Alicia M. Waters

Preclinical evaluation of engineered oncolytic herpes simplex virus for the treatment of neuroblastoma.

Despite intensive research efforts and therapeutic advances over the last few decades, the pediatric neural crest tumor, neuroblastoma, continues to be responsible for over 15% of pediatric cancer deaths. Novel therapeutic options are needed for this tumor. Recently, investigators have shown that mice with syngeneic murine gliomas treated with an engineered, neuroattenuated oncolytic herpes simplex virus-1 (oHSV), M002, had a significant increase in survival. M002 has deletions in both copies of the γ134.5 gene, enabling replication in tumor cells but precluding infection of normal neural cells. We hypothesized that M002 would also be effective in the neural crest tumor, neuroblastoma. We showed that M002 infected, replicated, and decreased survival in neuroblastoma cell lines. In addition, we showed that in murine xenografts, treatment with M002 significantly decreased tumor growth, and that this effect was augmented with the addition of ionizing radiation. Importantly, survival could be increased by subsequent doses of radiation without re-dosing of the virus. Finally, these studies showed that the primary entry protein for oHSV, CD111 was expressed by numerous neuroblastoma cell lines and was also present in human neuroblastoma specimens. We concluded that M002 effectively targeted neuroblastoma and that this oHSV may have potential for use in children with unresponsive or relapsed neuroblastoma.

Preclinical Evaluation of Engineered Oncolytic Herpes Simplex Virus for the Treatment of Pediatric Solid Tumors

Recently, investigators showed that mice with syngeneic murine gliomas that were treated with a neuroattenuated oncolytic herpes simplex virus-1 (oHSV), M002, had a significant increase in survival. M002 has deletions in both copies of the γ134.5 gene, enabling replication in tumor cells but precluding infection of normal cells. Previous studies have shown antitumor effects of other oHSV against a number of adult tumors including hepatocellular carcinoma and renal cell carcinoma. The purpose of the current study was to investigate the oncolytic potential of M002 against difficult to treat pediatric liver and kidney tumors. We showed that the oHSV, M002, infected, replicated, and decreased cell survival in hepatoblastoma, malignant rhabdoid kidney tumor, and renal sarcoma cell lines. In addition, we showed that in murine xenografts, treatment with M002 significantly increased survival and decreased tumor growth. Finally, these studies showed that the primary entry protein for oHSV, CD111 (nectin-1) was present in human hepatoblastoma and malignant rhabdoid kidney tumor specimens. We concluded that M002 effectively targeted these rare aggressive tumor types and that M002 may have potential for use in children with unresponsive or relapsed pediatric solid tumors.

Pediatric cancer gone viral. Part I: strategies for utilizing oncolytic herpes simplex virus-1 in children

Progress for improving outcomes in pediatric patients with solid tumors remains slow. In addition, currently available therapies are fraught with numerous side effects, often causing significant life-long morbidity for long-term survivors. The use of viruses to kill tumor cells based on their increased vulnerability to infection is gaining traction, with several viruses moving through early and advanced phase clinical testing. The prospect of increased efficacy and decreased toxicity with these agents is thus attractive for pediatric cancer. In part I of this two-part review, we focus on strategies for utilizing oncolytic engineered herpes simplex virus (HSV) to target pediatric malignancies. We discuss mechanisms of action, routes of delivery, and the role of preexisting immunity on antitumor efficacy. Challenges to maximizing oncolytic HSV in children are examined, and we highlight how these may be overcome through various arming strategies. We review the preclinical and clinical evidence demonstrating safety of a variety of oncolytic HSVs. In Part II, we focus on the antitumor efficacy of oncolytic HSV in pediatric tumor types, pediatric clinical advances made to date, and future prospects for utilizing HSV in pediatric patients with solid tumors.

Preclinical Evaluation of a Novel RXR Agonist for the Treatment of Neuroblastoma

Neuroblastoma remains a common cause of pediatric cancer deaths, especially for children who present with advanced stage or recurrent disease. Currently, retinoic acid therapy is used as maintenance treatment to induce differentiation and reduce tumor recurrence following induction therapy for neuroblastoma, but unavoidable side effects are seen. A novel retinoid, UAB30, has been shown to generate negligible toxicities. In the current study, we hypothesized that UAB30 would have a significant impact on multiple neuroblastoma cell lines in vitro and in vivo. Cellular survival, cell-cycle analysis, migration, and invasion were studied using AlamarBlue assays, FACS, and Transwell assays, respectively, in multiple cell lines following treatment with UAB30. In addition, an in vivo murine model of human neuroblastoma was utilized to study the effects of UAB30 upon tumor xenograft growth and animal survival. We successfully demonstrated decreased cellular survival, invasion, and migration, cell-cycle arrest, and increased apoptosis after treatment with UAB30. Furthermore, inhibition of tumor growth and increased survival was observed in a murine neuroblastoma xenograft model. The results of these in vitro and in vivo studies suggest a potential therapeutic role for the low toxicity synthetic retinoid X receptor selective agonist, UAB30, in neuroblastoma treatment. Mol Cancer Ther; 14(7); 1559–69. ©2015 AACR.

