Our laboratory has developed a process for generating mAbs with selectivity to unique peptides in the context of MHC molecules. Recently, we reported that RL4B, an mAb that we have called a TCR mimic (TCRm) because it recognizes peptide in the context of MHC, has cytotoxic activity in vitro and prevented growth of tumor cells in a prophylactic setting. When presented in the context of HLA-A2, RL4B TCRm recognizes the peptide GVLPALPQV derived from human chorionic gonadotropin (hCG)-β. In this study, we show that RL4B TCRm has strong binding affinity for the GVLPALPQV peptide/HLA-A2 epitope and fine binding specificity for cells that express endogenous hCGβ Ag and HLA-A2. In addition, suppression of tumor growth with RL4B TCRm was observed in orthotopic models for breast cancer. Using two aggressive human tumor cell lines, MDA-MB-231 and MCF-7, we provide evidence that RL4B TCRm significantly retards tumor growth, supporting a possible role for TCRm agents in therapeutic settings. Moreover, tumors in mice responded to RL4B TCRm therapy in a dose-dependent manner, eliminating tumors at the highest dose. RL4B TCRm strongly detects the hCGβ peptide/HLA-A2 epitope in human primary breast tumor tissue, but does not react or reacts weakly with normal breast tissue from the same patient. These results further illustrate the selective nature of TCRm Abs and the clinical relevance of the GVLPALPQV peptide/HLA-A2 epitope expression in tumor cells, because they provide the first evidence that Abs that mimic the TCR can be used to markedly reduce and suppress tumor growth.

mAbs that recognize peptides presented on the cell surface by MHC class I molecules are potential therapeutic agents for cancer therapy. We have previously demonstrated that these Abs, which we termed TCR mimic mAbs (TCRm), reduce tumor growth in models of breast carcinoma. However, mechanisms of TCRm-mediated tumor growth reduction remain largely unknown. In this study, we report that these Abs, in contrast to several mAbs used currently in the clinic, destroy tumor cells independently of immune effector mechanisms such as Ab-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). We found that TCRm-mediated apoptosis of tumor cells was associated with selective and specific binding of these Abs to peptide/HLA class I complexes, which triggered the activation of JNK and intrinsic caspase pathways. This signaling was accompanied by the release of mitochondrial cytochrome c and apoptosis-inducing factor. TCRm-induced apoptosis in tumor cells was completely inhibited by soluble MHC tetramers loaded with relevant peptide as well as with inhibitors for JNK and caspases. Furthermore, mAbs targeting MHC class I, independent of the peptide bound by HLA, did not stimulate apoptosis, suggesting that the Ab-binding site on the MHC/peptide complex determines cytotoxicity. This study suggests the existence of mechanisms, in addition to ADCC and CDC, through which these therapeutic Abs destroy tumor cells. These mechanisms would appear to be of particular importance in severely immunocompromised patients with advanced neoplastic disease, since immune cell-mediated killing of tumor cells through ADCC and CDC is substantially limited in these individuals.

No abstract

Abstract. One 2-year-old, 7.5 months pregnant Aberdeen Angus out of a herd of 100 apparently healthy cows, died within 10 hours of hospitalization. At necropsy, multiple foci of mucosal hemorrhage and ulceration were observed in the spiral colon and cecum. Virus isolation from intestinal lesions yielded a cytopathic virus, which was revealed by electron microscopy to be an approximately 27 nm, nonenveloped virus. Further characterization by reverse transcription-polymerase chain reaction (RT-PCR), sequencing of the 5'UTR and partial VP1 coding region, and phylogenetic analysis classified the virus isolate as bovine enterovirus type 1 (BEV-1). No other significant pathogens were detected. This is the first report of BEV-1 isolated in the USA from an animal with fatal enteric disease in more than 20 years. Further investigation is required to determine the prevalence of BEV in North America and to establish the clinical relevance of this understudied virus.Key words: BEV; BEV-Oklahoma; Bovine; enterovirus; picornavirus; typhlocolitis.Bovine enterovirus (BEV) belongs to the family Picornaviridae (picornaviruses), which consists of small (18-30 nm), nonenveloped viruses with an icosahedral capsid that encloses a single copy of positive-sense RNA genome. BEV is in the genus Enterovirus, along with poliovirus, human enterovirus, coxsackieviruses, swine vesicular disease virus, echovirus 11, and others. Originally classified into several serotypes, only 2 serotypes, BEV-1 and BEV-2, are now recognized. 10,15,18,19 Because of the unavailability of type specific antisera or a commercially available diagnostic test, a genotypic classification, which supports previous recognized serological distinctions, has been proposed. 7 Despite the large volume of information available on other enteroviruses, very little documentation exists on the pathogenesis of BEV infections in cattle or on its prevalence in North America. Several case reports in the 1950s and 1970s document the isolation of BEV from feces and various tissues from apparently healthy animals or from animals with clinical signs that ranged from mild to moderate diarrhea to reproductive disease. 3,4,14,20 However, these older reports are difficult to interpret as they relied solely on serological assays or had identified more than one infectious agent. This report describes the isolation and partial characterization of BEV-1 from a fatal enteric disease case in a heifer. A 2-year-old, 455 kg, 7.5 months pregnant Aberdeen Angus heifer was presented to the Boren Veterinary Medical Teaching Hospital of Oklahoma State University with a 5-hour history of progressive abdominal pain of sudden onset. According to the owner, the heifer was showing signs of colic by kicking at her abdomen and repeatedly getting up and down; she had been acting and eating normally the day prior to presentation. The animal had been raised on the farm and considered healthy until the episode of colic. Her vaccination (CattleMasterH 4 + VL5: IBR-BVD-PI3-BRSV-Campylobacter fetus-Leptospira canicola...

Abstract. The brain from a 15-month-old, black female Angus, with a 48-hour history of central nervous system disease, was submitted to the Oklahoma Animal Disease Diagnostic Laboratory. Microscopic findings consisted of acute, multifocal meningoencephalitis, with neuronal degeneration and necrosis and gliosis. Viral isolation yielded noncytopathic bovine viral diarrhea virus (BVDV). Virus genotyping classified the virus as BVDV type 2. Immunohistochemical labeling for BVDV antigens with BVD MAb 3.12F1 clone was prominent in the cytoplasm of neurons, glial cells, ependymal epithelium, perivascular macrophages and spindle cells, smooth muscle cells, and intravascular monocytes of the cerebrum and brain stem. Laboratory results support that tissue alterations occurred as a result of BVDV type 2 infection. In the absence of other clinical signs related to BVDV infection and using the microscopic and laboratory evidence presented, we propose that the BVDV type 2 isolated from this case may represent a neurovirulent strain of the virus. To the best of our knowledge, this is the first report of brain lesions and neuronal viral antigen localization in BVDV genotype 2 viral infection, acquired either congenitally or postnatally.