The CIt protocol: A blueprint to potentiate the immunogenicity of immunoproteasome-reprogrammed mesenchymal stromal cells.

Immunoproteasome-reprogrammed mesenchymal stromal cells (IRMs) can surpass dendritic cells at eliciting tumor-specific immunity. However, the current IRM vaccination regimen remains clinically unsuitable due to the relatively high dose of IRMs needed. Since the administration of a lower IRM dose triggers a feeble anti-tumoral response, we aimed to combine this vaccination regimen with different modalities to fine-tune the potency of the vaccine. In a nutshell, we found that the co-administration of IRMs and interleukin-12 accentuates the anti-tumoral response, whereas the cross-presentation potency of IRMs is enhanced via intracellular succinate build-up, delayed endosomal maturation, and increased endosome-to-cytosol plasticity. Stimulating phagocyte-mediated cancer efferocytosis by blocking the CD47-SIRPα axis was also found to enhance IRM vaccine outcomes. Upon designing a single protocol combining the abovementioned strategies, 60% of treated animals exhibited a complete response. Altogether, this is the first IRM-based vaccination study, optimized to simultaneously target three vaccine-related pitfalls: T-cell response, antigen cross-presentation, and cancer phagocytosis.

CD146 Defines a Mesenchymal Stromal Cell Subpopulation with Enhanced Suppressive Properties.

Mesenchymal stromal cells (MSCs) are largely known for their immune-suppressive capacity, hence, their common use in the control of unwanted inflammation. However, novel concepts related to their biology, combined with the urgent need to identify MSC subpopulations with enhanced suppressive properties, drive the search for isolation protocols optimized for clinical applications. We show, in this study, that MSCs expressing high CD146 levels exhibit altered surface expression profiles of CD44 and secrete elevated levels of interleukin (IL)-6, amongst other factors. In addition, CD146hi MSCs surpass the polyclonal parental populations in inhibiting alloreactive T cells in vitro, in both a soluble- and cell-contact-dependent manner. Despite the lack of CD146hi MSC-mediated activation of peritoneal macrophages to release the suppressive factor IL-10 in vitro, their administration in animals with graft-versus-host disease alleviates inflammation and leads to 40% survival rate up to 7 weeks post-transplantation. This pronounced inhibitory property is driven by CD146-mediated in situ efferocytosis by myeloid cells. Altogether, this study provides the impetus to adopt an isolation protocol for MSCs based on a CD146 expression profile before their therapeutic use and suggests a major role played by CD146 as a novel "eat-me" signal, capable of enhancing MSC uptake by competent phagocytes.

LSD1 Inhibition Enhances the Immunogenicity of Mesenchymal Stromal Cells by Eliciting a dsRNA Stress Response.

Mesenchymal stromal cells (MSCs) are commonly known for their immune-suppressive abilities. However, our group provided evidence that it is possible to convert MSCs into potent antigen presenting cells (APCs) using either genetic engineering or pharmacological means. Given the capacity of UM171a to trigger APC-like function in MSCs, and the recent finding that this drug may modulate the epigenome by inhibiting the lysine-specific demethylase 1 (LSD1), we explored whether the direct pharmacological inhibition of LSD1 could instill APC-like functions in MSCs akin to UM171a. The treatment of MSCs with the LSD1 inhibitor tranylcypromine (TC) elicits a double-stranded (ds)RNA stress response along with its associated responsive elements, including pattern recognition receptors (PRRs), Type-I interferon (IFN), and IFN-stimulated genes (ISGs). The net outcome culminates in the enhanced expression of H2-Kb, and an increased stability of the cell surface peptide: MHCI complexes. As a result, TC-treated MSCs stimulate CD8 T-cell activation efficiently, and elicit potent anti-tumoral responses against the EG.7 T-cell lymphoma in the context of prophylactic vaccination. Altogether, our findings reveal a new pharmacological protocol whereby targeting LSD1 in MSCs elicits APC-like capabilities that could be easily exploited in the design of future MSC-based anti-cancer vaccines.

Humoral Immunity to Allogeneic Immunoproteasome-Expressing Mesenchymal Stromal Cells Requires Efferocytosis by Endogenous Phagocytes.

