Biomarkers for Predicting Abiraterone Treatment Outcome and Selecting Alternative Therapies in Castration-Resistant Prostate Cancer.

Approximately one-third of patients with metastatic castration-resistant prostate cancer (CRPC) exhibited primary abiraterone resistance. To identify alternative treatment for abiraterone nonresponders, we performed drug discovery analyses using the L1000 database using differentially expressed genes identified in tumor biopsies and patient-derived xenograft (PDX) tumors between abiraterone responders and nonresponders enrolled in PROMOTE trial. This approach identified 3 drugs, including topoisomerase II (TOP2) inhibitor mitoxantrone, CDK4/6 inhibitor palbociclib, and pan-CDK inhibitor PHA-793887. These drugs significantly suppressed the growth of abiraterone-resistant cell lines and PDX models. Moreover, we identified 11 genes targeted by all 3 drugs that were associated with worse outcomes in both the PROMOTE and Stand Up To Cancer cohorts. This 11-gene panel might also function as biomarkers to select the 3 alternative therapies for this subgroup of patients with CRPC, warranting further clinical investigation.

Retrospective Review of Surgical Outcomes and Pair-housing Success in Vasectomized Rhesus Macaques (Macaca mulatta).

Providing social housing for adult male macaques can be challenging. One successful strategy for long-term social housing of adult male macaques is to pair them with adult females; however, unwanted breeding must be prevented by sterilization of the male or female. Vasectomy is a simple, highly effective, and minimally invasive method of contraception that is used at our institution to facilitate social housing. We performed a retrospective review to analyze the surgical outcomes and rate of postoperative complications after vasectomy of adult rhesus macaques at our research facility. In addition, we evaluated the success rate of pairing vasectomized macaques with female partners. Over 10 y, 16 macaques were vasectomized, of which 5 developed postoperative complications such as orchitis, epididymitis, or surgical site infection. These complications resolved completely and without incident after antibiotic and analgesic therapy; an additional male had postoperative incisional swelling that resolved quickly after NSAID treatment. This complication rate is consistent with that in humans by surgeons who perform open vasectomies relatively infrequently. In addition, 5 of the vasectomized macaques (31%) developed sperm granulomas, which are a common and generally benign complication in humans and have been reported to develop in 40% of macaques after vasectomy. Successful pair housing with a female partner was achieved for 13 of 16 (81%) of the vasectomized macaques. We conclude that surgical vasectomy is a safe and simple procedure that can be used as a highly effective method to facilitate social housing of adult male rhesus macaques in research facilities.

Endotracheal Intubation of Rabbits Using a Polypropylene Guide Catheter.

Endotracheal intubation in rabbits can be challenging due to their unusual anatomy. Achieving a patent airway during anesthesia is critical for the avoidance of airway obstruction, prevention of gastric tympany, and to allow ventilatory support. Due to the difficulty of intubation, alternative methods such as the use of laryngeal mask airways or laryngeal tubes have been explored. However, these methods do not result in direct access to the trachea and thus may present a risk for development of complications. In addition, lack of direct intubation of the trachea can result in personnel exposure to waste anesthetic gases. Numerous methods for endotracheal intubation have been described, including blind placement, use of a fiberoptic laryngoscope or endoscope, and cricoid placement. Despite these numerous publications, many still struggle to achieve success. Here we provide a detailed description of an intubation technique that can be taught with minimal training with a short time to proficiency. Briefly, after administration of injectable anesthesia and proper positioning of the rabbit, a polypropylene catheter is placed into the trachea by direct visualization using a laryngoscope. The catheter is then used as a guide to direct the endotracheal tube into the trachea. This method allows for intubation without the need for expensive equipment and can be performed by a single individual without the need for an assistant. In conclusion, this technique can be easily taught and performed at very little cost in any clinical or research setting.

Fatal Meningitis in Swine after Intrathecal Administration of Adeno-associated Virus Expressing Syngeneic Interleukin-10.

