Background Interleukins (ILs)-12 and -2 are recognized as potent anti-tumor molecules; yet, balancing effective dosing while mitigating systemic toxicity presents a significant hurdle for clinical use. A targeted delivery system that provides sustained local cytokine levels in the tumor microenvironment, while minimizing systemic exposure and its associated toxicities, may effectively tip the balance to overcome the recognized limitations of IL-12 and -2 therapies. Using its clinically validated replication-defective herpes simplex virus type 1 (HSV-1)-derived platform technology, Krystal Biotech, Inc. has developed KB707, a vector encoding human IL-12 and -2 for the treatment of solid tumors.

Methods For preclinical development, vectors were constructed to express murine Il12 and Il2, termed KB703 and KB704, respectively, as the human cytokines are only partially cross-reactive in mice. Transgene bioactivity was confirmed in vitro prior to animal studies. Efficacy of localized IL-12 and -2 delivery was then tested in the K7M2 Osteosarcoma lung metastases model. BALB/c mice were inoculated with K7M2 tumors IV on day 0, and cohorts were treated via inhalation with vehicle, single, or combinatorial vector therapy on days 14, 21, and 28.

Results In vivo, once weekly inhalation of combined KB703/KB704 was well tolerated in healthy mice, and IL-12 and IL-2 were detected in lung tissue and bronchoalveolar lavage fluid. Systemic cytokine exposure was also compared between inhaled vector recipients and animals treated intravenously (IV) with recombinant proteins at clinically relevant doses. Peak vector-derived IL-12 and IL-2 serum concentrations were considerably reduced compared to IV recombinant protein therapy, suggesting limited systemic cytokine exposure. In K7M2 studies, single agents demonstrated some efficacy over vehicle alone, yet combined KB703/KB704 therapy resulted in 100% overall survival at day 100 (p<0.01 compared to vehicle control). Histological analysis of lungs from KB703/KB704-treated animals revealed no tumor burden, with moderate peri-vascular and peri-bronchial immune cell infiltrate, suggesting immune-mediated control of tumor growth. A rechallenge study was then conducted where surviving KB703/KB704 dosed animals were reinoculated with K7M2 tumors IV 75 days after their first inoculation. Untreated, tumor recipient animals demonstrated a median survival of 36 days, while vector treated, rechallenged animals had a median survival >66 days post-rechallenge, suggesting a durable anti-tumor memory response. Additionally, a therapeutic benefit of KB703/KB704 therapy has been observed in a model of primary lung tumors.

Conclusions Overall, these results highlight the potential of Krystal’s HSV-1-based platform for sustained delivery of IL-12 and IL-2 as a novel immunotherapy for the treatment of primary and secondary lung tumors.

Ethics Approval The studies described here were performed in an AAALAC accredited facility, and protocols were IACUC approved prior to initiation.