The extensive use of mesenchymal stromal cells (MSCs) over the last decade has revolutionized modern medicine. From the delivery of pharmacological proteins to regenerative medicine and immune modulation, these cells have proven to be highly pleiotropic and responsive to their surrounding environment. Nevertheless, their role in promoting inflammation has been fairly limited by the questionable use of interferon-gamma, as this approach has also been proven to enhance the cells' immune-suppressive abilities. Alternatively, we have previously shown that de novo expression of the immunoproteasome (IPr) complex instills potent antigen cross-presentation capabilities in MSCs. Interestingly, these cells were found to express the major histocompatibility class (MHC) II protein, which prompted us to investigate their ability to stimulate humoral immunity. Using a series of in vivo studies, we found that administration of allogeneic ovalbumin (OVA)-pulsed MSC-IPr cells elicits a moderate antibody titer, which was further enhanced by the combined use of pro-inflammatory cytokines. The generated antibodies were functional as they blocked CD4 T-cell activation following their co-culture with OVA-pulsed MSC-IPr and mitigated E.G7 tumor growth in vivo. The therapeutic potency of MSC-IPr was, however, dependent on efferocytosis, as phagocyte depletion prior to vaccination abrogated MSC-IPr-induced humoral responses while promoting their survival in the host. In contrast, antibody-mediated neutralization of CD47, a potent "do not eat me signal", enhanced antibody titer levels. These observations highlight the major role played by myeloid cells in supporting antibody production by MSC-IPr and suggest that the immune outcome is dictated by a net balance between efferocytosis-stimulating and -inhibiting signals.

Bone marrow aspirate concentrate versus platelet-rich plasma for treating knee osteoarthritis: a one-year non-randomized retrospective comparative study.

BACKGROUND: Knee osteoarthritis (OA) is a debilitating condition affecting human body biomechanics and quality of life. Current standard care for knee OA leads to trivial improvement and entails multiple adverse effects or complications. Recently, investigational cell therapies injected intra-articularly, such as bone marrow aspirate concentrate (BMAC) and platelet-rich plasma (PRP), have shown safety and therapeutic potency providing patients with pain relief. In the current retrospective comparative study, we investigated the differences in pain and functional improvements in patients with symptomatic knee OA receiving intra-articular injections of BMAC vs PRP. METHODS: Pain and functionality scores were measured at baseline and at different time points post-injection over 12 months, using 3 self-administered, clinically validated questionnaires: the visual analogue scale (VAS) for assessing pain intensity, the knee injury and osteoarthritis outcome score (KOOS) for evaluating functionality and knee-related quality of life, and the Western Ontario and McMaster Universities Arthritis Index (WOMAC) for evaluating physical function. The repeated-measures general linear model with Sidak test for pairwise comparisons was used to investigate the influence of the treatment on the score evolution within groups (between baseline and each time point) and between groups (overall). RESULTS: The BMAC group (n = 26 knees) significantly improved in VAS, KOOS, and WOMAC scores between baseline and 12 months (57.4, 75.88, and 73.95% mean score improvement, respectively). In contrast, the PRP group (n = 13 knees) witnessed nonsignificant improvement in all scores. BMAC, in comparison to PRP, induced significant improvement in outcomes by 29.38% on the VAS scale, 53.89% on the KOOS scale, and 51.71% on the WOMAC scale (P < .002, P < .01, P < .011, respectively). CONCLUSIONS: Intra-articular autologous BMAC injections are safe, effective in treating pain, and ameliorate functionality in patients with symptomatic knee OA to a greater extent than PRP injections. Intra-articular autologous BMAC therapy is safe and provides more relief to patients with symptomatic knee osteoarthritis compared to PRP therapy.

UM171A-induced ROS promote antigen cross-presentation of immunogenic peptides by bone marrow-derived mesenchymal stromal cells.

BACKGROUND: Mesenchymal stromal cells (MSCs) have been extensively used in the clinic due to their exquisite tissue repair capacity. However, they also hold promise in the field of cellular vaccination as they can behave as conditional antigen presenting cells in response to interferon (IFN)-gamma treatment under a specific treatment regimen. This suggests that the immune function of MSCs can be pharmacologically modulated. Given the capacity of the agonist pyrimido-indole derivative UM171a to trigger the expression of various antigen presentation-related genes in human hematopoietic progenitor cells, we explored the potential use of UM171a as a means to pharmacologically instill and/or promote antigen presentation by MSCs. METHODS: Besides completing a series of flow-cytometry-based phenotypic analyses, several functional antigen presentation assays were conducted using the SIINFEKL-specific T-cell clone B3Z. Anti-oxidants and electron transport chain inhibitors were also used to decipher UM171a's mode of action in MSCs. Finally, the potency of UM171a-treated MSCs was evaluated in the context of therapeutic vaccination using immunocompetent C57BL/6 mice with pre-established syngeneic EG.7T-cell lymphoma. RESULTS: Treatment of MSCs with UM171a triggered potent increase in H2-Kb cell surface levels along with the acquisition of antigen cross-presentation abilities. Mechanistically, such effects occurred in response to UM171a-mediated production of mitochondrial-derived reactive oxygen species as their neutralization using anti-oxidants or Antimycin-A mitigated MSCs' ability to cross-present antigens. Processing and presentation of the immunogenic ovalbumin-derived SIINFEKL peptide was caused by de novo expression of the Psmb8 gene in response to UM171a-triggered oxidative stress. When evaluated for their anti-tumoral properties in the context of therapeutic vaccination, UM171a-treated MSC administration to immunocompetent mice with pre-established T-cell lymphoma controlled tumor growth resulting in 40% survival without the need of additional supportive therapy and/or standard-of-care. CONCLUSIONS: Altogether, our findings reveal a new immune-related function for UM171a and clearly allude to a direct link between UM171a-mediated ROS induction and antigen cross-presentation by MSCs. The fact that UM171a treatment modulates MSCs to become antigen-presenting cells without the use of IFN-gamma opens-up a new line of investigation to search for additional agents capable of converting immune-suppressive MSCs to a cellular tool easily adaptable to vaccination.

