Harold Atkins

Exploiting tumor-specific defects in the interferon pathway with a previously unknown oncolytic virus.

Interferons are circulating factors that bind to cell surface receptors, activating a signaling cascade, ultimately leading to both an antiviral response and an induction of growth inhibitory and/or apoptotic signals in normal and tumor cells. Attempts to exploit the ability of interferons to limit the growth of tumors in patients has met with limited results because of cancer-specific mutations of gene products in the interferon pathway. Although interferon-non-responsive cancer cells may have acquired a growth/survival advantage over their normal counterparts, they may have simultaneously compromised their antiviral response. To test this, we used vesicular stomatitis virus (VSV), an enveloped, negative-sense RNA virus exquisitely sensitive to treatment with interferon. VSV rapidly replicated in and selectively killed a variety of human tumor cell lines even in the presence of doses of interferon that completely protected normal human primary cell cultures. A single intratumoral injection of VSV was effective in reducing the tumor burden of nude mice bearing subcutaneous human melanoma xenografts. Our results support the use of VSV as a replication-competent oncolytic virus and demonstrate a new strategy for the treatment of interferon non-responsive tumors.

VSV strains with defects in their ability to shutdown innate immunity are potent systemic anti-cancer agents

Ideally, an oncolytic virus will replicate preferentially in malignant cells, have the ability to treat disseminated metastases, and ultimately be cleared by the patient. Here we present evidence that the attenuated vesicular stomatitis strains, AV1 and AV2, embody all of these traits. We uncover the mechanism by which these mutants are selectively attenuated in interferon-responsive cells while remaining highly lytic in 80% of human tumor cell lines tested. AV1 and AV2 were tested in a xenograft model of human ovarian cancer and in an immune competent mouse model of metastatic colon cancer. While highly attenuated for growth in normal mice, both AV1 and AV2 effected complete and durable cures in the majority of treated animals when delivered systemically.

Characterization of Transgenic Mice with Targeted Disruption of the Catalytic Domain of the Double-stranded RNA-dependent Protein Kinase, PKR

The interferon-inducible, double-stranded RNA-dependent protein kinase PKR has been implicated in anti-viral, anti-tumor, and apoptotic responses. Others have attempted to examine the requirement of PKR in these roles by targeted disruption at the amino terminal-encoding region of the Pkr gene. By using a strategy that aims at disruption of the catalytic domain of PKR, we have generated mice that are genetically ablated for functional PKR. Similar to the other mouse model of Pkr disruption, we have observed no consequences of loss of PKR on tumor suppression. Anti-viral response to influenza and vaccinia also appeared to be normal in mice and in cells lacking PKR. Cytokine signaling in the type I interferon pathway is normal but may be compromised in the erythropoietin pathway in erythroid bone marrow precursors. Contrary to the amino-terminal targeted Pkr mouse, tumor necrosis factor α-induced apoptosis and the anti-viral apoptosis response to influenza is not impaired in catalytic domain-targetedPkr-null cells. The observation of intact eukaryotic initiation factor-2α phosphorylation in these Pkr-null cells provides proof of rescue by another eukaryotic initiation factor-2α kinase(s).

The therapeutic potential of mesenchymal stem cell transplantation as a treatment for multiple sclerosis: consensus report of the International MSCT Study Group

Current therapies for multiple sclerosis effectively reduce inflammation, but do little in terms of repair to the damaged central nervous system. Cell-based therapies may provide a new strategy for bolstering regeneration and repair through neuro-axonal protection or remyelination. Mesenchymal stem cells modulate pathological responses in experimental autoimmune encephalitis, alleviating disease, but also stimulate repair of the central nervous system through the release of soluble factors. Autologous and allogeneic mesenchymal stem cells have been safely administered to individuals with hemato-oncological diseases and in a limited number of patients with multiple sclerosis. It is therefore reasonable to move mesenchymal stem cells transplantation into properly controlled human studies to explore their potential as a treatment for multiple sclerosis. Since it is likely that the first such studies will probably involve only small numbers of patients in a few centers, we formed an international panel comprising multiple sclerosis neurology and stem cell experts, as well as immunologists. The aims were to derive a consensus on the utilization of mesenchymal stem cells for the treatment of multiple sclerosis, along with protocols for the culture of the cells and the treatment of patients. This article reviews the consensus derived from our group on the rationale for mesenchymal stem cell transplantation, the methodology for generating mesenchymal stem cells and the first treatment protocol for multiple sclerosis patients.

Chemical targeting of the innate antiviral response by histone deacetylase inhibitors renders refractory cancers sensitive to viral oncolysis

Intratumoral innate immunity can play a significant role in blocking the effective therapeutic spread of a number of oncolytic viruses (OVs). Histone deacetylase inhibitors (HDIs) are known to influence epigenetic modifications of chromatin and can blunt the cellular antiviral response. We reasoned that pretreatment of tumors with HDIs could enhance the replication and spread of OVs within malignancies. Here, we show that HDIs markedly enhance the spread of vesicular stomatitis virus (VSV) in a variety of cancer cells in vitro, in primary tumor tissue explants and in multiple animal models. This increased oncolytic activity correlated with a dampening of cellular IFN responses and augmentation of virus-induced apoptosis. These results illustrate the general utility of HDIs as chemical switches to regulate cellular innate antiviral responses and to provide controlled growth of therapeutic viruses within malignancies. HDIs could have a profoundly positive impact on the clinical implementation of OV therapeutics.

