Tumour Evolution.

<p>A hypothetical pathway of tumour evolution from a normal cell to an advanced-stage cancer. Mutations in key regulatory genes lead to changes in cell physiology that favour tumour growth. Over time, these genetic defects accumulate to confer many of the known hallmarks of cancer <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003836#ppat.1003836-Hanahan2" target="_blank">[131]</a>. The sequence of these events and the timing represented here is only one example of how this might occur.</p

Evolution of viral fitness and cytotoxicity in MEFs.

<p>VSV was evolved in p53−/− MEFs for 40 serial passages (MOI = 0.05) and, for each evolved lineage (L1–L4), fitness and cytotoxicity were assayed in p53−/− MEFs (a, b) as well as in isogenic p53+/+ MEFs (c, d). Fitness was determined in four independent competition assays (24 hpi) at a 1∶1 input ratio against a common competitor isogenic to the WT. Cytotoxicity was calculated as the fraction of live cells at 18 hpi relative to uninfected cultures using the MTS assay. For each line, changes in viral fitness and cytotoxicity were evaluated using a one-sample t-test against the WT value (*: <i>P</i><0.05). The horizontal dashed lines indicate WT values, and error bars indicate the SEM.</p

Half-maximal effective dose (EC₅₀) of the evolved VSV lines L2 and L3, VSV WT and the VSV mutant MΔ51 in 4T1 and CT-26 cells.

<p>Cells were inoculated at the indicated MOI and viability was measured after 48 h using the Alamar Blue assay. EC₅₀ values and 95% confidence intervals were inferred from non-linear least-squares regression of cell viability against the MOI, as detailed in the Methods section.</p

Oncolytic Viruses Are Designed to Grow in the Tumour Niche.

<p>There are at least six key critical features of tumour cell growth that can be targeted by oncolytic viruses. These include changes in the expression of viral-host cell receptors, the antiviral response, nucleotide and protein synthesis, cell proliferation, and apoptosis. A number of engineered or selected oncolytic viruses exist that can exploit one or more of these malignant characteristics.</p

Western blot analysis of p53 expression and function.

<p>CT-26, Vero or 4T1 cells were irradiated with UV at the indicated doses to induce DNA damage and tested 16 h post-irradiation for expression of p53, p21, and the constitutively expressed GAPDH. The Western blot shows that p53 protein is expressed in all cells but is functional only in CT-26 and Vero cells, as judged by p21 levels.</p

Genetic changes found in the four evolved lines.

<p>For each gene, the nucleotide and amino acid substitutions are shown. Blue and red squares indicate synonymous and non-synonymous substitutions, respectively. The C10224U substitution is in hatchet to indicate that this position was polymorphic.</p

Effect of VSV L3 and WT on the growth of syngeneic Balb/c 4T1 and CT-26 tumors.

<p>Viruses were injected intratumorally 8 days (4T1) and 11 days (CT-26) after engraftment, and a second dose was administered 7 days later to 4T1-bearing mice. The experiment was terminated based on end-point criteria on days 14 (4T1) and 7 (CT-26). Top panels (a, c) show tumor growth throughout the course of the experiment for mice treated with the VSV L3 (black circles), VSV WT (grey circles) and the mock-inoculated control group (white squares). Bottom panels (b, d) indicate the tumor growth rate calculated between each viral dose and the first subsequent tumor measurement. The tumor growth rates of each group were compared using t-tests (**: <i>P</i><0.01; *: <i>P</i><0.05; n.s.: <i>P</i>>0.05). Error bars indicate the SEM.</p

DataSheet_1_Unlocking the potential of dimethyl fumarate: enhancing oncolytic HSV-1 efficacy for wider cancer applications.pdf

Immunotherapy and specifically oncolytic virotherapy has emerged as a promising option for cancer patients, with oncolytic herpes simplex virus-1 (oHSV-1) expressing granulocyte macrophage colony stimulating factor being the first OV to be approved by the FDA for treatment of melanoma. However, not all cancers are sensitive and responsive to oncolytic viruses (OVs). Our group has demonstrated that fumaric and maleic acid esters (FMAEs) are very effective in sensitizing cancer cells to OV infection. Of note, these FMAEs include dimethyl fumarate (DMF, also known as Tecfidera®), an approved treatment for multiple sclerosis and psoriasis. This study aimed to assess the efficacy of DMF in combination with oncolytic HSV-1 in preclinical cancer models. We demonstrate herewith that pre-treatment with DMF or other FMAEs leads to a significant increase in viral growth of oHSV-1 in several cancer cell lines, including melanoma, while decreasing cell viability. Additionally, DMF was able to enhance ex vivo oHSV-1 infection of mouse-derived tumor cores as well as human patient tumor samples but not normal tissue. We further reveal that the increased viral spread and oncolysis of the combination therapy occurs via inhibition of type I IFN production and response. Finally, we demonstrate that DMF in combination with oHSV-1 can improve therapeutic outcomes in aggressive syngeneic murine cancer models. In sum, this study demonstrates the synergistic potential of two approved therapies for clinical evaluation in cancer patients.</p

Additional file 2: Figure S2. of Combination of Paclitaxel and MG1 oncolytic virus as a successful strategy for breast cancer treatment

PAC blocks IFNβ production by infected tumor cells. A Microscopy pictures of EMT6, 4 T1 and EO771 tumor cells pre-treated or not for 4 h with recombinant IFNβ. B The IFNβ released from MG1-infected cells in the presence or absence of PAC was quantified by ELISA. Samples were analysed in triplicates and statistical significance was calculated using the unpaired two-tailed t test with Welch’s correction; *p < 0.05, **p < 0.01, ***p < 0.001. (TIF 6796 kb

Additional file 4: Figure S4. of Combination of Paclitaxel and MG1 oncolytic virus as a successful strategy for breast cancer treatment

PAC and MG1 synergistically kill breast cancer cell lines. Quantification of the signal obtained for triplicates or quadruplicates Coomassie Blue staining of EMT6, 4 T1 or EO771 cells infected or not with MG1-GFP and co-treated with 2 uM PAC for 48 h from Fig. 3. Statistical significance was calculated using the unpaired two-tailed t test with Welch’s correction; *p < 0.05, **p < 0.01, ***p < 0.001. (TIF 2942 kb