Clustered regularly interspaced short palindromic repeats (CRISPR) ni uburyo bwo guhindura genome bw’ikinyejana cya gatatu bwahinduye isi bitewe n’umusaruro wabwo mwinshi. Bwakoreshejwe mu kuvura indwara zitandukanye z’ibinyabuzima n’indwara zandura. Bagiteri zitandukanye n’izindi prokaryote (nka archaea) nazo zifite sisitemu ya CRISPR/Cas9 yo kwirinda phages. Byatangajwe ko ingamba zishingiye kuri CRISPR/Cas9 zishobora kubuza ikura n’iterambere rya kanseri y’ibere ya triple negative (TNBC) hifashishijwe imiterere y’uturemangingo dushobora guhinduka, transcription, na epigenetic regulation. Ubu buryo bwo kuvura bushobora gufasha gukemura ibibazo bikomeye nko kudakora imiti bigaragara no muri TNBC. Muri iki gihe, hari uburyo butandukanye bukoreshwa mu kugeza CRISPR/Cas9 ku turemangingo, nko mu buryo bw’umubiri (microinjection, electroporation na hydrodynamic modes), virusi (virusi ifitanye isano na adeno na lentivirus) n’ututari twa virusi (liposomes na lipid nanoparticles). Nubwo hakozwe uburyo butandukanye bwo kwiga impamvu za molekile zitera TNBC, kutagira uburyo bworoshye kandi bwibanda ku gutanga ibikoresho byo guhindura genome mu mubiri bigabanya ikoreshwa ryabyo mu buvuzi. Muri make, iri suzuma risuzuma byimbitse intambwe, imbogamizi, imbogamizi, n'amahirwe yo kuvura CRISPR/Cas9 kuri TNBC hashingiwe ku bimenyetso bihari. Turagaragaza kandi uburyo guhuza ubwenge bw'ubukorano n'ubumenyi bw'imashini bishobora kunoza ingamba za CRISPR/Cas9 mu kuvura TNBC.
Kanseri irangwa no kwigabanyamo ibice bitagira igaruriro, guhindagurika kw'igenzura ry'uturemangingo, no guhinduka kw'uturemangingo duto dutuma turwara kanseri (TSGs) (Matthews et al., 2022). Mu moko atandukanye ya kanseri, kanseri y'ibere ni yo kanseri ikunze kugaragara cyane mu bagore kandi ifite igipimo kinini cy'impfu ku isi yose (Waks na Winer, 2019). Kanseri y'ibere ni indwara itandukanye ifite imiterere itandukanye, nko mu miterere y'ibinyabuzima n'imiterere yabyo, uko indwara igaragara n'imyitwarire yabyo, ndetse n'uko bifatwa (Weigelt et al., 2010). Gushyira mu byiciro kanseri y'ibere bitanga ubumenyi nyabwo ku bijyanye no gusuzuma kanseri y'ibere no kumenya uko kanseri imera. Gukoresha ibimenyetso bisanzwe n'imiterere y'indwara byabaye ikintu nyamukuru gishingirwaho mu gushyira mu byiciro kanseri y'ibere (Tsang na Tse, 2019). Uburyo kanseri y'ibere ifatwa n'uko ifatwa biterwa n'ibintu byinshi, birimo kuba hari estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2/neu), n'urwego rw'imiterere y'umubiri, ubwoko bw'indwara, n'urwego rw'umubiri. Ubunini n'ubugari bw'uturemangingo tw'amaraso (Al-Tubaiti, 2020). Habonetse ubwoko butanu bw'ingirabuzimafatizo za kanseri y'ibere, harimo luminal A, luminal B, HER2-enriched, basal-like, na claudin-low (Prat et al., 2015). TNBC ni ubwoko bwa kanseri butarimo ER, PR, na HER2 zose (Yin et al., 2020). Ibi bintu bitera indwara bifitanye isano na TNBC, byemeza ko itera imbere vuba kandi ikaba ikomeye kurusha ubundi bwoko bwose bwa kanseri y'ibere (Feng et al., 2018). Byongeye kandi, Peru et al (2000) bakoresheje ikoranabuhanga rya microarray kugira ngo bongere gushyira mu byiciro kanseri y'ibere no kumenya ubwoko butanu bw'ingirabuzimafatizo za kanseri y'ibere (Cadenas, 2012). Kanseri y'ibere isa n'iya mbere ni ubwoko bwa kanseri y'ibere ifite imiterere itatu mibi kandi ifitanye isano no gutera imbere vuba. Byakagombye kumenyekana ko kanseri zose z'ibere zisa n'iz'ibanze zikunze gufatwa nabi nka TNBC, ariko 77% gusa ni zo TNBC; Ibinyuranye n'ibyo, 71-91% bya TNBC zisa n'iz'ibanze, bigaragaza ko ubwo bwoko bubiri bwa kanseri y'ibere buhurirana kandi bugaragaza ibyiciro bitandukanye (Wang D.-Y. et al., 2019). Ibi bituma hakenewe kugaragazwa itandukaniro rya TNBC kugira ngo hamenyekane uburyo bwo kurwara no kumenya ibisubizo bishobora guterwa n'ubuvuzi buriho n'ubutaha. Byongeye kandi, TNBC igizwe na 15-20% by'abarwayi bose ba kanseri y'ibere kandi ikunze kugaragara cyane mu bagore bari munsi y'imyaka 50. Impinduka za BRCA1 cyangwa BRCA2 zagaragaye muri 20% by'abarwayi ba TNBC (Xie et al., 2017; Tzikas et al., 2017). , 2020). Ubushakashatsi bwagaragaje kandi ko TNBC ifite imiterere yihariye y’ubudahangarwa bw’umubiri irimo urugero rwo hejuru rw’ibintu bikura mu mitsi, macrophages zifitanye isano na kanseri (TAMs), lymphocytes zinjira mu maraso (TILs), n’izindi molekile zigira uruhare mu gukura no kwimuka kwa kanseri. Kubwibyo, gusobanukirwa imiterere y’imiterere ya TNBC ni ingenzi cyane mu gutanga isuzuma no kuvura (Fan na He, 2022).
Kugira ngo hamenyekane uko TNBC ihagaze no kwemeza ko ivura neza, ni ngombwa gusuzuma neza, ahanini hashingiwe ku buvuzi bw’umubiri (IHC) kugira ngo hamenyekane ER, PR na HER2, na mammography kugira ngo hamenyekane ibimenyetso by’ibere, ni ingenzi. Ariko, mammography ntishobora kugaragaza neza ibimenyetso byo mu nda y’umurwayi nka necrosis na fibrosis (Deepak Singh et al., 2021). Kubera ko uburyo bubi bwo gusuzuma no gusuzuma indwara bubuza abaganga gutanga imiti ikwiye (Chaudhary, 2020), hari ingamba zitandukanye zikoreshwa mu kunoza ubuvuzi ku barwayi ba TNBC. Kuri ubu, imiti ibiri, doxorubicin na cyclophosphamide, ikoreshwa ku barwayi ba TNBC kandi yagaragaje umusaruro mwiza. Byongeye kandi, hari indi miti ya platine ikoreshwa, nka carboplatin na cisplatin (Sikov et al., 2015). Byongeye kandi, imiti igabanya PARP nka Olaparib, Velaparib, na PF-01367338 yakoreshejwe nk'imiti ishobora gukoreshwa mu kuvura TNBC (Ishino et al., 2018). Kuva aho byatangajwe ko inzira zo gutanga ibimenyetso za Wnt/b-Catenin, NOTCH, na Hedgehog zigira uruhare mu kubaho no gukomeza kwa TNBC, kwibanda ku miti muri izi nzira bishobora kuba ingamba y'ingenzi (Aysola et al., 2013). Nubwo kubaga, ubuvuzi bushingiye ku mirasire na chimiotherapie ari byo bikomeje kuba ingenzi mu kuvura TNBC muri iki gihe, hari intambwe ikomeye yatewe mu guteza imbere uburyo bushya bwo kuvura harimo uburyo bwo kuvura hifashishijwe ikoranabuhanga, ubuvuzi bw'imyororokere, ibikoresho bitandukanye bijyanye no guhindura imiterere y'uturemangingo bifitanye isano na CRISPR nka Cas9n, dCas9., CRISPR/Cas12, prime editing, na CRISPR/Cas9-targetted gene therapy. Aha tuzibanda ku bikoresho bitandukanye bijyanye no guhindura imiterere y'uturemangingo bikoreshwa mu kuvura TNBC. Muri byo, CRISPR/Cas9 yakunze kwitabwaho cyane.
