Alkylating anticancer agents and their relations to microRNAs.

Introduction

Alkylating agents represent an important tool for the daily fight against cancer. Interestingly, their introduction into clinical application based on chemical warfare. Like a knife that can kill in the hands of an assassin and heal in the hands of a surgeon, the potential of nitrogen and sulfur mustards as anticancer agents was identified after incidents with poison gas (reduction of white blood cells) during World War II and dates back to observations after poison gas attacks during World War I [ 1 , 2 ] . Rational chemical modifications led to less lethal anticancer drug candidates such as melphalan, chlorambucil, and cyclophosphamide (all of them are nitrogen mustards). Alkylation of bionucleophiles (proteins, nucleic acids) mainly occurs at N , O , and S sites with free electron pairs. In most cases DNA was identified as the main target of alkylating agents and both mono-functional (reaction with only one DNA strand) and bifunctional alkylating agents (reaction with two strands leading to DNA crosslinks) are known [ 2 ] . Pertinent drug resistance strategies of cancers in order to cope with alkylating drugs include elevated glutathione levels, enhanced DNA repair and modified DNA damage signaling, as well as expression of multidrug resistance proteins such as membrane transporters [ 2 ] . The prodrug temozolomide is a highlight of research concerning alkylating agents because it can penetrate the blood-brain-barrier and is clinically applied for the therapy of glioblastoma [ 3 ] . Meanwhile, nature has also provided very potent alkylating agents with promising anticancer activities such as mitomycins, cyclopropyl-indoles, and illudins [ 4-6 ] . Mitomycin C is applied for the treatment of various solid tumors [ 4 ] . Trabectedin is the latest natural alkylating agent that has been approved for the therapy of soft tissue sarcoma and platinum-resistant ovarian cancer [ 7 ] .

MicroRNAs (miRNAs) are small RNA molecules of 22-23 nucleotides and regulate numerous genes involved in cell differentiation, cell proliferation, and cell death [ 8 ] . Mature miRNAs usually bind to the 3’-untranslated region of target messenger RNAs (mRNAs) and one miRNA is able to bind to various mRNAs [ 9-11 ] . MiRNAs have become valuable tools for the diagnosis and prognosis of cancer diseases due to abnormal expression profiles in cancers [ 12 , 13 ] . In addition, miRNAs regulate metastasis formation (epithelial-to-mesenchymal transition) and survival of cancer stem-like cells [ 14 , 15 ] . Concerning cancer research, tumor suppressor miRNAs and oncogenic miRNAs (oncomirs) are of particular interest and played crucial roles for the sensitivity and resistance of various tumors to applied drugs [ 16 ] . Prominent miRNAs with great relevance to cancer disease are represented by the tumor suppressor let-7 family and the oncomir miR-21 [ 17 , 18 ] .

This review intends to give an overview of clinically approved alkylating agents and their interactions with miRNAs in cancer diseases concerning drug activity and resistance.

Synthetic alkylating agents and their interactions with miRNAs

Synthetic alkylating agents are widely applied for the therapy of solid tumors and of leukemia/lymphoma diseases. Relevant synthetic alkylating agents that are dealt with in this review can be subdivided into the following compound classes: mustards (e.g., nitrogen mustards such as mechlorethamine, melphalan, chlorambucil, bendamustine, cyclophosphamide, estramustine), diazomethane forming prodrugs (e.g., dacarbazine, temozolomide), and N -nitrosoureas (e.g., BCNU/carmustine). The influence of miRNAs on the activity of these alkylating drugs is discussed below.

Nitrogen mustards, miRNAs and cancer

As mentioned above, anticancer active nitrogen mustards were developed since 1942 from highly toxic poison gas applied or produced in both World Wars of the 20th century. In order to reduce the systemic toxicity of nitrogen mustard poison gas and of initially applied mustard drugs like mechlorethamine, anilin derivatives such as chlorambucil and melphalan were designed with reduced activity due to their aromatic amine system [Figure 1] . Soon later, bendamustine and the prodrug cyclophosphamide were developed as potent anticancer drugs [Figure 1] [ 19 ] . In addition, the alkylating estrogen-conjugate estramustine was disclosed [Figure 1] [ 19 ] . Recent efforts in the field of nitrogen mustard-based anticancer research to increase selectivity and reduce side effects included DNA-targeting strategies, brain-targeting strategies, antibody-directed enzyme prodrug therapy and gene-directed enzyme prodrug therapy strategies, and nitrogen mustard prodrugs activated by glutathione transferase [ 19 ] . The alkylation mode of action of these 2-chloroethylamino derivatives includes an intramolecular reaction to an aziridium intermediate that readily reacts with bionucleophiles [ 19 ] . Aside cytotoxic effects on non-malignant cells, genotoxic mutagenic effects were identified for nitrogen mustards as well.

Chlorambucil has been the standard treatment for chronic lymphocytic leukemia (CLL) for decades and the drug is still recommended as a mainstay for the currently widely applied antibody-based chemoimmunotherapy for CLL [ 20 ] . Chemoresistance of CLL is often mediated by the p53-signaling pathway which plays a crucial role for the cellular response to DNA damage [ 21 ] . The tumor suppressor microRNA miR-34a is connected with p53-signaling and low expression of miR-34a led to chemoresistance in CLL [ 22 ] . A considerable number of PCLBCL-LT (primary cutaneous diffuse large B-cell lymphomas, leg-type) patients did not respond to chlorambucil first-line therapy and upregulated expression of the oncogenic miR-17-92 cluster seems responsible leading to suppression of PTEN (phosphatase and TENsin homolog) [ 23 ] . A miR-210 targeting/inhibiting chlorambucil conjugate was synthesized, which suppressed a cancer relevant hypoxic mechanism and was active against hypoxic triple-negative breast cancer in mice [ 24 ] . A list of miRNAs associated with chlorambucil activity is provided in Table 1 .

Melphalan is a close analog of chlorambucil and has been applied for the treatment of multiple myeloma (MM) since the 1960’s [ 25 ] . From MM patients treated with melphalan circulating exosomal let-7b and miR-18a were identified as prognostic factors (i.e., improved survival) [ 26 ] . Suppression of the oncogenes Myc, Ras and CCND1 (cyclin D1) by let-7b may play a role as well as inhibition of the HIF1α-pathway [hypoxia-inducible factor (HIF)] by miR-18a [ 26 ] . IL-6-mediated downregulation of the tumor suppressor miR-15a/16 expression increased resistance of myeloma cells to melphalan treatment [ 27 ] . The oncogenic miR-221-222 family suppressed p53 upregulated modulator of apoptosis (PUMA) in MM cells leading to drug resistance [ 28 ] . Inhibition of miR-221 by a 13 mer LNA-i-miR-221 inhibitor broke melphalan resistance in MM by modulation of PUMA (upregulation) and ATP binding cassette C1 (ABCC1) transporter (downregulation) [ 29 ] . In addition, it was suggested that suppression of miR-451 can enhance the activity of melphalan in multiple myeloma via downregulation of multidrug resistance gene 1 (MDR1) [ 30 ] . A list of miRNAs involved in melphalan activity in myeloma is given in Table 2 .

Cyclophosphamide is one of the most successful anticancer drugs which is still widely applied for the therapy of many cancer diseases 60 years now after its development in the late 1950’s [ 31 ] . The drug is applied for the treatment of breast cancer, childhood cancers, and lymphoma [ 31 ] . Cyclophosphamide is a prodrug, which can be activated in a chemical or metabolic way. Enzymatic oxidation of cyclophosphamide generates cytotoxic phosphoramide mustard molecules leading to interstrand and intrastrand crosslinks of DNA as well as toxic acrolein which is responsible for the side effects of cyclophosphamide [ 32 ] .

