Targeted T cells are emerging as effective non-toxic therapies for cancer. Multiple elements, however, contribute to the overall pathogenesis of cancer through both distinct and redundant mechanisms. Hence, targeting multiple cancer-specific markers simultaneously could result in better therapeutic efficacy. We created a functional chimeric antigen receptor—the TanCAR, a novel artificial molecule that mediates bispecific activation and targeting of T cells. We demonstrate the feasibility of cumulative integration of structure and docking simulation data using computational tools to interrogate the design and predict the functionality of such a complex bispecific molecule. Our prototype TanCAR induced distinct T cell reactivity against each of two tumor restricted antigens, and produced synergistic enhancement of effector functions when both antigens were simultaneously encountered. Furthermore, the TanCAR preserved the cytolytic ability of T cells upon loss of one of the target molecules and better controlled established experimental tumors by recognition of both targets in an animal disease model. This proof-of-concept approach can be used to increase the specificity of effector cells for malignant versus normal target cells, to offset antigen escape or to allow for targeting the tumor and its microenvironment.

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Glioblastoma (GBM) is the most common primary brain cancer in adults and is virtually incurable. Recent studies have shown that CMV is present in the majority of GBMs. To evaluate if the CMV antigens pp65 and IE1, which are expressed in GBMs, can be targeted with CMV-specific T cells, we measured the frequency of T cells targeting pp65 and IE1 in the peripheral blood of a cohort of 11 sequentially-diagnosed CMV-seropositive GBM patients, and evaluated whether it was feasible to expand autologous CMV-specific T cells for future clinical studies. All 11 CMV-seropositive GBM patients had T cells specific for pp65 and IE1 in their peripheral blood assessed by IFNγ ELIspot assay. However, the precursor frequency of pp65-specific T cells was decreased in comparison to healthy donors (p=0.001). We successfully reactivated and expanded CMV-specific T cells from 6 out of 6 GBM patients using antigen presenting cells transduced with an adenoviral vector encoding pp65 and IE1. CMV-specific T-cell lines contained CD4-positive as well as CD8-positive T cells, recognized pp65- and IE1-positive targets and killed CMV-infected autologous GBM cells. Infusion of such CMV-specific T-cell lines may extend the benefits of T-cell therapy to patients with CMV-positive GBMs.

Primary cutaneous Ewing sarcoma is a very rare entity with less than 100 cases reported in the literature, sharing the same morphological and immunohistochemical characteristics as their osseous counterparts. Herein, to the best of our knowledge, we report the first case in English literature of a molecularly confirmed Ewing sarcoma with diffuse and strong SOX10 immunoreactivity. This exceedingly rare immunohistochemical finding along with the rarity of this tumor could easily lead to a misdiagnosis with significant repercussions. Our case highlights the difficulty in diagnosing primary cutaneous Ewing sarcoma as well as the pivotal role molecular diagnostics can play in specific scenarios.

Bispecific T cell engagers have demonstrated clinical efficacy; however, their use can be accompanied by severe toxicity. Mechanistic understanding of these toxicities is limited by a lack of suitable immunocompetent preclinical models. In this study, we describe an immunocompetent mouse tumor model that exhibits bispecific T cell engager-induced toxicity and recapitulates key features similar to those in human cytokine release syndrome. In this study, toxicity occurred between the second and fourth injections of an NK Group 2D bispecific T cell engager protein. Symptoms were transient, peaking 3-4 h after treatment and resolving by 8 h. Mice developed weight loss, elevated plasma cytokines, a significant reduction in spleen white pulp, and lymphocyte infiltration in the liver. Systemic cellular immune changes also occurred; notably, an increase in CD8 + T cell activation, an increase in myeloid cells in the blood, and a population of Ly-6C int monocytes (CD11b + Ly-6G 2 F4/80 2 ) emerged in the liver and spleens of bispecific protein-treated mice. IFN-g was primarily produced by CD8 + T cells in the spleen and was required for the observed changes in both T cell and myeloid populations. Rag deficiency, IFN-g deficiency, or depletion of either CD4 + or CD8 + T cells prevented toxicity, whereas perforin deficiency, GM-CSF deficiency, or modulation of the myeloid population through clodronate-mediated depletion showed a partial abrogation of toxicity. Together, these findings reveal that T cell activation by a bispecific T cell engager leads to changes in the host myeloid cell population, both of which contribute to treatment induced toxicity in immunocompetent mice.

The Editors posted an Expression of Concern for this article following notification that two images in Figure 9D were subsequently published as distinct samples in another paper ( 1). An institutional review of the primary data supports that the images in the JCI article are correct and that no corrective action is required.

Metanephric adenoma (MA) is a rare benign renal neoplasm that shares morphologic and immunophenotypic overlap with epithelial predominant Wilms tumor (e-WT) and with the solid variant of papillary renal cell carcinoma (s-PRCC). Cadherin 17 (CDH17) is expressed primarily in the normal intestine and digestive tract tumors and has not been detected in tumors from other sites including the kidney. We investigated the diagnostic utility of CDH17 in differentiating between MA, e-WT, and s-PRCC. Immunohistochemistry for CDH17, CD57, AMACR, WT-1, and CDX2 was performed on 17 e-WTs, 15 s-PRCCs and 21 MAs and assessed based on a combined score of extent and intensity. Normal adult kidney parenchyma was negative for CDH17 staining. CDH17 was expressed in the late stages of fetal kidney development at the junction of the glomerular space and proximal nephron. The majority of MAs (81%) demonstrated membranous CDH17 immunoreactivity in all components (acinar, tubular, and papillary), while all cases of e-WTs and s-PRCCs were negative (p<0.0001). WT-1 was negative in s-PRCC and was positive in all cases of e-WT and MA. All MAs were strongly positive for CD57; however, this marker was also moderate to strongly positive in 6 (35%) e-WTs and 2 (13%) s-PRCCs. AMACR was strongly positive in all s-PRCCs, but moderate reactivity was seen in 3 (17%) e-WTs and 2 MA (10%). CDH17 is a sensitive (81%) and highly specific (100%) marker for MA, and should be considered in the IHC panel for distinguishing MA from its mimics.

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