Cas9n, izwi kandi nka Cas9 nickase, ni ubwoko bwa poroteyine ya Cas9 yahinduwe mu buryo bwa CRISPR/Cas9 (Gupta et al., 2019). Mu miterere yayo y'umwimerere, poroteyine ya Cas9 irimo imiterere ibiri ya nuclease: RuvC na HNH, akazi kayo gakomeye ni ugucika kw'imirongo yombi ya ADN. Ariko, muri Cas9n, imwe muri imiterere ya nuclease, HNH, ihinduka mu buryo bwa genetiki kandi ntikora. Kubwibyo, imiterere ya HNH ya Cas9n iguma idakora. Urwego rwa RuvC gusa nirwo ruguma rukora, rutuma Cas9n igabanya cyangwa igatera imvune ku mirongo imwe ya ADN (Trevino na Zhang, 2014). Ugereranyije na Cas9, Cas9n ishobora kugabanya ingaruka zitari ku ntego no kunoza neza uburyo bwo gusana uturemangingo. Cas9n ishobora gukoreshwa mu gukemura ibibazo bitandukanye, nko gutuma imvune zihagarara ahantu runaka muri ADN ifite imirongo ibiri. Muri iyi ntego, molekile ebyiri za Cas9n zahujwe zikoreshwa hamwe (Yee, 2016). Mixed lineage kinase 3 (MLK3) ni poroteyine kinase ikora mitogen ikora nk'urwego rw'ingenzi mu gihe cya metastasis ya TNBC (Cronan et al., 2012).
MLK3 ishobora gukora inzira nyinshi zo gutanga ibimenyetso biganisha kuri metastasis ya TNBC. Urugero, inzira ya c-Jun N-terminal kinase (JNK) igenga motility y'uturemangingo no kwangirika kwa matrix yo hanze y'uturemangingo, naho MLK3 igakora inzira ya JNK, bityo yongera motility y'uturemangingo kandi igatanga imiterere y'ibintu biyitera (Rattanasinchai na Gallo, 2016). MLK3 inagenga EMT. Kugira ngo ibi bigerweho, ikora ibintu byo kwandika nk'ibinyabutabire, Slug na Twist. Ibi bintu bibuza igaragara ry'ibimenyetso bya epithelial kandi bigateza imbere igaragara ry'ibimenyetso bya mesenchymal, bigatuma habaho ubwoko bwa metastasis (Casalino et al., 2023). MLK3 yagaragaye ko igira uruhare mu kuvugurura ECM no gukora poroteyine nka matrix metalloproteinases (MMPs) zangiza ECM. Ibi byoroshya ukwinjira kw'uturemangingo twa kanseri no gukwirakwira ahantu ha kure (Katari et al., 2019). Bityo, ubushakashatsi bwabanje bwagaragaje ko MLK3 igira uruhare runini muri TNBC. Rattanasinchai na Gallo bakoresheje moderi za TNBC kugira ngo biga uruhare rwa MLK3 kandi basanze itera imbere kanseri binyuze mu nzira zihariye zo gutanga ibimenyetso. Bakoresheje CRISPR/Cas9n kugira ngo bahindure MLK3 kandi babonye igabanuka rikomeye rya metastasis ya TNBC (Rattanasinchai na Gallo, 2016).
dCas9, ikunze kwitwa Cas9 idakora, ni ubwoko bwahinduwe bwa poroteyine ya Cas9. Bitandukanye na Cas9 ikora, dCas9 nta gikorwa cya endonuclease, bigatuma idashobora gutera kuvunika kwa ADN inshuro ebyiri. Kubwibyo, dCas9 ishobora gukoreshwa mu kwibasira neza uturere twihariye twa genome nta mpinduka cyangwa impinduka ku ruhererekane rwa ADN (Wang et al., 2016). Muri dCas9, poroteyine ebyiri za endonuclease, RuvC na HNH, zishyirwaho ingufu mu guhagarika ibisigazwa by'ingenzi bya aside amine. (Richter et al., 2016). Nubwo idafite igikorwa cyo gutandukanya ADN, dCas9 igira uruhare runini mu bushakashatsi ku ngirabuzimafatizo no mu ikoranabuhanga rya biotechnology. Urugero, yemerera kwerekana uturere twihariye twa genome, yoroshya uburyo bwo kwandika, kandi igateza imbere impinduka za epigenetic (Brocken et al., 2018).
Igikoresho cya transcription ZEB1 (zinc finger E-box binding homeobox 1) gikora uruhare runini kandi rw’ingenzi mu guhuza epithelial-mesenchymal transition (EMT), inzira y’uturemangingo ibaho mu gihe cy’iterambere ry’urusoro, gusana ingirabuzimafatizo, no kwiyongera kwa kanseri. Bikubiyemo guhindura uturemangingo twa epithelial mo uturemangingo twa mesenchymal, bigatera impinduka mu miterere y’uturemangingo, kugenda, kwivanga, nibindi. (Wu et al., 2020). ZEB1 ibuza ibimenyetso byinshi bya epithelial, nka E-cadherin na Occludin, bishinzwe kubungabunga uburyo selile zifatana n’uturemangingo ndetse no kuba uturemangingo twa epithelial turi muri TNBC (Moreno-Bueno et al., 2008). Byongeye kandi, ZEB1 ikora ibimenyetso bimwe na bimwe nka N-cadherin, vimentin na fibronectin (Konradi et al., 2014) kandi inagenzura imiterere y’uturemangingo tugira uruhare mu kuvugurura cytoskeletal nka Rho GTPases na matrix metalloproteinases (MMPs) (Huang). et al., 2014). 2022), byongeye kandi, bigira ingaruka ku nzira zitandukanye zo gutanga ibimenyetso nko guhindura ubwiyongere bw'umubiri (TGF-β), Wnt signaling, nibindi, bityo bigatera kwibasira ikibyimba no kwangirika kwacyo muri TNBC (Chen et al., 2016). ). Ubushakashatsi bwinshi n'ibimenyetso bya siyansi bigaragaza ko ZEB1 ifite ubushobozi bukomeye nk'ikintu cy'agaciro mu gupima no kuvura TNBC. Mu bushakashatsi buherutse gukorwa, Waryah et al. bakoresheje icyitegererezo cya TNBC maze bagera ku gukuraho burundu ZEB1 na dCas9. Bityo, babonye umwihariko mwinshi cyane no gukumira hafi ya byose ZEB1 mu miterere y'umubiri (Waryah et al., 2023).
CRISPR/Cas12 ni igikoresho cyo guhindura ingirabuzimafatizo cyitwa CRISPR/Cpf1. Kiva muri sisitemu ya CRISPR/Cas, aho ijambo Cas12 risobanura poroteyine 12 ifitanye isano na CRISPR (Bharathkumar et al., 2022), isa neza na Cas9. Itandukaniro ryonyine ni uko irimo poroteyine ya Cas12. Ugereranyije na Cas9, Cas12 ifite imirimo idasanzwe nko gukora impera zifatana mu gihe cyo guhindura ingirabuzimafatizo, mu gihe Cas9 ikora impera zidafite aho zihuriye (Wang et al., 2021). Iyi miterere ya Cas12 igira uruhare mu ikoranabuhanga ryayo ryihariye ryo guhindura ADN. Nk'poroteyine ishobora kumenya neza no guca ADN y'ingenzi, ifite ubushobozi budasanzwe bwo gukora ibintu bitandukanye, bigatuma iba igikoresho cyiza mu bikorwa bitandukanye, harimo no guhindura ingirabuzimafatizo (Pickar-Oliver na Gersbach, 2019).
Kimwe n'izindi CRISPR, CRISPR/Cas12 nayo ishobora gukuraho cyangwa gukora jeni muri TNBC, yibanda ku jeni zigira uruhare runini mu gutera kwazo cyangwa igisubizo ku buvuzi (Yang na Zhang, 2023). Kugira ngo iki gikorwa kigerweho, hakozwe RNA y'ubuyobozi (gRNA) iyobora Cas12 kuri jeni y'ingenzi. Iyo Cas12 imaze gufatana n'intego yayo, ishyiramo ibice bibiri muri ADN kandi igakoresha uburyo bwo gusana ADN. Mu gihe cyo gusana, nucleotides zitari zo zishobora gushyirwamo, bigatuma habaho impinduka mu jeni no kubura imikorere (Zhang et al., 2021a). Byongeye kandi, verisiyo nziza ya enzyme ya Cas12 (yitwa dCas12) nayo yakoreshejwe mu gukora jeni. Mu kuyihuza na transcription activator, birashoboka gutera jeni zimwe na zimwe, bityo zigatangira gukora. Iri koranabuhanga rifite ubushobozi bukomeye mu guteza imbere gukora kwa jeni zigabanya kanseri (Sultan et al., 2022).
