Small molecule targeting of transcription-replication conflict for selective chemotherapy.

Targeting transcription replication conflicts, a major source of endogenous DNA double-stranded breaks and genomic instability could have important anticancer therapeutic implications. Proliferating cell nuclear antigen (PCNA) is critical to DNA replication and repair processes. Through a rational drug design approach, we identified a small molecule PCNA inhibitor, AOH1996, which selectively kills cancer cells. AOH1996 enhances the interaction between PCNA and the largest subunit of RNA polymerase II, RPB1, and dissociates PCNA from actively transcribed chromatin regions, while inducing DNA double-stranded breaks in a transcription-dependent manner. Attenuation of RPB1 interaction with PCNA, by a point mutation in RPB1's PCNA-binding region, confers resistance to AOH1996. Orally administrable and metabolically stable, AOH1996 suppresses tumor growth as a monotherapy or as a combination treatment but causes no discernable side effects. Inhibitors of transcription replication conflict resolution may provide a new and unique therapeutic avenue for exploiting this cancer-selective vulnerability.

Glioma-targeted delivery of exosome-encapsulated antisense oligonucleotides using neural stem cells.

Tropism of neural stem cells (NSCs) to hypoxic tumor areas provides an opportunity for the drug delivery. Here, we demonstrate that NSCs effectively transport antisense oligonucleotides (ASOs) targeting oncogenic and tolerogenic signal transducer and activator of transcription 3 (STAT3) protein into glioma microenvironment. To enable spontaneous, scavenger receptor-mediated endocytosis by NSCs, we used previously described CpG-STAT3ASO conjugates. Following uptake and endosomal escape, CpG-STAT3ASO colocalized with CD63+ vesicles and later with CD63+CD81+ exosomes. Over 3 days, NSCs secreted exosomes loaded up to 80% with CpG-STAT3ASO. Compared to native NSC exosomes, the CpG-STAT3ASO-loaded exosomes potently stimulated immune activity of human dendritic cells or mouse macrophages, inducing nuclear factor κB (NF-κB) signaling and interleukin-12 (IL-12) production. Using orthotopic GL261 tumors, we confirmed that NSC-mediated delivery improved oligonucleotide transfer from a distant injection site into the glioma microenvironment versus naked oligonucleotides. Correspondingly, the NSC-delivered CpG-STAT3ASO enhanced activation of glioma-associated microglia. Finally, we demonstrated that NSC-mediated CpG-STAT3ASO delivery resulted in enhanced antitumor effects against GL261 glioma in mice. Peritumoral injections of 5 × 105 NSCs loaded ex vivo with CpG-STAT3ASO inhibited subcutaneous tumor growth more effectively than the equivalent amount of oligonucleotide alone. Based on these results, we anticipate that NSCs and NSC-derived exosomes will provide a clinically relevant strategy to improve delivery and safety of oligonucleotide therapeutics for glioma treatment.

Multiple Treatment Cycles of Neural Stem Cell Delivered Oncolytic Adenovirus for the Treatment of Glioblastoma.

Tumor tropic neural stem cells (NSCs) can improve the anti-tumor efficacy of oncovirotherapy agents by protecting them from rapid clearance by the immune system and delivering them to multiple distant tumor sites. We recently completed a first-in-human trial assessing the safety of a single intracerebral dose of NSC-delivered CRAd-Survivin-pk7 (NSC.CRAd-S-pk7) combined with radiation and chemotherapy in newly diagnosed high-grade glioma patients. The maximum feasible dose was determined to be 150 million NSC.CRAd-Sp-k7 (1.875 × 1011 viral particles). Higher doses were not assessed due to volume limitations for intracerebral administration and the inability to further concentrate the study agent. It is possible that therapeutic efficacy could be maximized by administering even higher doses. Here, we report IND-enabling studies in which an improvement in treatment efficacy is achieved in immunocompetent mice by administering multiple treatment cycles intracerebrally. The results imply that pre-existing immunity does not preclude therapeutic benefits attainable by administering multiple rounds of an oncolytic adenovirus directly into the brain.

Large, Anionic Liposomes Enable Targeted Intraperitoneal Delivery of a TLR 7/8 Agonist To Repolarize Ovarian Tumors' Microenvironment.