Cell Therapy: Types, Regulation, and Clinical Benefits.

Cell therapy practices date back to the 19th century and continue to expand on investigational and investment grounds. Cell therapy includes stem cell- and non-stem cell-based, unicellular and multicellular therapies, with different immunophenotypic profiles, isolation techniques, mechanisms of action, and regulatory levels. Following the steps of their predecessor cell therapies that have become established or commercialized, investigational and premarket approval-exempt cell therapies continue to provide patients with promising therapeutic benefits in different disease areas. In this review article, we delineate the vast types of cell therapy, including stem cell-based and non-stem cell-based cell therapies, and create the first-in-literature compilation of the different "multicellular" therapies used in clinical settings. Besides providing the nuts and bolts of FDA policies regulating their use, we discuss the benefits of cell therapies reported in 3 therapeutic areas-regenerative medicine, immune diseases, and cancer. Finally, we contemplate the recent attention shift toward combined therapy approaches, highlighting the factors that render multicellular therapies a more attractive option than their unicellular counterparts.

Autologous BMAC Therapy Improves Spinal Degenerative Joint Disease in Lower Back Pain Patients.

Spinal degenerative joint disease (DJD) is associated with lower back pain (LBP) arising from the degeneration of intervertebral discs (IVD), facet joints, intertransversarii muscles, and interspinous ligaments among other anatomical structures. To circumvent the socioeconomic burdens and often-problematic surgical options imposed by DJD therapy, cell-based biologic modalities like bone marrow aspirate concentrate (BMAC) have been investigated in pre-clinical and clinical settings, mostly for IVD degeneration (IDD), with encouraging outcomes. In this study, we evaluated the differences in therapeutic benefits of BMAC between IVD- and facet joint-originating chronic LBP. Eighteen patients diagnosed with chronic LBP met the selection criteria. Following discography and provocation testing, 13 patients tested positive and were assigned into IDD-associated LBP (1st arm), while the remaining 5 tested negative and were assigned into facetogenic LBP (2nd arm). Autologous BMAC was injected intradiscally in the 1st arm, while the 2nd arm received posterior spinal chain injections. No procedure-related serious events ensued. Clinical improvement was evaluated over 12 months based on pain and functionality questionnaires (VAS, BPI, RAND-36), opioid use, and changes in disc parameters assessed by magnetic resonance imaging (MRI). Ameliorated VAS and BPI scores differed significantly between both arms in favor of IDD patients who also took significantly less opioids. Average RAND-36 scores showed no significant difference between groups albeit a trend suggesting improvement was observed in IDD patients. MRI scans conducted on IDD patients demonstrated marked elevation in disc height and spinal canal space size without worsening disc quality. Overall, this is the first study investigating the potency of BMAC as an IDD treatment in Canada and the first globally for addressing facetogenic pain using cellular therapy.

Engineering immunoproteasome-expressing mesenchymal stromal cells: A potent cellular vaccine for lymphoma and melanoma in mice.

Dendritic cells (DCs) excel at cross-presenting antigens, but their effectiveness as cancer vaccine is limited. Here, we describe a vaccination approach using mesenchymal stromal cells (MSCs) engineered to express the immunoproteasome complex (MSC-IPr). Such modification instills efficient antigen cross-presentation abilities associated with enhanced major histocompatibility complex class I and CD80 expression, de novo production of interleukin-12, and higher chemokine secretion. This cross-presentation capacity of MSC-IPr is highly dependent on their metabolic activity. Compared with DCs, MSC-IPr hold the ability to cross-present a vastly different epitope repertoire, which translates into potent re-activation of T cell immunity against EL4 and A20 lymphomas and B16 melanoma tumors. Moreover, therapeutic vaccination of mice with pre-established tumors efficiently controls cancer growth, an effect further enhanced when combined with antibodies targeting PD-1, CTLA4, LAG3, or 4-1BB under both autologous and allogeneic settings. Therefore, MSC-IPr constitute a promising subset of non-hematopoietic antigen-presenting cells suitable for designing universal cell-based cancer vaccines.