Interleukin-10 (IL-10) delivered by intrathecal (i.t.) gene vectors is a candidate investigational new drug (IND) for several chronic neurological disorders such as neuropathic pain. We performed a preclinical safety study of IL-10. A syngeneic large animal model was used delivering porcine IL-10 (pIL-10) to the i.t. space in swine by adeno-associated virus serotype 8 (AAV8), a gene vector that was previously found to be nontoxic in the i.t. space. Unexpectedly, animals became ill, developing ataxia, seizures, and an inability to feed and drink, and required euthanasia. Necropsy demonstrated lymphocytic meningitis without evidence of infection in the presence of normal laboratory findings for body fluids and normal histopathology of peripheral organs. Results were replicated in a second animal cohort by a team of independent experimenters. An extensive infectious disease and neuropathology workup consisting of comprehensive testing of tissues and body fluids in a specialized research veterinary pathology environment did not identify a pathogen. These observations raise the concern that i.t. IL-10 therapy may not be benign, that previously used xenogeneic models testing the human homolog of IL-10 may not have been sensitive enough to detect toxicity, and that additional preclinical studies may be needed before clinical testing of IL-10 can be considered.

Demodex musculi Infestation in Genetically Immunomodulated Mice.

Demodex musculi, a prostigmatid mite that has been reported infrequently in laboratory mice, has been identified with increasing frequency in contemporary colonies of immunodeficient mice. Here we describe 2 episodes of D. musculi infestation with associated clinical signs in various genetically engineered mouse strains, as well as treatment strategies and an investigation into transmissibility and host susceptibility. The first case involved D. musculi associated with clinical signs and pathologic lesions in BALB/c-Tg(DO11.10)Il13(tm) mice, which have a defect in type 2 helper T cell (Th2) immunity. Subsequent investigation revealed mite transmission to both parental strains (BALB/c-Tg[DO11.10] and BALB/c-Il13(tm)), BALB/c-Il13/Il4(tm), and wild-type BALB/c. All Tg(DO11.10)Il13(tm) mice remained infested throughout the investigation, and D. musculi were recovered from all strains when they were cohoused with BALB/c-Tg(DO11.10)Il13(tm) index mice. However, only Il13(tm) and Il13/Il4(tm) mice demonstrated persistent infestation after index mice were removed. Only BALB/c-Tg(DO11.10)Il13(tm) showed clinical signs, suggesting that the phenotypic dysfunction of Th2 immunity is sufficient for persistent infestation, whereas clinical disease associated with D. musculi appears to be genotype-specific. This pattern was further exemplified in the second case, which involved NOD.Cg-Prkdc(scid)Il2r(tm1Wjl)/SzJ (NSG) and C;129S4 Rag2(tm1.1Flv) Il2rg(tm1.1Flv)/J mice with varying degrees of blepharitis, conjunctivitis, and facial pruritis. Topical amitraz decreased mite burden but did not eliminate infestation or markedly ameliorate clinical signs. Furthermore, mite burden began to increase by 1 mo posttreatment, suggesting that topical amitraz is an ineffective treatment for D. musculi. These experiences illustrate the need for vigilance regarding opportunistic and uncommon pathogens in rodent colonies, especially among mice with immunologic deficits.

Use of (18)F-fluorodeoxyglucose positron emission tomography-computed tomography to aid in diagnosing intestinal adenocarcinoma in 2 rhesus macaques (Macaca mulatta).

Two aged female rhesus macaques (Macaca mulatta) presented with weight loss and intermittent inappetence. The signalment and constellation of clinical signs led clinicians to suspect the presence of intestinal adenocarcinoma. Because of each animal's advanced age and inconclusive radiographic findings, a noninvasive diagnostic tool was preferred over exploratory laparotomy to assist in determining a diagnosis. Consequently, 2-[(18)F]fluoro-2-deoxy-d-glucose (FDG) positron emission tomography-CT (FDG-PET-CT) was chosen to aid in confirming a suspicion of gastrointestinal adenocarcinoma in both animals. FDG is a glucose analogue labeled with fluorine-18 and is taken up by highly metabolically active cells, as observed in many cancers. Tomography revealed an annular constriction of the small intestine with focal FDG uptake in one animal, and an FDG avid transmural mass in the ascending colon of the second animal. Necropsy later confirmed both sites to be adenocarcinomas. This report supports the use of FDG-PET-CT as an adjunct to conventional radiography in the diagnosis of intestinal adenocarcinoma in nonhuman primates.