Clinical sensitivity to cyclophosphamide is strongly correlated with the ability of cancer cells to induce apoptosis upon DNA damage [ 33 ] . Due to its significance in cancer therapy the relations between miRNAs and cyclophosphamide is well studied, in particular, in lymphoma [ 34 , 35 ] . Upregulation of circulating oncogenic miR-125b and miR-130a was determined in B-cell lymphoma samples from patients treated with cyclophosphamide-based chemotherapy [ 36 ] . In diffuse large B-cell lymphoma (DLBCL, the most common non-Hodgkin lymphoma type) treated with R-CHOP (rituximab, cyclophosphamide, adriamycin, vincristine, and prednisone), increased miR-181a expression prolonged progression free survival (PFS) by suppression of FOXP1 and O 6-methylguanine DNA methyltransferase (MGMT) while increased miR-18a levels led to shorter overall survival (OS) and high miR-222 expression to shorter PFS [ 37 ] . MiR-93 (targets: p21, BCL2L11), miR-221 and miR-222 (target: p27) were upregulated in DLBCL patients with poor outcomes after cyclophosphamide-based therapies [ 38 ] . Knockdown of the well-known oncomir miR-21 sensitized DLBCL cells to CHOP treatment by upregulation of PTEN [ 39 ] . Shorter OS was observed from DLBCL patients with high miR-21 expression in the tumor tissue, while high miR-21 levels in the serum promoted relapse free survival [ 40 ] . Downregulated miR-199a/b also shortened progression free survival time [ 40 ] . In addition, CHOP or R-CHOP treated DLBCL patients with upregulated miR-200c expression displayed shorter OS than patients with suppressed miR-200c [ 41 ] . Induced miR-155 expression was connected with R-CHOP resistance in DLBCL patients although miR-155 sensitized patients to AKT (protein kinase B, ak thymoma) signaling probably by targeting p85α and SHIP1 [ 42 ] . In contrast to that, upregulated expression of miR-129-5p in DLBCL patients treated with CHOP or R-CHOP led to much longer median survival than in patients with downregulated miR-129-5p [ 43 ] . The tumor suppressor miR-146a was downregulated in R-CHOP-treated DBCL patients who displayed drug resistance [ 44 ] . Suppressed expression of the tumor suppressors miR-146b-5p and miR-320d led to shorter progression free survival in CHOP-treated DLBCL patients [ 45 ] . Higher serum levels of miR-33a and miR-455-3p indicated better response while high levels of miR-224 (target: CD59), miR-520d-3p and miR-1236 were connected with worse R-CHOP response in DLBCL patients [ 35 , 46 ] . Suppressed miR-224 expression in the tumor tissue also indicated poor survival [ 39 ] . In addition, a higher R-CHOP response rate and longer survival was observed from DLBCL patients with 7q gain, which was attributed to the upregulated expression of miR-25, miR-96, miR-182, and miR-589 [ 47 ] . Lists of miRNAs involved in cyclophosphamide activity in lymphoma are given in Tables 3 and 4 .

MicroRNAs also seem to play a role for drug resistance in breast cancer patients receiving cyclophosphamide as part of the TFAC therapy (paclitaxel, 5-fluorouracil, adriamycin, cyclophosphamide) [ 48 ] . The tumor suppressor miR-9 bound to the mRNA of human epidermal growth factor 2 (HER2) and increased the response to cyclophosphamide [ 49 ] . Low expression of let-7, miR-10b, miR-34, miR-155, miR-200c, miR-205, miR-451, and miR-3200 as well as high expression of miR-21, miR-195, and miR-221 were observed after cyclophosphamide treatment [ 50 , 51 ] . In particular, expression of the tumor suppressor miR-205 sensitized breast cancer cells to TAC treatment (docetaxel, doxorubicin, cyclophosphamide) by suppression of vascular endothelial growth factor A and fibroblast growth factor 2 [ 52 ] . In contrast to that, miR-663 overexpression was associated with resistance to cyclophosphamide in MDA-MB-231/ADM breast cancer cells (cells resistant to adriamycin) by suppression of heparin sulfate proteoglycan 2 [ 53 ] . The MDA-MB-231 cell line is a widely applied model for triple-negative breast cancer (TNBC, i.e., no or reduced expression of estrogen receptor, HER2/neu and progesterone receptor), which is a very problematic form of breast cancer showing chemotherapy resistance and poor prognosis. Samples from cyclophosphamide-treated TNBC patients with good response to chemotherapy exhibited higher miR-200b-3p (possible targets: PLCB1, MYCN, CCND2, RERG) and miR-190a (possible targets: BCL11A, CALCR, FOXP2, HOXC5) as well as lower miR-512-5p (possible targets: BCL2L2, POLD3, c-Myc) expression when compared with TNBC patients displaying weak chemotherapy response [ 54 ] . The microRNAs miR-30a, miR-9-3p, miR-770 and miR-143-5p were also identified as markers for chemotherapy response by TNBC patients. Chemotherapy-responding TNBC patients revealed upregulated miR-30a (affected transcriptional regulation in cancer) and miR-9-3p (affected mTOR/mammalian target of rapamycin and TGF/transforming growth factor-β signaling) as well as suppressed miR-770-5p (affected B and T cell receptor signaling) and miR-143-5p (affected B and T cell receptor signaling as well as mTOR signaling) [ 55 ] . A list of miRNAs involved in cyclophosphamide activity in breast cancer is given in Table 5 .

The relations between miRNAs and cyclophosphamide were also investigated in further cancer diseases. VAC treatment (vindesine, doxorubicin, cyclophosphamide) is applied as a second-line treatment of small cell lung cancer. The structurally simple HDAC inhibitor valproic acid increased the activity of VAC treatment both in vitro and in vivo via induction of miR-589 and suppression of miR-575 expression [ 56 ] . In addition, overexpression of the oncomirs miR-27b and miR-298 led to cyclophosphamide resistance in Panc-1 pancreatic cancer cells by inhibition of vitamin D receptor and cytochrome P3A4 (CYP3A4) [ 57 ] . It became evident that CYP3A4 plays a crucial role for the activation of cyclophosphamide in these cancer cells. A list of miRNAs involved in cyclophosphamide activity in various cancers is given in Table 6 .

Estramustine phosphate is applied for the treatment of advanced prostate cancer and combines the DNA-damaging N -mustard scaffold with a tubulin polymerization inhibiting steroid (estradiol-17β-phosphate) released upon metabolization of the drug [ 58 ] . In prostate cancer cells, estramustine phosphate induced apoptosis via suppression of the oncomir miR-31 [ 59 ] . In addition, downregulation of the tumor suppressor miR-4319, which suppresses HER2 expression, was associated with poor chemotherapy response in prostate cancer patients and induced miR-4319 expression sensitized prostate cancer cells to estramustine treatment [ 60 ] . A list of microRNAs involved in estramustine phosphate anticancer activity is given in Table 7 .

Dacarbazine and temozolomide, miRNAs and cancer

Dacarbazine and temozolomide [Figure 2] are valuable anticancer drugs for the treatment of melanoma and of brain tumors such as glioblastomas (GBMs) [ 61 , 62 ] . Their mode of action implies the generation of highly toxic DNA-alkylating diazomethane molecules that kill the cancer cells [ 63 ] .

Growth inhibition of melanoma cells by dacarbazine was mediated by induction of the expression of miR-200 family miRNAs (miR-141, miR-200a/b/c) which regulate apoptosis [ 64 ] . Dacarbazine is also widely applied for the treatment of lymphomas as part of the ABVD therapy (adriamycin, bleomycin, vinblastine, dacarbazine). Complete response to ABVD therapy was achieved by classical Hodgkin lymphoma patients with reduced miR-494 and miR-1973 plasma levels [ 65 ] . A list of miRNAs involved in dacarbazine activity in various cancers is given in Table 8 .

