Therapeutic Strategies Targeting the Exosomes in Multiple Myeloma

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1		Hong Chang	--	1847	2023-04-10 18:31:48	\|
2	format	Jason Zhu	-1 word(s)	1846	2023-04-11 05:08:39	\|

This entry is adapted from the peer-reviewed paper 10.3390/cells12071030

Multiple myeloma (MM) is a malignancy of plasma cells in the bone marrow and is characterized by the clonal proliferation of B-cells producing defective monoclonal immunoglobulins. Despite the developments in treatment, drug resistance remains one of the major challenges in the therapy of MM. The crosstalk between MM cells and other components within the bone marrow microenvironment (BME) is the major determinant of disease phenotypes. Exosomes have emerged as the critical drivers of this crosstalk by allowing the delivery of informational cargo comprising multiple components from miniature peptides to nucleic acids. Such material transfers have now been shown to perpetuate drug-resistance development and disease progression in MM. MicroRNAs(miRNAs) specifically play a crucial role in this communication considering their small size that allows them to be readily packed within the exosomes and widespread potency that impacts the developmental trajectory of the disease inside the tumor microenvironment (TME).

multiple myeloma exosome exosomal miRNA

1. Introduction

Drug resistance is the main challenge in management in a broad range of cancers, especially in Multiple myeloma (MM). In the process of myelomagenesis, the bone marrow microenvironment provides an intricate network of cellular communication to favor the tumor niche formation ^[1]. Targeting this intracellular network provides new opportunities for the therapy of the disease. Regarding the importance of exosomal miRNAs in the communications within the bone marrow microenvironment (BME) in developing drug resistance and the progress of the disease, they are considered as desirable therapeutic targets in MM treatment interventions ^[1]. Exosome-based strategies in cancer therapy or diagnosis have currently achieved a high level of interest, crucially focusing upon (1) inhibiting exosome-mediated crosstalks by reducing the production of exosomes and/or blocking their uptake, (2) their usage as therapeutic tools for the delivery of drugs, and (3) their role as potential biomarkers for early diagnosis, prognosis, or response to therapy.

2. Exosome Inhibitors

Regarding the role of exosome crosstalks in cancer progression, the inhibition of exosome release/uptake has been an attractive target for cancer treatments. In a study by Zheng et al., pharmacological inhibitors, including heparin, wortmannin, dynasore, and omeprazole have been used to block different uptake routes of specific exosomes and inhibit the tumor progression ^[2]. Furthermore, blocking exosome endocytosis by chemical inhibitors sensitized MM tumors to bortezomib treatment ^[3]. Many efforts have been made so far to design efficient exosome inhibitors. Most of these inhibitors are derived from synthetic compounds and are currently considered drugs.

SST0001 is a chemically modified heparin with anti-heparinase activity and has shown the potential to suppress exosome secretion in MM cells ^[4]. Heparinase activity in tumor cells enhances exosome secretion and alters the tumor exosomal cargo with tumorigenic factors which promote tumor progression ^[4]. The inhibition of the heparinase activity by SST0001 suppressed MM progression in vivo, even when confronted with an aggressively growing tumor within the human bone ^[5].

GW4869, another inhibitor of exosome release, is a neutral sphingomyelinase and inhibits exosome release from the plasma membrane and has shown cytotoxic effects on the MM cells ^[6] Interestingly, GW4869 reduced osteolysis and led to a reduction in tumor growth and angiogenesis in 5TGM1 mice ^[7].

Furthermore, the inhibition of the MM-exosome uptake by BMSCs using the endocytosis inhibitors suppressed exosome-induced changes in these cells and inhibited tumor-cell survival and proliferation ^[2].

Interestingly, it has been shown that some of the anticancer and anti-angiogenesis natural compounds can reduce exosome secretion and alter exosomal miRNA contents ^[8]. D Rhamnose β-hederin (DRβ-H), an active component isolated from a natural Chinese plant, reduced the secretion of exosomes and subsequently inhibited the growth and promoted the apoptosis of breast cancer cells by decreasing the level of tumorigenic encapsulated miRNAs ^[9]^[10]. DRβ-H also reversed the chemoresistance of breast cancer cells by reducing the secretion of exosomes from drug-resistant cells as well as the regulation of exosomal miRNA expression ^[11]. In addition, Shikonin (SK) inhibited exosome release and reduced the exosomal miR-128 level ^[8].