Ovarian cancer is the most lethal gynecological malignancy in the United States. Current standard of treatment includes surgical debulking and chemotherapy, such as cisplatin and paclitaxel. However, the patients' response rate for chemotherapy in ovarian cancer is not optimal, and they often develop chemoresistance and suffer from side effects. Current clinical trials make extensive use of immune checkpoint blockade (ICB) as a novel cancer immunotherapeutic strategy against ovarian tumors. However, the response rates for ICB antibodies remain limited to 10-20% of treated ovarian cancer patients despite the success of this approach in melanoma, renal, head and neck, and nonsmall cell lung cancers. This lack of efficacy is often attributed to the "cold" immune status of ovarian tumors, as these tumors often have a low number of tumor-infiltrating lymphocytes (TILs) but a high number of suppressive immune cells, including tumor-associated macrophages (TAMs), myeloid-derived suppressor cells (MDSCs), or regulatory T cells (Tregs). Repolarizing TAMs could be a promising strategy to reshape the tumor immune microenvironment and promote antitumor activity when combined with ICBs. Toll-like receptor (TLR) 7 and 8 agonists, such as imiquimod and resiquimod, are potent immunostimulatory molecules with potential to repolarize macrophages. However, these small molecules have poor pharmacokinetic profiles and can induce severe side effects when administered systemically. Previously, our group demonstrated that various large, anionic nanomaterials (silica, PLGA, and polystyrene) specifically target TAMs when administered intraperitoneally (IP) to ovarian tumor-bearing mice. In the present study, we demonstrate that large, anionic liposomes administered IP also efficiently localize to TAMs and can be used to target the delivery of resiquimod. Resiquimod delivered in this targeted fashion promoted activation of M1 macrophages and T cell infiltration, while reducing the percentage of Tregs in the tumor microenvironment. Finally, liposome-formulated resiquimod significantly enhanced the efficacy of PD1 blockade against syngeneic ovarian tumors. We anticipate that further optimization of our liposomal delivery strategy can generate a clinically relevant strategy for more effective and safer immunotherapy for ovarian cancer patients.

Neural stem cell delivery of an oncolytic adenovirus in newly diagnosed malignant glioma: a first-in-human, phase 1, dose-escalation trial.

BACKGROUND: Malignant glioma is the most common and lethal primary brain tumour, with dismal survival rates and no effective treatment. We examined the safety and activity of NSC-CRAd-S-pk7, an engineered oncolytic adenovirus delivered by neural stem cells (NSCs), in patients with newly diagnosed high-grade glioma. METHODS: This was a first-in-human, open-label, phase 1, dose-escalation trial done to determine the maximal tolerated dose of NSC-CRAd-S-pk7, following a 3 + 3 design. Patients with newly diagnosed, histologically confirmed, high-grade gliomas (WHO grade III or IV) were recruited. After neurosurgical resection, NSC-CRAd-S-pk7 was injected into the walls of the resection cavity. The first patient cohort received a dose starting at 6·25 × 1010 viral particles administered by 5·00 × 107 NSCs, the second cohort a dose of 1·25 × 1011 viral particles administered by 1·00 × 108 NSCs, and the third cohort a dose of 1·875 × 1011 viral particles administered by 1·50 × 108 NSCs. No further dose escalation was planned. Within 10-14 days, treatment with temozolomide and radiotherapy was initiated. Primary endpoints were safety and toxicity profile and the maximum tolerated dose for a future phase 2 trial. All analyses were done in all patients who were included in the trial and received the study treatment and were not excluded from the study. Recruitment is complete and the trial is finished. The trial is registered with ClinicalTrials.gov, NCT03072134. FINDINGS: Between April 24, 2017, and Nov 13, 2019, 12 patients with newly diagnosed, malignant gliomas were recruited and included in the safety analysis. Histopathological evaluation identified 11 (92%) of 12 patients with glioblastoma and one (8%) of 12 patients with anaplastic astrocytoma. The median follow-up was 18 months (IQR 14-22). One patient receiving 1·50 × 108 NSCs loading 1·875 × 1011 viral particles developed viral meningitis (grade 3) due to the inadvertent injection of NSC-CRAd-S-pk7 into the lateral ventricle. Otherwise, treatment was safe as no formal dose-limiting toxicity was reached, so 1·50 × 108 NSCs loading 1·875 × 1011 viral particles was recommended as a phase 2 trial dose. There were no treatment-related deaths. The median progression-free survival was 9·1 months (95% CI 8·5-not reached) and median overall survival was 18·4 months (15·7-not reached). INTERPRETATION: NSC-CRAd-S-pk7 treatment was feasible and safe. Our immunological and histopathological findings support continued investigation of NSC-CRAd-S-pk7 in a phase 2/3 clinical trial. FUNDING: US National Institutes of Health.