Additive Effects of Mechanical Marrow Ablation and PTH Treatment on de Novo Bone Formation in Mature Adult Rats.

Mechanical ablation of bone marrow in young rats induces rapid but transient bone growth, which can be enhanced and maintained for three weeks by the administration of parathyroid hormone (PTH). Additionally, marrow ablation, followed by PTH treatment for three months leads to increased cortical thickness. In this study, we sought to determine whether PTH enhances bone formation after marrow ablation in aged rats. Aged rats underwent unilateral femoral marrow ablation and treatment with PTH or vehicle for four weeks. Both femurs from each rat were analyzed by X-ray and pQCT, then analyzed either by microCT, histology or biomechanical testing. Marrow ablation alone induced transient bone formation of low abundance that persisted over four weeks, while marrow ablation followed by PTH induced bone formation of high abundance that also persisted over four weeks. Our data confirms that the osteo-inducive effect of marrow ablation and the additive effect of marrow ablation, followed by PTH, occurs in aged rats. Our observations open new avenues of investigations in the field of tissue regeneration. Local marrow ablation, in conjunction with an anabolic agent, might provide a new platform for rapid site-directed bone growth in areas of high bone loss, such as in the hip and wrist, which are subject to fracture.

Fructosamine reference ranges in rhesus macaques (Macaca mulatta).

Naturally occurring diabetes mellitus (DM) is common in several species of Old and New World nonhuman primates. Fructosamine values provide important information about recent glycemic control and can be useful in the diagnosis and management of DM. However, despite an abundance of reports in the literature describing spontaneous and induced DM in monkeys, few reference ranges are available for fructosamine. Reference ranges have been published for woolly monkeys (Lagothrix lagotricha), cynomolgus macaques (Macaca fascicularis), and stumptail macaques (Macaca arctoides) but currently are not available for rhesus macaques. At our institution, DM is a common diagnosis in aging rhesus macaques. Here we report a reference range for fructosamine in rhesus macaques. The overall range was 157 to 230 µmol/L, with male rhesus and macaques 10 y or older having significantly higher values than do female rhesus and macaques younger than 10 y, respectively. This range provides clinical veterinarians with an additional tool for evaluating glycemic control in rhesus macaques.

Maropitant citrate for treatment of ulcerative dermatitis in mice with a C57BL/6 background.

Murine ulcerative dermatitis (UD) is a common progressive condition of mice with a C57BL/6 background. Typically, mice present with scabs and crusts on the skin of the dorsal neck and ears, and are often severely pruritic. Animals tend to scratch the lesions, causing additional trauma to the already ulcerated and inflamed skin. Therapeutic intervention largely has been unsuccessful, in part due to the lack of a known cause for the disease. Though the exact etiology of UD has not been elucidated, substance P (SP) has recently been demonstrated as an important neuropeptide linked to the itch-scratch cycle. SP functions at the tachykinin neurokinin 1 (NK1) receptor. We hypothesized that inhibition of SP binding to the NK1 receptor would decrease the itch sensation, thus decreasing scratching behavior and subsequent skin trauma. The purpose of this study was to evaluate the effectiveness of an NK1 receptor antagonist, maropitant citrate, as a treatment for murine UD. Treatment with 1 mg/kg maropitant citrate significantly reduced the size of UD lesions in mice.

Use of p63, a myoepithelial cell marker, in determining the invasiveness of spontaneous mammary neoplasia in a rhesus macaque (Macaca mulatta).