The development of temozolomide as an anticancer drug started in the 1980’s and is a nice example of the productive interplay between chemists, pharmacists and clinicians during the development process [ 63 ] . Temozolomide is a prodrug and decomposes in aqueous solution. Initially CO 2 is released from the molecule whereupon diazomethane is formed that reacts with bionucleophiles [ 63 ] . The interplay between temozolomide and miRNAs in cancer was thoroughly studied over the last years. Initial studies revealed that temozolomide-induced apoptosis was inhibited by upregulated miR-21 accompanied by decreased Bax/Bcl-2 ratio and caspase-3 activity in GBM cells [ 66 , 67 ] . Treatment of GBM cells with temozolomide led to increased miR-21 expression and GBM cells were sensitized to temozolomide by downregulation of miR-21 and miR-17 [ 68-70 ] . In particular, antisense miR-21 formulated with PLGA [poly(lactic-co-glycolic acid)] nanoparticle carriers increased the anticancer activity of temozolomide against GBM cells [ 71 ] . Temozolomide-resistant GBM cells also exhibited increased miR-10a*, miR-195 (also in cancer stem cells of GMB tumors via mothers against decapentaplegic homolog 2 targeting), and miR-455-3p expression [ 72 , 73 ] . However, in melanoma cells, miR-195 functioned as a tumor suppressor by suppression of prohibitin 1 [ 74 ] . In addition, chemoresistance to temozolomide was associated with upregulation of miR-9 (targeting/patched homolog 1) in CD133-positive GBM cells and of Bcl-2-interacting mediator of cell death - targeting miR-138 [ 75-77 ] . Downregulation of the tumor suppressor miR-16 increased Bcl-2 activity and led to temozolomide resistance in glioma cells [ 78 ] . Suppressed miR-29c expression associated with upregulated reversionless 3-like (REV3L) led to temozolomide resistance in glioma cells [ 79 ] . In addition, miR-29c enhanced temozolomide activity by indirect suppression of MGMT [ 80 ] . MiR-30a expression also enhanced temozolomide activity against U251 GBM cells by inhibition of autophagy and suppression of beclin 1 [ 81 ] . Upregulation of the tumor suppressor miR-136 led to higher temozolomide activity in glioma cells by suppression of astrocyte elevated gene 1 (AEG-1) [ 82 ] . Similarly, induced miR-143 expression increased temozolomide activity against GBM cells via neuroblastoma rat sarcoma oncogene targeting [ 83 ] . The oncomir miR-93 led to temozolomide resistance of glioma cells by upregulation of p21 [ 84 ] . MiR-125b-2 induced temozolomide resistance in glioblastoma stem cells was associated with downregulation of Bax and upregulation of Bcl-2 [ 85 ] . Downregulation of the oncomirs miR-221/miR-222 refurnished p53 signaling pathway and promoted apoptosis of GBM cells treated with temozolomide [ 86 ] . MiR-141-3p is another p53-targeting oncomir leading to temozolomide resistance in glioma cells [ 87 ] . MAP kinase/extracellular-signal regulated kinase (ERK) signaling was induced by the oncomir miR-299-5p, which targeted golgi phosphoprotein 3 and, thus, led to temozolomide resistance in GBM cells [ 88 ] . Temozolomide resistance in GBM basing on active GSK3β was overcome by upregulated miR-101 [ 89 ] . Further to this, miR-128 mediated temozolomide-induced cell death in glioma cells via inhibition of mTOR signaling and suppression of insulin-like growth factor 1, phosphoinositide-3-kinase regulatory subunit 1, rapamycin-insensitive companion of mTOR and mTOR [ 90 ] . High expression of miR-130a sensitized glioma cells to temozolomide via apurinic/apyrimidinic endonuclease 1 suppression and was upregulated in GBM patients with better response to temozolomide [ 91 ] . A direct influence on temozolomide resistance was elucidated for miR-181d and miR-603, which suppressed the DNA repair enzyme MGMT leading to greater sensitivity to temozolomide [ 92 ] . In addition, miR-142-3p promoted temozolomide activity in GBM cells by suppression of MGMT [ 93 ] . MiR-648 and miR-767-3p were identified as further MGMT targeting/suppressing miRNAs, which enhanced the anticancer activity of temozolomide in T98G GBM cells [ 94 ] . MiR-182 expression also increased temozolomide activity in GBM cells by apoptosis promotion via c-Met, HIF2A and BCL2L12 suppression [ 95 ] . However, expression of miR-132 caused temozolomide resistance in GBM cells (U78MG) via downregulation of tumor suppressor candidate 3 [ 96 ] . Temozolomide-resistant GBM cells and tissues exhibited suppressed miR-370-3p expression levels, which is a tumor suppressor responsible for downregulated MGMT expression and blocked DNA repair [ 97 ] . In contrast to that, expression of the oncomir miR-423-5p led to temozolomide resistance in glioma cells by targeting ING-4 [ 98 ] . Glioblastoma samples from patients treated with temozolomide revealed upregulated miR-629-3p expression (targeting genes involved in translation and RNA processing) in case of good responders with prolonged OS [ 99 ] . The tumor suppressor miR-1268 also sensitized glioma cells (T98G) to temozolomide treatment [ 100 ] . In addition, miR-1294 suppressed targeting protein for Xenopus kinesin-like protein 2 in glioma cells leading to enhanced temozolomide activity [ 101 ] . Some miRNAs are special “Janus-type” cases here. Although miR-181b/c are reported as tumor suppressors in GBM, reduced expression of these miRNAs was associated with better temozolomide response [ 102 ] . And although miR-221 and miR-222 suppress MGMT, their upregulation led to weaker responses to temozolomide [ 103 ] . In a cancer type dependent way, miR-195 acted either as a tumor suppressor (melanoma) or as an oncomir (GBM, see above). Last but not least, the combination of temozolomide with miRNA-regulating drugs appears promising. For instance, curcumin (a polyphenol isolated from turmeric, Curcuma longa ) was able to sensitize GBM cells (C6) to temozolomide via suppression of miR-10b [ 104 ] . Lists of miRNAs involved in temozolomide anticancer activity are given in Tables 9 and 10 .

N -nitrosoureas, miRNAs and cancer

Anticancer active N -nitrosoureas were developed in the course of a screening program initiated by the National Cancer Institute. Starting from the hit compound 1-methyl-3-nitro-1-nitrosoguanidine further analogs were prepared leading to 1-methyl-1-nitrosourea which was active against intracerebrally implanted murine leukemia [ 105 ] . Further fine-tuning of this compound finally led to carmustine/BCNU (bis-chloroethylnitrosourea, Figure 3 ) which entered clinical trials in 1964 and was approved by the FDA in 1977 for the treatment of brain tumors (BCNU passes the blood-brain-barrier because of its lipophilicity), lymphomas and myeloma [ 106 ] . A newer study recommends the application of BCNU for the treatment of recurrent GBM [ 107 ] . BCNU is a prodrug, which decomposes to afford alkylating chloroethyl moieties that can form DNA interstrand crosslinks [ 108 ] . Carbamoylation of nucleoprotein lysine residues via isocyanate intermediates can also play a role for the anticancer mode of action of BCNU [ 109 ] .

Expression analysis of BCNU-treated glioma cells led to dysregulation of let-7b (tumor suppressor), miR-125b-2 (oncomir), miR-133a-1 (tumor suppressor/oncomir), and miR-183 (oncomir) [ 110 ] . It was also shown that miR-21 expression induced BCNU-resistance in glioma cells via downregulation of Spry2 (sprout homolog 2) [ 111 ] . Although miR-181d was identified as a tumor suppressor and temozolomide-sensitizing factor (see above), GBM patients with implanted BCNU wafers displayed prolonged overall and progression-free survival in case of suppressed miR-181d expression [ 112 ] . In addition, high expression of the oncomir miR-221 suppressed PTEN and led to PI3K/Akt activation and resistance to BCNU [ 113 ] . A list of miRNAs involved in BCNU anticancer activity is given in Table 11 .

Natural alkylating agents and their interactions with miRNAs

Natural alkylating agents were investigated for anticancer activity since the 1950’s [ 114 , 115 ] . Meanwhile, natural alkylating agents are widely applied for the therapy of various cancer diseases. The alkaloids mitomycin C and trabectedin were approved for the therapy of cancer [Figure 4] . The influence of miRNAs on the activity of these natural alkylating drugs is discussed below.

