Development of Allogeneic Stem Cell-Based Platform for Delivery and Potentiation of Oncolytic Virotherapy.

We describe the repurposing and optimization of the TK-positive (thymidine kinase) vaccinia virus strain ACAM1000/ACAM2000™ as an oncolytic virus. This virus strain has been widely used as a smallpox vaccine and was also used safely in our recent clinical trial in patients with advanced solid tumors and Acute Myeloid Leukemia (AML). The vaccinia virus was amplified in CV1 cells and named CAL1.

Invariant NKT cell-augmented GM-CSF-secreting tumor vaccine is effective in advanced prostate cancer model.

Invariant natural killer T cells (iNKT cells) express a semi-invariant T cell receptor that recognizes certain glycolipids (including α-galactosylceramide, αGC) bound to CD1d, and can induce potent antitumor responses. Here, we assessed whether αGC could enhance the efficacy of a GM-CSF-producing tumor cell vaccine in the transgenic SV40 T antigen-driven TRAMP prostate cancer model. In healthy mice, we initially found that optimal T cell responses were obtained with αGC-pulsed TRAMP-C2 cells secreting GM-CSF and milk fat globule epidermal growth factor protein-8 (MFG-E8) with an RGD to RGE mutation (GM-CSF/RGE TRAMP-C2), combined with systemic low dose IL-12.

Ivermectin converts cold tumors hot and synergizes with immune checkpoint blockade for treatment of breast cancer.

We show that treatment with the FDA-approved anti-parasitic drug ivermectin induces immunogenic cancer cell death (ICD) and robust T cell infiltration into breast tumors. As an allosteric modulator of the ATP/P2X4/P2X7 axis which operates in both cancer and immune cells, ivermectin also selectively targets immunosuppressive populations including myeloid cells and Tregs, resulting in enhanced Teff/Tregs ratio. While neither agent alone showed efficacy in vivo, combination therapy with ivermectin and checkpoint inhibitor anti-PD1 antibody achieved synergy in limiting tumor growth (p = 0.

Purinergic Signaling Within the Tumor Microenvironment.

Accumulating studies have clearly demonstrated high concentrations of extracellular ATP (eATP) within the tumor microenvironment (TME). Implications of these findings are multifold as ATP-mediated purinergic signaling has been shown to mediate a variety of cancer-related processes, including cell migration, resistance to cytotoxic therapy, and immune regulation. Broad roles of ATP within the tumor microenvironment are linked to the abundance of ATP-regulated purinergic receptors on cancer and stromal and various immune cell types, as well as on the importance of ATP release and signaling in the regulation of multiple cellular processes.

Consensus guidelines for the definition, detection and interpretation of immunogenic cell death.

Cells succumbing to stress via regulated cell death (RCD) can initiate an adaptive immune response associated with immunological memory, provided they display sufficient antigenicity and adjuvanticity. Moreover, multiple intracellular and microenvironmental features determine the propensity of RCD to drive adaptive immunity. Here, we provide an updated operational definition of immunogenic cell death (ICD), discuss the key factors that dictate the ability of dying cells to drive an adaptive immune response, summarize experimental assays that are currently available for the assessment of ICD in vitro and in vivo, and formulate guidelines for their interpretation.

First-in-human study of TK-positive oncolytic vaccinia virus delivered by adipose stromal vascular fraction cells.

Background: ACAM2000, a thymidine kinase (TK)-positive strain of vaccinia virus, is the current smallpox vaccine in the US. Preclinical testing demonstrated potent oncolytic activity of ACAM2000 against several tumor types. This Phase I clinical trial of ACAM2000 delivered by autologous adipose stromal vascular fraction (SVF) cells was conducted to determine the safety and feasibility of such a treatment in patients with advanced solid tumors or acute myeloid leukemia (AML).

Correction to: Toward a comprehensive view of cancer immune responsiveness: a synopsis from the SITC workshop.

Following publication of the original article [1], the author reported that an author name, Roberta Zappasodi, was missed in the authorship list.

Toward a comprehensive view of cancer immune responsiveness: a synopsis from the SITC workshop.

Tumor immunology has changed the landscape of cancer treatment. Yet, not all patients benefit as cancer immune responsiveness (CIR) remains a limitation in a considerable proportion of cases. The multifactorial determinants of CIR include the genetic makeup of the patient, the genomic instability central to cancer development, the evolutionary emergence of cancer phenotypes under the influence of immune editing, and external modifiers such as demographics, environment, treatment potency, co-morbidities and cancer-independent alterations including immune homeostasis and polymorphisms in the major and minor histocompatibility molecules, cytokines, and chemokines.

Delivery of oncolytic vaccinia virus by matched allogeneic stem cells overcomes critical innate and adaptive immune barriers.

Background: Previous studies have identified IFNγ as an important early barrier to oncolytic viruses including vaccinia. The existing innate and adaptive immune barriers restricting oncolytic virotherapy, however, can be overcome using autologous or allogeneic mesenchymal stem cells as carrier cells with unique immunosuppressive properties.