Early Trauma-Induced Coagulopathy is Associated with Increased Ventilator-Associated Pneumonia in Spinal Cord Injury Patients.

Introduction: Early trauma-induced coagulopathy may increase susceptibility to nosocomial infections such as ventilator-associated pneumonia. However, the relationship between trauma- induced coagulopathy and the development of ventilator-associated pneumonia in spinal cord injury patients has not been evaluated. Methods: We conducted a 5-year retrospective study of 300 spinal cord injury patients admitted to Level 1 trauma center. Standard coagulation studies were evaluated upon arrival, prior to fluid resuscitative efforts, and at 24 h after admission. Based on these studies, three groups of patients were identified: no coagulopathy, latent coagulopathy, and admission coagulopathy. Ventilator- associated pneumonia was identified utilizing Centers for Disease Control and Prevention criteria. Since we used the data in the trauma registry and did not have the information on FiO2 and PEEP, we elected to use the VAP terminology and not the VAE sequence. Demographic, injury, and clinical characteristics were compared among no coagulopathy, latent coagulopathy, and admission coagulopathy groups using chi-square test and ANOVA for categorical and continuous variables, respectively. A logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for the association between coagulopathy and both ventilator-associated pneumonia and mortality. Results: The incidence of ventilator-associated pneumonia was 54.5% (OR 4.01, 95% CI 1.76–9.15) in spinal cord injury patients with admission coagulopathy, compared with the 17.5% in spinal cord injury patients with no coagulopathy. Mortality was significantly higher in spinal cord injury patients with admission coagulopathy than in spinal cord injury patients with no coagulopathy (OR 6.14, 95% CI 1.73–21.73). After adjusting for age, race, injury mechanism, Injury Severity Score, base deficit at admission, the number of pRBC units transfused in the first 24 h, and hospital stay, only the association of ventilator-associated pneumonia among those with admission coagulopathy remained significant (OR 3.51, 95% CI 1.48–8.32). Compared with those with no coagulopathy, patients with admission coagulopathy had a higher odds of death (4.10, 95% CI 1.53–11.02), though this association lost significance after adjustment (OR 3.56, 95% CI 0.90–14.12). There was no statistical difference in mortality for latent coagulopathy compared with no coagulopathy patients. Conclusion: Coagulopathy on admission in patients with spinal cord injury is associated with a statistically significant increase in ventilator-associated pneumonia incidence. Additional research is warranted to further characterize this association.

Safety and utility of scheduled nonnarcotic analgesic medications in children undergoing craniotomy for brain tumor.

BACKGROUNDWe have reported that a scheduled nonnarcotic analgesic regimen after dorsal lumbar rhizotomy and Chiari I malformation decompression is efficacious in managing postoperative pain in children. To date, this regimen has not been analyzed in children after brain tumor biopsy or resection. OBJECTIVETo elucidate the safety and utility of such an analgesic protocol in these patients. PATIENTS AND METHODSA database review was conducted to identify children who received a scheduled dose of alternating acetaminophen and ibuprofen after craniotomy for tumor biopsy or resection, and postoperative imaging was evaluated. RESULTSFifty-one children who met the inclusion criteria were identified. On postoperative imaging, 17.67% had routine, postoperative blood in the resection cavity according to both radiology and neurosurgical review. One patient had moderate postoperative bleeding in the tumor cavity. Overall, 44 of the 51 patients (86.3%) required no or minimal narcotic medication for pain. CONCLUSIONA scheduled regimen of nonsteroidal antiinflammatory drugs given in alternating doses immediately after craniotomy for tumor biopsy or resection and throughout hospitalization did not result in any significant postoperative hemorrhage in our patient series.

