**5. Safety Considerations**

To date, clinical experience with virotherapy-enhancing combinations is limited and our current understanding on the synergism of select combinations has been based on extensive preclinical studies. Twenty years of clinical testing of OV's in monotherapy settings have underlined their excellent safety profile with grade 1 and 2 being the most commonly reported adverse events [5]. To what extent some small molecule combinations may compromise such a safety profile or adversely affect the overall therapeutic efficacy of oncolytic viruses is currently, in large part, subject to conjecture and should therefore be carefully addressed in pre-clinical settings. For example, dimethyl fumarate potentiates replication and oncolysis induced by VSV∆M51 [66], but lowers leukocyte counts and can result in reactivation of JC virus, leading to multifocal leukoencephalopathy (PML). Some HDIs have also been shown to reactivate latent HIV [292], EBV and HSV-1 [293]. The risk that such compounds may reactivate a second virus, with that virus' interactions with the initial oncolytic virus being unknown, should not be underestimated. The specific inhibition profiles of the particular small molecule, as well as the OV in question, will also determine the outcome of an OV/drug combination. While enhancing OV replication, inhibition of certain HDACs (HDAC 2, 6, 11) may enhance Treg function [294], so choosing a drug with a favorable profile, selection of patients with low tumor Treg counts or careful scheduling of the drug and OV may enhance the final anti-tumor synergy. In addition, some virotherapy-enhancing combinations may also potentially enhance the safety profile. For example, ruxolitinib has long been proposed to enhance activity of numerous OVs due to countering the antiviral JAK/STAT signaling and no toxicities have been reported in different preclinical studies [44,49]. However, its combination with an interferon-armed VSV-hIFN-NIS in two current clinical trials (see Table 3) may also act to offset potential toxicities caused by excessive production of the interferon transgene in particularly permissive tumors.

**Table 3.** Currently active \* clinical trials with oncolytic virus and small molecule compound combinations.



**Table 3.** *Cont*.

AdV, adenovirus; GM-CSF, granulocyte-macrophage colony-stimulating factor; hIFN, human interferon; HSV-1, herpes simplex virus type 1; ICP, infected cell protein; TMZ-CD40L, trimerized membrane-bound CD40 ligand; VSV, vesicular stomatitis virus; VV, vaccinia virus. \* clinicaltrials.org accessed on 23 June 2021; search term "oncolytic"; filters "recruiting" and "active, not recruiting".