Prime editing ni ikoranabuhanga rishya kandi rigezweho ryo gutunganya genome. Rishobora guhindura ADN y'ibinyabuzima neza cyane (Chen na Liu, 2023). Prime editing igerwaho binyuze mu guhuza ibice bibiri by'ingenzi: enzyme ya CRISPR/Cas9 yahinduwe na transcriptase isubira inyuma. Uruhare rwa enzyme ya CRISPR/Cas9 ni ukwibanda ku hantu nyaho muri genome, mu gihe reverse transcriptase ifasha guhindura neza ADN aho yagenewe (Hassan et al., 2021). Mu buryo bwo gutunganya primer, intambwe ya mbere irimo gukora RNA y'ubuyobozi bwo gutunganya primer (pegRNA) (Standage-Beier et al., 2021). Irimo urutonde rw'icyerekezo n'icyitegererezo cya RNA bihuye n'ahantu hifuzwa ho gutunganya muri ADN y'icyerekezo. Hanyuma pegRNA yinjizwa mu turemangingo tw'icyerekezo hamwe na primer editing nuclease (PE2). PE2 ni poroteyine ivanze igizwe na enzyme ya Cas9, reverse transcriptase, na primer editing adaptateur (Martín-Alonso et al., 2021). Muri selile, pegRNA na PE2 complexes bishakisha ADN yihariye bashaka guhindura. Enzyme ya Cas9 ikata ADN igakora icyitegererezo kigizwe n'imirongo imwe (Choi et al., 2022). Reverse transcriptase ikoresha iyi template kugira ngo itegure ADN yo kuyihindura. Hamwe no gukoporora ADN, amabwiriza yo guhindura icyitegererezo cya RNA nayo arimo. Amaherezo, umugozi wa ADN mushya ukoreshwa nk'icyitegererezo cyo gusana icyuho muri ADN, bigatuma habaho impinduka za ADN zirimo impinduka zifuzwa (Ochoa-Sanchez et al., 2021). Uburyo bwo guhindura bwa primer bushobora gutuma habaho impinduka nyinshi muri genes. Ishobora kwibasira impinduka z'ingingo, gushyiramo, gusiba, ndetse no gusimbuza genes (Anzalone et al., 2019; Chen na Liu, 2023). Prime editing itanga inyungu nyinshi ugereranyije n'ikoranabuhanga rya kera ryo guhindura genome. Ibi birimo kwiyongera k'ubunyangamugayo, kugabanya ingaruka zitari iz'ingenzi, no gushobora guhindura ADN utishingikirije ku gucika kw'imirongo ibiri ya ADN (Anzalone et al., 2020).
Ikoranabuhanga rya CRISPR/Cas9 ryabanje gukorwa kugira ngo ririnde bagiteri kwimura plasmide no kwandura phage, nyuma rigakoreshwa nk'igikoresho cyiza cyo kwibanda kuri ADN gishingiye kuri RNA mu guhindura genome (Jiang na Doudna, 2017). Byongeye kandi, sisitemu ya CRISPR/Cas9 byavuzwe ko iboneka muri 50% na 87% bya genome za bagiteri na archaeal, uko bikurikirana (Ishino et al., 2018). CRISPR/Cas9 ni igikoresho gishobora gusiba, gushyiramo no gukosora uruhererekane rw'uturemangingo rudasanzwe hakoreshejwe uburyo bwa "in vivo" na "in vitro" (Sabit et al., 2021). Byongeye kandi, CRISPR/Cas9 byagaragaye ko ari igice cy'ubudahangarwa bw'umubiri bushobora guhinduka bitewe n'uburyo bwihariye bwo kwibanda ku ngirabuzimafatizo (Chen na Zhang, 2018).
CRISPR/Cas9 igizwe na Cas9 na RNA imwe y’ubuyobozi (sgRNA). Cas9 ni endonuclease igizwe n’ibice byinshi bya poroteyine. Byongeye kandi, Cas9 ifite imiterere yihariye y’imiterere n’imiterere (Pacesa et al., 2022) kuko igizwe n’uduce tubiri: kumenya (REC) na nuclease (NUC). Agace ka REC gagabanyijemo uduce dutatu: REC1, REC2, na bridge helices (Cromwell et al., 2018). NUC igizwe n’uduce dutatu, ari two RuvC (RuvC I, RuvC II, RuvC III), HNH na protospacer adjointe motif (PAM) interaction domain (Ishusho ya 1) (Song et al., 2016). sgRNA igizwe n’ibice bibiri, birimo CRISPR RNA (crRNA) na transcoding small RNA (tracer RNA) (Ishusho ya 1). SgRNA ihuza poroteyine ya Cas9 na connexins kugira ngo ikore complex active, izwi kandi nka effector complex (Richter et al., 2012). CrRNA ni 18-20 nucleotide base pair ifite uruhare runini mu kumenya imiterere ya ADN. Byongeye kandi, crRNA ihuza na ADN target, kandi tracrRNA ikora nk'igisenge cya Cas9 nuclease kugira ngo ihuze ADN target (Manghwar et al., 2019). Ku rundi ruhande, PAM sequence ifite nucleotides 3, yemeza, igena kandi igahuza uburyo effector complex ihuzwa na ADN. Cas9 protein complex subunits RuvC na HNH zifite catalytic activity (Richter et al., 2012; Asmamaw na Zawdie, 2021). HNH nuclease domaine ya Cas9 subunit ica umugozi wa ADN ujyanye na crRNA. Urusobe rw'imitsi ya RuvC ruca izindi mitsi ya ADN rugakora imitsi ibiri (DSBs), hanyuma uburyo bubiri butandukanye bwo gusana imitsi ya ADN bugatangira gukoreshwa (Jiang na Doudna, 2017) (Ishusho ya 1).
Ishusho ya 1. Incamake ya CRISPR/Cas9 (A). Ibice bigize sisitemu ya CRISPR/Cas9: (i). Cas9 endonuclease ishinzwe gukata urukurikirane rwa ADN, (ii) RNA imwe y’ubuyobozi (sg) ikomoka ku kwivanga kwa crRNA na tra-crRNA chimeras. (biri). Cas9 irimo ibice byinshi nka Rec I, Rec II, NUC lobe (HNH na Ruv C ni ibice bito) na PAM interaction domaine (C) hamwe n’imikorere ijyanye nayo. CRISPR/Cas9 protein complex igabanya urukurikirane rwa ADN mu buryo butari ubwuzuzanye kandi bwuzuzanye (D). CRISPR/Cas9 ihindura genome mu byiciro bitatu: kumenya, gukata no gusana. Sg-RNA yakozwe iyobora Cas9 kandi ikamenya urukurikirane rwifuzwa binyuze muri crRNA yuzuzanye. Cas9 imenya urukurikirane rwa PAM kuri 5′-NGG-3′ kandi ishongesha ADN, ikora hybrid ya ADN-RNA kandi igatuma habaho cleavage. Umuyoboro wa HNH wa Cas9 uca umugozi wuzuzanya, naho umuyoboro wa RuvC ugaca umugozi udafite inyongera. CRISPR/Cas9 isana dsDNA iyicamo ibice mu buryo bubiri: guhuza impera non-homologous (NHEJ) no gusana homology-directed repair (HDR). NHEJ isana ADN ifite imigozi ibiri mu gihe nta ADN y’amahanga ifite homologous binyuze mu buryo bwa enzyme, uburyo bushobora gukora amakosa bushobora kwinjiza cyangwa gukuraho uruhererekane rwa ADN rutunguranye. HDR ni yihariye cyane kandi isaba ADN ifite imigozi idafite inyongera.
Inzira zo gusana zikoreshwa na CRISPR/Cas9 zirimo uburyo bwo guhuza impera n’inyuma butari bumwe (NHEJ) n’uburyo bwo gusana buyobowe na homology (HDR). NHEJ ni inzira ikunze kwibasirwa n’amakosa kuko ikubiyemo gushyiramo cyangwa gusiba kandi ntisaba imiterere iyo ari yo yose mu gihe cyo gusana. Ikoresha nucleotides zidasanzwe kugira ngo ikore poroteyine zisanzwe (Abbasi et al., 2021). Ubu buryo bwo gusana bugizwe n’ibice bine nka KU complex, protein complex type 4 (XRCC-4), enzyme itunganya DNA end, na protein kinase DNA-PKcs (Abbasi et al., 2021). Poroteyine ya KU complex ifite utunyangingo tubiri, Ku 70 na Ku 80, kandi igira uruhare runini mu buryo bwa NHEJ, kubera ko uburyo bwo gusana butangirana no gufatanya utunyangingo tubiri (Ku 70 na Ku 80) ku mpera z’utunyangingo cyangwa hafi ya tworoshye za ADN (Abbasi et al., 2021), ikaba nk'aho ikoreshwa mu guteranya ibindi bintu bifitanye isano na NHEJ aho byakomeretse (Yang et al., 2020). XRCC-4 na ADN ligase bigizwe na aside amine 334 na 911, kandi XRCC4-DNA ligase IV complex itera imbaraga zo gufatanya udunyangingo twa ADN (Chatterjee et al., 2015). Enzyme itunganya ADN end, izwi kandi nka polynucleotide kinase 3′-phosphate, ni enzyme itunganya ADN end. Ishobora gukuraho itsinda rya 3′P muri ADN no gufosforisa itsinda rya 5′OH mu gihe cyo gusana DSB. Bikubiyemo kandi gusana imiyoboro imwe (SSBs) hakoreshejwe inzira yo gusana SSB (Chatterjee et al., 2015). Protein kinase DNA-PKcs ni poroteyine kinase ishingiye kuri ADN igizwe n'igice cya catalytic cya PIKKs (phosphatidylinositol 3-kinase-related kinases) n'umuryango wa ataxia telangiectasia mutated (ATM), hamwe na ATM na Rad3-related ATRs. imiyoboro imwe (DSBs) n'imiyoboro imwe (Yue et al., 2020; Peng et al., 2016).
HDR ni uburyo bwo gusana bufite ishingiro kandi bukwiye kuko amakuru akorerwa kopi hakoreshejwe imiterere ya ADN duplex isanzwe, nubwo bisaba ko habaho chromatids zindi. Ibi bibaho mu gihe cya S/G2 cy'ingengabihe y'uturemangingo tw'inyamabere. HDR iboneka cyane cyane mu bwoko bw'umusemburo, ariko NHEJ ni ingenzi mu nyamabere (Burma et al., 2006; Abbasi et al., 2021). Uburyo bwose bwo gusana bugaragara ku Ishusho ya 1.