Mitomycins, miRNAs and cancer

Mitomycins are bacterial indole alkaloids. The first mitomycins A and B were isolated in 1956 before mitomycin C was obtained as blue-violet crystals from Streptomyces caespitosus by Japanese groups in 1958 [ 116 , 117 ] . Mitomycin C turned out to be the most anticancer active derivative of this group of mitomycins and entered clinical trials in Japan shortly after its discovery [ 118 ] . Currently, it is applied for the treatment of localized bladder cancer, anal cancer, head-and-neck cancer, and breast cancer (palliative 2nd or 3rd line treatment) [ 119 ] . Mitomycin C is a prodrug and activated mitomycin C forms DNA crosslinks that are highly lethal to cancer cells [ 120 ] . Activation of mitomycin C occurs via reduction of the benzoquinone scaffold to a hydroquinone (a reaction catalyzed by enzymes such as DT-diaphorase). Subsequent elimination of methanol forms a reactive aziridine system, which reacts with DNA via aziridine ring opening. Elimination of carbamate enables a second alkylating step leading to DNA crosslinking [ 120 ] .

Transfection of HT-29 colon cancer cells with let-7-1 suppressed COP9 signalosome expression and increased mitomycin C activity [ 121 ] . Expression of the tumor suppressor miR-31 enhanced the tumor growth inhibition of urothelial bladder cancer by mitomycin C both in vitro and in vivo [ 122 ] . MiR-31 inhibited integrin α5 directly and suppressed Akt and ERK signaling [ 122 ] . In addition, miR-34a expression sensitized medulloblastoma to mitomycin C treatment by downregulation of melanoma antigen A and upregulation of p53 [ 123 ] . In contrast to that, inhibition of miR-191-5p expression led to higher mitomycin C activity against breast cancer cells via induction of apoptosis, SRY-box 4, and p53 [ 124 ] . A slight increase in mitomycin C resistance was observed for Snail-expressing mesenchymal MCF-7 breast cancer cells, which displayed suppressed miR-200 family expression [ 125 ] . Mitomycin C was shown to induce senescence in human mesenchymal stem cells via upregulation of the tumor suppressor aminoacyl-tRNA synthetase-interacting multifunctional protein 3/p18, and suppression of AIMP3/p18 was observed for miR-543 and miR-590-3p [ 126 ] . This mechanism may also play a role for the anticancer activity of mitomycin C. Oxaliplatin-resistant HCT-116/l-OHP colon cancer cells transfected with miR-1915 mimics revealed enhanced mitomycin C sensitivity by suppression of Bcl-2 [ 127 ] . A list of miRNAs involved in mitomycin C anticancer activity is given in Table 12 .

Since mitomycin C is a DNA-damaging drug, its long-term genotoxic effects and the inheritable aberration of miRNA expression induced by mitomycin C were investigated. Indeed, the treatment of HeLa cells with mitomycin C exhibited upregulated inherited expression of oncomirs such as miR-19b-3p, miR-21-3p, miR-30a-3p, miR-30e-3p, and miR-182-5p as well as suppressed inherited expression of the tumor suppressors miR-23b-3p, miR-29b-3p, miR-99a-5p, miR-99b-5p, miR-100-5p, miR-148a-3p, miR-193a-3p,,iR-340-5p, and miR-365-3p [ 128 ] . It is possible that new tumors can form basing on the long-term effects of mitomycin C and the observed aberrant miRNA expression.

Trabectedin, miRNAs and cancer

Trabectedin (ecteinascidin 743, Yondelis ® ) is a rather new natural alkylating agent that belongs to the class of tetrahydroisoquinoline alkaloids. It was isolated from the Caribbean tunicate Ecteinascidia turbinata , which is only the host of the trabectedin-producing symbiont Endoecteinascidia frumentensis [ 129 ] . The high anticancer activity of trabectedin led to its clinical approval for the treatment of soft tissue sarcoma [ 129 ] . The unique DNA-damaging mechanism of trabectedin includes binding to nitrogen-N2 of guanine DNA bases in the minor groove. This DNA-trabectedin adduct interacts with DNA repair proteins of the transcription-coupled nucleotide excision repair DNA-repair system which causes cell death via the formation of double-strand breaks mainly in homologous recombination-deficient cells [ 129 ] . This mechanism is almost unique among known alkylating agents and only illudins have revealed a similar mode of action [ 130 ] . Trabectedin also blocked the transcriptional activity of fused in sarcoma-C/EBP-homologous protein (FUS-CHOP) [ 131 ] . The difference in miRNA expression between trabectedin-sensitive and trabectedin-resistant myxoid liposarcoma cells (402-91 sensitive and 402-91/ET trabectedin-resistant cells) was investigated and the resistant cells showed two-fold higher miR-21 expression (target: PDCD4/programmed cell death 4) as well as three-fold lower let-7e expression (targets: CCND1, E2F5, SEMA4C) [ 132 ] . The oncomir miR-7 was also upregulated while the tumor suppressors miR-98, miR-130a and miR-192 were suppressed in the resistant 402-91/ET cells [ 132 ] . The miRNAs miR-7, miR-21 and miR-130a probably act via FUS-CHOP since these miRNAs have CHOP-binding motifs [ 132 ] . In cholangiocarcinoma, trabectedin treatment led to upregulation of the oncomir miR-494-3p [ 133 ] . In addition, the tumor suppressors let-7c and miR-214-3p were suppressed [ 133 ] . Interestingly, trabectedin downregulated the oncomirs miR-21-3p, miR-21-5p, and miR-331-3p (oncomir in HCC), and upregulated the tumor suppressors miR-375 (tumor suppressor in colon and pancreatic cancer) and miR-4284 (tumor suppressor in glioblastoma), which may be a reason for the relatively high activity of trabectedin in this cancer model [ 133 ] . A list of miRNAs involved in trabectedin anticancer activity is given in Table 13 .

Conclusion

Alkylating drugs still play a crucial role for the therapy of various cancer diseases. While some examples are only applied for the treatment of special tumors (e.g., estramustine for the treatment of prostate cancer), other drugs (e.g., cyclophosphamide) are widely applied. The anticancer activity of these alkylating agents is regulated by various cellular factors. Aside proteins, small RNA molecules called miRNAs revealed a crucial role for the outcome of therapies based on alkylating drugs. Vice versa, alkylating drugs can also regulate miRNA expression leading to enhanced sensitivity of the affected cancer. Thus, a detailed understanding of the interplay between alkylating drugs and miRNAs is crucial for the development of new and improved cancer therapies. In particular, combination therapies with alkylating agents should be carefully checked in view of the corresponding miRNAs involved in alkylating drug response and resistance, and the co-drugs for combination therapies with these alkylating agents should be selected accordingly. Prolonged survival and improved quality of life would be possible and conceivable prospects for many cancer patients.

Declarations

Authors’ contributions

The author contributed solely to the article.

Availability of data and materials

Not applicable.

Financial support and sponsorship

None.

Conflicts of interest

The author declared that there are no conflicts of interest.

Ethical approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Copyright

© The Author(s) 2019.

1 Smith SL War! What is it good for? Mustard gas medicine. CMAJ 2017 189 E321 2 10.1503/cmaj.161032 28246228 PMC5325736

2 Ralhan R Kaur J Alkylating agents and cancer therapy. Expert Opin Ther Patents 2007 17 1061 75 10.1517/13543776.17.9.1061

3 Baker SD Wirth M Statkevich P Reidenberg P Alton K Absorption, metabolism, and excretion of 14C-temozolomide following oral administration to patients with advanced cancer. Clin Cancer Res 1999 5 309 17 10037179

4 Bradner WT Mitomycin C: a clinical update. Cancer Treat Rev 2001 27 35 50 10.1053/ctrv.2000.0202 11237776

5 Ghosh N Sheldrake HM Searcey M Pors K Chemical and biological explorations of the family of CC-1065 and the duocarmycin natural products. Curr Topics Med Chem 2009 9 1494 524 10.2174/156802609789909812 19903166

6 Schobert R Knauer S Seibt S Biersack B Anticancer active illudins: recent developments of a potent alkylating compound class. Curr Med Chem 2011 18 790 807 10.2174/092986711794927766 21182482

7 Carter NJ Keam SJ Trabectedin: a review of its use in soft tissue sarcoma and ovarian cancer. Drugs 2010 70 355 76 10.2165/11202860-000000000-00000 20166769

8 Guarnieri DJ DiLeone RJ MicroRNAs: a new class of gene regulators. Ann Med 2008 40 197 208 10.1080/07853890701771823 18382885