Here we describe a case of mammary gland ductal carcinoma in an aged rhesus macaque. Tumors were diagnosed based on routine hematoxylin and eosin staining. Invasiveness was further characterized by p63 immunohistochemistry. p63 is a p53 homolog that strongly and specifically stains nuclei of myoepithelial cells in human and canine mammary tissue. Because p63 has an affinity for the nucleus of myoepithelial cells, it is readily visible. Staining of mammary tissue from the monkey for p63 revealed that multiple foci of neoplastic cells had breached the myoepithelial cell layer surrounding ducts, suggesting the potential for local invasion of the tumor. Regional metastasis was confirmed at necropsy. To our knowledge, this is the first documented use of p63 for effectively determining the invasive nature of a mammary tumor in a nonhuman primate and the first use of p63 as an effective means of staining myoepithelial cells in a mammary ductal carcinoma in a nonhuman primate. Because nonhuman primates are important animal models for human diseases, including neoplasia, this method may prove useful for both diagnostic and research purposes.

Poliomyelitis in MuLV-infected ICR-SCID mice after injection of basement membrane matrix contaminated with lactate dehydrogenase-elevating virus.

The arterivirus lactate dehydrogenase-elevating virus (LDV) causes life-long viremia in mice. Although LDV infection generally does not cause disease, infected mice that are homozygous for the Fv1(n) allele are prone to develop poliomyelitis when immunosuppressed, a condition known as age-dependent poliomyelitis. The development of age-dependent poliomyelitis requires coinfection with endogenous murine leukemia virus. Even though LDV is a common contaminant of transplantable tumors, clinical signs of poliomyelitis after inadvertent exposure to LDV have not been described in recent literature. In addition, LDV-induced poliomyelitis has not been reported in SCID or ICR mice. Here we describe the occurrence of poliomyelitis in ICR-SCID mice resulting from injection of LDV-contaminated basement membrane matrix. After exposure to LDV, a subset of mice presented with clinical signs including paresis, which was associated with atrophy of the hindlimb musculature, and tachypnea; in addition, some mice died suddenly with or without premonitory signs. Mice presenting within the first 6 mo after infection had regions of spongiosis, neuronal necrosis and astrocytosis of the ventral spinal cord, and less commonly, brainstem. Axonal degeneration of ventral roots prevailed in more chronically infected mice. LDV was identified by RT-PCR in 12 of 15 mice with typical neuropathology; positive antiLDV immunolabeling was identified in all PCR-positive animals (n = 7) tested. Three of 8 mice with neuropathology but no clinical signs were LDV negative by RT-PCR. RT-PCR yielded murine leukemia virus in spinal cords of all mice tested, regardless of clinical presentation or neuropathology.

Tracheal stenosis and adenocarcinoma in an olive baboon (Papio cynocephalus anubis).

An adult female baboon (Papio cynocephalus anubis) presented for progressive difficulty in endotracheal intubation. Over a 7-y period prior to presentation, she was anesthetized and intubated 67 times for imaging by using single-photon emission computed tomography or positron emission tomography. Laryngoscopic examination revealed tracheal stenosis. Because of increased anesthetic risk and lack of alternative use, she was euthanized, and partial necropsy focusing on the larynx, trachea, and associated structures was performed. Gross examination revealed rigidity and functional fusion of the proximal 5 or 6 tracheal rings and narrowing of the lumen. Histology revealed ossification of tracheal rings and fibrosis of overlying tissue. In addition, a transmural umbilicated mass was present midway down the cervical trachea on its dorsolateral aspect. Histology of the tracheal mass identified a relatively well-circumscribed transmural adenocarcinoma. The combination of overall histologic pattern, evidence of anaplasia, and results of immunohistochemical staining was consistent with a diagnosis of adenoid cystic carcinoma. Anterior tracheal stenosis is a reported complication of intubation in humans and animals. Primary tracheal neoplasms are rare in domestic and research animals and, to our knowledge, have not previously been reported to occur in nonhuman primates.