Kubera ko TNBC iterwa n’ibitagenda neza mu buryo bw’uturemangingo no mu buryo bw’impinduka mu maraso, gukosora ibitagenda neza mu buryo bw’uturemangingo hakoreshejwe CRISPR/Cas9 bishobora kuba uburyo bwiza bwo kuvura (Chen et al., 2019). Byongeye kandi, hari ibintu bimwe na bimwe byo kwandika bifitanye isano n’uburyo bw’impinduka mu maraso bishobora kugaragaza imiterere yihariye y’uturemangingo twa kanseri, bigaragaza ko uburyo bwo kwandika maraso bushobora kuba uburyo bwiza bwo kuvura kanseri (Drost et al., 2017). Gukoresha iyi miterere y’uturemangingo tw’uturemangingo, nk’ibisembwa mu maraso, mu maraso no mu maraso, mu guteza imbere imiti bishobora kunoza umusaruro w’ubuvuzi no kugabanya ikiguzi cyo gusuzuma. CRISPR ni igikoresho gishobora gukoreshwa mu guhindura imiterere y’uturemangingo, kitabasha gusa kumenya no kumenya intego za gene zitera kanseri, ahubwo gishobora no gukoreshwa mu guhindura, gukumira no guhindura imiterere y’uturemangingo mu turemangingo tw’abantu (Imbonerahamwe ya 1) (Ahmed et al., 2021). Hakozwe ubushakashatsi butandukanye bwa CRISPR kugira ngo hashakishwe imiterere y’uturemangingo ifitanye isano n’ibintu bigabanya kanseri, oncogene, no kudakoresha imiti. Ikoreshwa rya CRISPR/Cas9 mu guhindura oncogene zitandukanye za TNBC rigaragazwa ku Ishusho ya 2.
Ishusho ya 2. Guhindura imiterere y’uturemangingo tw’uturemangingo twa TNBC hakoreshejwe CRISPR/Cas9 bigatuma ikibyimba kigabanuka kandi kikagabanuka. Izi ngirangingo zirimo CDK7, NAT1, UBR5, YTHDF2, ITGA9, CXCR4 na CXCR7, Crypto1, ROR1 na ST8SIA, zigira uruhare mu kubaho no kugabanuka kwa TNBC.
Kwimuka kw'uturemangingo twa kanseri, kwinjira, no guhinduka kwa epithelial-mesenchymal (EMT) bifitanye isano na ITGA9. Ubushakashatsi bwabanje bwagaragaje ko ITGA9 ari ingenzi mu nzira ya Notch kandi igira uruhare runini mu kwangirika kwa rhabdomyosarcoma (Molist et al., 2020). Byongeye kandi, ITGA9 yagaragaye ko ifitanye isano rya hafi n'ingaruka z'uburwayi bw'abarwayi mu bwoko butandukanye bwa kanseri, harimo na kanseri y'ibere (Wang Z. et al., 2019). Byongeye kandi, isesengura rya bioinformatics rya ITGA9 ryagaragaje ko imiterere yayo muri TNBC yari hejuru cyane ugereranyije n'izindi kanseri z'ibere. Igipimo cya ITGA9 cyo hejuru gifitanye isano no kwangirika kwa kanseri no kongera kugaragara mu barwayi ba TNBC. Guhagarika ITGA9 na CRISPR/Cas9 byatumye imiterere isa n'iya kanseri (CSC), angiogenesis ya kanseri, gukura kwa kanseri, no kugabanya kwangirika kwa kanseri binyuze mu gutuma β-catenin igabanuka muri TNBC (Wang Z. et al., 2019).
Crypto-1 ni kimwe mu bigize umuryango wa TGF-β kandi ni ingenzi cyane mu ivuka ry’urusoro, kubungabunga uturemangingo tw’urusoro, no kugabanuka kwa kanseri (Ishii et al., 2021). Izwi kandi nka Tdgf-1, ni poroteyine y’ibimenyetso ya GPI ifasha mu kugenzura ikorwa ry’umurongo w’ibanze, mesoderm na endoderm, ndetse no gushyiraho uburyo bwo kudahuza ibumoso n’iburyo mu iterambere ry’ingingo z’umubiri mu gihe cyo kuremwa k’urusoro (Zhang et al., 2021b). Byongeye kandi, Cripto-1 byagaragaye ko igira uruhare mu ihinduka rya epithelial-mesenchymal (EMT) nk’ikimenyetso cy’uturemangingo tw’urusoro (Zhang et al., 2021b). EMT ni ingenzi cyane atari gusa ku mikorere itandukanye nko gukura k’urusoro, fibrosis no gukira kw’ibikomere, ahubwo no mu kwinjira kwa kanseri no kugabanuka kwayo. Byongeye kandi, Cripto-1 byagaragaye ko ikorana n’uduce tune twa Notch kandi ikarushaho kongera ubushobozi bwayo bwo gukura nyuma yo guhinduka (Brandstadter na Maillard, 2019). Uburyo bwo gutanga ibimenyetso bya Notch buzwiho kugira uruhare mu kubungabunga uturemangingo twa kanseri y'ibere ku muntu. Ubushakashatsi bwagaragaje ko CRISPR/Cas9-itera Crypto-1 gucika burundu bigabanya ukwiyongera kwa kanseri no kwangirika kwayo. Kubwibyo, Crypto-1 ishobora kuba intego ikomeye yo kuvura TNBC (Castro et al., 2015).
Poroteyine ya XCL12 hamwe na receptor yayo ya chemokine CXC (CXCR4 na CXCR7) bigira uruhare rutandukanye mu kwiyongera kw'uturemangingo twa kanseri, gukura, kwimuka no kwinjira (Wu et al., 2015). Izi chemoreceptors zagiye zifitanye isano n'iterambere rya TNBC binyuze mu nzira nyinshi zo gutanga ibimenyetso haba muri vivo na in vitro models (Wu et al., 2015). Byongeye kandi, gukora kwa CXCR4 na CXCR7 bifitanye isano no kuba hashobora kubaho kwandura indwara no kudakomeza kubaho neza muri TNBC (Karn et al., 2022). Kubwibyo, gucika kwa CXCR4 na CXCR7 bishobora kuba gene nziza ku miti yo kuvura kanseri y'ibere, harimo na TNBC. Ubushakashatsi bwakozwe na Yang et al. Yang et al (2019) bwakoresheje CRISPR/Cas9 kugira ngo bufatanye mu gukuraho gene za CXCR4 na CXCR7 kandi bwasanze kwiyongera, gukura, kwimuka no kwinjira byagabanutse cyane (Yang et al., 2019).
Ubwiyongere bw'urugero rwa miR-3662, oncogene ya TNBC, bwagaragaye mu turemangingo twa kanseri y'ibere (Yi et al., 2022). Kwangirika kwa miR-3662 byagaragaye ko bibuza gukura kwa kanseri y'ibere no kwangirika kwa kanseri haba mu mubiri no mu mubiri (Agarwal na Gupta, 2021). HBP-1 ni ikintu gikomeye kibuza Wnt/-catenin signaling kandi gishobora kuba nyirabayazana w'ikura rya miR-3662 ry'uturemangingo twa TNBC. Vuba aha, Yi et al. bavumbuye ko miR-3662-HBP1 axis igenga inzira ya Wnt/-catenin signaling mu turemangingo twa TNBC (Yi et al., 2022). Bitewe n'uko ikibyimba kigaragara, miR-3662 ishobora kuba intego yo kuvura TNBC. Bityo, kwangirika kwa miR-3662 hakoreshejwe CRISPR/Cas9 bishobora kuba uburyo bwiza bwo gukora imiti mishya yo kuvura TNBC.
UBR5 ni nucleophosphoprotein ya 300 kDa yagaragaye nk'ikintu cy'ingenzi mu kugenzura ubukana bwa tumori, metastasis, n'uburyo ubudahangarwa bw'umubiri butera kanseri zitandukanye (Shearer et al., 2015; Fu et al., 2023). Byatangajwe ko UBR5 yiyongera cyane mu bipimo bya TNBC kandi ikangura imikorere ya ERα binyuze mu gutuma habaho kwiyongera binyuze mu bikorwa bya ubiquitin ligase (Bolt et al., 2015). Inyigo zose zo gukurikirana exome ku bipimo by'ibanze bya TNBC nazo zagaragaje ko UBR5 yiyongera, bigaragaza uruhare rwayo mu iterambere rya TNBC. Byongeye kandi, gusiba UBR5 bishingiye kuri CRISPR/Cas9 byagaragaje ko TNBC yagabanyije cyane metastasis no gukura mu bipimo by'imbeba zagerageje. Byongeye kandi, gushyira UBR5 mu gishushanyo cy'imbeba zo mu bwoko bwa wild-type byasubije imikorere yayo yose, mu gihe iyi ngaruka itagaragaye mu bwoko bwa mutant butakora (Liao et al., 2017). Kubura UBR5 bifitanye isano no kwiyongera kwa apoptosis, necrosis, no kubuza gukura kwa kanseri muri TNBC bitewe na angiogenesis mbi. Bitewe no kubura UBR5, gukwirakwira kwa kanseri mu ngingo za kure biragabanuka, kandi UBR5 itera EMT idasanzwe ahanini binyuze mu kugabanya E-cadherin (Zhang na Weinberg, 2018). Vuba aha, UBR5 byagaragaye ko ari ikintu cy'ingenzi cyane mu guhindura IFN-γ-induced PDL1 transcription muri TNBC bitewe no kutagira E3 ubiquitination activity. Isesengura rya RNA transcriptome ryagaragaje ko UBR5 ishobora kugira ingaruka ku miterere y'uturemangingo dufitanye isano n'inzira ya IFN-γ kandi igateza imbere transactivation ya PDL1 binyuze mu kongera urugero rw'imikorere ya protein kinase RNA (PKR) hamwe na transducers zayo na activators. transcription 1 (STAT1) na interferon regulatory factor 1 (IRF1). Ariko, gukuraho kwa UBR5 na PD-L1 hakoreshejwe CRISPR/Cas9 bifitanye isano n’uburyo bwo kuvura kurusha uko byakorwa mu buryo bwonyine, bikagira ingaruka zikomeye ku bidukikije bya kanseri (Wu et al., 2022). Bityo, CRISPR/Cas9 ishobora kuba igikoresho cy'ingenzi cyo kubuza imikorere ya UBR5, bityo bigahagarika ikwirakwira rya TNBC no gukura kwa kanseri.