Thermal analysis of laser irradiation-gold nanorod combinations at 808 nm, 940 nm, 975 nm and 1064 nm wavelengths in breast cancer model.

BACKGROUND: Photothermal therapy is currently under the spotlight to improve the efficacy of minimally invasive thermal treatment of solid tumors. The interplay of several factors including the radiation wavelengths and the nanoparticle characteristics underlie the thermal outcome. However, a quantitative thermal analysis in in vivo models embedding nanoparticles and under different near-infrared (NIR) wavelengths is missing. PURPOSE: We evaluate the thermal effects induced by different combinations of NIR laser wavelengths and gold nanorods (GNRs) in breast cancer tumor models in mice. MATERIALS AND METHODS: Four laser wavelengths within the therapeutic window, i.e., 808, 940, 975, and 1064 nm were employed, and corresponding GNRs were intratumorally injected. The tissue thermal response was evaluated in terms of temperature profile and time constants, considering the step response of a first-order system as a model. RESULTS: The 808 nm and 1064 nm lasers experienced the highest temperature enhancements (>24%) in presence of GNRs compared to controls; conversely, 975 nm and 940 nm lasers showed high temperatures in controls due to significant tissue absorption and the lowest temperature difference with and without GNRs (temperature enhancement <10%). The presence of GNRs resulted in small time constants, thus quicker laser-induced thermal response (from 67 s to 33 s at 808 nm). CONCLUSIONS: The thermal responses of different GNR-laser wavelength combinations quantitatively validate the widespread usage of 808 nm laser for nanoparticle-assisted photothermal procedures. Moreover, our results provide insights on other usable wavelengths, toward the identification of an effective photothermal treatment strategy for the removal of focal malignancies.

Allogeneic human neural stem cells for improved therapeutic delivery to peritoneal ovarian cancer.

BACKGROUND: Immortalized, clonal HB1.F3.CD 21 human neural stem/progenitor cells (NSCs), loaded with therapeutic cargo prior to intraperitoneal (IP) injection, have been shown to improve the delivery and efficacy of therapeutic agents in pre-clinical models of stage III ovarian cancer. In previous studies, the distribution and efficacy of the NSC-delivered cargo has been examined; however, the fate of the NSCs has not yet been explored. METHODS: To monitor NSC tropism, we used an unconventional method of quantifying endocytosed gold nanorods to overcome the weaknesses of existing cell-tracking technologies. RESULTS: Here, we report efficient tumor tropism of HB1.F3.CD 21 NSCs, showing that they primarily distribute to the tumor stroma surrounding individual tumor foci within 3 h after injection, reaching up to 95% of IP metastases without localizing to healthy tissue. Furthermore, we demonstrate that these NSCs are non-tumorigenic and non-immunogenic within the peritoneal setting. CONCLUSIONS: Their efficient tropism, combined with their promising clinical safety features and potential for cost-effective scale-up, positions this NSC line as a practical, off-the-shelf platform to improve the delivery of a myriad of peritoneal cancer therapeutics.

Neural stem cells secreting bispecific T cell engager to induce selective antiglioma activity.

Glioblastoma (GBM) is the most lethal primary brain tumor in adults. No treatment provides durable relief for the vast majority of GBM patients. In this study, we've tested a bispecific antibody comprised of single-chain variable fragments (scFvs) against T cell CD3ε and GBM cell interleukin 13 receptor alpha 2 (IL13Rα2). We demonstrate that this bispecific T cell engager (BiTE) (BiTELLON) engages peripheral and tumor-infiltrating lymphocytes harvested from patients' tumors and, in so doing, exerts anti-GBM activity ex vivo. The interaction of BiTELLON with T cells and IL13Rα2-expressing GBM cells stimulates T cell proliferation and the production of proinflammatory cytokines interferon γ (IFNγ) and tumor necrosis factor α (TNFα). We have modified neural stem cells (NSCs) to produce and secrete the BiTELLON (NSCLLON). When injected intracranially in mice with a brain tumor, NSCLLON show tropism for tumor, secrete BiTELLON, and remain viable for over 7 d. When injected directly into the tumor, NSCLLON provide a significant survival benefit to mice bearing various IL13Rα2+ GBMs. Our results support further investigation and development of this therapeutic for clinical translation.

