Systemic Delivery of a Dual PI3K/mTOR Inhibitor More Effective than Topical Delivery in Preventing Anal Carcinogenesis in an HPV Transgenic Mouse Model

Abstract

Introduction: Anal dysplasia is a growing health concern that over time can result in squamous cell carcinoma (SqCC) of the anus. In this study, we compare a topical versus systemic (oral) administration of LY3023414, a dual PI3K/mTOR inhibitor, to prevent anal carcinogenesis in a Human Papillomavirus (HPV) mouse model of anal cancer.

Materials and methods: K14E6/E7 transgenic mice were used to model HPV-induced anal carcinogenesis. Mice with varying starting anal histologies (normal histology, low-grade, and high-grade anal dysplasia) were treated topically at the anus or systemically via oral gavage with LY3023414 with or without topical carcinogen for 20 weeks. Mice were monitored for overt anal tumor development and anal tissue was assessed for histology and markers of PI3K and mTOR activity (pAKT and pS6, respectively).

Results: LY3023414 treatment, regardless of the mode of delivery, significantly decreased overt tumor development in mice starting with normal histology and low-grade anal dysplasia. Systemic LY3023414 treatment was more effective in delaying tumor onset than topical treatment. Mice treated with systemic LY3023414 had significantly reduced rates of anal SqCC when starting with normal and low-grade anal dysplasia compared to topical treatment. Topical treatment was only effective in reducing SqCC in the setting of low-grade dysplasia. LY3023414 inhibition of pAKT and pS6 expression varied with starting histology. Neither treatment mode was effective in the setting of high-grade anal dysplasia.

Conclusion: Systemic LY3023414 treatment was more effective than topical application in delaying the progression of normal anal histology and low-grade dysplasia to anal cancer in HPV-associated mice.

Keywords: Anal cancer; Anal dysplasia; Dual PI3K/mTOR inhibitor; LY3023414; Squamous cell carcinoma of the anus.

Figure 1:

The percentage of K14E6/E7 mice in each treatment group that developed overt anal tumors at any point during the 20-week period treatment. The notation * represents statistical significance (p <0.05) between groups. A. Mice that began treatment at 5 weeks of age/ normal anal histology. B. Mice that began treatment at 15 weeks of age/ low-grade anal dysplasia. C. Mice that began treatment at 25 weeks of age/ high-grade anal dysplasia.

Figure 2:

Tumor-free survival as assessed by Kaplan Meyer survival analysis over the 20-week treatment period. A. Mice that began treatment at 5 weeks of age/ normal anal histology. The green-purple-black line shows where no treatment, topical LY3023414-only, and systemic LY3023414-only groups overlap. Statistical significance (p <0.05) was achieved between groups: DMBA-only and systemic LY3023414 + DMBA (red and yellow lines, respectively) and DMBA-only and topical LY3023414 + DMBA groups (red and blue lines, respectively). B. Mice that began treatment at 15 weeks of age/ low-grade anal dysplasia. The green-black line shows where no treatment and systemic LY3023414-only groups overlap. Statistical significance (p <0.05) was achieved between groups: DMBA-only and systemic LY3023414 + DMBA (red and yellow lines, respectively) and DMBA-only and topical LY3023414 + DMBA groups (red and blue lines, respectively) C. Mice that began treatment at 25 weeks of age/ high-grade anal dysplasia. The purple-black line shows where topical LY3023414-only and systemic LY3023414-only groups overlap.

Figure 3:

The percentage of K14E6/E7 mice that developed squamous cell carcinoma (SqCC) of the anus in each treatment group at the end of the 20-week treatment period as assessed by histology: H&E-stained anal tissue. The notation * represents statistical significance (p < 0.05) between groups. A. Mice that began treatment at 5 weeks of age/ normal anal histology. B. Mice that began treatment at 15 weeks of age/ low-grade anal dysplasia. C. Mice that began treatment at 25 weeks of age/high-grade anal dysplasia.

Figure 4:

Mean pAKT and pS6 protein expression, markers of PI3K and mTOR activity respectively, quantified via immunohistochemical staining sections of K14E6/E7 mice anal tissue. Mean ± standard deviation values are reported in the RawIntDen/Area (raw integrated density/area). The notation * represents statistical significance (p < 0.05). A, B. Mice that began treatment at 5 weeks of age/ normal anal histology. C, D. Mice that began treatment at 15 weeks of age/ low-grade anal dysplasia. E, F. Mice that began treatment at 25 weeks of age/ high-grade anal dysplasia.