ROR1 ni poroteyine yo mu bwoko bwa I transmembrane igaragara mu gihe cya kanseri no mu mikurire y’umwana uri mu nda kandi yagaragaye ko ari poroteyine yo mu nda (Nicholas Borcherding, 2014). Imyitwarire y’abantu ba kanseri zitandukanye ifitanye isano no kwiyongera kwa ROR1. Ibisubizo byiza byabonetse mu bushakashatsi bwakozwe mu buryo bwa "in vivo" no mu buryo bwa "in vitro" bugamije imiti ivura ROR1 (Chien et al., 2016). Ingano ya ROR1 mRNA mu biopsies by’ibere nabyo byagaragaye ko bifitanye isano n’ibibyimba by’ibere bisa n’ibisanzwe (BL) ndetse no kwimuka kwabyo mu bindi bice by’umubiri. Byongeye kandi, kwiyongera kwa ROR1 byagaragaye ko ari ikimenyetso cy’uko TNBC ishobora gutera imbere (Chien et al., 2016). Ariko, gucecekesha ROR1 hakoreshejwe CRISPR/Cas9 kugira ngo ugabanye gukura kwa TNBC no kwangirika kw’uturemangingo byaba ari ingamba nziza.
Uburyo bwo gukwirakwiza sialyl busaba kongeramo aside sialic muri glycoconjugates, bikongerwamo na sialyltransferases (STs). ST8SIA1 iri mu muryango wa ST kandi igira uruhare runini mu gutera indwara zitandukanye nka leukemia ya lymphocytic na kanseri y'inkondo y'umura (Chang et al., 2018). Isesengura rya RNA ryerekana ko ST8SIA1 igaragara cyane mu ngingo z'ibere z'abarwayi ba TNBC kandi ko ifitanye isano ryiza n'impinduka mu ngirabuzimafatizo ya tumor p53, ishobora kugira uruhare mu gutera kwa TNBC (Battula et al., 2017). Byongeye kandi, ST8SIA1 igira uruhare mu gutera no kongera kwa TNBC, bigaragaza ko igira uruhare runini mu kubaho no gukura kwa TNBC. Mu buryo bwa TNBC bwakozwe muri vitro, byagaragaye ko ST8SIA1 yagabanutse na CRIPSR/Cas9 ibuza gukura no gutera (Battula et al., 2017). Ibi bigaragaza ko CRISPR/Cas9 ishobora kuba igikoresho cy'ingenzi mu kubuza imikorere ya oncogene ya ST8SIA1 mu kuvura TNBC.
NAT1 ni enzyme ikora mu buryo bwa metabolisme itera imbaraga mu ikorwa ry’ibinyabutabire bya xenobiotic mu cyiciro cya kabiri kandi igaragara hafi mu ngingo zose z’umuntu. NAT1 ishobora gukoresha cofactor folate kugira ngo yibasire acetyl-CoA (acetyl-CoA) nubwo nta amine substrate ihari (Stepp et al., 2015; Laurieri et al., 2014). Ubushakashatsi bwagaragaje ko NAT1 igenzura imikorere ya matrix metalloproteinase 9 (MMP9) mu turemangingo twa kanseri y’ibere kandi ikarinda ubwoko bwa ogisijeni bukora (ROS) mu gihe cyo kwiyima isukari (Wang et al., 2018). Gukuraho NAT1 byagaragaye ko bibuza pyruvate dehydrogenase complex, bigatera imikorere mibi ya mitochondrial (Wang L. et al., 2019). Byongeye kandi, izindi raporo zitandukanye zagaragaje ko gukumira NAT1 hakoreshejwe molecule nto no gucecekesha siRNA bishobora kugabanya ukwiyongera no kwiyongera kw’uturemangingo twa kanseri y’ibere (Stepp et al., 2018). Vuba aha, CRISPR/Cas9 yakoreshejwe mu gukumira NAT1 mu ngirabuzimafatizo za kanseri y'ibere MDA-MB-231, bigira ingaruka ku mikorere y'uturemangingo bitewe n'urwego rw'imikorere yatwo. Ubu bushakashatsi bwagaragaje kandi ko NAT-1 ari ingenzi cyane mu iterambere no kwangirika kwa TNBC (Carlisle et al., 2020).
Guhinduranya uburyo oncogenes zikora bigengwa na super enhancers, transcription factors na cofactors (Hnisz et al., 2015). Byongeye kandi, itsinda rya cyclin-dependent kinases (CDKs), nka CDK7, CDK8, CDK9, CDK12, na CDK13, birakenewe kugira ngo hagenzurwe uburyo transcriptions zikorwa. Muri zo, CDK7 igira uruhare mu gukwirakwiza fosfori ya RNA polymerase II, ikaba ari ingenzi cyane mu gutangiza no kwagura transcriptions za oncogene mu ndwara ya TNBC. Mu bushakashatsi bw’ibanze bwakozwe, gusiba CDK7 na CRISPR/Cas9 byabujije TNBC, bigaragaza ko pathogenesis ya TNBC iterwa na CDK7 (Wang Y. et al., 2015).
Ubushakashatsi bwagaragaje ko impinduka muri MYC na RBP (poroteyine ifatanya na RNA) zishobora gutera apoptosis, mu gihe impinduka za gene imwe muri MYC cyangwa RBP zitagira ingaruka ku mikurire y'uturemangingo twa kanseri (Einstein et al., 2021). Hasuzumwe RBP zirenga 1000 muri genome y'umuntu hakoreshejwe isomero rya CRISPR/Cas9. Muri zo, RBP 57 zagaragaye ko ari ingenzi mu mikurire y'uturemangingo twa kanseri dufite urugero rwa MYC ruri hejuru cyane (Wheeler et al., 2020). Byongeye kandi, YTHDF2 ni ingenzi mu gukomeza gukura kw'uturemangingo twa TNBC kandi ikagabanya ingano y'inyandiko za methyl mu gihe cyo kwandika no guhindura mu turemangingo twa kanseri dufite MYC nyinshi. Byongeye kandi, YTHDF2 si ingenzi ku turemangingo twa kanseri, tudashingiye cyane ku rugero rwa MYC ruri hejuru kugira ngo abarwayi ba TNBC bakomeze kubaho (Einstein et al., 2021), bigaragaza ko ishobora kuba intego yo kuvura imiti ishobora gutsinda TNBC.
Poroteyine z'urutoki rwa zinc (ZNF) zigize hafi 1% by'uturemangingo tw'umubiri w'umuntu. Ubushakashatsi bwagaragaje ko ZNF ishobora kugenzura ukwiyongera kw'uturemangingo muri kanseri zitandukanye, nka kanseri y'umwijima, kanseri y'ibere na kanseri y'inkondo y'umura (Zhang W. et al., 2021; Li et al., 2017). Amasomero yakozwe na CRISPR knockout yakoreshejwe mu gusuzuma imiterere itandukanye y'uturemangingo (TSGs) mu turemangingo twa kanseri y'ibere (Shalem et al., 2014). Kuva icyo gihe, inyigo ya transcriptomic ku turemangingo twa kanseri y'ibere twa CRISPR/Cas9 ZNF319 twakuweho yagaragaje ko ZNF319 ari imiterere y'uturemangingo igabanya ukwiyongera kwa kanseri y'ibere bityo ikaba igira uruhare mu buryo butandukanye bwo gutanga ibimenyetso n'indi mirimo ya biyolojiya (Wang L. et al., 2022).
Ferroptosis ni ubwoko bw'urupfu rw'uturemangingo rwateguwe bitewe n'uko hari icyuma. Birazwi ko iterambere rya ferroptosis riterwa n'uko hari lipid peroxides. Byongeye kandi, byatangajwe ko PKCβII igaragaza peroxidation y'amavuta hakiri kare, kandi kwiyongera kwa peroxidation y'amavuta bifitanye isano na ferroptosis. Gusuzuma isomero ry'ibice birwanya kinase bya CRISPR / Cas9 byagaragaje ko PKCβII igira uruhare mu gikorwa cyo peroxidation y'amavuta, ibi bikaba ari ingenzi kuri ferroptosis mu turemangingo twa MDA-MB-231 (Zhang et al., 2022). Kubwibyo, ibi bigaragaza ko gukurwaho kwa PKCβII na CRISPR / Cas9 bishobora kuba intego y'uturemangingo duto two kurwanya kanseri mu kuvura indwara ziterwa na ferroptosis (Zhang et al., 2022).
