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Patients with multiple myeloma (MM) have an increased risk of sepsis due to underlying disease- and treatment-related immunosuppression. However, data on sepsis incidence, causative pathogens, and impact on outcomes in newly diagnosed MM (NDMM) are limited. We conducted a retrospective observational study of 92 NDMM patients who developed sepsis between 2022 and 2023 at a tertiary care center in Italy. Patient characteristics, sepsis criteria [Quick Sequential Organ Failure Assessment, Systemic Inflammatory Response Syndrome (SIRS)], microbiology results, and associations with progression-free survival (PFS) were analyzed. In this cohort of 92 critically-ill patients, pathogenic organisms were identified via microbiological culture in 74 cases. However, among the remaining 18 culture-negative patients, 9 exhibited a SIRS score of 2 and another 9 had a SIRS score of 4, suggestive of a clinical presentation consistent with sepsis despite negative cultures. Common comorbidities included renal failure (60%), anemia (71%), and bone disease (83%). Gram-negative (28%) and Gram-positive (23%) bacteria were frequent causative organisms, along with fungi (20%). Cox Univariate analyses for PFS showed statically significant HR in patients with albumin ≥ 3.5 vs < 3.5 (HR = 5.04, p < 0.001), Karnofsky performance status ≥ 80 vs < 80 (HR = 2.01, p = 0.002), and early-stage vs late-stage disease by International Staging System (HR = 4.76 and HR = 12.52, both p < 0.001) and Revised International Staging System (R-ISS III vs R-ISS I, HR = 7.38, p < 0.001). Sepsis is common in NDMM and associated with poor outcomes. Risk stratification incorporating sepsis severity, comorbidities, and disease stage may help guide preventive strategies and optimize MM management., (© 2024. The Author(s).)

Multiple myeloma (MM) progression and drug resistance depend on the crosstalk between MM cells and bone marrow (BM) fibroblasts (FBs). During monoclonal gammopathy of undetermined significance (MGUS) to MM transition, MM cell-derived exosomes (EXOs) reprogram the miRNA (miR) profile of FBs, inducing the overexpression miR-23b-3p, miR-27b-3p, miR-125b-5p, miR-214-3p, and miR-5100. Here, we demonstrate that the miR content of MM FB-derived EXOs (FB-EXOs) overlaps the miR profile of parental FBs by overexpressing comparable levels of miR-23b-3p, miR-27b-3p, miR-125b-5p, miR-214-3p, and miR-5100. Recipient MM cells co-cultured with MM FB-EXOs selectively overexpress only miR-214-3p and miR-5100 but not miR-23b-3p, miR-27b-3p, and miR-125b-5p, suggesting a putative selective transfer. MM cells express HOTAIR, TOB1-AS1, and MALAT1 lncRNAs. Transient transfection of MM cells with lnc·siRNAs demonstrates that HOTAIR, TOB1-AS1, and MALAT1 lncRNAs are sponges for miR-23b-3p, miR-27b-3p, and miR-125b-5p. Indeed, lncRNA knockdown significantly increased miR levels in U266 MM cells co-cultured with MM FB-EXOs. Selective miR-214-3p and miR-5100 overexpression modulates MAPK, PI3K/AKT/mTOR, and p53 pathways in MM cells. Interrogation using the DIANA tools algorithm and transient overexpression using miR mimic probes confirmed the involvement of miR-214-3p and miR-5100 and their target genes, PTEN and DUSP16, respectively, in the modulation of these intracellular pathways. Finally, the uptake of EXOs as well as miR-214-3p and miR-5100 overexpression increase MM cell proliferation and resistance to bortezomib-induced apoptosis by switching the balance between pro-/anti-apoptotic proteins. Overall, these data show that MM cells are not simply a container into which EXOs empty their cargo. On the contrary, tumour cells finely neutralize exosomal miRs via lncRNA expression to ensure their survival. © 2021 The Pathological Society of Great Britain and Ireland., (© 2021 The Pathological Society of Great Britain and Ireland.)

Multiple myeloma (MM) progression closely depends on the bidirectional crosstalk between tumor cells and the surrounding microenvironment, which leads to the creation of a tumor supportive niche. Extracellular vesicles (EVs) have emerged as key players in the pathological interplay between the malignant clone and near/distal bone marrow (BM) cells through their biologically active cargo. Here, we describe the role of EVs derived from MM and BM cells in reprogramming the tumor microenvironment and in fostering bone disease, angiogenesis, immunosuppression, drug resistance, and, ultimately, tumor progression. We also examine the emerging role of EVs as new therapeutic agents for the treatment of MM, and their potential use as clinical biomarkers for early diagnosis, disease classification, and therapy monitoring.

