Publications

J Clin Invest. 2018 Jun 1;128(6):2487-2499. doi: 10.1172/JCI88169.

Despite significant advances in the treatment of multiple myeloma (MM), most patients succumb to disease progression. One of the major immunosuppressive mechanisms that is believed to play a role in myeloma progression is the expansion of regulatory T cells (Tregs). In this study, we demonstrate that myeloma cells drive Treg expansion and activation by secreting type 1 interferon (IFN). Blocking IFN α and β receptor 1 (IFNAR1) on Tregs significantly decreases both myeloma-associated Treg immunosuppressive function and myeloma progression. Using syngeneic transplantable murine myeloma models and bone marrow (BM) aspirates of MM patients, we found that Tregs were expanded and activated in the BM microenvironment at early stages of myeloma development. Selective depletion of Tregs led to a complete remission and prolonged survival in mice injected with myeloma cells. Further analysis of the interaction between myeloma cells and Tregs using gene sequencing and enrichment analysis uncovered a feedback loop, wherein myeloma-cell-secreted type 1 IFN induced proliferation and expansion of Tregs. By using IFNAR1-blocking antibody treatment and IFNAR1-knockout Tregs, we demonstrated a significant decrease in myeloma-associated Treg proliferation, which was associated with longer survival of myeloma-injected mice. Our results thus suggest that blocking type 1 IFN signaling represents a potential strategy to target immunosuppressive Treg function in MM.

JCI Insight. 2017 Nov 2;2(21). pii: 91599. doi: 10.1172/jci.insight.91599.

Tregs hold great promise as a cellular therapy for multiple immunologically mediated diseases, given their ability to control immune responses. The success of such strategies depends on the expansion of healthy, suppressive Tregs ex vivo and in vivo following the transfer. In clinical studies, levels of transferred Tregs decline sharply in the blood within a few days of the transfer. Tregs have a high rate of apoptosis. Here, we describe a new mechanism of Treg self-inflicted damage. We show that granzymes A and -B (GrA and GrB), which are highly upregulated in human Tregs upon stimulation, leak out of cytotoxic granules to induce cleavage of cytoplasmic and nuclear substrates, precipitating apoptosis in target cells. GrA and GrB substrates were protected from cleavage by inhibiting granzyme activity in vitro. Additionally, we show – by using cytometry by time of flight (CYTOF) – an increase in GrB-expressing Tregs in the peripheral blood and renal allografts of transplant recipients undergoing rejection. These GrB-expressing Tregs showed an activated phenotype but were significantly more apoptotic than non-GrB expressing Tregs. This potentially novel finding improves our understanding of Treg survival and suggests that manipulating Gr expression or activity might be useful for designing more effective Treg therapies.

ACS Nano. 2017 Nov 28;11(11):11041-11046. doi: 10.1021/acsnano.7b05083. Epub 2017 Oct 25.

Kidney transplant patients require life-long surveillance to detect allograft rejection. Repeated biopsy, albeit the clinical gold standard, is an invasive procedure with the risk of complications and comparatively high cost. Conversely, serum creatinine or urinary proteins are noninvasive alternatives but are late markers with low specificity. We report a urine-based platform to detect kidney transplant rejection. Termed iKEA (integrated kidney exosome analysis), the approach detects extracellular vesicles (EVs) released by immune cells into urine; we reasoned that T cells, attacking kidney allografts, would shed EVs, which in turn can be used as a surrogate marker for inflammation. We optimized iKEA to detect T-cell-derived EVs and implemented a portable sensing system. When applied to clinical urine samples, iKEA revealed high level of CD3-positive EVs in kidney rejection patients and achieved high detection accuracy (91.1%). Fast, noninvasive, and cost-effective, iKEA could offer new opportunities in managing transplant recipients, perhaps even in a home setting.

Cell Rep. 2016 May 10;15(6):1202-13. doi: 10.1016/j.celrep.2016.04.007. Epub 2016 Apr 28.

Active-targeted delivery to lymph nodes represents a major advance toward more effective treatment of immune-mediated disease. The MECA79 antibody recognizes peripheral node addressin molecules expressed by high endothelial venules of lymph nodes. By mimicking lymphocyte trafficking to the lymph nodes, we have engineered MECA79-coated microparticles containing an immunosuppressive medication, tacrolimus. Following intravenous administration, MECA79-bearing particles showed marked accumulation in the draining lymph nodes of transplanted animals. Using an allograft heart transplant model, we show that targeted lymph node delivery of microparticles containing tacrolimus can prolong heart allograft survival with negligible changes in tacrolimus serum level. Using MECA79 conjugation, we have demonstrated targeted delivery of tacrolimus to the lymph nodes following systemic administration, with the capacity for immune modulation in vivo.

Nat Commun. 2017 Nov 22;8(1):1692. doi: 10.1038/s41467-017-01760-5.

Proteasome inhibitors benefit patients with multiple myeloma and B cell-dependent autoimmune disorders but exert toxicity from inhibition of proteasomes in other cells. Toxicity should be minimized by reversible inhibition of the immunoproteasome β5i subunit while sparing the constitutive β5c subunit. Here we report β5i-selective inhibition by asparagine-ethylenediamine (AsnEDA)-based compounds and present the high-resolution cryo-EM structural analysis of the human immunoproteasome. Despite inhibiting noncompetitively, an AsnEDA inhibitor binds the active site. Hydrophobic interactions are accompanied by hydrogen bonding with β5i and β6 subunits. The inhibitors are far more cytotoxic for myeloma and lymphoma cell lines than for hepatocarcinoma or non-activated lymphocytes. They block human B-cell proliferation and promote apoptotic cell death selectively in antibody-secreting B cells, and to a lesser extent in activated human T cells. Reversible, β5i-selective inhibitors may be useful for treatment of diseases involving activated or neoplastic B cells or activated T cells.