Development and Preliminary Clinical Activity of PD-1-Guided CTLA-4 Blocking Bispecific DART Molecule.

Combination immunotherapy with antibodies directed against PD-1 and CTLA-4 shows improved clinical benefit across cancer indications compared to single agents, albeit with increased toxicity. Leveraging the observation that PD-1 and CTLA-4 are co-expressed by tumor-infiltrating lymphocytes, an investigational PD-1 x CTLA-4 bispecific DART molecule, MGD019, is engineered to maximize checkpoint blockade in the tumor microenvironment via enhanced CTLA-4 blockade in a PD-1-binding-dependent manner. In vitro, MGD019 mediates the combinatorial blockade of PD-1 and CTLA-4, confirming dual inhibition via a single molecule. MGD019 is well tolerated in non-human primates, with evidence of both PD-1 and CTLA-4 blockade, including increases in Ki67+CD8 and ICOS+CD4 T cells, respectively. In the ongoing MGD019 first-in-human study enrolling patients with advanced solid tumors (NCT03761017), an analysis undertaken following the dose escalation phase revealed acceptable safety, pharmacodynamic evidence of combinatorial blockade, and objective responses in multiple tumor types typically unresponsive to checkpoint inhibitor therapy.

Development of MGD007, a gpA33 x CD3-Bispecific DART Protein for T-Cell Immunotherapy of Metastatic Colorectal Cancer.

We have developed MGD007 (anti-glycoprotein A33 x anti-CD3), a DART protein designed to redirect T cells to target gpA33 expressing colon cancer. The gpA33 target was selected on the basis of an antibody-based screen to identify cancer antigens universally expressed in both primary and metastatic colorectal cancer specimens, including putative cancer stem cell populations. MGD007 displays the anticipated-bispecific binding properties and mediates potent lysis of gpA33-positive cancer cell lines, including models of colorectal cancer stem cells, through recruitment of T cells. Xenograft studies showed tumor growth inhibition at doses as low as 4 µg/kg. Both CD8 and CD4 T cells mediated lysis of gpA33-expressing tumor cells, with activity accompanied by increases in granzyme and perforin. Notably, suppressive T-cell populations could also be leveraged to mediate lysis of gpA33-expressing tumor cells. Concomitant with CTL activity, both T-cell activation and expansion are observed in a gpA33-dependent manner. No cytokine activation was observed with human PBMC alone, consistent with the absence of gpA33 expression on peripheral blood cell populations. Following prolonged exposure to MGD007 and gpA33 positive tumor cells, T cells express PD-1 and LAG-3 and acquire a memory phenotype but retain ability to support potent cell killing. In cynomolgus monkeys, 4 weekly doses of 100 µg/kg were well tolerated, with prolonged PK consistent with that of an Fc-containing molecule. Taken together, MGD007 displays potent activity against colorectal cancer cells consistent with a mechanism of action endowed in its design and support further investigation of MGD007 as a potential novel therapeutic treatment for colorectal cancer. Mol Cancer Ther; 17(8); 1761-72. ©2018 AACR.

Identification of an autoantibody panel to separate lung cancer from smokers and nonsmokers.

BACKGROUND: Sera from lung cancer patients contain autoantibodies that react with tumor associated antigens (TAAs) that reflect genetic over-expression, mutation, or other anomalies of cell cycle, growth, signaling, and metabolism pathways. METHODS: We performed immunoassays to detect autoantibodies to ten tumor associated antigens (TAAs) selected on the basis of previous studies showing that they had preferential specificity for certain cancers. Sera examined were from lung cancer patients (22); smokers with ground-glass opacities (GGOs) (46), benign solid nodules (55), or normal CTs (35); and normal non-smokers (36). Logistic regression models based on the antibody biomarker levels among the high risk and lung cancer groups were developed to identify the combinations of biomarkers that predict lung cancer in these cohorts. RESULTS: Statistically significant differences in the distributions of each of the biomarkers were identified among all five groups. Using Receiver Operating Characteristic (ROC) curves based on age, c-myc, Cyclin A, Cyclin B1, Cyclin D1, CDK2, and survivin, we obtained a sensitivity = 81% and specificity = 97% for the classification of cancer vs smokers(no nodules, solid nodules, or GGO) and correctly predicted 31/36 healthy controls as noncancer. CONCLUSION: A pattern of autoantibody reactivity to TAAs may distinguish patients with lung cancer versus smokers with normal CTs, stable solid nodules, ground glass opacities, or normal healthy never smokers.