Bivugwa ko kuba imiti idakora neza ari yo nyirabayazana w’impfu zigera kuri 90% mu barwayi ba kanseri kandi ni imwe mu mbogamizi zikomeye mu kuvura kanseri (Bukowski et al., 2020). Ibisubizo bigaragaza ko umubare uhagije w’uturemangingo tujyanye no gusohoka kw’imiti, gusana ADN, apoptosis n’inzira zitandukanye zo gutanga ibimenyetso ku turemangingo bifitanye isano no kudakora neza imiti (Haider et al., 2020). Muri byo, uturemangingo twinshi twibasiwe n’ibikoresho bya CRISPR/Cas9 kandi twagaragaje umusaruro mwiza mu kugabanya kudakora neza imiti no kongera imikorere myiza y’imiti irwanya kanseri (Vaghari-Tabari et al., 2022). Byongeye kandi, amasomero yo gusuzuma gene ya CRISPR/Cas9 afite ubushobozi bwo kuzamuka cyane yagize uruhare mu guhindura imikorere y’uturemangingo dushobora kuba dufite ubushobozi bwo kudakora neza imiti (Shalem et al., 2015). Kugira ngo hamenyekane imiterere y’uturemangingo twa paclitaxel, hakoreshejwe uburyo bwa RNA sequencing hamwe n’isuzuma rya sgRNA muri rusange rya genome kugira ngo hamenyekane imiterere umunani y’uturemangingo, harimo histone deacetylase 9 (HDAC9), ifitanye isano no kudakora imiti ku barwayi ba TNBC isubira (B et al., 2020). Mu bundi bushakashatsi, hagaragaye ukwiyongera k’ubwiyongere bwa diserine/threonine na tyrosine protein kinase (DSTYK) mu gihe abarwayi ba TNBC bakira imiti irwanya kanseri. Byongeye kandi, kugwa kwa DSTYK hakoreshejwe CRISPR/Cas9 byazamuye cyane apoptosis y’uturemangingo twa kanseri turwanya imiti muri vitro (uturemangingo twa SUM102PT na MDA-MB-468) ndetse n’icyitegererezo cya TNBC muri vivo (Ogbu et al., 2021).
Nubwo TNBC ikora ku buryo budasanzwe inzira ya MAPK, ingaruka z’ubuvuzi zishingiye kuri MEK ni mbi cyane. Isuzuma ryakozwe ku bubiko bwa genome rya CRISPR/Cas9 ryagaragaje ko gukumira PSMG2 (proteome assembly chaperone 2) bituma uturemangingo twa TNBC BT549 na MB468 tugera ku musemburo wa MEK AZD6244. Gutakaza PSMG2 na CRISPR/Cas9 byahinduye imikorere isanzwe ya proteasome, bituma PDPK1 ivanwaho, bityo bikarushaho kunoza uturemangingo twa kanseri mu buryo bwa TNBC bw’imbeba buterwa na AZD6244 (MEK inhibitor) na MG132 (proteasome inhibitor). Bityo, proteasome na MAP kinase (MEK) ikumirwa bishobora gutangwa mu buryo buhuriweho kugira ngo bigabanye ukwiyongera kw’uturemangingo twa kanseri (Wang X. et al., 2022).
CRISPR/Cas9 yanakoreshejwe mu gusuzuma niba hari ikibazo cy’imikorere y’uturemangingo tw’umubiri (gene) bitera TNBC resistance (Shu et al., 2020). Ge et al basobanuye uburyo uturemangingo twa kanseri twavuwe na JQ1, BET bromodomain inhibitor (BBDI), muri TNBC. Bakoresheje CRISPR/Cas9, basanze gusiba gene ya rb1 byatumye TNBC inanirwa kuvura kanseri JQ1. Kubwibyo, imikorere ya rb1 ni ingenzi mu gusubiza imiti ya JQ1 muri TNBC. Banavuze ko paclitaxel ari CDK4/6 kinase/microtubule inhibitor, kandi ko kuyihuza na BBDI nka JQ1 bishobora gutanga ibisubizo byiza ku buvuzi bwa TNBC idashobora kuvura kanseri (Ge et al., 2020).
Imiterere y’inyandiko ya RNA ndende idakora (lncRNA) yavuzwe ko ari yo ituma habaho ubudahangarwa ku buvuzi bwa neoadjuvant muri TNBC. Ubu bushakashatsi bwerekana ko inyandiko eshanu zitandukanye za MALAT1 lncRNA zigaragara cyane muri TNBC. Byongeye kandi, gusiba MALAT1 hakoreshejwe CRISPR/Cas9 byongera ubushobozi bw’uturemangingo twa TNBC BT-549 kuri paclitaxel na doxorubicin, bigaragaza ko bishobora kugira uruhare mu kudahangarwa kwa MALAT1 muri TNBC (Shaath et al., 2021).
Impinduka muri BRCA1 (BRCA1m) ziratandukanye bityo zikaba zigoye kuzimenya. Kwibanda kuri PARP1 (poly (ADP-ribose) polymerase), umufatanyabikorwa w’ubukorano wa BRCA1, bishobora kongera ubushobozi bwo kwandura kanseri mu miti ya TNBC. Gukoresha CRISPR/Cas9, gukuraho PARP1 byongera ubushobozi bwo kwandura imiti irwanya kanseri nka doxorubicin, gemcitabine na docetaxel ku turemangingo twa TNBC twa mBRCA1, bigaragaza ko PARP1 nayo igira uruhare mu kudakira imiti muri TNBC (Vaghari-Tabari et al., 2022). Impinduka muri gene zigabanya kanseri BRCA1 cyangwa BRCA2 zizwiho kongera amahirwe yo kurwara kanseri y'ibere mu bantu. Kuvura aba barwayi bisaba gukoresha imiti igabanya PARP. Byongeye kandi, byagaragaye ko gukuraho kwa nucleotide salvage factor DNPH1 hakoreshejwe CRISPR/Cas9 bishobora gukuraho nucleotide 5-hydroxymethyldeoxyuridine (hmdU) monophosphate irimo uburozi, bityo bikongera uburyo uturemangingo tudafite BRCA twakira PARP inhibitors (Fugger). Bityo, CRISPR/Cas9 na PARP1 inhibitors bishobora kuba ingamba y'ingenzi yo kuvura TNBC.
Ingirabuzimafatizo za glycoprotein (P-gp) ni ngirabuzimafatizo zirwanya imiti myinshi, muri rusange zikaba ziyongera ku kigero cya 41% by'imiti yose ya TNBC (Sun et al., 2020). Isohoka ry'imiti iterwa na P-gp ryagaragaye nk'ikintu cy'ingenzi kigenzura ubudahangarwa bw'imiti muri kanseri y'ibere. Imiti irwanya P-gp yagaragaje ko ishobora kongera ubushobozi bwo gufata imiti irwanya kanseri muri kanseri y'ibere (Famta et al., 2021). Bityo, gukuraho cyangwa guhagarika imiti irwanya P-gp na P-gp hakoreshejwe CRISPR/Cas9 bishobora kuba uburyo bw'ingenzi bwo gutsinda ubudahangarwa bw'imiti muri TNBC.
ATP-binding cassette transporter G2 (ABCG2) izwiho gutera ubudahangarwa bw'imiti muri TNBC (Palasuberniam et al., 2015), nubwo nta raporo zihari ku isano iri hagati ya CRISPR/Cas9 na ABCG2 silencing no kudakora kwa gene suppressor. PTEN ishobora kongera gukora kwa ABCG2 (Palasuberniam et al., 2015), (Deepak Singh et al., 2021). Kubwibyo, gukoresha CRISP/Ca9 kugira ngo ukureho ABCG2 no kuyivanga n'umuti ubuza ABCG2 bishobora kuba uburyo bwiza bwo kuvura ubudahangarwa bw'imiti muri TNBC. Imbonerahamwe ya 2 ivuga CRISPR/Cas9 yibanda ku ngirabuzimafatizo zose zirwanya ubudahangarwa.
Imbonerahamwe ya 2. CRISPR/Cas9 igamije ubwoko butandukanye bw’uturemangingo durwanya TNBC kugira ngo itume uturemangingo twitabira imiti irwanya kanseri.
Amasomero ya CRISPR/Cas9 ni ibikoresho bishobora gukoreshwa mu gusuzuma impinduka za gene zijyanye n’indwara ya kanseri (Chan et al., 2022). Ubu buryo bwo gusuzuma bukubiyemo intambwe enye nko (a) kubaka isomero, (b) gutanga lentiviral, (c) gusuzuma imiterere y’uturemangingo, na (d) gusesengura gene target. Nubwo TNBC ifite uburyo budasanzwe bwo gukora inzira ya MAPK, imiterere y’ubuvuzi bwa MEK ku barwayi ba TNBC bafite impinduka mu ngirabuzimafatizo zigabanya kanseri nka PTEN, RB1, na TP53 ni mbi cyane.