Methods: To test the ability of mesenchymal stem cells to overcome innate and adaptive immune barriers and to successfully deliver oncolytic vaccinia virus to tumor cells, we performed flow cytometry and virus plaque assay analysis of ex vivo co-cultures of stem cells infected with vaccinia virus in the presence of peripheral blood mononuclear cells from healthy donors.

The ratio of ADSCs to HSC-progenitors in adipose tissue derived SVF may provide the key to predict the outcome of stem-cell therapy.

Background: Stromal vascular fraction (SVF) represents an attractive source of adult stem cells and progenitors, holding great promise for numerous cell therapy approaches. In 2017, it was reported that 1524 patients received autologous SVF following the enzymatic digestion of liposuction fat. The treatment was safe and effective and patients showed significant clinical improvement.

Modulation of P2X4/P2X7/Pannexin-1 sensitivity to extracellular ATP via Ivermectin induces a non-apoptotic and inflammatory form of cancer cell death.

Overexpression of P2X7 receptors correlates with tumor growth and metastasis. Yet, release of ATP is associated with immunogenic cancer cell death as well as inflammatory responses caused by necrotic cell death at sites of trauma or ischemia-reperfusion injury. Using an FDA-approved anti-parasitic agent Ivermectin as a prototype agent to allosterically modulate P2X4 receptors, we can switch the balance between the dual pro-survival and cytotoxic functions of purinergic signaling in breast cancer cells.

IAP inhibitors enhance co-stimulation to promote tumor immunity.

The inhibitor of apoptosis proteins (IAPs) have recently been shown to modulate nuclear factor κB (NF-κB) signaling downstream of tumor necrosis factor (TNF) family receptors, positioning them as essential survival factors in several cancer cell lines, as indicated by the cytotoxic activity of several novel small molecule IAP antagonists. In addition to roles in cancer, increasing evidence suggests that IAPs have an important function in immunity; however, the impact of IAP antagonists on antitumor immune responses is unknown. In this study, we examine the consequences of IAP antagonism on T cell function in vitro and in the context of a tumor vaccine in vivo.

Milk fat globule EGF-8 promotes melanoma progression through coordinated Akt and twist signaling in the tumor microenvironment.

The pathogenesis of malignant melanoma involves the interplay of tumor cells with normal host elements, but the underlying mechanisms are incompletely understood. Here, we show that milk fat globule EGF-8 (MFG-E8), a secreted protein expressed at high levels in the vertical growth phase of melanoma, promotes disease progression through coordinated alpha(v)beta(3) integrin signaling in the tumor microenvironment. In a murine model of melanoma, MFG-E8 enhanced tumorigenicity and metastatic capacity through Akt-dependent and Twist-dependent pathways.

Block of T cell development in P53-deficient mice accelerates development of lymphomas with characteristic RAG-dependent cytogenetic alterations.

Mice deficient in the DNA damage sensor P53 display normal T cell development but eventually succumb to thymic lymphomas. Here, we show that inactivation of the TCR beta gene enhancer (E beta) results in a block of T cell development at stages where recombination-activating genes (RAG) are expressed. Introduction of the E beta mutation into p53-/- mice dramatically accelerates the onset of lethal thymic lymphomas that harbor RAG-dependent aberrant rearrangements, chromosome 14 and 12 translocations, and amplification of the chromosomal region 9A1-A5.

The T-cell receptor beta variable gene promoter is required for efficient V beta rearrangement but not allelic exclusion.

To investigate the role of promoters in regulating variable gene rearrangement and allelic exclusion, we constructed mutant mice in which a 1.2-kb region of the V beta 13 promoter was either deleted (P13(-/-)) or replaced with the simian virus 40 minimal promoter plus five copies of Gal4 DNA sequences (P13(R/R)). In P13(-/-) mice, cleavage, rearrangement, and transcription of V beta 13, but not the flanking V beta gene segments, were significantly inhibited.

The T cell receptor beta enhancer promotes access and pairing of Dbeta and Jbeta gene segments during V(D)J recombination.

The precise function of cis elements in regulating V(D)J recombination is still controversial. Here, we determined the effect of inactivation of the TCRbeta enhancer (Ebeta) on cleavage and rearrangement of Dbeta1, Dbeta2, Jbeta1, and Jbeta2 gene segments in CD4-CD8- [double-negative (DN)] and CD4+CD8+ [double-positive (DP)] thymocytes. In Ebeta-deficient mice, (i) Dbeta1 rearrangements were more severely impaired than Dbeta2 rearrangements; (ii) most of the Dbeta and Jbeta cleavages and rearrangements occurred in DP, rather than in DN, thymocytes; and (iii) most of the 3' Dbeta1 cleavages were coupled to 5' Dbeta2 cleavages instead of to Jbeta cleavages, resulting in nonstandard Dbeta1-Dbeta2-Jbeta2 joints.