Autophagy: autophagy is a crucial process that allows the recycling of important cellular components in response to stress conditions such as those induced by cisplatin treatment. Indeed, regulation of autophagy has been widely associated with cisplatin resistance phenotype. Rescue of miR-1-3p increases the sensitivity of cisplatin resistance cells by inhibiting ATG3, a key autophagic protein
[38]. Furthermore, exosomal transfer of miR-425-3p was found to increase autophagic flux and chemoresistance by inhibiting AKT1 in the targeted NSCLC cells
[39].
-
4. Beyond Targeting NSCLC Cells: The Role of miRNA in Regulating Immune Response to Cisplatin Treatment
Besides targeting tumor cells, platinum compounds can also elicit an immune response against cancer cells through different mechanisms, i.e., by augmenting CD8
+ T cells tumor infiltration, inducing maturation of Antigen-presenting cells (APCs), by downmodulating regulatory T cells (Tregs) and decreasing Myeloid-derived suppressor cells (MDSCs) which compose the so-called ‘immune permissive environment’
[40][41]. Moreover, cisplatin was also described to induce PD-L1 expression in vitro and in vivo
[42][43][44][45][46], a co-inhibitory factor of the immune response which in turn augments the sensitivity of cancer cells to therapies using immune checkpoint inhibitors (ICIs) against PD-L1 [
[47][48]. In keeping with this, Fujita et al., demonstrated that miR-197-3p modulated cisplatin resistance by targeting the cyclin-dependent kinase CKS1B and, indirectly, the transcription factor STAT3, which ultimately leads to increased PD-L1 expression thus sensitizing PD-L1
high tumor cells to cisplatin
[42]. Alternatively, an independent report showed that LncRNA MALAT1 induces chemoresistance of NSCLC cells through direct regulation of p120 catenin by competing with miR-197-3p
[49].
Other miRNAs were found functionally involved in modulating cisplatin resistance though the exact molecular mechanisms were unclear
[17][50][51][52][53]
5. miRNA Regulation of Platinum-Based Therapy Response
Cisplatin is the standard first-line treatment for advanced-stage NSCLC. It can be administered in combination with third-generation anticancer agents such as Gemcitabine, Docetaxel, Paclitaxel, or Vinorelbine, thus also called platinum-based doublet chemotherapy aka P-doublet
[54]. As shown in
Figure 1, a lower number of miRNAs was found involved in regulating responses to P-doublet in NSCLC, which is mainly ascribable to a lower number of research studies that analyzed miRNAs roles in P-doublet therapy. However, Lin et al. showed that miR-30-5p expression impairs resistance to cisplatin alone or in combination with pemetrexed
[55] while it enhances paclitaxel sensitivity in an independent study
[56]. Cai et al. found that miR-128-3p upregulation increased resistance to multiple drugs though used alone (cisplatin, gemcitabine, or paclitaxel;
[57]), probably suggesting a similar effect when combined therapy would be otherwise used. Other miRNAs were also identified to regulate sensitivity to both cisplatin and paclitaxel (i.e., miR-186-5p
[58][59]; miR-17-5p
[17][60][61]; miR-34c-3p
[62]) or cisplatin and docetaxel (miR-141-3p
[63][64]; and miR-379-5p
[65],
[66]).
6. miRNAs Which Modulate Response to EGFR Inhibitors
Mutations in EGFR are frequent in NSCLC patients (~10–20% of Caucasian, and up to 60% of South-East Asian patients) and cause the constitutive ligand-independent activation of EGFR receptor thus promoting cell growth and survival
[67]. EGFR tyrosin kinase inhibitors (TKIs) are small molecules that bind the adenosine triphosphate (ATP) pocket of EGFR and inhibit its autophosphorylation and downstream signal transduction
[68], including RAS–MAPK, PI3K–AKT, and JAK–STAT signaling. Initially, first- (gefitinib, erlotinib) and second-generation TKIs (afatinib, dacomitinib) have been developed and showed a better efficacy vs. platinum-based therapy alone in EGFR-mutated patients
[69][[70][71]. More recently, third-generation TKIs, such as Osimertinib, have shown a significant improvement in progression-free survival and overall survival
[72], thus becoming the front-line therapy for patients with EGFR-mutated NSCLC
[73]. Despite the initial response, resistance to EGFR TKIs inevitably occurs through both EGFR-target-dependent and independent mechanisms
[74]. Some miRNAs were shown to modulate sensitivity to Gefitinib, Erlotinib, Afatinib, and Osimertinib with a high level of evidence (score ≥ 4 or 5) (
Figure 1). In this context, transcriptome rewiring of TKIs resistant cells upon miRNA induction would activate alternative signaling pathways to bypass EGF inhibition
[74]. For example, the rescue of miR-19a expression reverses gefitinib resistance in NSCLC by directly targeting 3′UTR of c-MET
[75] which is one of the most altered pathways implicated in EGFR-TKIs inhibition
[3][74]. Loss of PTEN was associated with both primary and acquired EGFR-TKIs resistance due to an increased PI3K signaling. Accordingly, miR-21 overexpression is associated with acquired resistance to gefitinib via inhibition of PTEN and PDCD4 and induction of PI3K/AKT
[28][76]. Notably, increased expression of miR-21 was associated with afatinib resistance in both in vitro experimental models and in patients
[77].