Multiple myeloma (MM) progression closely depends on bone marrow (BM) angiogenesis. Several factors sustain angiogenesis, including cytokines, growth factors, and cell-to-cell interactions. Herein, BM thrombopoietin (TPO) was shown to support angiogenesis and disease progression in MM. Patients with MM at different progression phases had higher levels of BM and circulating TPO than monoclonal gammopathy of undetermined significance/smoldering MM patients, suggesting that TPO correlates with disease progression and prognosis. Endothelial cells from patients with monoclonal gammopathy of undetermined significance (MGECs) and endothelial cells from MM (MMECs) expressed TPO receptor, and the TPO treatment triggered their angiogenic capabilities in vitro. Indeed, TPO-treated MGECs and MMECs showed enhanced angiogenesis on Matrigel and spontaneous cell migration and chemotaxis by acting as a chemotactic agent. TPO also had an angiogenic activity in vivo in the chorioallantoic membrane assay system. Finally, TPO treatment increased the release of active matrix metalloproteinase (MMP)-9 and MMP-2 in MGECs and of MMP-2 in MMECs and affected the balance between angiogenic/antiangiogenic factors in the MM BM. Our results support the angiogenic activity of TPO, and suggest that it may have a critical role in promoting the angiogenic switch during MM progression. Accordingly, TPO may be envisaged as a new angiogenic and prognostic factor in patients with MM., (Copyright © 2021 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Multiple myeloma, the second most common hematologic malignancy, frequently relapses because of chemotherapeutic resistance. Fibroblast growth factors (FGF) act as proangiogenic and mitogenic cytokines in multiple myeloma. Here, we demonstrate that the autocrine FGF/FGFR axis is essential for multiple myeloma cell survival and progression by protecting multiple myeloma cells from oxidative stress-induced apoptosis. In keeping with the hypothesis that the intracellular redox status can be a target for cancer therapy, FGF/FGFR blockade by FGF trapping or tyrosine kinase inhibitor impaired the growth and dissemination of multiple myeloma cells by inducing mitochondrial oxidative stress, DNA damage, and apoptotic cell death that were prevented by the antioxidant vitamin E or mitochondrial catalase overexpression. In addition, mitochondrial oxidative stress occurred as a consequence of proteasomal degradation of the c-Myc oncoprotein that led to glutathione depletion. Accordingly, expression of a proteasome-nondegradable c-Myc protein mutant was sufficient to avoid glutathione depletion and rescue the proapoptotic effects due to FGF blockade. These findings were confirmed on bortezomib-resistant multiple myeloma cells as well as on bone marrow-derived primary multiple myeloma cells from newly diagnosed and relapsed/refractory patients, including plasma cells bearing the t(4;14) translocation obtained from patients with high-risk multiple myeloma. Altogether, these findings dissect the mechanism by which the FGF/FGFR system plays a nonredundant role in multiple myeloma cell survival and disease progression, and indicate that FGF targeting may represent a therapeutic approach for patients with multiple myeloma with poor prognosis and advanced disease stage. SIGNIFICANCE: This study provides new insights into the mechanisms by which FGF antagonists promote multiple myeloma cell death. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/11/2340/F1.large.jpg., (©2020 American Association for Cancer Research.)

MicroRNAs (miRNAs, or miRs) are single-strand short non-coding RNAs with a pivotal role in the regulation of physiological- or disease-associated cellular processes. They bind to target miRs modulating gene expression at post-transcriptional levels. Here, we present an overview of miRs deregulation in the pathogenesis of multiple myeloma (MM), and discuss the potential use of miRs/nanocarriers association in clinic. Since miRs can act as oncogenes or tumor suppressors, strategies based on their inhibition and/or replacement represent the new opportunities in cancer therapy. The miRs delivery systems include liposomes, polymers, and exosomes that increase their physical stability and prevent nuclease degradation. Phase I/II clinical trials support the importance of miRs as an innovative therapeutic approach in nanomedicine to prevent cancer progression and drug resistance. Results in clinical practice are promising., Competing Interests: The authors declare no conflicts of interest.

Daratumumab (Dara) is the first-in-class human-specific anti-CD38 mAb approved for the treatment of multiple myeloma (MM). Although recent data have demonstrated very promising results in clinical practice and trials, some patients do not achieve a partial response, and ultimately all patients undergo progression. Dara exerts anti-MM activity via antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), complement-dependent cytotoxicity (CDC), and immunomodulatory effects. Deregulation of these pleiotropic mechanisms may cause development of Dara resistance. Knowledge of this resistance may improve the therapeutic management of MM patients.

