Amundarain, A. (Ane)

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    Aptamers, a new therapeutic opportunity for the treatment of multiple myeloma
    (2022) Amundarain, A. (Ane); Pastor, F. (Fernando); Aguirre-Ena, X. (Xabier); Prosper-Cardoso, F. (Felipe)
    Simple Summary Multiple Myeloma (MM) remains incurable due to high relapse rates and fast development of drug resistances. Monoclonal antibodies (mAb) have revolutionized MM treatment, opening the door to chemotherapy-free yet curative treatments. Nevertheless, antibody-based therapies face several difficulties which could be overcome by nucleic acid aptamers. Aptamers are short oligonucleotide ligands that bind their targets with great affinity and specificity, and can be easily conjugated to different cargoes for their cell-specific delivery. This review summarizes the aptamers that have been tested in MM so far with promising results, and proposes various strategies for the further development of aptamer-based strategies against MM. Multiple Myeloma (MM) remains an incurable disease due to high relapse rates and fast development of drug resistances. The introduction of monoclonal antibodies (mAb) has caused a paradigm shift in MM treatment, paving the way for targeted approaches with increased efficacy and reduced toxicities. Nevertheless, antibody-based therapies face several difficulties such as high immunogenicity, high production costs and limited conjugation capacity, which we believe could be overcome by the introduction of nucleic acid aptamers. Similar to antibodies, aptamers can bind to their targets with great affinity and specificity. However, their chemical nature reduces their immunogenicity and production costs, while it enables their conjugation to a wide variety of cargoes for their use as delivery agents. In this review, we summarize several aptamers that have been tested against MM specific targets with promising results, establishing the rationale for the further development of aptamer-based strategies against MM. In this direction, we believe that the study of novel plasma cell surface markers, the development of intracellular aptamers and further research on aptamers as building blocks for complex nanomedicines will lead to the generation of next-generation targeted approaches that will undoubtedly contribute to improve the management and life quality of MM patients.
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    Characterization of complete lncRNAs transcriptome reveals the functional and clinical impact of lncRNAs in multiple myeloma.
    (Springer Nature, 2021) Carrasco-León, A. (Arantxa); Ezponda, T. (Teresa); Meydan, C. (Cem); Valcárcel-García, L.V. (Luis Vitores); Ordoñez, R. (Raquel); Kulis, M. (Marta); Garate, L. (Leire); Miranda, E. (Estibaliz); Segura, V. (Víctor); Guruceaga, E. (Elizabeth); Vilas-Zornoza, A. (Amaia); Alignani, D. (Diego); Pascual, M. (Marien); Amundarain, A. (Ane); Castro-Labrador, L. (Laura); San Martín, P. (Patxi); El-Omri, H. (Halima); Taha, R.Y. (Ruba Y.); Calasanz-Abinzano, M.J. (Maria Jose); Planes-Pedreño, F.J. (Francisco Javier); Paiva, B. (Bruno); Mason, C. E. (Christopher, E.); San-Miguel, J.F. (Jesús F.); Martin-Subero, J.I. (Jose Ignacio); Melnick, A. (Ari); Prosper-Cardoso, F. (Felipe); Aguirre-Ena, X. (Xabier)
    Multiple myeloma (MM) is an incurable disease, whose clinical heterogeneity makes its management challenging, highlighting the need for biological features to guide improved therapies. Deregulation of specific long non-coding RNAs (lncRNAs) has been shown in MM, nevertheless, the complete lncRNA transcriptome has not yet been elucidated. In this work, we identified 40,511 novel lncRNAs in MM samples. lncRNAs accounted for 82% of the MM transcriptome and were more heterogeneously expressed than coding genes. A total of 10,351 overexpressed and 9,535 downregulated lncRNAs were identified in MM patients when compared with normal bone-marrow plasma cells. Transcriptional dynamics study of lncRNAs in the context of normal B-cell maturation revealed 989 lncRNAs with exclusive expression in MM, among which 89 showed de novo epigenomic activation. Knockdown studies on one of these lncRNAs, SMILO (specific myeloma intergenic long non-coding RNA), resulted in reduced proliferation and induction of apoptosis of MM cells, and activation of the interferon pathway. We also showed that the expression of lncRNAs, together with clinical and genetic risk alterations, stratified MM patients into several progression-free survival and overall survival groups. In summary, our global analysis of the lncRNAs transcriptome reveals the presence of specific lncRNAs associated with the biological and clinical behavior of the disease.
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    Gene expression derived from alternative promoters improves prognostic stratification in multiple myeloma.
    (Springer nature, 2021) Valcárcel-García, L.V. (Luis Vitores); Amundarain, A. (Ane); Kulis, M. (Marta); Charalampopoulou, S. (Stella); Melnick, A. (Ari); San-Miguel, J.F. (Jesús F.); Martin-Subero, J.I. (Jose Ignacio); Planes-Pedreño, F.J. (Francisco Javier); Agirre, X. (Xabier); Prosper-Cardoso, F. (Felipe)
    Clinical and genetic risk factors are currently used in multiple myeloma (MM) to stratify patients and to design specific therapies. However, these systems do not capture the heterogeneity of the disease supporting the development of new prognostic factors. In this study, we identified active promoters and alternative active promoters in 6 different B cell subpopulations, including bone-marrow plasma cells, and 32 MM patient samples, using RNA-seq data. We find that expression initiated at both regular and alternative promoters was specific of each B cell subpopulation or MM plasma cells, showing a remarkable level of consistency with chromatin-based promoter definition. Interestingly, using 595 MM patient samples from the CoMMpass dataset, we observed that the expression derived from some alternative promoters was associated with lower progression-free and overall survival in MM patients independently of genetic alterations. Altogether, our results define cancer-specific alternative active promoters as new transcriptomic features that can provide a new avenue for prognostic stratification possibilities in patients with MM.