Feasibility of intracerebrally administering multiple doses of genetically modified neural stem cells to locally produce chemotherapy in glioma patients.

Neural stem cells (NSCs) are tumor tropic and can be genetically modified to produce anti-cancer therapies locally in the brain. In a prior first-in-human study we demonstrated that a single dose of intracerebrally administered allogeneic NSCs, which were retrovirally transduced to express cytosine deaminase (CD), tracked to glioma sites and converted oral 5-fluorocytosine (5-FC) to 5-fluorouracil (5-FU). The next step in the clinical development of this NSC-based anti-cancer strategy was to assess the feasibility of administering multiple intracerebral doses of CD-expressing NSCs (CD-NSCs) in patients with recurrent high-grade gliomas. CD-NSCs were given every 2 weeks using an indwelling brain catheter, followed each time by a 7-d course of oral 5-FC (and leucovorin in the final patient cohort). Fifteen evaluable patients received a median of 4 (range 2-10) intracerebral CD-NSC doses; doses were escalated from 50 × 106 to 150 × 106 CD-NSCs. Neuropharmacokinetic data confirmed that CD-NSCs continuously produced 5-FU in the brain during the course of 5-FC. There were no clinical signs of immunogenicity, and only three patients developed anti-NSC antibodies. Our results suggest intracerebral administration of serial doses of CD-NSCs is safe and feasible and identified a recommended dose for phase II testing of 150 × 106 CD-NSCs.

Novel Chimeric Poxvirus CF17 Improves Survival in a Murine Model of Intraperitoneal Ovarian Cancer Metastasis.

Despite improvements in surgical techniques and chemotherapy, ovarian cancer remains the most lethal gynecologic cancer. Thus, there is an urgent need for more effective therapeutics, particularly for chemo-resistant peritoneal ovarian cancer metastases. Oncolytic virotherapy represents an innovative treatment paradigm; however, for oncolytic viruses tested from the last generation of genetically engineered viruses, the therapeutic benefits have been modest. To overcome these limitations, we generated a chimeric poxvirus, CF17, through the chimerization of nine species of orthopoxviruses. Compared with its parental viruses, CF17 has demonstrated superior oncolytic characteristics. Here, we report the oncolytic potential of CF17 in ovarian cancer. Replication of CF17 and its resulting cytotoxicity were observed at multiplicities of infection (MOIs) as low as 0.001 in human and mouse cancer cell lines in vitro. Furthermore, CF17 exerted potent antitumor effects in a syngeneic mouse model of ovarian cancer at doses as low as 6 × 106 plaque-forming units. Together, these data merit further investigation of the potential use of this novel chimeric poxvirus as an effective treatment for aggressive intraperitoneal ovarian cancer.

Neural Stem Cells Improve the Delivery of Oncolytic Chimeric Orthopoxvirus in a Metastatic Ovarian Cancer Model.

Oncolytic virotherapy represents a promising approach for treating recurrent and/or drug-resistant ovarian cancer. However, its successful application in the clinic has been hampered by rapid immune-mediated clearance, which reduces viral delivery to the tumor. Patient-derived mesenchymal stem cells that home to tumors have been used as viral delivery tools, but variability associated with autologous cell isolations limits the clinical applicability of this approach. We previously developed an allogeneic, clonal neural stem cell (NSC) line (HB1.F3.CD21) that can be used to deliver viral cargo. Here, we demonstrate that this NSC line can improve the delivery of a thymidine kinase gene-deficient conditionally replication-competent orthopoxvirus, CF33, in a preclinical cisplatin-resistant peritoneal ovarian metastases model. Overall, our findings provide the basis for using off-the-shelf allogeneic cell-based delivery platforms for oncolytic viruses, thus providing a more efficient delivery alternative compared with the free virus administration approach.