Byongeye kandi, isuzuma ryakozwe hifashishijwe CRISPR/Cas9 ryagerageje molekile ikomeye kandi itoranya, dehydrocatrol, iboneka cyane mu binyabutabire n'amababi bya Guatemala, kugira ngo humve ingaruka za dehydrocatrol kuri MDA-MB-231. Uburyo bwo guhumanya uturemangingo tw’ingirabuzimafatizo. Ibiranga imiterere ya mesenchymal y’ubwoko bwa TNBC. Iri suzuma rishingiye kuri CRISPR/Cas9 ryagaragaje kandi ko HSD17B11, gene ikora 17β-hydroxysteroid dehydrogenase type 11, igaragara cyane mu tunyabutabire twa MDA-MB-231 kandi igatanga dehydrofalcarinol yihariye ku tunyabutabire twa MDA-MB-231 (Grant et. al., 2020). Bityo, ibi bigaragaza ko isuzuma rya CRISPR/Cas9 rifite ubushobozi bukomeye mu kumenya uburyo bw’imiterere y’ibintu karemano bishobora kurwanya kanseri.
Byongeye kandi, kuba TNBC ishobora kurwara kanseri byakorewe ubushakashatsi hakoreshejwe uburyo bwo gusuzuma CRISPR/Cas9 mu buryo budafite aho bubogamiye, bwagaragaje isano iri hagati y’inzira za oncogenic n’izigabanya kanseri. Ibice by’ingenzi by’inzira za mTOR na Hippo byavuzwe ko bigira uruhare runini mu kugenzura kanseri muri TNBC. Byongeye kandi, ubushakashatsi bwagaragaje ko kubuza mTORC1/2 na YAP oncoprotein mu buryo bw’imiti bibuza neza indwara za TNBC hakoreshejwe uburyo bwa in vitro drug-matrix synergy model hamwe na xenografts zikomoka ku barwayi mu mubiri. Byongeye kandi, kubuza mTORC1/2 kwa Torin-1 kongera imbaraga za macropinocytosis, mu gihe kubuza YAP kwa vertebrorfin biganisha ku rupfu rw’uturemangingo twa TNBC. Ibyo bisubizo byose hamwe bigaragaza imbaraga n’ubudahangarwa bw’isuzuma rya CRISPR mu mubiri kugira ngo hamenyekane uburyo bushya kandi bufite akamaro bwo kuvura TNBC (Dai et al., 2021).
Kugabanuka k'ubudahangarwa bw'umubiri ni ikintu cy'ingenzi mu gutuma habaho kanseri. Utunyangingo twa kanseri twirinda ko ubudahangarwa bw'umubiri bushobora kunyura mu nzira zo kwirinda, harimo no kubangamira imikorere y'utunyangingo tw'umubiri mu bidukikije bya kanseri no kwangiza ubudahangarwa bw'umubiri. Kubwibyo, guteza imbere ubudahangarwa bw'umubiri bushobora kuba uburyo bw'ingenzi bwo kurwanya kanseri. Gukoresha CRISPR/Cas9-based gene modification bikemura ibibazo byinshi bijyanye n'imikorere mibi y'ubudahangarwa bw'umubiri mu buryo butandukanye. CRISPR/Cas9 yakoreshejwe mu kunoza ubudahangarwa bw'umubiri bwo kurwanya kanseri y'ibere binyuze mu nzira zikurikira (Ishusho ya 3).
Ishusho ya 3. Ubuvuzi bwa CRISPR/Cas9 bwibasira uturemangingo twa TNBC. Gutakaza CDK5 no gutakaza PDL1 na CD155 byongera ubudahangarwa bw'umubiri. Mu buryo nk'ubwo, gutakaza A2AR no gutakaza TAA nka HER2, mucin 1 na TEM8 byongereye ubushobozi bwa CAR-T mu kwica uturemangingo twa kanseri. Isuzuma rya T cells riyobowe na CRISPR/Cas9 ryagaragaje ko guhungabana kwa p38 kinase byongera imikorere ya T cells antitumor. Uturemangingo twa T twahinduwe na CRISPR/Cas9 twongera imikorere ya TCR (receptor ya T cells), bigatuma uturemangingo twa T tugira imikorere ya antitumor.
Intego yo kuvura indwara z’ubudahangarwa bw’umubiri ni ugukangura ubudahangarwa bw’umubiri kugira ngo burwanye uturemangingo twa kanseri. Ubusanzwe poroteyine nyinshi zo kugenzura ubudahangarwa bw’umubiri zirinda ingaruka z’ubwiyongere bw’umubiri ariko zishobora gufasha kanseri kuzirinda (Topalian et al., 2015). Izi poroteyine zirinda ingaruka z’ubudahangarwa bw’umubiri zifatanye n’uduce tw’umubiri ku buso bw’uturemangingo tw’umubiri. Poroteyine nyinshi zo kugenzura (nka CD155 na PD-L1) zireba agace k’umubiri k’umubiri k’umubiri kandi zigaragara muri kanseri y’ibere, cyane cyane TNBC (Li Y.-C. et al., 2020). Gukuraho PD-L1 cyangwa agace k’umubiri kayo hakoreshejwe CRISPR/Cas9 bishobora gutuma ubudahangarwa bw’umubiri bwibasira uturemangingo twa TNBC (Yahata et al., 2019). Byongeye kandi, kugabanya igaragara rya PD-L1 binyuze mu gukuraho CDK5 hakoreshejwe CRISPR/Cas9 byagaragaye ko bibuza gukura kwa kanseri mu buryo bwa vitro no mu mubiri (Deng et al., 2020). Kugabanya gukura kwa kanseri y'ibere mu buryo bwa "in vitro" na "in vivo" bigaragaza ko kugabanya CD155 binyuze muri shRNA bishobora kugira ingaruka ku ndwara ya kanseri y'ibere (Gao et al., 2018).
Uturemangingo twa CAR T dutanga CARs zimenya antigens zifitanye isano na kanseri (TAAs) kandi zishobora gukoresha poroteyine zo kugenzura kugira ngo zongere imikorere yazo (Li C. et al., 2020). Hari ingamba zitandukanye zishobora guterwa no kuvura uturemangingo twa CAR T muri kanseri y'ibere, harimo na TAA nyinshi nka HER2, mucin1, na TEM8 (Bajgain et al., 2018). Hari ibimenyetso bigaragaza ko uturemangingo twa CAR T twibasira mesothelin (igaragara cyane mu turemangingo twa TNBC BT-459) zigira akamaro kanini mu kurwanya kanseri iyo PD-1 ikuweho hakoreshejwe CRISPR/Cas9 (Hu et al., 2019). Ariko, gutandukanya uturemangingo twa T n'abarwayi hanyuma ukazihindura ex vivo ni igikorwa gitwara igihe kinini kandi gitwara igihe kinini. Nubwo uturemangingo twa T twose dushobora kugabanya ibisabwa mu kwitandukanya, uturemangingo twa T dutanga uturemangingo tugaragaza antigen ya leukocyte y'umuntu (HLA) yo mu cyiciro cya I na receptor ya T (TCR) tugomba gukurwaho kugira ngo hirindwe ko habaho ubusembwa bwa graft-versus-host (Ren et al., 2017). , 2017a). Hakoreshejwe CRISPR/Cas9, HDR ishobora gukuraho TCR na HLA icyarimwe no gukuraho jeni ikora CAR. Ubushakashatsi bwagaragaje ko CRISPR/Cas9 ishobora gukoreshwa mu kwinjiza anti-CD19 CAR mu gace ka TCR, bityo igatanga CAR neza nta gutakaza uturemangingo twa T (Dimitri et al., 2022). Ikoranabuhanga rya Multiplex ryakozwe kugira ngo rikore uturemangingo twa CAR T dukoresheje uburyo bwa allogeneic binyuze mu gukuraho TCR, beta-2-microglobulin (B2M), HLA-I subunit n'izindi poroteyine nka PD-1 na CTLA-4, zishobora kugira ubushobozi bwo kurwanya kanseri kurusha TNBC (Eyquem et al., 2017; Liu et al., 2017; Ren et al., 2017b; Dimitri et al., 2022).
Gukoresha CRISPR/Cas9 bishobora kunoza imikorere y’uturemangingo twa CAR-T. Adenosine izwiho kugira ubushobozi bwo gukumira ubudahangarwa bw’umubiri kandi ishobora kugabanya ubudahangarwa bw’umubiri mu kurwanya kanseri binyuze mu guhagarika imikorere y’uturemangingo twa T no gukora adenosine A2A receptors (A2AR) (Vigano et al., 2019). Gucecekesha A2AR hakoreshejwe CRISPR/Cas9 byazamuye cyane imikorere y’uturemangingo twa CAR-T mu mubiri (Giuffrida et al., 2021). Uturemangingo twa T tuzwiho kugaragaza imiterere itandukanye ya phenotypic nko kwaguka kw’uturemangingo, gutandukanya uturemangingo, stress ya oxidative na stress ya genomic. Gusuzuma uturemangingo twa T bishingiye kuri CRISPR/Cas9 byagaragaje ko habayeho ihungabana ry’uturemangingo 25 twa T cell receptor kinase. Muri two, gusiba p38 kinase byagaragaye ko byongera imikorere y’uturemangingo twa T antitumor, bigaragaza ko p38 kinase ari ingenzi mu kugenzura imikorere y’uturemangingo twa CAR-T (Gurasamy et al., 2020).