9 Bartels DB MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 2004 116 281 98 10.1016/S0092-8674(04)00045-5 14744438

10 Wu S Huang J Ding Y Zhao L Liang L Multiple microRNAs modulate p21Cip1/Waf1 expression by directly targeting its 3’ untranslated region. Oncogene 2010 29 2302 8 10.1038/onc.2010.34 20190813

11 Calin GA Croce CM MicroRNA signatures in human cancers. Nat Rev Cancer 2006 6 857 66 10.1038/nrc1997 17060945

12 Mallick R Patnaik SK Yendumuri S Micro RNAs and lung cancer: biology and prognosis in diagnosis and prognosis. J Carcinog 2010 9 8 10.4103/1477-3163.67074 20808843 PMC2925280

13 Rothe F Ignatiadis M Chaboteaux C Haibe-Kains B Kheddoumi N Global microRNA expression profiling identifies miR-210 associated with tumor proliferation, invasion and poor clinical outcome in breast cancer. PLoS One 2011 6 e20980 10.1371/journal.pone.0020980 21738599 PMC3126805

14 Shimono Y Zabala M Cho RW Lobo N Dalerba P Downregulation of miRNA-200c links breast cancer stem cells with normal cells. Cell 2009 138 592 603 10.1016/j.cell.2009.07.011 19665978 PMC2731699

15 Gregory PA Bracken CP Bert AG Godall GJ MicroRNAs as regulators of epithelial-mesenchymal transition. Cell Cycle 2008 7 3112 8 10.4161/cc.7.20.6851 18927505

16 Biersack B Interactions between anticancer active platinum complexes and non-coding RNAs/microRNAs. Non-coding RNA Res 2017 2 1 17 10.1016/j.ncrna.2016.10.001 30159416 PMC6096430

17 Su JL Chen PS Johansson G Kuo ML Function and regulation of let-7 microRNAs. MicroRNA 2012 1 34 9 25048088 10.2174/2211536611201010034

18 Feng YH Tsao CJ Emerging role of microRNA-21 in cancer. Biomed Rep 2016 5 395 402 10.3892/br.2016.747 27699004 PMC5038362

19 Singh RK Kumar S Prasad DN Bhardwai TR Therapeutic journey of nitrogen mustards alkylating anticancer agents: historic to future perspectives. Eur J Med Chem 2018 151 401 33 10.1016/j.ejmech.2018.04.001 29649739

20 Goede V Eichhorst B Fischer K Wendtner CM Hallek M Past, present and future role of chlorambucil in the treatment of chronic lymphocytic leukemia. Leuk Lymphoma 2015 56 1585 92 10.3109/10428194.2014.963077 25219593

21 Zenz T Mohr J Edelmann J Sarno A Hoth P Treatment resistance in chronic lymphocytic leukemia: the role of the p53 pathway. Leuk Lymphoma 2009 50 510 3 10.1080/10428190902763533 19347737

22 Zenz T Mohr J Eldering E Kater AP Bühler A MiR-34a as part of the resistance network in chronic lymphocytic leukemia. Blood 2009 113 3801 8 10.1182/blood-2008-08-172254 18941118

23 Battistella M Romero M Castro-Vega LJ Gapihan G Bouhidel F The high expression of the microRNA 17-92 cluster and its paralogs, and the downregulation of the target gene PTEN, is associated with primary cutaneous B-cell lymphoma progression. J Invest Dermatol 2015 135 1659 67 10.1038/jid.2015.27 25634356

24 Costales MG Haga CL Velagapudi SP Childs-Disney JL Phinney DG Small molecule inhibition of microRNA-210 reprograms an oncogenic hypoxic circuit. J Am Chem Soc 2017 139 3446 55 10.1021/jacs.6b11273 28240549 PMC5810126

25 Zweegman S Huijgens PC Treatment of myeloma: recent developments. Anticancer Drugs 2002 13 339 51 11984079 10.1097/00001813-200204000-00002

26 Manier S Liu CJ Avet-Loiseau H Park J Shi J Prognostic role of circulating exosomal miRNAs in multiple myeloma. Blood 2017 129 2429 36 10.1182/blood-2016-09-742296 28213378 PMC5409448

27 Hao M Zhang L An G Sui W Yu Z Suppressing miRNA-15a/16 expression by interleukin-6 enhances drug-resistance in myeloma cells. J Hematol Oncol 2011 4 37 10.1186/1756-8722-4-37 21936961 PMC3189173

28 Zhao JJ Chu ZB Hu Y Lin J Wang J Targeting the miR-221-222/PUMA/BAK/BAX pathway abrogates dexamethasone resistance in multiple myeloma. Cancer Res 2015 75 4384 97 10.1158/0008-5472.CAN-15-0457 26249174 PMC4609291

29 Gullá A Di Martino MT Cantafio MEG Morelli E Amodio N A 13 mer LNA-i-miR-221 inhibitor restores drug sensitivity in melphalan-refractory multiple myeloma cells. Clin Cancer Res 2015 22 1222 33 10.1158/1078-0432.CCR-15-0489 26527748 PMC4775414

30 Du J Liu S He J Liu X Qu Y MicroRNA-451 regulates stemness of side population cells via PI3K/Akt/mTOR signaling pathway in multiple myeloma. Oncotarget 2015 6 14993 5007 10.18632/oncotarget.3802 25915427 PMC4558131

31 Emadi A Jones RJ Brodsky RA Cyclophosphamide and cancer: golden anniversary. Nat Rev Clin Oncol 2009 6 638 47 10.1038/nrclinonc.2009.146 19786984

32 Boddy AV Yule SM Metabolism and pharmacokinetics of oxazaphosphorines. Clin Pharmacokinet 2000 39 291 304 10.2165/00003088-200038040-00001 10803453

33 Zhang J Tian Q Chan SY Duan W Zhou S Insights into oxaphosphorine resistance and possible approaches to its circumvention. Drug Resist Updat 2005 8 271 97 10.1016/j.drup.2005.08.003 16154799

34 Cho WCS Great potential of microRNAs as predictive and prognostic markers for cancer. Expert Rev Mol Diagn 2012 12 315 8 10.1586/erm.12.21 22616694

35 Yu X Li Z New insights into microRNAs involves in drug resistance in diffuse large B cell lymphoma. Am J Transl Res 2015 7 2536 42 26885255 PMC4731655

36 Yuan WX Gui YX Na WN Chao J Yang X Circulating microRNA-125b and microRNA-130a expression profiles predict chemoresistance to R-CHOP in diffuse large B-cell lymphoma patients. Oncol Lett 2016 11 423 32 10.3892/ol.2015.3866 26870228 PMC4726930

37 Alencar AJ Malumbres R Kozloski GA Advani R Talreja N MicroRNAs are independent predictors of outcome in diffuse large B-cell lymphoma patients treated with R-CHOP. Clin Cancer Res 2011 17 4125 35 10.1158/1078-0432.CCR-11-0224 21525173 PMC3117929

38 Montes-Moreno S Martinez N Sanchez-Espiridión B Uriarte RD Rodriguez ME MiRNA expression in diffuse large B-cell lymphoma treated with chemoimmunotherapy. Blood 2011 118 1034 40 10.1182/blood-2010-11-321554 21633089

39 Bai H Wei J Deng C Yang X Wang C MicroRNA-21 regulates the sensitivity of diffuse large B-cell lymphoma cells to the CHOP chemotherapy regimen. Int J Hematol 2013 97 223 31 10.1007/s12185-012-1256-x 23275230

40 Zheng Z Li X Zhu Y Gu W Xie X Prognostic significance of miRNA in patients with diffuse large B-cell lymphoma: a meta-analysis. Cell Physiol Biochem 2016 39 1891 904 10.1159/000447887 27771712

41 Berglund M Hedstrom G Amini RM Enblad G Thunberg U High expression of microRNA-200c predicts poor clinical outcome in diffuse large B-cell lymphoma. Oncol Rep 2013 29 720 4 10.3892/or.2012.2173 23232598

42 Iqbal J Shen Y Huang X Liu Y Wake L Global microRNA expression profiling uncovers molecular markers for classification and prognosis in aggressive B-cell lymphoma. Blood 2015 125 1137 45 10.1182/blood-2014-04-566778 25498913 PMC4326773