Specific targeting of ovarian tumor-associated macrophages by large, anionic nanoparticles.

Immunotherapy is emerging as one of the most effective methods for treating many cancers. However, immunotherapy can still introduce significant off-target toxicity, and methods are sought to enable targeted immunotherapy at tumor sites. Here, we show that relatively large (>100-nm) anionic nanoparticles administered intraperitoneally (i.p.) selectively accumulate in tumor-associated macrophages (TAMs). In a mouse model of metastatic ovarian cancer, fluorescently labeled silica, poly(lactic-co-glycolic acid), and polystyrene nanoparticles administered i.p. were all found to selectively accumulate in TAMs. Quantifying silica particle uptake indicated that >80% of the injected dose was in TAMs. Particles that were smaller than 100 nm or cationic or administered intravenously (i.v.) showed no TAM targeting. Moreover, this phenomenon is likely to occur in humans because when freshly excised human surgical samples were treated with the fluorescent silica nanoparticles no interaction with healthy tissue was seen but selective uptake by TAMs was seen in 13 different patient samples. Ovarian cancer is a deadly disease that afflicts ∼22,000 women per year in the United States, and the presence of immunosuppressive TAMs at tumors is correlated with decreased survival. The ability to selectively target TAMs opens the door to targeted immunotherapy for ovarian cancer.

Erratum: Enhanced Delivery of Oncolytic Adenovirus by Neural Stem Cells for Treatment of Metastatic Ovarian Cancer.

[This corrects the article DOI: 10.1016/j.omto.2018.12.003.].

Developing Implantable Scaffolds to Enhance Neural Stem Cell Therapy for Post-Operative Glioblastoma.

Pre-clinical and clinical studies have shown that engineered tumoricidal neural stem cells (tNSCs) are a promising treatment strategy for the aggressive brain cancer glioblastoma (GBM). Yet, stabilizing human tNSCs within the surgical cavity following GBM resection is a significant challenge. As a critical step toward advancing engineered human NSC therapy for GBM, we used a preclinical variant of the clinically utilized NSC line HB1.F3.CD and mouse models of human GBM resection/recurrence to identify a polymeric scaffold capable of maximizing the transplant, persistence, and tumor kill of NSC therapy for post-surgical GBM. Using kinetic bioluminescence imaging, we found that tNSCs delivered into the mouse surgical cavity wall by direct injection persisted only 3 days. We found that delivery of tNSCs into the cavity on nanofibrous electrospun poly-l-lactic acid scaffolds extended tNSC persistence to 8 days. Modifications to fiber surface coating, diameter, and morphology of the scaffold failed to significantly extend tNSC persistence in the cavity. In contrast, tNSCs delivered into the post-operative cavity on gelatin matrices (GEMs) persisted 8-fold longer as compared to direct injection. GEMs remained permissive to tumor-tropic homing, as tNSCs migrated off the scaffolds and into invasive tumor foci both in vitro and in vivo. To mirror envisioned human brain tumor trials, we engineered tNSCs to express the prodrug/enzyme thymidine kinase (tNSCstk) and transplanted the therapeutic cells in the post-operative cavity of mice bearing resected orthotopic patient-derived GBM xenografts. Following administration of the prodrug ganciclovir, residual tumor volumes in mice receiving GEM/tNSCs were reduced by 10-fold at day 35, and median survival was extended from 31 to 46 days. Taken together, these data begin to define design parameters for effective scaffold/tNSC composites and suggest a new approach to maximizing the efficacy of tNSC therapy in human patient trials.

NSCs are permissive to oncolytic Myxoma virus and provide a delivery method for targeted ovarian cancer therapy.

Despite the development of many anticancer agents over the past 20 years, ovarian cancer remains the most lethal gynecologic malignancy. Due to a lack of effective screening, the majority of patients with ovarian cancer are diagnosed at an advanced stage, and only ~20% of patients are cured. Thus, in addition to improved screening methods, there is an urgent need for novel anticancer agents that are effective against late-stage, metastatic disease. Oncolytic virotherapy is a promising approach; unfortunately, systemic delivery of viruses to tumors remains a major challenge. In this regard, neural stem/progenitor cells (NSCs) with well-established tumor-homing properties may serve as an effective delivery platform for oncolytic viruses. In this study, we tested the efficacy of myxoma virus (MYXV), a rabbit-specific poxvirus that has demonstrated efficacy against a variety of tumors, using human and mouse ovarian cancer cell lines. We showed that MYXV effectively lysed ovarian cancer cells in vitro, reducing their viability. We also demonstrated that MYXV can infect human NSCs, specifically the clonal HB1.F3.CD21 NSC line. Taken together, these results suggest that NSC-mediated delivery of MYXV may be a promising strategy for achieving more selectively targeted anti-tumor efficacy.

