CD44 and TNFSF14 Identified as Key Immune Targets in Glioblastoma

April 23, 2025 Author Summary 0

CD44 and TNFSF14 Identified as Key Immune Targets in Glioblastoma

New Insights into Glioblastoma’s Immune Evasion: CD44 and TNFSF14 Take Center Stage

Glioblastoma (GBM) remains one of the most aggressive and lethal cancers, with a median survival of just 15 months. Despite intensive treatment that combines surgical removal, radiation therapy, and chemotherapy (most often with temozolomide), progress has been limited. This lack of therapeutic breakthroughs is largely due to the tumor’s complexity—its heterogeneity, ability to recur, and the protective nature of the blood–brain barrier, which hinders drug delivery.

GBM is not only comprised of rapidly dividing tumor cells but also includes a population of cancer stem cells that fuel resistance and recurrence. Beyond the cancer cells themselves, the tumor microenvironment (TME) plays a critical role. It is a mix of immune and non-immune cells, including regulatory T cells (Tregs), natural killer (NK) cells, and macrophages, which together create an immunosuppressive environment that helps the tumor thrive.

Although immunotherapy has transformed treatment for several cancer types, its success in GBM has been modest. One reason is the tumor’s ability to suppress immune responses through mechanisms such as the PD-1/PD-L1 checkpoint pathway. PD-L1 is a protein that helps tumor cells evade immune detection by disabling T cell function. While PD-L1 is present in GBM, its expression varies, and clinical trials targeting it have had disappointing results.

In light of these challenges, a new study published in Biomolecules and Biomedicine by Zottel et al. sought to explore the PD-L1 interaction network in GBM to uncover alternative immune targets. The research identified two genes—CD44 and TNFSF14—as potential contributors to the tumor’s immunosuppressive behavior.

Mapping the PD-L1 Network

The study began by constructing a network of genes associated with PD-L1 using bioinformatics tools including Cytoscape and STRING. Over-representation analysis revealed that the genes connected to PD-L1 are heavily involved in immune-related processes, such as PD-1 signaling, T cell activation, and allograft rejection. These results confirmed the central role of immune regulation in GBM’s pathology.

From this network, the researchers focused on 15 genes that were consistently overexpressed in GBM compared to lower-grade glioma (LGG) and normal brain tissue in at least two major datasets—TCGA, CGGA, and Rembrandt. They further narrowed the list to three genes—CD44, TNFSF14, and HOXD13—which also correlated with worse patient survival.

Experimental Validation Confirms Overexpression

To support the in silico findings, the research team validated gene expression using qPCR on human tissue samples, including GBM, LGG, and healthy brain tissues. The results showed that CD44 and TNFSF14 were significantly overexpressed in GBM compared to both LGG and normal tissue. These genes were also able to distinguish GBM from normal brain with high sensitivity and specificity, achieving ROC curve AUC values of 0.961 and 0.834, respectively. HOXD13, while overexpressed, showed lower diagnostic value (AUC = 0.553).

Subtype Specificity: A Link to Mesenchymal GBM

GBM can be classified into molecular subtypes, including classical, neural, proneural, and mesenchymal (ME). The mesenchymal subtype is linked to worse prognosis, greater immune suppression, and more aggressive tumor behavior.

CD44 and TNFSF14 were found to be more strongly expressed in the ME subtype, suggesting a potential role in driving the subtype’s immunosuppressive features. This connection is particularly important, as recurrence in GBM often involves a transition from proneural to mesenchymal subtype, further complicating treatment.

Ties to the Immune Landscape

Using TIMER 2.0, a tool that estimates immune cell infiltration from gene expression data, the authors found that CD44 and TNFSF14 expression positively correlated with regulatory T cells (Tregs) and resting NK cells—both indicators of an immune-suppressive environment. Notably, both genes negatively correlated with activated NK cells, which are crucial for anti-tumor immunity.

Additionally, CD44 was positively associated with resting memory CD4+ T cells and negatively with M2 macrophages, further hinting at its role in immune modulation.

The study also showed a moderate positive correlation between CD44 and PD-L1 expression, while TNFSF14 showed a weaker positive association. This implies that these genes may participate in the same immunoregulatory pathways or contribute to PD-L1 expression regulation.

Broader Context: Pan-Cancer Gene Expression

To assess the relevance of these findings in other cancer types, the team analyzed gene expression using the GEPIA database. CD44 was broadly overexpressed in many tumors, while TNFSF14 generally showed higher expression in normal tissues, with a few exceptions including GBM. HOXD13 had variable expression across tumors, with limited utility as a GBM marker.

What This Means for GBM Research

This study highlights CD44 and TNFSF14 as important players in GBM’s immune landscape. Their consistent overexpression in GBM, especially in the ME subtype, and association with immune evasion mechanisms suggests they could serve as biomarkers for prognosis or therapeutic targets.