Regarding the potential of these components in the regulation of exosome release as well as the exosomal miRNA content, they may be practical in controlling disease progression or therapy resistance in MM patients. DHA could alter exosome secretion and exosomal miRNA contents and inhibit tumor growth ^[12]. In addition, the treatment of breast cancer cells with Epigallocatechin gallate (EGCG) increased miR-16 exosomal encapsulation and inhibited macrophage infiltration and M2 polarization ^[13]. Regarding the important role of miR-16 in MM tumor cells, it would be worth to evaluate the role of these neutral components in the inhibition of exosomal crosstalk and MM progression.

3. Exosomal miRNA Delivery as Therapeutic Strategy

There is some evidence for using engineered exosomes for the specific targeting of tumor cells and delivery of synthetic miRNA to inhibit the tumor cell growth. miR-21 is one of the main exosomal miRNAs in BME during myelomagenesis, and its targeting has been reported with therapeutic effects ^[14]^[15]. Ibrutinib suppresses the expression of miR-21 expression in MM cells by inhibiting nuclear factor-κB and STAT3 signaling pathways and was suggested as a promising potential treatment for this disease ^[16]. In addition, the attenuation of miR-21 with the NL101 drug suppressed the growth of B-cell lymphoma by targeting the c-Myc/Mxd1 loop ^[15].

The glioblastoma-targeted delivery of antisense miR-21 using engineered exosomes was performed by Kim et al. ^[17]. The intravenous injection of this synthetic drug into glioblastoma rat models attenuated the expression of miR-21 and promoted the expression of PDCD4 and PTEN in animals and suppressed tumor growth ^[17].

In addition, there is a great deal of evidence on the therapeutic effects of the artificial inhibition/overexpression of the miRNAs in MM to prevent tumor progression (Table 1).

Table 1. The therapeutic effects of artificial inhibition/overexpression of the miRNAs in MM.

	Therapeutic Cargo	Biological Activities	Key Findings	Refs
1	miR-29b	Targeting the epigenetics modifiers including DNMTs	Synthetic miR-29b mimics improved the aberrant expression of DNMTs in MM cells	^[18]
2	miR-15 and-16	Targeting AKT serine/threonine-protein-kinase (AKT3)	Overexpression of miRNA-15a and -16 had showed anti-MM effects (in vitro and in vivo)	^[19]
3	miR-324-5p	Targeting Hedgehog (Hh) signaling pathway	Overexpression of miR-324-5p functionally reduced cell growth and cell survival in MM and improved resistance to bortezomib in vitro and in vivo	^[20]
4	miR338-3p	Targeting Cyclin-dependent kinases	Overexpression of this miRNA suppressed proliferation and increased the apoptosis of MM cells	^[21]
5	miR-152	Targeting DKK1	Over expression of miR-152 improved DR, and inhibited the bone disruption in an intrabone MM mouse model	^[22]
8	miR-125b	Targeting IRF4 and BLIMP-1	miR-125b overexpression had an inhibition effect on the proliferation and survival of MM cells and also enhanced apoptosis and cell death in these cells	^[23]
9	miR-137/197 synthetic mimics	Targeting MCL-1	Increased the apoptosis and exerted an inhibition effect on the proliferation, colony formation, and migration ability in MM tumor cells	^[24]
10	miR-34a mimics	Targeting CDK6, BCL-2, and NOTCH1	Enhanced the apoptosis of MM cells and inhibited the proliferation in these cells	^[25]
11	Anti-miR 221/222	Upregulation of PTEN, PUMA, p27Kip1, and p57Kip2.	Induced the antiproliferative effects in MM cells	^[26]

The overexpression of miR-29 in the BME impaired the differentiation and activation of osteoclasts ^[27]. In addition, miR-29b modified Th1 differentiation by targeting interferon-ɣ and exerted potent antimyeloma activity ^[28].