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    The Role of lncRNAs in the Pathobiology and Clinical Behavior of Multiple Myeloma
    (2021) Amundarain, A. (Ane); Carrasco-León, A. (Arantxa); Prosper-Cardoso, F. (Felipe); Gómez-Echarte, N. (Nahia); Aguirre-Ena, X. (Xabier)
    Simple Summary Multiple myeloma (MM), the second most common hematological neoplasm, is still considered an incurable disease. Long non-coding RNAs (lncRNAs), genes that do not encode proteins, participate in numerous biological processes, but their deregulation, like that of coding genes, can contribute to carcinogenesis. Increasing evidence points to the relevant role of lncRNAs in the development of human tumors, such that they emerge as attractive biomarkers and therapeutic targets for cancer treatment, including MM. Here we review the oncogenic or tumor-suppressor functions of lncRNAs in MM and provide an overview of novel therapeutic approaches based on lncRNAs that will help to improve the management of these patients. MM is a hematological neoplasm that is still considered an incurable disease. Besides established genetic alterations, recent studies have shown that MM pathogenesis is also characterized by epigenetic aberrations, such as the gain of de novo active chromatin marks in promoter and enhancer regions and extensive DNA hypomethylation of intergenic regions, highlighting the relevance of these non-coding genomic regions. A recent study described how long non-coding RNAs (lncRNAs) correspond to 82% of the MM transcriptome and an increasing number of studies have demonstrated the importance of deregulation of lncRNAs in MM. In this review we focus on the deregulated lncRNAs in MM, including their biological or functional mechanisms, their role as biomarkers to improve the prognosis and monitoring of MM patients, and their participation in drug resistance. Furthermore, we also discuss the evidence supporting the role of lncRNAs as therapeutic targets through different novel RNA-based strategies.
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    LncRNAs: Novel therapeutic targets to treat Multiple Myeloma with RNA-based therapies
    (Universidad de Navarra, 2023-11-16) Amundarain, A. (Ane); Prosper-Cardoso, F. (Felipe); Prosper, F. (Felipe); Aguirre-Ena, X. (Xabier)
    Multiple Myeloma (MM) is a hematologic neoplasm caused by the clonal proliferation of aberrant plasma cells in the bone marrow (BM). Hyperdiploidy and IGH translocations are the primary genetic events in MM, each affecting approximately 50% of patients. However, there are cases where the translocation partner of IGH is still unknown. Moreover, beyond genetics, epigenomic and transcriptomic studies have uncovered the extensive chromatin activation of regulatory elements in MM, from where thousands of long non-coding RNAs (lncRNAs) are expressed. We hypothesize that these lncRNAs could be involved in translocations occurring in MM. Furthermore, the inhibition of one of these lncRNA, named SMILO, triggers MM cell death, suggesting the potential of lncRNAs as novel therapeutic targets in MM. To unravel the contribution of lncRNAs to translocations in MM, we identified fusion transcripts (FTs) from strand specific RNA-seq data in samples from different B cell subpopulations and MM patient samples. After a stringent computational filtering, we detected FTs in all studied subpopulations, implying that FTs present in healthy samples may be contributing to physiological processes. Nevertheless, FTs present in healthy samples were filtered to detect MM-specific FTs. As hypothesized, the 30% of MM-specific FTs occurred with lncRNAs (lncFTs), and interestingly, an important percentage of MM-specific FTs were transcription readthroughs (RT) between adjacent genes, therefore leading to the identification of novel RNA transcript classes with unknown functional and prognostic potential in this disease. Finally, the contribution of lncFTs to the prognosis of MM patients was assessed by multivariate survival studies performed in the CoMMpass dataset including 599 MM patient samples, demonstrating that the combination of the presence of certain lncFTs with classical genomic high-risk markers can improve the prognostic stratification of MM patients. On the other hand, we propose the generation of aptamer-siRNA chimeras as RNA-based therapies that will enable the targeted silencing of oncogenic coding and non-coding genes, including lncRNAs and FTs, specifically in the pathological plasma cells of MM. The silencing is achieved with therapeutic siRNA molecules which are delivered specifically to pathological plasma cells by MM-specific RNA aptamers. To identify the MM-specific aptamers we applied a Cell-SELEX approach, where a random RNA aptamer library is subjected to iterative binding rounds with MM cell lines to enrich sequences able to bind target cells specifically. After 10 selection rounds, we identified 10 enriched aptamer clusters, from which aptamer 1 (APT1), aptamer 1 (APT3) and aptamer 1 (APT9) were selected for validation studies. Flow cytometry and confocal microscopy studies showed that all aptamers bind preferentially and internalize into MM cell lines, besides, APT3 and APT9 were minimally toxic to MM cell lines up to 5 M, suggesting they could be used as siRNAs carriers for targeted delivery without inherent toxic effects. Likewise, we identified effective siRNAs able to inhibit both SMILO lncRNA and RRM1 coding gene with known oncogenic effects in MM, so they were selected for chimera generation with the APT3 to obtain the therapeutic silencing of these genes in the MM cell. In conclusion, this doctoral thesis aimed at elucidating the involvement and the prognostic significance of lncRNAs in fusion transcripts occurring in MM, as well as to develop RNA-based therapeutic approaches that will enable the targeted silencing of these novel transcript classes together with coding genes in a cell-specific fashion.