Aberrant microRNA (miR) expression has an important role in tumour progression, but its involvement in bone marrow fibroblasts of multiple myeloma patients remains undefined. We demonstrate that a specific miR profile in bone marrow fibroblasts parallels the transition from monoclonal gammopathy of undetermined significance (MGUS) to myeloma. Overexpression of miR-27b-3p and miR-214-3p triggers proliferation and apoptosis resistance in myeloma fibroblasts via the FBXW7 and PTEN/AKT/GSK3 pathways, respectively. Transient transfection of miR-27b-3p and miR-214-3p inhibitors demonstrates a cooperation between these two miRNAs in the expression of the anti-apoptotic factor MCL1, suggesting that miR-27b-3p and miR-214-3p negatively regulate myeloma fibroblast apoptosis. Furthermore, myeloma cells modulate miR-27b-3p and miR-214-3p expression in fibroblasts through the release of exosomes. Indeed, tumour cell-derived exosomes induce an overexpression of both miRNAs in MGUS fibroblasts not through a simple transfer mechanism but by de novo synthesis triggered by the transfer of exosomal WWC2 protein that regulates the Hippo pathway. Increased levels of miR-27b-3p and miR-214-3p in MGUS fibroblasts co-cultured with myeloma cell-derived exosomes enhance the expression of fibroblast activation markers αSMA and FAP. These data show that the MGUS-to-myeloma transition entails an aberrant miRNA profile in marrow fibroblasts and highlight a key role of myeloma cells in modifying the bone marrow microenvironment by reprogramming the marrow fibroblasts' behaviour. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd., (Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Interactions of multiple myeloma (MM) cells with endothelial cells (ECs) enhance angiogenesis and MM progression. Here, we investigated the role of Notch signaling in the cross talk between ECs and MM cells enabling angiogenesis. MMECs showed higher expression of Jagged1/2 ligands, of activated Notch1/2 receptors, and of Hes1/Hey1 Notch target genes than ECs from monoclonal gammopathy of undetermined significance patients, suggesting that homotypic activation of Notch pathway occurs in MM. MM cells co-cultured with MMECs triggered Notch activation in these cells through a cell-to-cell contact-dependent way via Jagged1/2, resulting in Hes1/Hey1 overexpression. The angiogenic effect of Notch pathway was analyzed through Notch1/2·siRNAs and the γ-secretase inhibitor MK-0752 by in vitro (adhesion, migration, chemotaxis, angiogenesis) and in vivo (Vk12598/C57B/6 J mouse model) studies. Activated Notch1/2 pathway was associated with the overangiogenic MMEC phenotype: Notch1/2 knockdown or MK-0752 treatment reduced Hes1/Hey1 expression, impairing in vitro angiogenesis of both MMECs alone and co-cultured with MM cells. MM cells were unable to restore angiogenic abilities of treated MMECs, proving that MMEC angiogenic activities closely rely on Notch pathway. Furthermore, Notch1/2 knockdown affected VEGF/VEGFR2 axis, indicating that the Notch pathway interferes with VEGF-mediated control on angiogenesis. MK-0752 reduced secretion of proangiogenic/proinflammatory cytokines in conditioned media, thus inhibiting blood vessel formation in the CAM assay. In the Vk12598/C57B/6 J mouse, MK-0752 treatment restrained angiogenesis by reducing microvessel density. Overall, homotypic and heterotypic Jagged1/2-mediated Notch activation enhances MMECs angiogenesis. Notch axis inhibition blocked angiogenesis in vitro and in vivo, suggesting that the Notch pathway may represent a novel therapeutic target in MM., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

We have previously demonstrated that recombinant human erythropoietin (rHuEpo) is involved in the regulation of the angiogenic response in multiple myeloma (MM) through a direct effect on macrophages and endothelial cells isolated from the bone marrow of patients with MM. The aim of the present study was designed to determine the effects of rHuEpo on cancer-associated fibroblasts (CAFs) from monoclonal gammopathy of undetermined significance (MGUS) and MM patients by means of in vitro and in vivo assays. rHuEpo treatment reduces the expression of mRNA levels of fibroblast activation markers, namely alpha smooth actin (αSMA) and fibroblast activation protein (FAP) in MGUS and MM CAFs, and of pro-inflammatory and pro-angiogenic cytokines, including interleukin (IL)-6 and IL-8, vascular endothelial growth factor-A (VEGF-A), fibroblast growth factor-2 (FGF-2), and hepatocyte growth factor (HGF) in MM CAFs. Moreover, rHuEpo inhibits the proliferative activity of MM CAFs and increased the apoptosis of MGUS and MM CAFs. Overall, these data suggest that rHu-Epo down-regulates CAFs pro-tumorigenic activity. Moreover, these results are not suggestive for a pro-angiogenic activity of rHuEpo on CAFs. In fact, rHuEpo pre-treatment induces a low angiogenic response in vivo in the chorioallantoic membrane (CAM) assay of MGUS and MM CAFs conditioned medium, not comparable to that of a well-known angiogenic cytokine, VEGF-A, tested in the same assay.

