LyP-1-Modified Oncolytic Adenoviruses Targeting Transforming Growth Factor β Inhibit Tumor Growth and Metastases and Augment Immune Checkpoint Inhibitor Therapy in Breast Cancer Mouse Models

Abstract

We report here the development of oncolytic adenoviruses (Ads) that have reduced toxicity, enhanced tumor tropism, produce strong antitumor response, and can overcome resistance to immune checkpoint inhibitor therapy in breast cancer. We have shown that LyP-1 receptor (p32) is highly expressed on the surface of breast cancer cells and tumors from cancer patients, and that increased stromal expression of transforming growth factor b-1 (TGFb-1) is associated with triple-negative breast cancer. Therefore, we constructed oncolytic Ads, AdLyp.sT and mHAdLyp.sT, in which the p32-binding LyP-1 peptide was genetically inserted into the adenoviral fiber protein. Both AdLyp.sT and mHAdLyp.sT express sTGFbRIIFc, a TGFb decoy that can inhibit TGFb pathways. mHAdLyp.sT is an Ad5/48 chimeric hexon virus in which hypervariable regions (HVRs 1-7) of Ad5 are replaced with the corresponding Ad48 HVRs. AdLyp.sT and mHAdLyp.sT exhibited better binding, replication, and produced higher sTGFbRIIFc protein levels in breast cancer cell lines compared with Ad.sT or mHAd.sT control viruses without LyP-1 peptide modification. Systemic delivery of mHAdLyp.sT in mice resulted in reduced hepatic/systemic toxicity compared with Ad.sT and AdLyp.sT. Intravenous delivery of AdLyp.sT and mHAdLyp.sT elicited a strong antitumor response in a human MDA-MB-231 bone metastasis model in mice, as indicated by bioluminescence imaging, radiographic tumor burden, serum TRACP 5b and calcium, and body weight analyses. Furthermore, intratumoral delivery of AdLyp.sT in 4T1 model in immunocompetent mice inhibited tumor growth and metastases, and augmented anti-PD-1 and anti-CTLA-4 therapy. Based on these studies, we believe that AdLyp.sT and mHAdLyp.sT can be developed as potential targeted immunotherapy agents for the treatment of breast cancer.

The Role of the Tumor Microenvironment in Developing Successful Therapeutic and Secondary Prophylactic Breast Cancer Vaccines

“…Oncolytic viruses (OV) transform immunotherapy from a sterile immunity to a pathogen-associated-like immunity, as these viruses replicate preferentially in tumor cells and release TAAs and PAMPS [ 132 , 133 ]. In this emerging in situ vaccination strategy in breast cancer models, pox- [ 134 , 135 ], reo- [ 136 ], herpes simplex [ 137 , 138 , 139 ], adeno- [ 140 , 141 , 142 , 143 ], Newcastle disease [ 144 , 145 ], Vesicular stomatitis [ 146 ], and measles [ 147 , 148 , 149 ] viruses have been employed to make breast tumors more immunogenic and more responsive to immunotherapies by priming the TME to signs of infection and releasing TAAs [ 150 ]. Moreover, these viruses can be engineered to express various TAAs to further prime an immunological response from TME.…”

“…Moreover, these viruses can be engineered to express various TAAs to further prime an immunological response from TME. In the last couple years, the field of oncolytic viruses in BC models has been burgeoning and opened new avenues for combination immunotherapies, such as vaccines [ 146 , 151 ], suppressor cell targeting [ 152 ], and checkpoint inhibitors [ 134 , 140 , 146 , 150 , 152 , 153 , 154 , 155 , 156 ]. Moreover, several BC clinical trials are currently ongoing (see Table 2 ).…”

“…In other instances, researchers have engineered viral-like nanoparticles with immune stimulatory molecules such as IL-33 [ 158 ]. Recently, a group of investigators engineered an oncolytic adenovirus expressing a TGF-β decoy (sTGFβRIIFc), which inhibited pro-tumorigenic signaling from fibroblasts in the TME of TNBC models [ 140 ]. This strategy was combined with checkpoint inhibitors in a 4T1 Balb/c model, which led to significant tumor regression.…”

“…Of interest, based on observation of CD8+ T-cell infiltration, Niavarani et al combined their vaccine-OV approach with anti-PD-1 treatment, which improved overall survival compared to PD-1 or vaccine-OV alone in a 4T1-Balb/c model [ 146 ]. This might be an indication of a wider trend in BC-OV research, as multiple groups have seen BC tumor regression and increased survival by combining OVs with checkpoint inhibitors [ 134 , 140 , 146 , 150 , 152 , 153 , 154 , 155 , 156 ]. Chon et al, for instance, used antibodies blocking both CTLA-4 and PD-1 in a MMTV-PyMT transgenic mouse model administration of mJX-594 (JX), a vaccinia virus engineered to express GM-CSF and is attenuated via viral thymidine kinase disruption [ 153 ].…”

Viroimmunotherapy for breast cancer: promises, problems and future directions

“…The virus was found to effectively kill HER2 + BrCa cells in vitro and resulted in a significant anti-tumor effect against a HER2 + tumor in mice. Another oncolytic adenovirus for BrCa was studied by Xu et al [50], in which the fiber protein was altered to include Lyp-1 peptide to reduce hepatic toxicity and increase infection of BrCa cells, which usually express high levels of Lyp-1 receptor. Furthermore, the virus was engineered to encode a decoy of transforming growth factor-beta (TGF-β).…”