43 Hedstrom G Thunberg U Berglund M Simonsson M Amini RM Low expression of microRNA-129-5p predicts poor clinical outcome in diffuse large B cell lymphoma (DLBCL). Int J Hematol 2013 97 465 71 10.1007/s12185-013-1303-2 23463124

44 Huang H Gu J Yao S Yao Z Zhao Y MicroRNAs are related to rituximab with cyclophosphamide, doxorubicin, vincristine, and prednisone resistance in patients with diffuse large B-cell lymphoma. Cancer Transl Med 2015 1 6 10 10.4103/2395-3977.151470

45 Wu PY Zhang XD Zhu J Guo XY Wang JF Low expression of microRNA-146b-5p and microRNA-320d predicts poor outcome of large B-cell lymphoma treated with cyclophosphamide, doxorubicin, vincristine, and prednisone. Hum Pathol 2014 45 1664 73 10.1016/j.humpath.2014.04.002 24931464

46 Song G Gu L Li J Tang Z Liu H Serum microRNA expression profiling predict response to R-CHOP treatment in diffuse large B cell lymphoma patients. Ann Hematol 2014 93 1735 43 10.1007/s00277-014-2111-3 24858372

47 Chigrinova E Mian M Shen Y Greiner TC Chan WC Integrated profiling of diffuse large B-cell lymphoma with 7q gain. Br J Haematol 2011 153 499 503 10.1111/j.1365-2141.2011.08628.x 21418177

48 Salter KH Acharya CR Walters KS Redman R Anguiano A An integrated approach t the prediction of chemotherapeutic response in patients with breast cancer. PLoS One 2008 3 e1908 10.1371/journal.pone.0001908 18382681 PMC2270912

49 Sun G Sun L Liu Y Xing H Wang K Her-2 expression regulated by downregulation of miR-9 and which affects chemotherapeutic effect in breast cancer. Cancer Gene Ther 2017 24 194 202 10.1038/cgt.2014.82 28361910

50 Gezer U Keskin S Igci A Tükenmez M Tiryakioglu D Abundant circulating microRNAs in breast cancer patients fluctuate considerably during neoadjuvant chemotherapy. Oncol Lett 2014 8 845 8 10.3892/ol.2014.2188 25009660 PMC4081437

51 Al-Khanbashi M Caramuta S Alajmi AM Al-Haddabi I Al-Riyami M Tissue and serum miRNA profile in locally advanced breast cancer (LABC) in response to neo-adjuvant chemotherapy (NAC) treatment. PLoS One 2016 11 e152032 10.1371/journal.pone.0152032 27064979 PMC4827834

52 Hu Y Qiu Y Yagüe E Ji W Liu J MiRNA-205 targets VEGFA and FGF2 and regulates resistance to chemotherapeutics in breast cancer. Cell Death Dis 2016 7 e2291 10.1038/cddis.2016.194 27362808 PMC5108343

53 Hu H Li S Cui X Lv X Jiao Y The overexpression of hypomethylated miR-663 induces chemotherapy resistance in human breast cancer cells by targeting heparin sulfate proteoglycan 2 (HSPG2). J Biol Chem 2013 288 10973 85 10.1074/jbc.M112.434340 23436656 PMC3630848

54 Kolacinska A Morawiec J Fendler W Malachowska B Morawiec Z Association of microRNAs and pathogenic response to preoperative chemotherapy in triple negative breast cancer: preliminary report. Mol Biol Rep 2014 41 2851 7 10.1007/s11033-014-3140-7 24469723 PMC4013446

55 García-Vazquez R Ruiz-Garzía E García AM Astudillo-de la Vega H Lara-Medina F A microRNA signature associated with pathological complete response to novel neoadjuvant therapy regimen in triple-negative breast cancer. Tumor Biol 2017 39 1010428317702899 10.1177/1010428317702899 28621239

56 Hubaux R Vandermeers F Cosse JP Crisanti C Kapoor V Valproic acid improves second-line regimen of small cell lung carcinoma in preclinical models. ERJ Open Res 2015 1 00028 2015 10.1183/23120541.00028-2015 27730151 PMC5005116

57 Pan YZ Gao W Yu AM MicroRNAs regulate CYP3A4 expression via direct and indirect targeting. Drug Metab Dispos 2009 37 2112 7 10.1124/dmd.109.027680 19581388 PMC2769037

58 Ravery V Fizazi K Oudard S Drouet L Eymard JC The use of estramustine phosphate in the modern management of advanced prostate cancer. BJU Int 2011 108 1782 6 10.1111/j.1464-410X.2011.10201.x 21756277

59 Wei C Pan Y Huang H Li YP Estramustine phosphate induces prostate cancer cell line PC3 apoptosis by down-regulating miR-31 levels. Eur Rev Med Pharmacol Sci 2018 22 40 5 10.26355/eurrev_201801_14098 29364469

60 Lin X Wang Y Re-expression of microRNA-4319 inhibits growth of prostate cancer via Her-2 suppression. Clin Transl Oncol 2018 20 1400 7 10.1007/s12094-018-1871-y 29633185

61 Teimouri F Nikfar S Abdollahi M Efficacy and side effects of dacarbazine in comparison with temozolomide in the treatment of malignant melanoma: a meta-analysis consisting of 1314 patients. Melanoma Res 2013 23 381 9 10.1097/CMR.0b013e3283649a97 23880781

62 Zhu W Zhou L Qian JQ Qiu TZ Shu YQ Temozolomide for the treatment of brain metastases: a review of 21 clinical trials. World J Clin Oncol 2014 5 19 27 10.5306/wjco.v5.i1.19 24527399 PMC3920177

63 Newlands ES Stevens MFG Wedge SR Wheelhouse RT Brock C Temozolomide: a review of its discovery, chemical properties, pre-clinical development and clinical trials. Cancer Treat Rev 1997 23 35 61 10.1016/S0305-7372(97)90019-0 9189180

64 Chen YN Dacarbazine inhibits proliferation of melanoma FEMX-1 cells by up-regulating expression of miRNA-200. Eur Rev Med Pharmacol Sci 2017 21 1191 7 28387914

65 Jones K Nourse JP Keane C Bhatnagar A Gandhi MK Plasma microRNA are disease response biomarkers in classical Hodgkin lymphoma. Clin Cancer Res 2013 20 253 64 10.1158/1078-0432.CCR-13-1024 24222179

66 Shi L Chen J Yang J Pan T Zhang S MiR-21 protected human glioblastoma U78MG cells from chemotherapeutic drug temozolomide induced apoptosis be decreasing Bax/Bcl-2 ratio and caspase-3 activity. Brain Res 2010 1352 255 64 10.1016/j.brainres.2010.07.009 20633539

67 Zhang S Wan Y Pan T Gu X Qian C MicroRNA-21 inhibitor sensitizes human glioblastoma U251 stem cells to chemotherapeutic drug temozolomide. J Mol Neurosci 2012 47 346 56 10.1007/s12031-012-9759-8 22528454

68 Wong ST Zhang XQ Zhuang JT Chan HL Li CH MicroRNA-21 inhibition enhances in vitro chemosensitivity of temozolomide-resistant glioblastoma cells. Anticancer Res 2012 32 2835 41 22753745

69 Gwak HS Kim TH Jo GH Kim YJ Kwak HJ Silencing of microRNA-21 confers radio-sensitivity through inhibition of the PI3K/AKT pathway and enhancing autophagy in malignant glioma cell lines. PLoS One 2012 7 e47449 10.1371/journal.pone.0047449 23077620 PMC3471817

70 Comincini S Allavena G Palumbo S Morini M Durando F MicroRNA-17 regulates the expression of ATG7 and modulates the autophagy process, improving the sensitivity to temozolomide and low-dose ionizing radiation treatments in human glioblastoma cells. Cancer Biol Ther 2013 14 574 86 10.4161/cbt.24597 23792642 PMC3742487

71 Ananta JS Paulmurugan R Massoud TF Nanoparticle-delivered antisense microRNA-21 enhances the effects of temozolomide on glioblastoma. Mol Pharmaceutics 2015 12 4509 17 10.1021/acs.molpharmaceut.5b00694 26559642