Furthermore, the inhibition of IRF4 by synthetic miR-125b-5p induced the antitumor activity against MM tumors in vitro and in vivo. IRF4 plays a crucial role in the biology of Treg and Th17 cells ^[29]. On the other hand, Di Martino et al. reported evidence of a tumor-suppressor function of miR-34a in MM. They have shown that the synthetic miR-34a mimics inhibit growth and promote the apoptosis of MM cells by the suppression of CDK6, BCL-2, and NOTCH1 expression ^[25].

Although many studies have confirmed the usage of miRNAs as promising therapeutic agents for MM, no miRNA molecules have been conducted in clinical trials for MM patients. There are some challenges to be overcome to achieve an efficient and optimized therapy method, including maintenance of the stability of miRNA molecules or the efficiency of delivery methods.

For solving these problems, several methods have been tried, for instance, the conjunction of miRNAs with liposome nanoparticles. A liposomal formulation of miR-34a (MRX34) is in phase 1 clinical trials for the treatment of patients with solid tumors ^[30]. MRX34 is a miRNA drug that contains a double-stranded miR-34a mimic, which is encapsulated in a liposomal nanoparticle ^[30]. The antitumor activity of miR-34a is important in hematologic malignancies, and it is downregulated in MM patients ^[31]^[32]. Therefore, the miRNA drug MRX34 may offer new therapeutic approaches for MM.

In addition, miR-16 mimics are now in a Phase I clinical trial for patients with malignant pleural mesothelioma. TargomiRs are minicells loaded with miR-16-based mimic miRNA and targeted to EGFR that are designed to improve the loss of the miR-15 and miR-16 family miRNAs in tumor cells ^[33]. Regarding the role of miR-15 and -16 deficiency in the progression of MM malignancy and drug resistance, this drug may be practical for this disease.

4. Potential of Exosomal miRNAs as Biomarkers

Regarding the importance of monitoring the patients in the primary stage of MM for developing malignancy plasma cells or end-stage tissue damage, there are no reliable biomarkers to predict which MGUS patients may develop malignancy and who will remain stable.

Exosomal miRNAs can be easily isolated from almost all the bio-fluids and are interested as a noninvasive way to obtain information about the status of the disease ^[34]. On the other hand, the high quantity of exosome production in MM patients and their differential miRNA contents in the different stages of myelomagenesis potentiate them as promising biomarkers for early diagnosis, patient prognosis, and monitoring for developing drug resistance (Figure 1).

Figure 1. The applications of the exosomal miRNAs in the clinical practice: Exosomal miRNAs are potential diagnostic biomarkers for early detection of MM progress and they are examined for their prognostic potential. In addition, exosomes can be engineered for delivery of therapeutic miRNAs to increase their expression or targeting them to improve the immune response or induction of MM cell apoptosis by reprograming gene expression in the target cells (blue downward arrow shows downregulation and orange upward arrow shows upregulation of miRNAs).

In a study by Manier et al. on 156 patients with newly diagnosed MM, two circulating exosomal miRNAs, let-7b and miR-18a, were associated with improved survival in patients with MM ^[35]. Zhang et al. observed that the downregulation of exosomal miRNAs miR-16-5p, miR-15a-5p, miR-20a-5p, and miR-17-5p was correlated with resistance to bortezomib ^[35]. Furthermore, a meta-analysis study reported that the upregulation of miR-92a and downregulation of miR-16, -25, -744, -15a, let-7e, and -19b are associated with poor prognosis in MM ^[36].

Although there are plenty of studies on the role of exosomal miRNAs as potential biomarkers in the different stages of MM, there is a lack of consistency in the results. This inconsistency may be attributed to the presence of technical issues in the different array of methods or using platforms. In addition, miRNA profiling in biofluid samples may be affected by the sample size or the technical issues in pre-analytical and analytical steps. Therefore, for confirming the candidate miRNAs as biomarkers for usage in clinical practice, they require to be further validated in larger cohorts.

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