Cell adhesion in the multiple myeloma (MM) microenvironment has been recognized as a major mechanism of MM cell survival and the development of drug resistance. Here we addressed the hypothesis that the protein junctional adhesion molecule-A (JAM-A) may represent a novel target and a clinical biomarker in MM. We evaluated JAM-A expression in MM cell lines and in 147 MM patient bone marrow aspirates and biopsies at different disease stages. Elevated JAM-A levels in patient-derived plasma cells were correlated with poor prognosis. Moreover, circulating soluble JAM-A (sJAM-A) levels were significantly increased in MM patients as compared with controls. Notably, in vitro JAM-A inhibition impaired MM migration, colony formation, chemotaxis, proliferation and viability. In vivo treatment with an anti-JAM-A monoclonal antibody (αJAM-A moAb) impaired tumor progression in a murine xenograft MM model. These results demonstrate that therapeutic targeting of JAM-A has the potential to prevent MM progression, and lead us to propose JAM-A as a biomarker in MM, and sJAM-A as a serum-based marker for clinical stratification.

Multiple myeloma (MM) drug resistance (DR) is a multistep transformation process based on a powerful interplay between bone marrow stromal cells and MM cells that allows the latter to escape anti-myeloma therapies. Here we present an overview of the role of the bone marrow microenvironment in both soluble factors-mediated drug resistance (SFM-DR) and cell adhesion-mediated drug resistance (CAM-DR), focusing on the role of new players, namely miRNAs, exosomes and cancer-associated fibroblasts.

Bortezomib (bort) has improved overall survival in patients with multiple myeloma (MM), but the majority of them develop drug resistance. In this study, we demonstrate that bone marrow (BM) fibroblasts (cancer-associated fibroblasts; CAFs) from bort-resistant patients are insensitive to bort and protect the RPMI8226 and patients' plasma cells against bort-induced apoptosis. Bort triggers CAFs to produce high levels of interleukin (IL)-6, IL-8, insulin-like growth factor (IGF)-1 and transforming growth factor (TGF) β. Proteomic studies on CAFs demonstrate that bort resistance parallels activation of oxidative stress and pro-survival autophagy. Indeed, bort induces reactive oxygen species in bort-resistant CAFs and activates autophagy by increasing light chain 3 protein (LC3)-II and inhibiting p62 and phospho-mammalian target of rapamycin. The small-interfering RNA knockdown of Atg7, and treatment with 3-methyladenine, restores bort sensitivity in bort-resistant CAFs and produces cytotoxicity in plasma cells co-cultured with CAFs. In the syngeneic 5T33 MM model, bort-treatment induces the expansion of LC3-II(+) CAFs. TGFβ mediates bort-induced autophagy, and its blockade by LY2109761, a selective TβRI/II inhibitor, reduces the expression of p-Smad2/3 and LC3-II and induces apoptosis in bort-resistant CAFs. A combination of bort and LY2109761 synergistically induces apoptosis of RPMI8226 co-cultured with bort-resistant CAFs. These data define a key role for CAFs in bort resistance of plasma cells and provide the basis for a novel targeted therapeutic approach.

Regulatory T cells (Tregs) are essential for maintenance of self-tolerance; however, tumor cells can exploit the tolerance to escape the immune system. We investigated the Tregs frequency in patients with multiple myeloma (MM) and in those with monoclonal gammopathy of undetermined significance (MGUS), and found that CD4(+) FoxP3(+) and CD8(+) FoxP3(+) Tregs were significantly increased in patients with MM and correlated with the active phase. Both Tregs subsets were expanded in cocultures of CD3(+) lymphocytes and fresh CD138(+) MM plasma cells or RPMI8226 and U266 cell lines and functioned as natural (n) and inducible (i) Tregs insofar as they inhibited the proliferation of stimulated CD3 lymphocytes via contact-dependent and contact-independent pathways. Induction of Tregs by MM plasma cells required a contact-dependent pathway, implying antigen recognition by T cells. MM plasma cells acted as immature and tolerogenic antigen-presenting cells (APCs), in that they displayed low CD80/CD86 expression associated with a phagocytic activity. By acting as immature APCs, MM plasma cells plausibly expand (n)Tregs and (i)Tregs both through conversion of CD3(+) FoxP3(-) into CD3(+) FoxP3(+) T cells and proliferation of CD3(+) FoxP3(+) T cells, which may suppress the anti-MM immune response., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)