A let-7 MicroRNA-sensitive Vesicular Stomatitis Virus Demonstrates Tumor-specific Replication

Creation of potent oncolytic viruses (OVs) suitable for the clinic may require new strategies in virus design. Replication-competent viruses facilitate a variety of approaches to achieving tumor specificity. Altered expression of microRNAs is a common hallmark of cancer that we demonstrate can be used to alter expression of a potent wild-type viral gene to achieve tumor-specific replication of an engineered vesicular stomatitis virus (VSV). Incorporation of let-7 microRNA complementary sequences within VSV eliminates undesirable replication and associated toxicity in normal cells but permits growth in cancer cells in vitro and in vivo. This is proof of concept that viruses designed to exploit the differential microRNA expression in cancer cells is a viable approach, potentially useful in optimizing oncolytic viral gene expression for maximal antitumor activity and safety.

Differential genomic targeting of the transcription factor TAL1 in alternate haematopoietic lineages

TAL1/SCL is a master regulator of haematopoiesis whose expression promotes opposite outcomes depending on the cell type: differentiation in the erythroid lineage or oncogenesis in the T‐cell lineage. Here, we used a combination of ChIP sequencing and gene expression profiling to compare the function of TAL1 in normal erythroid and leukaemic T cells. Analysis of the genome‐wide binding properties of TAL1 in these two haematopoietic lineages revealed new insight into the mechanism by which transcription factors select their binding sites in alternate lineages. Our study shows limited overlap in the TAL1‐binding profile between the two cell types with an unexpected preference for ETS and RUNX motifs adjacent to E‐boxes in the T‐cell lineage. Furthermore, we show that TAL1 interacts with RUNX1 and ETS1, and that these transcription factors are critically required for TAL1 binding to genes that modulate T‐cell differentiation. Thus, our findings highlight a critical role of the cellular environment in modulating transcription factor binding, and provide insight into the mechanism by which TAL1 inhibits differentiation leading to oncogenesis in the T‐cell lineage.

The global landscape of stem cell clinical trials

AIM To provide a comprehensive analysis of clinical trials (CTs) listed in worldwide registries involving new applications for stem cell-based treatments and account for the role of industry. MATERIALS & METHODS We developed a data set of 4749 stem cell CTs up to 2013 in worldwide registries. We defined 1058 novel CTs (i.e., trials that were not observational in nature; did not involve an established stem cell therapy for an established indication, such as hematopoietic stem cells for leukemia; and did not investigate supportive measures). Based on trial descriptions, we manually coded these for eight additional elements. RESULTS Our analysis details the characteristics of novel stem cell CTs (e.g., stem cell types being tested, disease being targeted, and whether interventions were autologous or allogeneic), geotemporal trends, and private sector involvement as sponsor or collaborator. CONCLUSION The field is progressing at a steady pace with emerging business models for stem cell therapeutics. However, therapeutic rhetoric must be tempered to reflect current clinical and research realities.

Identification of Genetically Modified Maraba Virus as an Oncolytic Rhabdovirus

To expand our current array of safe and potent oncolytic viruses, we screened a variety of wild-type (WT) rhabdoviruses against a panel of tumor cell lines. Our screen identified a number of viruses with varying degrees of killing activity. Maraba virus was the most potent of these strains. We built a recombinant system for the Maraba virus platform, engineered a series of attenuating mutations to expand its therapeutic index, and tested their potency in vitro and in vivo. A double mutant (MG1) strain containing both G protein (Q242R) and M protein (L123W) mutations attenuated Maraba virus in normal diploid cell lines, yet appeared to be hypervirulent in cancer cells. This selective attenuation was mediated through interferon (IFN)-dependent and -independent mechanisms. Finally, the Maraba MG1 strain had a 100-fold greater maximum tolerable dose (MTD) than WT Maraba in vivo and resulted in durable cures when systemically administered in syngeneic and xenograft models. In summary, we report a potent new oncolytic rhabdovirus platform with unique tumor-selective attenuating mutations.

Targeting Tumor Vasculature With an Oncolytic Virus

Oncolytic viruses (OVs) have been engineered or selected for cancer cell-specific infection however, we have found that following intravenous administration of vesicular stomatitis virus (VSV), tumor cell killing rapidly extends far beyond the initial sites of infection. We show here for the first time that VSV directly infects and destroys tumor vasculature in vivo but leaves normal vasculature intact. Three-dimensional (3D) reconstruction of infected tumors revealed that the majority of the tumor mass lacks significant blood flow in contrast to uninfected tumors, which exhibit relatively uniform perfusion. VSV replication in tumor neovasculature and spread within the tumor mass, initiates an inflammatory reaction including a neutrophil-dependent initiation of microclots within tumor blood vessels. Within 6 hours of intravenous administration of VSV and continuing for at least 24 hours, we observed the initiation of blood clots within the tumor vasculature whereas normal vasculature remained clot free. Blocking blood clot formation with thrombin inhibitors prevented tumor vascular collapse. Our results demonstrate that the therapeutic activity of an OV can go far beyond simple infection and lysis of malignant cells.