Both genes are involved in immune regulation:

  • CD44 is known to mark GBM stem cells and is implicated in cancer cell invasion and resistance.
  • TNFSF14 (also known as LIGHT) influences T cell infiltration and interacts with immune checkpoint molecules.

The research aligns with other studies suggesting these genes may be useful in guiding immunotherapy or monitoring immune activity in GBM. Importantly, their association with PD-L1 expression supports further investigation into combination therapies or immune checkpoint modulation strategies.

Future Directions

While the current findings are promising, the authors emphasize the need for functional studies. This includes co-culture experiments with immune cells, gene knockout models, and single-cell sequencing to understand the precise roles of CD44 and TNFSF14 in tumor immunity.

Such studies could help develop more effective immunotherapies that overcome the challenges posed by GBM’s complex microenvironment and heterogeneity.

Conclusion

This research underscores the potential of CD44 and TNFSF14 as biomarkers and immune regulators in GBM. By connecting gene expression data to immune suppression and tumor subtype, the study lays the groundwork for more targeted and responsive treatment strategies. For researchers and clinicians in the field, these insights could help steer the development of next-generation immunotherapies for one of the most challenging cancers.

 

The translation of the preceding English text in Slovenian:

 

Nove ugotovitve o imunski izmikanju glioblastoma: CD44 in TNFSF14 v ospredju

Glioblastom (GBM) ostaja eden najagresivnejših in najbolj smrtonosnih rakov, s povprečnim preživetjem le 15 mesecev. Kljub intenzivnemu zdravljenju, ki vključuje kirurško odstranitev, radioterapijo in kemoterapijo (najpogosteje s temozolomidom), je napredek omejen. Razlog za pomanjkanje terapevtskih prebojev tiči predvsem v kompleksnosti tumorja — njegovi heterogenosti, sposobnosti ponovitve in zaščitni vlogi krvno-možganske pregrade, ki otežuje dostavo zdravil.

GBM ne sestavljajo le hitro delujoče tumorske celice, ampak tudi populacija rakavih matičnih celic, ki prispevajo k odpornosti in ponovitvi. Poleg teh celic ima pomembno vlogo tudi tumorsko mikrookolje (TME). To je mešanica imunskih in neimunskih celic, vključno z regulatornimi T celicami (Treg), naravnimi celicami ubijalkami (NK) in makrofagi, ki skupaj ustvarjajo imunosupresivno okolje, ki tumorju omogoča uspevanje.

Čeprav je imunoterapija spremenila zdravljenje številnih vrst raka, so bili njeni uspehi pri GBM skromni. Eden od razlogov je sposobnost tumorja, da zavira imunske odzive z mehanizmi, kot je PD-1/PD-L1 pot. PD-L1 je beljakovina, ki tumorju pomaga uiti zaznavi imunskega sistema z zaviranjem delovanja T celic. Čeprav je PD-L1 prisoten pri GBM, je njegova izraženost spremenljiva, klinične študije, usmerjene vanj, pa niso prinesle zadovoljivih rezultatov.

Zaradi teh izzivov je nova študija, objavljena v reviji Biomolecules and Biomedicine avtorjev Zottel et al., raziskovala interakcijsko mrežo PD-L1 pri GBM z namenom odkriti alternativne imunske tarče. Raziskava je identificirala dve potencialni ključni geni — CD44 in TNFSF14, ki naj bi prispevali k imunosupresivnemu vedenju tumorja.

Kartiranje PD-L1 mreže

Študija se je začela z oblikovanjem genske mreže, povezane s PD-L1, s pomočjo bioinformatičnih orodij, kot sta Cytoscape in STRING. Analiza prenasičenosti je pokazala, da so geni, povezani s PD-L1, močno vključeni v imunske procese, kot so PD-1 signalizacija, aktivacija T celic in zavrnitev presadkov. Ti rezultati potrjujejo osrednjo vlogo imunske regulacije v patogenezi GBM.

Iz te mreže so raziskovalci izbrali 15 genov, ki so bili dosledno prekomerno izraženi v GBM v primerjavi z gliomi nižjega razreda (LGG) in normalnim možganskim tkivom, v vsaj dveh večjih bazah podatkov — TCGA, CGGA in Rembrandt. Seznam so nadalje zožili na tri gene — CD44, TNFSF14 in HOXD13, ki so bili povezani tudi s slabšim preživetjem bolnikov.

Eksperimentalna potrditev prekomerne izraženosti

Za podporo in silico ugotovitvam je raziskovalna skupina preverila izraženost genov z metodo qPCR na človeških vzorcih tkiv, vključno z GBM, LGG in zdravim možganskim tkivom. Rezultati so pokazali, da sta bila CD44 in TNFSF14 izrazito prekomerno izražena v GBM v primerjavi z LGG in normalnim tkivom. Ta dva gena sta bila sposobna ločiti GBM od zdravega tkiva z visoko občutljivostjo in specifičnostjo (AUC 0,961 in 0,834). HOXD13 je imel nižjo diagnostično vrednost (AUC = 0,553).