72 Ujifuku K Mitsutake N Takakura S Matsuse M Saenko V MiR-195, miR-455-3p and miR-10a(*) are implicated in acquired temozolomide resistance in glioblastoma multiforme cells. Cancer Lett 2010 296 241 8 10.1016/j.canlet.2010.04.013 20444541

73 Tezcan G Tunca B Bekar A Preusser M Berghoff AS MicroRNA expression pattern modulates temozolomide response in GBM tumors with cancer stem cells. Cell Mol Neurobiol 2014 34 679 92 10.1007/s10571-014-0050-0 24691539 PMC11488951

74 Cirilo PDR de Sousa Andrade LN Corrêa BRS Qiao M Furuya TK MicroRNA-195 acts as an anti-proliferative miRNA in human melanoma cells by targeting prohibitin 1. BMC Cancer 2017 17 750 10.1186/s12885-017-3721-7 29126391 PMC5681823

75 Munoz JL Rodriguez-Cruz V Rameshwar P High expression of miR-9 in CD133(+) glioblastoma cells in chemoresistance to temozolomide. J Cancer Stem Cell Res 2015 3 e1003 10.14343/JCSCR.2015.3e1003 27347493 PMC4917210

76 Munoz JL Rodriguez-Cruz V Ramkissoon SH Ligon KL Greco SJ Temozolomide resistance in glioblastoma occurs by miRNA-9-targeted PTCH1, independent of sonic hedgehog level. Oncotarget 2015 6 1190 201 10.18632/oncotarget.2778 25595896 PMC4359226

77 Stojcheva N Schechtmann G Sass S Roth P Florea AM MicroRNA-138 promotes acquired alkylator resistance in glioblastoma by targeting the Bcl-2-interacting mediator BIM. Oncotarget 2016 7 12937 50 10.18632/oncotarget.7346 26887050 PMC4914333

78 Han J Chen Q MiR-16 modulate temozolomide resistance by regulating BCL-2 in human glioma cells. Int J Clin Exp Pathol 2015 8 12698 707 26722459 PMC4680404

79 Luo H Chen Z Wang S Zhang R Qiu W C-Myc-miR-29c-REV3L signaling pathway drives the acquisition of temozolomide resistance in glioblastoma. Brain 2015 138 3654 72 10.1093/brain/awv287 26450587

80 Xiao S Yang X Qiu X Lv R Liu J MiR-29c contribute to glioma cells temozolomide sensitivity by targeting O6-methylgunine-DNA methyltransferases indirectly. Oncotarget 2016 7 50229 38 10.18632/oncotarget.10357 27384876 PMC5226579

81 Xu J Huang H Peng R Ding X Jiang B MicroRNA-30a increases the chemosensitivity of U251 glioblastoma cells to temozolomide by directly targeting beclin 1 and inhibiting autophagy. Exp Ther Med 2018 15 4798 804 10.3892/etm.2018.6007 29805498 PMC5952084

82 Wu H Liu Q Cai T Chen YD Liao F MiR-136 modulates glioma cell sensitivity to temozolomide by targeting astrocyte elevated gene-1. Diagn Pathol 2014 9 173 10.1186/s13000-014-0173-0 25266957 PMC4195982

83 Wang L Shi ZM Jiang CF Liu X Chen Q MiR-143 acts as a tumor suppressor by targeting N-RAS and enhances temozolomide-induced apoptosis in glioma. Oncotarget 2014 5 5416 27 10.18632/oncotarget.2116 24980823 PMC4170647

84 Chen R Liu H Cheng Q Jiang B Peng R MicroRNA-93 promotes the malignant phenotypes of human glioma cells and induces their chemoresistance to temozolomide. Biol Open 2016 5 669 77 10.1242/bio.015552 27185265 PMC4920179

85 Shi L Zhang S Feng K Wu F Wan Y MicroRNA-125b-2 confers human glioblastoma stem cell resistance to temozolomide through the mitochondrial pathway of apoptosis. Int J Oncol 2012 40 119 29 10.3892/ijo.2011.1179 21879257

86 Chen L Zhang J Han L Zhang A Zhang C Downregulation of miR-221/222 sensitizes glioma cells to temozolomide by regulating apoptosis independently of the p53 status. Oncol Rep 2012 27 854 60 10.3892/or.2011.1535 22075712

87 Zhou X Wu W Zeng A Nie E Jin X MicroRNA-141-3p promotes glioma cell growth and temozolomide resistance by directly targeting p53. Oncotarget 2017 8 71080 94 10.18632/oncotarget.20528 29050344 PMC5642619

88 Peng Y He X Chen H Duan H Shao B Inhibition of microRNA-299-5p sensitizes glioblastoma cells to temozolomide via the MAPK/ERK signaling pathway. Biosci Rep 2018 38 BSR20181051 10.1042/BSR20181051 30061180 PMC6131327

89 Tian T Mingyi M Qiu X Qiu Y MicroRNA-101 reverses temozolomide resistance by inhibition of GSK3b in glioblastoma. Oncotarget 2016 7 79584 95 10.18632/oncotarget.12861 27792996 PMC5346737

90 Chen PH Cheng CH Shih CM Ho KH Lin CW The inhibition of microRNA-128 on IGF-1-activating mTOR signaling involves in temozolomide-induced glioma cell apoptotic death. PLoS One 2016 11 e0167096 10.1371/journal.pone.0167096 27893811 PMC5125683

91 Chen H Li X Zheng H MiR-130a can predict response to temozolomide in patients with glioblastoma multiforme, independently of O6-methylguanine-DANN methyltransferase. J Transl Med 2015 13 69 10.1186/s12967-015-0435-y 25890369 PMC4345002

92 Kushwaha D Ramakrishnan V Ng K Steed T Nguyen T A genome-wide miRNA screen revealed miR-603 as a MGMT-regulating miRNA in glioblastomas. Oncotarget 2014 5 4026 39 10.18632/oncotarget.1974 24994119 PMC4147303

93 Lee YY Yarmishyn AA Wang ML Chen HY Chiou SH MicroRNA-142-3p is involved in regulation of MGMT expression in glioblastoma cells. Cancer Management Res 2018 10 775 85 10.2147/CMAR.S157261 29695934 PMC5903834

94 Kreth S Limbeck E Hinske LC Schütz SV Thon N In human glioblastomas transcript elongation by alternative polyadenylation and miRNA targeting is a potent mechanism of MGMT silencing. Acta Neuropathol 2013 125 671 81 10.1007/s00401-013-1081-1 23340988

95 Kouri FM Hurley FA Daniel WL Day ES Hua Y MiR-182 integrates apoptosis, growth, and differentiation programs in glioblastoma. Genes Dev 2015 29 732 45 10.1101/gad.257394.114 25838542 PMC4387715

96 Cheng ZX Yin WB Wang ZY MicroRNA-132 induces temozolomide resistance and promotes the formation of cancer stem cell phenotypes by targeting tumor suppressor candidate 3 in glioblastoma. Int J Mol Med 2017 40 1307 14 10.3892/ijmm.2017.3124 28901390 PMC5627876

97 Gao YT Chen XB Liu HL Up-regulation of miR-370-3p restores glioblastoma multiforme sensitivity to temozolomide by influencing MGMT expression. Sci Rep 2016 6 32972 10.1038/srep32972 27595933 PMC5011744

98 Li S Zeng A Hu Q Yan W Liu Y MiR-423-5p contributes to a malignant phenotype and temozolomide chemoresistance in glioblastomas. Neuro Oncol 2017 19 55 65 10.1093/neuonc/now129 27471108 PMC5193021

99 Chen HY Li WB Zhang CB Zhang W Zheng HY MiR-629-3p level significantly predicts prognosis in glioblastoma patients with temozolomide chemotherapy. Curr Signal Transd Ther 2014 9 9 14 10.2174/1574362409999140707103609

100 Li XB Wang J Xu AD Huang JM Meng LQ The microRNA-1268 rs28599926 polymorphism modified diffusely infiltrating astrocytoma risk and prognosis. Int J Clin Exp Med 2016 9 21615 24 10.3390/ijms15010250 24378850 PMC3907808