Inositol 1,4,5-trisphosphate 3-kinase B is a negative regulator of BCR signaling that controls B cell selection and tolerance induction.

Inositol 1,4,5-trisphosphate 3-kinase B (or Itpkb) converts inositol 1,4,5-trisphosphate to inositol 1,3,4,5-tetrakisphosphate upon Ag receptor activation and controls the fate and function of lymphocytes. To determine the role of Itpkb in B cell tolerance, Itpkb(-/-) mice were crossed to transgenic mice that express a BCR specific for hen egg lysozyme (IgHEL). B cells from Itpkb(-/-) IgHEL mice possess an anergic phenotype, hypoproliferate in response to cognate Ag, and yet they exhibit enhanced Ag-induced calcium signaling. In IgHEL transgenic mice that also express soluble HEL, lack of Itpkb converts anergy induction to deletion. These data establish Itpkb as a negative regulator of BCR signaling that controls the fate of developing B cells and tolerance induction.

Preferential humoral immune response in prostate cancer to cellular proteins p90 and p62 in a panel of tumor-associated antigens.

BACKGROUND: Cytoplasmic p90 autoantigen was recently cloned from a cDNA expression library using serum antibody from a cancer patient. The humoral immune response to p90 in prostate cancer and benign prostatic hyperplasia (BPH) was examined. METHODS: An antigenic fragment of recombinant p90 protein and several other tumor-associated antigens (TAAs) were used in ELISA and Western blotting to detect antibodies in sera from patients with prostate cancer, BPH, and other controls. RESULTS: Autoantibodies to p90 were detected in 30.8% of 133 prostate cancer patients versus 1.5% in 68 BPH patients. When a selected panel of six TAAs including p90 were used for immunoscreening, the cumulative positive reactions in prostate cancer sera reached 92.5%, significantly higher than in BPH and other control sera. Antibodies to p90 showed the highest frequency in prostate cancer (30.8%), followed by antibodies to p62 (22.6%). CONCLUSIONS: A panel of six selected TAAs was shown to have high sensitivity and specificity as immunodiagnostic markers in prostate cancer.

Effect of combined T- and B-cell depletion of allogeneic HLA-mismatched bone marrow graft on the magnitude and kinetics of Epstein-Barr virus load in the peripheral blood of bone marrow transplant recipients.

Recipients of T-cell-depleted bone marrow (BM) transplants (BMT) frequently develop Epstein-Barr virus (EBV)-associated post-transplant lymphoproliferative disease (PTLD) preceded by a rapid and prominent increase of EBV load in the peripheral blood. The level of this increase positively correlates with the incidence of PTLD. Using a semiquantitative PCR assay we compared the blood levels of EBV-DNA in patients transplanted with either T-cell or T- and B-cell-depleted human leukocyte antigen (HLA)-mismatched BM grafts. Combined T- and B-cell depletion correlated with significantly lower maximal levels of EBV load, which were reached with slower kinetics. These data indicate that B-cell depletion of BM can be used for prophylaxis of PTLD in BM transplant recipients and can affect the long-term balance between EBV and its host.

Epstein-Barr virus inhibits the development of dendritic cells by promoting apoptosis of their monocyte precursors in the presence of granulocyte macrophage-colony-stimulating factor and interleukin-4.

Epstein-Barr virus (EBV) is a tumorigenic human herpesvirus that persists for life in healthy immunocompetent carriers. The viral strategies that prevent its clearance and allow reactivation in the face of persistent immunity are not well understood. Here we demonstrate that EBV infection of monocytes inhibits their development into dendritic cells (DCs), leading to an abnormal cellular response to granulocyte macrophage-colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4) and to apoptotic death. This proapoptotic activity was not affected by UV inactivation and was neutralized by EBV antibody-positive human sera, indicating that binding of the virus to monocytes is sufficient to alter their response to the cytokines. Experiments with the relevant blocking antibodies or with mutated EBV strains lacking either the EBV envelope glycoprotein gp42 or gp85 demonstrated that interaction of the trimolecular gp25-gp42-gp85 complex with the monocyte membrane is required for the effect. Our data provide the first evidence that EBV can prevent the development of DCs through a mechanism that appears to bypass the requirement for viral gene expression, and they suggest a new strategy for interference with the function of DCs during the initiation and maintenance of virus-specific immune responses.