Integrative Transcriptome Analyses Empower the Anti-COVID-19 Drug Arsenal.

The beginning of the 21st century has been marked by three distinct waves of zoonotic coronavirus outbreaks into the human population. The COVID-19 (coronavirus disease 2019) pandemic is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and emerged as a global threat endangering the livelihoods of millions worldwide. Currently, and despite collaborative efforts, diverse therapeutic strategies from ongoing clinical trials are still debated. To address the need for such an immediate call of action, we leveraged the largest dataset of drug-induced transcriptomic perturbations, public SARS-CoV-2 transcriptomic datasets, and expression profiles from normal lung transcriptomes. Most importantly, our unbiased systems biology approach prioritized more than 50 repurposable drug candidates (e.g., corticosteroids, Janus kinase and Bruton kinase inhibitors). Further clinical investigation of these FDA-approved candidates as monotherapy or in combination with an antiviral regimen (e.g., remdesivir) could lead to promising outcomes in patients with COVID-19.

Intralesional Injection of Bone Marrow Aspirate Concentrate for the Treatment of Osteonecrosis of the Knee Secondary to Systemic Lupus Erythematosus: A Case Report.

Case: An 18-year-old female patient with Systemic Lupus Erythematosus (SLE) and corticosteroid-associated extensive bilateral symptomatic knee Osteonecrosis (ON) (Ficat IV), treated with sequential intralesional injections of autologous bone marrow aspirate concentrate (BMAC) under ultrasound guidance. At 3 months, pain was almost absent (VAS) and KOOS/WOMAC showed significant improvement sustained up to 24 months. At 12 months MRI indicated bone maturation, significantly reduced BM edema and subchondral fluid volume, and no collapse/fragmentation signs. Discussion: The clinical and imaging significant improvement observed in this patient suggests that BMAC intralesional injections effectively restored the compromised bone structure. After larger studies, this technique can become an alternative to decompressing surgery for ON cases.

Mesenchymal Stem Cells Beyond Regenerative Medicine.

Mesenchymal stem cells (MSCs) are competent suitors of cellular therapy due to their therapeutic impact on tissue degeneration and immune-based pathologies. Additionally, their homing and immunomodulatory properties can be exploited in cancer malignancies to transport pharmacological entities, produce anti-neoplastic agents, or induce anti-tumor immunity. Herein, we create a portfolio for MSC properties, showcasing their distinct multiple therapeutic utilities and successes/challenges thereof in both animal studies and clinical trials. We further highlight the promising potential of MSCs not only in cancer management but also in instigating tumor-specific immunity - i.e., cancer vaccination. Finally, we reflect on the possible reasons impeding the clinical advancement of MSC-based cancer vaccines to assist in contriving novel methodologies from which a therapeutic milestone might emanate.

Thymoproteasome-Expressing Mesenchymal Stromal Cells Confer Protective Anti-Tumor Immunity via Cross-Priming of Endogenous Dendritic Cells.

Proteasomes are complex macromolecular structures existing in various forms to regulate a myriad of cellular processes. Besides degrading unwanted or misfolded proteins (proteostasis), distinct immune functions were ascribed for the immunoproteasome and thymoproteasome (TPr) complexes. For instance, antigen degradation during ongoing immune responses mainly relies on immunoproteasome activity, whereas intrathymic CD8 T-cell development requires peptide generation by the TPr complex. Despite these substantial differences, the functional contribution of the TPr to peripheral T-cell immunity remains ill-defined. We herein explored whether the use of mesenchymal stromal cells (MSCs) engineered to exhibit altered proteasomal activity through de novo expression of the TPr complex can be exploited as a novel anti-cancer vaccine capable of triggering potent CD8 T-cell activation. Phenotypic and molecular characterization of MSC-TPr revealed a substantial decrease in MHCI (H2-Kb and H2-Dd) expression along with elevated secretion of various chemokines (CCL2, CCL9, CXCL1, LIX, and CX3CL1). In parallel, transcriptomic analysis pinpointed the limited ability of MSC-TPr to present endogenous antigens, which is consistent with their low expression levels of the peptide-loading proteins TAP, CALR, and PDAI3. Nevertheless, MSC-TPr cross-presented peptides derived from captured soluble proteins. When tested for their protective capacity, MSC-TPr triggered modest anti-tumoral responses despite efficient generation of effector memory CD4 and CD8 T cells. In contrast, clodronate administration prior to vaccination dramatically enhanced the MSC-TPr-induced anti-tumoral immunity clearly highlighting a refractory role mediated by phagocytic cells. Thus, our data elute to a DC cross-priming-dependant pathway in mediating the therapeutic effect of MSC-TPr.