Selile T zishobora guhindurwa mu buryo bw’uturemangingo hakoreshejwe CRISPR/Cas9 kugira ngo zikore TCR nyinshi. Kwimura selile T zahinduwe mu buryo bw’uturemangingo ku barwayi byagaragaje ko zirwanya kanseri cyane kurusha selile T zahinduwe mu buryo bw’uturemangingo. Kubwibyo, TCR zishobora guhangana na TCR zahinduwe mu buryo bw’uturemangingo ku barwayi, bishobora kugira ingaruka ku bushobozi bwo kuvura kanseri. Kugira ngo iki kibazo gikemuke, gukoresha CRISPR/Cas9 gukuraho TCR-β mu turemangingo twakira hanyuma bigahita byohereza selile T za TCR-β ku barwayi ba kanseri bishobora kugaragaza ubudahangarwa bw’umubiri bwiza bwo kurwanya kanseri hatabayeho gusiganwa ku mibonano mpuzabitsina na TCR (Fan et al. 2018). Bityo, usibye selile T zahinduwe na CRISPR, TCR ziba zifite ubushobozi bwo kwiyumvamo antigens za kanseri inshuro igihumbi kurusha selile T zahinduwe na TCR. Byongeye kandi, mu ndwara zitandukanye za leukemia, selile T zahinduwe zakozwe na γδ TCR+ CRISPR zigaragaza uburyo selile T za CD4+ na CD8+ zigaragara cyane kurusha selile TCR zisanzwe (Legut et al., 2018). Bityo, uturemangingo twa T twahinduwe twakozwe na CRISPR/Cas9 dushobora kuba uburyo bwiza bwo kuvura indwara z’ubudahangarwa mu kurwanya TNBC.
Integrins ni molekile zifatana mu turemangingo ziboneka mu turemangingo kandi zigatuma uturemangingo dufatana na matrix yo hanze y’uturemangingo (ECM) (Hamidi na Ivaska, 2018). Kugabanuka kw’uturemangingo bijyana no gukura no kwimuka kwa kanseri binyuze mu guhindura matrix yo hanze y’uturemangingo, bigatuma uturemangingo twa kanseri tubaho mu miyoboro y’amaraso (Hamidi na Ivaska, 2018). CRISPR/Cas9-mediated integrin knockdown igabanya iterambere rya kanseri, ihindagurika ry’uturemangingo, no kwiyongera kwa colonne muri TNBC. Kugabanuka kwa integrin a5 (ITGA5) byavuzwe ko bigabanya kwimuka no kwiyongera kw’uturemangingo mu zindi kanseri nka kanseri y’ibihaha (Ju et al., 2017), bigaragaza ko integrin a5 ishobora kuba ikintu cy’ingenzi mu gutera kwa TNBC.
Gukora imbeba zo mu bwoko bwa "knockout" hakoreshejwe uburyo bwa gakondo bw'uturemangingo tw'inda (ES) ni igikorwa gisaba imbaraga nyinshi, gitwara igihe kinini, kandi kidakora neza. Bifata amezi n'imyaka kwibasira uturemangingo twa ES binyuze mu kongera guhuza uturemangingo, korora imbeba za chimeric, hanyuma ukazihuza n'imbeba za heterozygous kugira ngo zibyare abana ba homozygous. Guhuza amoko y'imbeba birakenewe kugira ngo habeho imbeba zifite impinduka nyinshi za gene. Ariko, hari ibibazo byinshi byavutse iyo hakoreshwa imbeba zaturutse ku gukuraho uturemangingo twa ES hakoreshejwe CRISPR/Cas9 cyangwa gushyira ibice bya CRISPR/Cas9 mu magi y'ingirabuzima fatizo imwe. Urugero, impinduka zikunze kubaho kuri biallelic loci kandi ntabwo ari gene yihariye. Biherutse kuvugwa ko uturemangingo twa ES dukoresha ikoranabuhanga rya CRISPR/Cas9 dushobora gushyiramo impinduka za biallelic icyarimwe mu turemangingo tugera kuri 5 (Nishizono et al., 2021). Kuri iyi ntego, Cas9 mRNA na gRNA eshanu zihariye za gene byashyizwe icyarimwe mu turemangingo twa ES. Ibi bigaragaza icyizere n'akamaro k'ubu buryo, nubwo izi mpinduka zishobora kuba zaratanzwe igihe imirongo y'ibanze yavumburwaga kugira ngo hakorwe imbeba eshanu z'impinja. Ubushakashatsi kandi buvuga ko havumbuwe ikintu gitangaje: uturemangingo twa ES ntabwo tugikenewe mu gukora imbeba zahinduwe mu buryo bwa genetiki. Ahubwo, kugira ngo havanweho ibikomoka kuri genetiki runaka, Cas9 mRNA na gRNA byashyizwe mu ntanga z'uturemangingo tw'ingirabuzima fatizo imwe. Kubera iyo mpamvu, imirongo y'ibanze yakozwe ishobora gukoreshwa mu kwiga ingaruka zo gusiba gene mu mbeba (Qin et al., 2016). Bityo, CRISPR/Cas9 yemerera iremwa ry'imbeba zahinduwe mu buryo bwa genetiki kugira ngo zivure TNBC ku giciro gito ugereranyije n'ubuhanga busanzwe bwa genetiki. Hakoreshejwe sisitemu ya CRISPR/Cas, hakozwe icyitegererezo gishya cy'imbeba za knockout gishobora gutangiza neza impinduka mu ngingo muri gene imwe cyangwa nyinshi z'umwimerere muri TNBC. Hashingiwe kuri iki gitekerezo, ingero z'inyamaswa za TNBC zishobora kandi gukorwa hakoreshejwe CRISPR/Cas9-mediated deletion ya BRCA1 na p53, bigatera gutakaza HR repair, kudahinduka kwa genome, na fenicha ya mutant (Annunziato et al., 2020).
Muri iki gihe, uburyo butandukanye nka mammography, magnetic resonance imaging (MRI) na ultrasound ni byo bikoreshwa mu gusuzuma TNBC. Ariko, ubu buryo bufite imbogamizi zimwe na zimwe. Mammography ikoreshwa mu gusuzuma ingingo z'ibere aho gusuzuma metastases, aho uturemangingo twa kanseri twimukiye mu zindi ngingo. Ultrasonography si uburyo bwizewe bwo gusuzuma TNBC (Chen na Lee-Felker, 2023). MRI ifite ubushobozi bwo kumenya kurusha ultrasound na mammography, ariko uburyo bwo gusuzuma ni buke (Sha na Chen, 2022). Biopsy y'uturemangingo ni uburyo bwo kumenya uturemangingo twa kanseri. Ariko, rimwe na rimwe, biopsy ishobora kubura uturemangingo twa kanseri niba urushinge ruvuye mu gace rukeneye. Byongeye kandi, birahenze cyane kandi bishobora gutera ihungabana ku murwayi. TNBC ni ubwoko butandukanye bwa kanseri, bityo biopsy ishobora kudatanga amakuru ahagije ku bwoko bwa kanseri. Kubwibyo, uburyo bushya bwizewe bwo gusuzuma TNBC burakenewe byihutirwa. CRISPR/Cas9 ishobora gukora nk'ubundi buryo bworoshye cyane kandi buto bwo gusuzuma kanseri y'ibere. Muri urwo rwego, ingamba zishingiye kuri CRISPR/Cas9 zishobora gukoreshwa mu kunoza uburyo bwa PCR bwo gusuzuma TNBC. Ubwa mbere, poroteyine za Cas9 na cpf1 zo muri sisitemu ya CRISPR zikoreshwa mu gukuraho ADN itari yihariye, hanyuma izi poroteyine ebyiri (Cas9 na cpf1) zishobora kumenya umurongo wa PAM mbere yo gufatana na ADN yibandwaho (Deepak Singh et al., 2021). Bityo, PCR ishobora kumenya impinduka zijyanye n'iterambere rya kanseri. Ubushakashatsi bwinshi bwakoresheje iyi ngamba ishingiye kuri CRISPR kugira ngo hamenyekane impinduka zitandukanye muri kanseri zitandukanye (Safari et al., 2019). Bityo, uburyo bwa PCR bushingiye kuri CRISPR bushobora kugabanya kwishingikiriza ku buryo bwo gusuzuma TNBC ku buryo bwo kwibasira nka biopsy-based immunohistochemistry.
Impinduka mu miterere y’uturemangingo tw’imisemburo nazo zagaragaye muri TNBC (Chen na Russo, 2009). Ingamba za PCR zishingiye kuri CRISPR/Cas9 zikoresha microarrays mu gusuzuma indwara zishobora gukurikirana neza izi mpinduka (Hajian et al., 2019). Bishobora kandi guha abakora mu buvuzi amakuru ajyanye n’impinduka zihariye ku barwayi ba TNBC, bishobora gufasha kunoza uburyo bwabo bwo kuvura. Ariko, ubu buryo buhuriweho bufite imbogamizi. Kubwibyo, hakenewe ubushakashatsi bwinshi mbere yuko ubu buryo bwa CRISPR/Cas-PCR bukoreshwa mu buvuzi mu gusuzuma TNBC (Yang et al., 2019).
Igihe cyo kohereza: 14 Ukwakira 2024