Quantitative Evaluation of Intraventricular Delivery of Therapeutic Neural Stem Cells to Orthotopic Glioma.

Neural stem cells (NSCs) are inherently tumor-tropic, which allows them to migrate through normal tissue and selectively localize to invasive tumor sites in the brain. We have engineered a clonal, immortalized allogeneic NSC line (HB1.F3.CD21; CD-NSCs) that maintains its stem-like properties, a normal karyotype and is HLA Class II negative. It is genetically and functionally stable over time and multiple passages, and has demonstrated safety in phase I glioma trials. These properties enable the production of an "off-the-shelf" therapy that can be readily available for patient treatment. There are multiple factors contributing to stem cell tumor-tropism, and much remains to be elucidated. The route of NSC delivery and the distribution of NSCs at tumor sites are key factors in the development of effective cell-based therapies. Stem cells can be engineered to deliver and/or produce many different therapeutic agents, including prodrug activating enzymes (which locally convert systemically administered prodrugs to active chemotherapeutic agents); oncolytic viruses; tumor-targeted antibodies; therapeutic nanoparticles; and extracellular vesicles that contain therapeutic oligonucleotides. By targeting these therapeutics selectively to tumor foci, we aim to minimize toxicity to normal tissues and maximize therapeutic benefits. In this manuscript, we demonstrate that NSCs administered via intracerebral/ventricular (IVEN) routes can migrate efficiently toward single or multiple tumor foci. IVEN delivery will enable repeat administrations for patients through an Ommaya reservoir, potentially resulting in improved therapeutic outcomes. In our preclinical studies using various glioma lines, we have quantified NSC migration and distribution in mouse brains and have found robust migration of our clinically relevant HB1.F3.CD21 NSC line toward invasive tumor foci, irrespective of their origin. These results establish proof-of-concept and demonstrate the potential of developing a multitude of therapeutic options using modified NSCs.

Silica Coated Paclitaxel Nanocrystals Enable Neural Stem Cell Loading For Treatment of Ovarian Cancer.

Ovarian cancer is commonly diagnosed only after it has metastasized to the abdominal cavity (stage III). While the current standard of care of intraperitoneal (IP) administration of cisplatin and paclitaxel (PTX) combination chemotherapy has benefit, patient 5-year survival rates are low and have not significantly improved in the past decade. The ability to target chemotherapy selectively to ovarian tumors while sparing normal tissue would improve efficacy and decrease toxicities. We have previously shown that cisplatin-loaded nanoparticles (NPs) loaded within neural stem cells (NSCs) are selectively delivered to ovarian tumors in the abdominal cavity following IP injection, with no evidence of localization to normal tissue. Here we extended the capabilities of this system to also include PTX delivery. NPs that will be loaded into NSCs must contain a high amount of drug by weight but constrain the release of the drug such that the NSCs are viable after loading and can successfully migrate to tumors. We developed silica coated PTX nanocrystals (Si[PTX-NC]) meeting these requirements. Si[PTX-NC] were more effective than uncoated PTX-NC or Abraxane for loading NSCs with PTX. NSCs loaded with Si[PTX-NC] maintained their migratory ability and, for low dose PTX, were more effective than free PTX-NC or Si[PTX-NC] at killing ovarian tumors in vivo. This work demonstrates that NSC/NP delivery is a platform technology amenable to delivering different therapeutics and enables the pursuit of NSC/NP targeted delivery of the entire preferred chemotherapy regimen for ovarian cancer. It also describes efficient silica coating chemistry for PTX nanocrystals that may have applications beyond our focus on NSC transport.

Enhanced Delivery of Oncolytic Adenovirus by Neural Stem Cells for Treatment of Metastatic Ovarian Cancer.