Other pathways were implicated in modulating sensitivity to EGFR-TKIs such as the regulation of NF1 which is a GTPase that negatively regulates the Ras/MEK/ERK pathway and is targeted by miR-641 regulation
[78]. Yet, miR-326 impairs resistance to Gefitinib by direct inhibition of the type I interferon receptor IFNAR2
[79], which is relevant for JAK-STAT signaling activation
[80] and is associated with the response to EGFR-TKIs
[81]. Likewise, miR-762 is a potential downstream effector of the IL-6/STAT3 pathway and regulates gefitinib resistance via direct regulation of the target gene ABR
[82]. Yet, loss of expression of miR-206 induces the over-activation of STAT3 which leads to Gefitinib resistance
[83]. Furthermore, miR-206 have been reported to regulate response to gefinitib via other molecular mechanisms, including the direct regulation of the multidrug resistance protein ABCB1
[84] and by overcoming HGF activity (i.e., the ligand of Met) and promote MAPK and PI3K/AKT downstream pathways
[85].
Gefinitib activity can be also modulated by the regulation of miR-146b-5p/nuclear factor kB (NF-kB) axis which impacts the NF-kB-related IL-6 and IL-8 production and ultimately enhances gefitinib-induced apoptosis
[86]. Other miRNAs can influence Gefitinib response by regulating phenotypic traits of resistant cells. For example, Let-7 downregulation concomitantly increases in vitro self-renewal capability and Gefitinib resistance via direct regulation of MYC
[87]. Furthermore, miR-17-5p increases tumor sphere formation and Gefitinib resistance by targeting CDKN1A (aka p21)
[87][88]. This miRNA has also been associated with Erlotinib resistance although the molecular mechanisms have not been yet investigated
[89]. Modulation of YAP1 gene expression by miR-7 is another mechanism of Gefitinib resistance via induction of EMT. Indeed, YAP1 was found to be targeted by miR-7 also through exosomal miR-7 transfer
[90]. Similarly, miR-200c-3p inhibits downstream signaling pathways of EGFR and regulates both EMT and gefitinib resistance
[91][[92]. MiR-124-3p was found to be downregulated in gefitinib-resistant NSCLC patients, and its overexpression reversed EMT transformation and gefitinib resistance, at least in part, by the direct regulation of SNAI2 and STAT3
[93]. Contrariwise, miR-124-3p inhibition was associated with an increased sphere-forming efficiency and gefitinib resistance of EGFR-mutated cells through the unleashed expression of USP14
[94].
Other miRNAs were reported to regulate response to first-generation EGFR-TKIs through direct regulation of newly identified target genes including PELI3 by miR-365a-5p
[95]; ATG5 by miR-153-3p
[96]; TGFBR2 by miR-942-5p
[95] and LHX6 by miR-214
[97]. Lastly, exosomal miR-184 was recently proposed as a biomarker resistance to third-generation TKI such as Osimertinib, which was reported to alter AKT phosphorylation and Osimertinib-induced cell death in cooperation with miR-22-3p
[98][[99]
Further studies are urgently needed to better understand the role of miRNAs in the regulation of response to third-generation TKIs, with a particular focus on atypical
EGFR mutations that showed low response to EGFR-TKIs.
7. miRNAs Associated to Resistance to Radiotherapy
Radiotherapy is an important component of the multi-modality treatment for metastatic NSCLC, with ~50% of NSCLC patients undergoing radiotherapy either for a curative or palliative intent
[100]. Radiotherapy kills tumor cells either by directly inducing DNA damage, which leads to cell death as well as increases anti-tumor immune response
[101] or by indirectly altering tumor vasculature
[102]. As said before, a large fraction of miRNAs associated with radiotherapy was downregulated in radioresistant samples and vice versa upregulated in radiosensitive ones, for example:(i) miR-218-5p overexpression restored the sensitivity of radiation-resistant cells through direct regulation of PRKDC, a member of the non-homologous end-joining pathway in the DNA double-strand break repair response
[103]; (ii) rescue of miR-126-3p expression increased radiation-induced apoptosis by inactivation of the PI3K/AKT signaling
[104]. MiR-449a, a p53-responsive miRNA, was found to regulate LDHA expression, which in turn inhibited glycolysis and increased radiotherapy sensitivity
[105]; (iii) miR-375-3p and miR-513a-3p were respectively sponged by circ_0086720 and LINC00473, and their inhibition attenuated the knockdown-mediated radiosensitivity induced by these two ncRNAs
[106][[107]; (iv) re-expression of members of the let-7 family was functionally associated to increased radiosensitivity in NSCLC cells, although the molecular mechanisms were not fully investigated
[97]; (v) miR-21-5p was found overexpressed in radioresistant samples while its inhibition was able to increase the sensitivity to radiotherapy through several molecular mechanisms including direct regulation of PTEN and PDCD4 genes, inactivation of the PI3K/AKT signaling or through modulation of HIF1-alpha dependent metabolism regulation
[108][109][110][111][112]. Similarly, miR-25-3p was upregulated in radiation-resistant patients and was found to affect radiotherapy sensitivity by direct regulation of the newly identified target gene BTG2, a cell cycle modulator that acts as an effector of p53-induced cell cycle arrest
[113] .