101 Chen H Liu L Li X Shi Y Liu N MicroRNA-1294 inhibits the proliferation and enhances the chemosensitivity of temozolomide via the direct targeting of TPX2. Am J Cancer Res 2018 8 291 301 29511599 PMC5835696

102 Ahir BK Ozer H Engelhard HH Lakka SS MicroRNAs in glioblastoma pathogenesis and therapy: a comprehensive review. Crit Rev Oncol Hematol 2017 120 22 33 10.1016/j.critrevonc.2017.10.003 29198335

103 Quintavalle C Mangani D Roscigno G Romano G Diaz-Lagares A MiR-221/222 target the DNA methyltransferase MGMT in glioma cells. PLoS One 2013 8 e74466 10.1371/journal.pone.0074466 24147153 PMC3798259

104 Bassiouny AR Zaky A Synergistic effect of curcumin on temozolomide inhibition of cancer stem cell-like properties and reduced chemoresistance of glioblastoma C6. J Stem Cell Res Ther 2012 2 5 10.4172/2157-7633.S1.005

105 Schepartz SA Early history and development of the nitrosoureas. Cancer Treat Rep 1976 60 647 9 782693

106 Weiss RB Issell BF The nitrosoureas: carmustine (BCNU) and lomustine (CCNU). Cancer Treat Rev 1982 9 313 30 10.1016/S0305-7372(82)80043-1 6762924

107 Reithmeier T Graf E Piroth T Trippel M Pinsker MO BCNU for recurrent glioblastoma multiforme: efficacy, toxicity and prognostic factors. BMC Cancer 2010 10 30 10.1186/1471-2407-10-30 20122270 PMC2837009

108 Kohn KW Interstrand coss-linking of DNA by 1,3-bis(2-chloroethyl)-1-nitrosourea and other 1-(2-chloroethyl)-1-nitrosoureas. Cancer Res 1977 37 1450 4 851960

109 Panasci LC Green D Nagourney R Fox P Schein PS A structure-activity analysis of chemical and biological parameters of chloroethylnitrosoureas in mice. Cancer Res 1977 37 2615 8 141323

110 Winkler JA Hendricks WP Johnson D Xia J Scheck AC Global microRNA expression analysis in human malignant glioma cells reveals novel miRNA-mediated therapy-resistance mechanisms. Proc Amer Assoc Cancer Res 2005 46 Abstract No 5942

111 Wang GB Liu JH Hu J Xue K MiR-21 enhanced glioma cells resistance to carmustine via decreasing Spry2 expression. Eur Rev Med Pharmacol Sci 2017 21 5065 71 10.26355/eurrev_201711_13819 29228450

112 Sippl C Ketter R Bohr L Kim YJ List M MiRNA-181d expression significantly affects treatment responses to carmustine wafer implantation. Neurosurgery 2018 10.1093/neuros/nyy214 29846701

113 Xie Q Yan Y Huang Z Zhong X Huang L MicroRNA-221 targeting PI3-K/Akt signaling axis indces cell proliferation and BCNU resistance in human glioblastoma. Neuropathol 2014 34 455 64 10.1111/neup.12129 24780067

114 Anchel M Hervey A Robbins WJ Antibiotic substances from Basidiomycetes. VII. Clitocybe illudens. Proc Natl Acad Sci 1950 6 30 6 15417544 10.1073/pnas.36.5.300 PMC1063187

115 Crooke ST Bradner WT Mitomycin C: a review. Cancer Treat Rev 1976 3 121 39 10.1016/S0305-7372(76)80019-9 786455

116 Hata T Sugawara R Mitomycin, a new antibiotic from Streptomyces. II. Description of the strain. J Antibiot 1956 9 147 51 13385187

117 Wakaki S Marumo H Tomioka K Shimizu G Kato E Isolation of new fractions of antitumor mitomycins. Antibiotics Chemother 1958 8 228 40 24544727

118 Frank W Osterberg AE Mitomycin C: a evaluation of the Japanese reports. Cancer Chemother Rep 1960 9 114 9 13701377

119 Bradner WT Mitomycin C: a clinical update. Cancer Treat Rev 2001 27 35 50 10.1053/ctrv.2000.0202 11237776

120 Tomasz M Mitomycin C: small, fast and deadly (but very selective). Chem Biol 1995 2 575 9 10.1016/1074-5521(95)90120-5 9383461

121 Feist M Huang X Müller JM Rau B Dubiel W Can hyperthermic intraperitoneal chemotherapy efficiency be improved by blocking the DNA repair factor COP9 signalosome? Int J Colorectal Dis 2014 29 673 80 10.1007/s00384-014-1861-7 24728517

122 Xu T Qin L Zhu Z Wang X Liu Y MicroRNA-31 functions as a tumor suppressor and increases sensitivity to mitomycin-C in urothelial bladder cancer by targeting integrin α5. Oncotarget 2016 7 27445 57 10.18632/oncotarget.8479 27050274 PMC5053662

123 Weeraratne SD Amani V Neiss A Teider N Scott DK MiR-34a confers chemosensitivity through modulation of MAGE-A and p53 in medulloblastoma. Neuro Oncol 2011 13 165 75 10.1093/neuonc/noq179 21177782 PMC3064629

124 Sharma S Nagpal N Ghosh PC Kulshreshtha R p53-miR-191-SOX4 regulatory loop affects apoptosis in breast cancer. RNA 2017 23 1237 46 10.1261/rna.060657.117 28450532 PMC5513068

125 Mezencev R Matyunina LV Jabbari N McDonald JF Snail-induced epithelial-to-mesenchymal transition of MCF-7 breast cancer cells: systems analysis of molecular changes and their effect on radiation and drug sensitivity. BMC Cancer 2016 16 236 10.1186/s12885-016-2274-5 26988558 PMC4797178

126 Lee S Yu KR Ryu YS Oh YS Hong IS MiR-543 and miR-590-3p regulate human mesenchymal stem cell aging via direct targeting of AIMP3/p18. AGE 2014 36 9724 10.1007/s11357-014-9724-2 25465621 PMC4259092

127 Xu K Liang X Cui D Wu Y Shi W MiR-1915 inhibits Bcl-2 to modulate multidrug resistance by increasing drug-sensitivity in human colorectal carcinoma cells. Mol Carcinogen 2013 52 70 8 10.1002/mc.21832 22121083

128 Tarasov VA Matishov DG Shin EF Boyko NV Timoshkina NN Inheritable changes in miRNAs expression in HeLa cells after X-ray and mitomycin C treatment. Russ J Gen 2014 50 798 806 10.1134/S1022795414080092 25731020

129 Le VH Inai M Williams RM Kan T Ecteinascidins. A review of the chemistry, biology and clinical utility of potent tetrahydroisoquinoline antitumor antibiotics. Nat Prod Rep 2015 32 328 47 10.1039/c4np00051j 25273374 PMC4806878

130 Jaspers NG Raams A Kelner MJ Ng JM Yamashita YM Anti-tumour compounds illudin S and irofulven induce DNA lesions ignored by global repair and exclusively processed by transcription- and replication-coupled repair pathways. DNA Repair 2002 1 1027 38 10.1016/S1568-7864(02)00166-0 12531012

131 Forni C Minuzzo M Virdis E Tamborini E Simone M Trabectedin (ET-743) promotes differentiation in myxoid liposarcoma tumors. Mol Cancer Ther 2009 8 449 57 10.1158/1535-7163.MCT-08-0848 19190116

132 Uboldi S Calura E Beltrame L Nerini IF Marchini S A systems biology approach to characterize the regulatory networks leading to trabectedin resistance in an in vitro model of myxoid liposarcoma. PLoS One 2012 7 e35423 10.1371/journal.pone.0035423 22523595 PMC3327679

133 Neia CP Cavalloni G Chiorino G Ostano P Aglietta M Gene and microRNA modulation upon trabectedin treatment in a human intrahepatic cholangiocarcinoma paired patient derived xenograft and cell line. Oncotarget 2016 7 86766 80 10.18632/oncotarget.13575 27902465 PMC5349952