Oncolytic virotherapy is a promising approach for treating recurrent and/or drug-resistant ovarian cancer. However, its successful application in the clinic has been hampered by rapid immune-mediated clearance or neutralization of the virus, which reduces viral access to tumor foci. To overcome this barrier, patient-derived mesenchymal stem cells have been used to deliver virus to tumors, but variability associated with autologous cell isolations prevents this approach from being broadly clinically applicable. Here, we demonstrate the ability of an allogeneic, clonal neural stem cell (NSC) line (HB1.F3.CD21) to protect oncolytic viral cargo from neutralizing antibodies within patient ascites fluid and to deliver it to tumors within preclinical peritoneal ovarian metastases models. The viral payload used is a conditionally replication-competent adenovirus driven by the survivin promoter (CRAd-S-pk7). Because the protein survivin is highly expressed in ovarian cancer, but not in normal differentiated cells, viral replication should occur selectively in ovarian tumor cells. We found this viral agent was effective against cisplatin-resistant ovarian tumors and could be used as an adjunct treatment with cisplatin to decrease tumor burden without increasing toxicity. Collectively, our data suggest NSC-delivered CRAd-S-pk7 virotherapy holds promise for improving clinical outcome, reducing toxicities, and improving quality of life for patients with advanced ovarian cancer.

Concise Review: Neural Stem Cell-Mediated Targeted Cancer Therapies.

Cancer is one of the leading causes of morbidity and mortality worldwide, with 1,688,780 new cancer cases and 600,920 cancer deaths projected to occur in 2017 in the U.S. alone. Conventional cancer treatments including surgical, chemo-, and radiation therapies can be effective, but are often limited by tumor invasion, off-target toxicities, and acquired resistance. To improve clinical outcomes and decrease toxic side effects, more targeted, tumor-specific therapies are being developed. Delivering anticancer payloads using tumor-tropic cells can greatly increase therapeutic distribution to tumor sites, while sparing non-tumor tissues therefore minimizing toxic side effects. Neural stem cells (NSCs) are tumor-tropic cells that can pass through normal organs quickly, localize to invasive and metastatic tumor foci throughout the body, and cross the blood-brain barrier to reach tumors in the brain. This review focuses on the potential use of NSCs as vehicles to deliver various anticancer payloads selectively to tumor sites. The use of NSCs in cancer treatment has been studied most extensively in the brain, but the findings are applicable to other metastatic solid tumors, which will be described in this review. Strategies include NSC-mediated enzyme/prodrug gene therapy, oncolytic virotherapy, and delivery of antibodies, nanoparticles, and extracellular vesicles containing oligonucleotides. Preclinical discovery and translational studies, as well as early clinical trials, will be discussed. Stem Cells Translational Medicine 2018;7:740-747.

Long-term stability and computational analysis of migration patterns of L-MYC immortalized neural stem cells in the brain.

BACKGROUND: Preclinical studies indicate that neural stem cells (NSCs) can limit or reverse central nervous system (CNS) damage through delivery of therapeutic agents for cell regeneration. Clinical translation of cell-based therapies raises concerns about long-term stability, differentiation and fate, and absence of tumorigenicity of these cells, as well as manufacturing time required to produce therapeutic cells in quantities sufficient for clinical use. Allogeneic NSC lines are in growing demand due to challenges inherent in using autologous stem cells, including production costs that limit availability to patients. METHODS/PRINCIPAL FINDINGS: We demonstrate the long-term stability of L-MYC immortalized human NSCs (LM-NSC008) cells in vivo, including engraftment, migration, and absence of tumorigenicity in mouse brains for up to nine months. We also examined the distributions of engrafted LM-NSC008 cells within brain, and present computational techniques to analyze NSC migration characteristics in relation to intrinsic brain structures. CONCLUSIONS/SIGNIFICANCE: This computational analysis of NSC distributions following implantation provides proof-of-concept for the development of computational models that can be used clinically to predict NSC migration paths in patients. Previously, models of preferential migration of malignant tumor cells along white matter tracts have been used to predict their final distributions. We suggest that quantitative measures of tissue orientation and white matter tracts determined from MR images can be used in a diffusion tensor imaging tractography-like approach to describe the most likely migration routes and final distributions of NSCs administered in a clinical setting. Such a model could be very useful in choosing the optimal anatomical locations for NSC administration to patients to achieve maximum therapeutic effects.