below), hus indica ing ha
py op osis-inducing nano he apies a e p omising app oaches ha could
hal umo p og ession simul aneously by a acking umo cells and
enhancing immune esponse.
6. GSDM-media ed cell dea h: e ec s on he umo
mic oen i onmen and he cance esponse o immuno he apy
GSDM py op osis and he immune esponse a e engaged in a com-
plex bidi ec ional c oss alk. On one hand, cance cell lysis and he
subsequen elease o DAMPs and cy okines p omo es an immunos i-
mula o y umo mic oen i onmen ha could lead o cance ejec ion
[87]. On he o he hand, cy o oxic T cells and NK cells can di ec ly
igge py op osis by clea ing GSDME o GSDMB in cance cells ia
pe o in-media ed elease o GZMB and GZMA, espec i ely [35,37].
The e o e, py op osis and ac i a ed immunocy es could gene a e a
posi i e eedback loop in an icance immuni y (Fig. 2).
GSDME-media ed py op osis gene ally associa es wi h a s ong im-
mune an i umo esponse [87]. GSDME up egula ion in human cance s
co ela e posi i ely wi h immune in il a ion [96], and i s o e -
exp ession in mu ine b eas cance and melanoma xenog a s in
immunocompe en mice signi ican ly augmen ed he numbe o umo
in il a ing lymphocy es (TILs), mos ly CD8+T and NK cells. Ac i a ed
TIL cy o oxici y and inc eased phagocy osis by umo -associa ed mac-
ophages p o oked umo e adica ion [35]. Mo eo e , a e he apeu ic
challenge by BRAF–MEK inhibi o s GSDME-media ed py op osis in
mu ine melanoma models inc eased T cell in il a ion and imp o ed
ea men esponses in i o [85]. Consis en wi h he bene icial e ec o
GSDME py op osis in cance ea men , as commen ed be o e, he
GSDME a ge ed nano he apies LipoDDP-DCT and BNP induced a s ong
immune eac ion, including he elease o cy okines and p o-
in lamma o y molecules, ec ui men and ac i a ion o cy o oxic T
cells, and dend i ic cell ma u a ion, ha al oge he p o oked umo
e adica ion [91,92], Alike GSDME nano he apies, he e icacy o he NP-
GSDMA3 +Phe-BF3 nanomedicine depends on he immune an i- umo
eac ion. In ac , he py op o ic killing o a small ac ion o umo cells
(20%) was su icien o ac i a e, ia IL1-β sec e ion, a po en an i- umo
e ec (T cells, NK and mac ophages) esul ing in he elimina ion o he
bulk umo [95] (Fig. 2C).
Rema kably, he GSDME umo supp esso unc ion in bo h un-
ea ed and he apy-challenged umo s seem o ely on he p esence o
ac i a ed cy o oxic cells. Thus, deple ion o NK and CD8+T cells o
knocking-ou pe o in e e ses GSDME e ec s on umo g ow h o un-
ea ed cance s [35] and deple ion o CD4+/CD8+T-cells signi ican ly
dampens he esponse o xenog a ed melanoma cells o BRAF–MEK
inhibi o s. Consequen ly, immunocompe en (e.g., Balb/c) and no
immunode icien (nu/nu) mouse s ains should be used o un eil he ull
e ec s o GSDME on cance . In ac , di e ences in mouse s ains may
explain in pa he con lic ing esul s ob ained wi h GSDME-posi i e
cance xenog a s (discussed in sec ion 2).
O no e, while in i o GSDME ac i a ion in cance cells occu s ia
caspase-3 ( esul ing in nec osis seconda y o apop osis), in i o clea age
by kille -cell GZMA p o okes immunogenic cell dea h (Fig. 2), a o m o
cell dea h ha is su icien o ac i a e an adap i e immune esponse
[35,87]. Ac ually, he “ accine” inocula ion o immunocompe en mice
wi h GSDME-o e exp essing cance cells p oduces he e icien killing o
he seconda y umo implan a ion. Suppo ing his idea, GSDME-
media ed py op osis igge s an immune esponse agains esidual
cance , since Gsdme-silenced mu ine melanomas ecu ed mo e
equen ly han Gsdme-exp essing ones a e he in e up ion o BRAF-
MEK inhibi o s he apy [85].
In opposi ion o he in insic umo supp ession media ed by
GSDME, GSDMB p omo es di e se p o- umo e ec s in b eas cance
[55,56], and i s exogenous exp ession in mu ine colon and melanoma
cells does no a ec umo g ow h in immunocompe en mice [37]. In
ac , o expose he po en ial GSDMB an i umo e ec , he s imula ion o
he an i umo immune esponse by PD-1 immune checkpoin inhibi o s
is equi ed o e icien GZMA/GSDMB-media ed umo py op osis and
eg ession [37] (Fig. 2B). This shows ha igge ing GSDMB cy o oxici y
wi hin umo s may equi e addi ional signals om he umo mic oen-
i onmen in o de o enhance immune ecogni ion and cance killing.
In his sense, GSDMB exp ession in cance cells is up egula ed by cy o-
kines (IFNs and TNF-
α
) p oduced by TILs [37], and his may con ibu e
o main aining a posi i e eedback loop o py op osis (Fig. 2B). Mo e-
o e , his wo k indica es ha py op osis and immuno he apy could
syne gize o p oduce a p o ec i e immune eac ion. Suppo ing his
idea, Wang e al. 2020 p o ed ha he NP-GSDMA3 +Phe-BF
3
he apy
sensi ized 4T1 umo s o in i o an i-PD-1 ea men [95] (Fig. 2C).
The e o e, py op osis-induced in lamma ion, ia GSDM agonis s, can
coope a e wi h immune checkpoin inhibi o s, hus leading o an
imp o ed e ec i eness o cance immuno he apy. Based on hese da a,
he cu en hypo hesis is ha GSDM-media ed py op osis could con e
immunologically “cold” o “ho ” umo s, and likely espond be e o
immune check-poin inhibi o s.
7. Fine unning o GSDM-media ed cell dea h o oncologic
ea men : emaining ques ions
Al hough he da a desc ibed abo e e eal a c ucial ole o GSDMs as
no el he apeu ic a ge s and as key de e minan s o chemo he apeu ic
and immuno he apy ea men s he e a e s ill a numbe o ques ions o
be esol ed be o e GSDM-media ed oncologic he apies became a eali y
a he clinical le el.
7.1. Con olling he speci ici y, in ensi y and iming o GSDM-media ed
cance cell dea h
Cu en ly, p edic ing he inal e ec o py op osis in he apy
ou come is in ica e. In o de o exploi success ully GSDM-py op osis in
cance he apy, he e a e h ee key pa ame e s ha need o be inely
con olled (speci ici y, iming and in ensi y o cance lysis and
D. Sa i´
o e al.
BBA - Re iews on Cance 1876 (2021) 188635
16
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