Effect of Repeat Dosing of Engineered Oncolytic Herpes Simplex Virus on Preclinical Models of Rhabdomyosarcoma

Rhabdomyosarcoma (RMS), a tumor of skeletal muscle origin, is the most common sarcoma of childhood. Despite multidrug chemotherapy regimens, surgical intervention, and radiation treatment, outcomes remain poor, especially in advanced disease, and novel therapies are needed for the treatment of these aggressive malignancies. Genetically engineered oncolytic viruses, such as herpes simplex virus-1 (HSV), are currently being explored as treatments for pediatric tumors. M002, an oncolytic HSV, has both copies of the γ134.5 gene deleted, enabling replication in tumor cells but thwarting infection of normal, postmitotic cells. We hypothesized that M002 would infect human RMS tumor cells and lead to decreased tumor cell survival in vitro and impede tumor growth in vivo. In the current study, we demonstrated that M002 could infect, replicate in, and decrease cell survival in both embryonal (ERMS) and alveolar rhabdomyosarcoma (ARMS) cells. Additionally, M002 reduced xenograft tumor growth and increased animal survival in both ARMS and ERMS. Most importantly, we showed for the first time that repeated dosing of oncolytic virus coupled with low-dose radiation provided improved tumor response in RMS. These findings provide support for the clinical investigation of oncolytic HSV in pediatric RMS.

Pediatric cancer gone viral. Part II: potential clinical application of oncolytic herpes simplex virus-1 in children

Oncolytic engineered herpes simplex viruses (HSVs) possess many biologic and functional attributes that support their use in clinical trials in children with solid tumors. Tumor cells, in an effort to escape regulatory mechanisms that would impair their growth and progression, have removed many mechanisms that would have protected them from virus infection and eventual virus-mediated destruction. Viruses engineered to exploit this weakness, like mutant HSV, can be safely employed as tumor cell killers, since normal cells retain these antiviral strategies. Many preclinical studies and early phase trials in adults demonstrated that oncolytic HSV can be safely used and are highly effective in killing tumor cells that comprise pediatric malignancies, without generating the toxic side effects of nondiscriminatory chemotherapy or radiation therapy. A variety of engineered viruses have been developed and tested in numerous preclinical models of pediatric cancers and initial trials in patients are underway. In Part II of this review series, we examine the preclinical evidence to support the further advancement of oncolytic HSV in the pediatric population. We discuss clinical advances made to date in this emerging era of oncolytic virotherapy.

Targeting Focal Adhesion Kinase Suppresses the Malignant Phenotype in Rhabdomyosarcoma Cells

Despite the tremendous advances in the treatment of childhood solid tumors, rhabdomyosarcoma (RMS) continues to provide a therapeutic challenge. Children with metastatic or relapsed disease have a disease-free survival rate under 30%. Focal adhesion kinase (FAK) is a nonreceptor tyrosine kinase that is important in many facets of tumorigenesis. Signaling pathways both upstream and downstream to FAK have been found to be important in sarcoma tumorigenesis, leading us to hypothesize that FAK would be present in RMS and would impact cellular survival. In the current study, we showed that FAK was present and phosphorylated in pediatric alveolar and embryonal RMS tumor specimens and cell lines. We also examined the effects of FAK inhibition upon two RMS cell lines utilizing parallel approaches including RNAi and small molecule inhibitors. FAK inhibition resulted in decreased cellular survival, invasion, and migration and increased apoptosis. Furthermore, small molecule inhibition of FAK led to decreased tumor growth in a nude mouse RMS xenograft model. The findings from this study will help to further our understanding of the regulation of tumorigenesis in RMS and may provide desperately needed novel therapeutic strategies for these difficult-to-treat tumors.

Focal adhesion kinase and p53 synergistically decrease neuroblastoma cell survival.

Neuroblastoma is the most common extracranial solid tumor of childhood and is responsible for over 15% of pediatric cancer deaths. Focal adhesion kinase (FAK) is a nonreceptor tyrosine kinase that is important in many facets of neuroblastoma tumor development and progression. The p53 oncogene, although wild type in most neuroblastomas, lacks significant function as a tumor suppressor in these tumors. Recent reports have found that FAK and p53 interact in some tumor types. We have hypothesized FAK and p53 coordinately control each others expression and also interact in neuroblastoma. In the present study, we showed that not only do FAK and p53 interact but each one controls the expression of the other. In addition, we also examined the effects of FAK inhibition combined with p53 activation in neuroblastoma and showed that these two, in combination, had a synergistic effect on neuroblastoma cell survival. The findings from this present study help to further our understanding of the regulation of neuroblastoma tumorigenesis and may provide novel therapeutic strategies and targets for neuroblastoma and other pediatric solid tumors.