Specifičnost podtipov: povezava z mezinhimskim GBM

GBM lahko razdelimo na molekularne podtipe, vključno s klasičnim, nevronskim, pronevralnim in mezinhimskim (ME). Mezinhimski podtip je povezan s slabšo prognozo, večjo imunosupresijo in bolj agresivnim vedenjem tumorja.

CD44 in TNFSF14 sta bila močneje izražena v ME podtipu, kar kaže na možno vlogo pri oblikovanju imunosupresivnih lastnosti tega podtipa. Ta povezava je pomembna, saj se ponovitev GBM pogosto pojavi s prehodom iz pronevralnega v mezinhimski podtip, kar dodatno otežuje zdravljenje.

Povezava z imunskim okoljem

Z uporabo orodja TIMER 2.0, ki ocenjuje infiltracijo imunskih celic na podlagi izraženosti genov, so avtorji ugotovili, da izraženost CD44 in TNFSF14 pozitivno korelira z regulatornimi T celicami (Treg) in mirovanjem NK celic — kar sta kazalnika imunosupresivnega okolja. Opažena je bila tudi negativna korelacija z aktiviranimi NK celicami, ki so ključne za protitumorsko imunost.

Poleg tega je bil CD44 pozitivno povezan z mirovanjem spominskih CD4+ T celic in negativno z M2 makrofagi, kar dodatno kaže na njegovo vlogo v imunski modulaciji.

Študija je pokazala tudi zmerno pozitivno korelacijo med izraženostjo CD44 in PD-L1, medtem ko je bil pri TNFSF14 ta odnos šibkejši. To nakazuje, da lahko ti geni sodelujejo v enakih imunoregulacijskih poteh ali prispevajo k regulaciji izraženosti PD-L1.

Širši kontekst: izraženost genov pri drugih rakih

Za oceno pomembnosti teh ugotovitev pri drugih vrstah raka je skupina analizirala izraženost genov v bazi GEPIA. CD44 je bil splošno prekomerno izražen pri številnih tumorjih, medtem ko je bil TNFSF14 večinoma bolj izražen v zdravih tkivih, z nekaj izjemami, vključno z GBM. HOXD13 je imel spremenljivo izraženost med tumorji in omejeno uporabnost kot označevalec za GBM.

Kaj to pomeni za raziskave GBM

Ta študija poudarja CD44 in TNFSF14 kot pomembna dejavnika v imunskem okolju GBM. Njuna dosledna prekomerna izraženost, zlasti v ME podtipu, in povezava z mehanizmi izmikanja imunskemu sistemu nakazujeta, da bi lahko služila kot biološki označevalci prognoze ali terapevtske tarče.

Oba gena imata vlogo v imunski regulaciji:

  • CD44 označuje matične celice GBM in je povezan z invazijo ter odpornostjo rakavih celic.

  • TNFSF14 (znan tudi kot LIGHT) vpliva na infiltracijo T celic in interakcijo z imunskimi nadzornimi molekulami.

Raziskava sovpada z drugimi študijami, ki kažejo, da bi bila ta gena lahko uporabna pri vodenju imunoterapije ali spremljanju imunskega odziva pri GBM. Njuna povezanost z izraženostjo PD-L1 podpira nadaljnje raziskave o kombiniranih terapijah ali strategijah modulacije imunske kontrole.

Prihodnje usmeritve

Čeprav so trenutne ugotovitve obetavne, avtorji poudarjajo potrebo po funkcionalnih študijah. To vključuje co-culture poskuse z imunskimi celicami, modele z izbitjem genov (knockout) in enocelično sekvenciranje za razumevanje natančnih vlog CD44 in TNFSF14 v tumorski imunosti.

Takšne študije bi lahko pripomogle k razvoju učinkovitejših imunoterapij, ki bi presegle izzive kompleksnega mikrookolja GBM in njegove heterogenosti.

Zaključek

Raziskava poudarja potencial CD44 in TNFSF14 kot označevalcev in imunskih regulatorjev pri GBM. S povezovanjem izraženosti genov z imunosupresijo in tumorskim podtipom študija postavlja temelje za bolj ciljno usmerjene in učinkovite terapevtske pristope. Za raziskovalce in klinične zdravnike na tem področju te ugotovitve ponujajo smernice za razvoj naslednje generacije imunoterapij proti enemu najzahtevnejših rakov.

Reference:

Alja Zottel, Neja Šamec, Ivana Jovčevska

TNFSF14 and CD44 are overexpressed in glioblastoma and associated with immunosuppressive microenvironment.

Biomol Biomed [Internet]. 2025 Feb. 13 [cited 2025 Apr. 22];

Available from: https://www.bjbms.org/ojs/index.php/bjbms/article/view/11791

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