Combinational Immunotherapy with Allo-DRibble Vaccines and Anti-OX40 Co-Stimulation Leads to Generation of Cross-Reactive Effector T Cells and Tumor Regression

Abstract

It is well-known that vaccines comprising of irradiated whole tumor cells or tumor-derived heat shock proteins can generate tumor-specific immune responses. In contrast, we showed recently that vaccines composed of autophagosomes (DRibbles) derived from syngeneic sarcomas could induce cross-reactive T-cell responses and cross-protection against the tumor. This unusual property of DRibbles was related to the selective recruitment of defective ribosomal products (DRiPs) and other short-lived proteins (SLiPs) into autophagosomes via sequestosome (SQSTM1, p62) mediated association of ubiquitinated SLiPs to the autophagy gene product LC3. Here, we extend our observations to mammary carcinomas from mice of different genetic background. We demonstrated that combined of intranodal administration of autologous or allogeneic DRibbles together with anti-OX40 antibody led to robust proliferation, expansion, and differentiation of memory and effector T cells. We also showed that SLiPs is an excellent source of antigen for cross-priming of CD8+ T-cells that recognize shared tumor antigens in the context of host MHC class I molecules. Thus, our results provide a strong basis for novel clinical trials that combine allogeneic “off-the-shelf” DRibble vaccines together with antibodies against co-stimulatory molecules.

Targeting autophagy in cancer

“…Inhibition of autophagy at this stage prevents breakdown of DRiPs and SLiPs that have concentrated in the autophagosome, and allows for fractionation and collection of the DRibble corpuscles to provide the protein needed to create effective DRibble vaccines 89 . DRibble tumor vaccines developed from these proteins have been shown to induce cross-reactive T-cell responses and tumor antigen cross- protection 90 . Preliminary analysis of a Phase II trial evaluating the use of DRibble vaccines in patients with non-small cell lung cancer (NSCLC) demonstrated that at 12 weeks, PBMCs from treated patients had multiple induced and increased antibody responses 91 .…”

<p>Anti-Tumor Efficacy of an Adjuvant Built-In Nanovaccine Based on Ubiquitinated Proteins from Tumor Cells</p>

“…Vx3 protein was expressed and purified based on our previous work. 19,20 In brief, pUbiG101-Vx3(A7)-eGFP expressing plasmid was introduced into E. coli DH5-α competent cells (Invitrogen) to induce the expression of His-Vx3-eGFP fusion protein. Next, cells were harvested and treated with lysozyme (Sigma).…”

“…12 Recent studies have confirmed that DRibbles (defective ribosomal products-containing blebs) isolated from tumor cells with the induction of autophagy and inhibition of lysosomal/proteasomal activity are sufficient to stimulate dramatic T-cell activation and kill carcinoma cells in different tumor models such as melanoma, lung cancer, breast cancer and liver cancer. [13][14][15][16][17][18][19] Moreover, we have demonstrated that ubiquitinated proteins (UPs) are the critical TAA source of DRibbles which induce the antitumor efficacy. Therefore, different strategies for UPs enrichment have been developed to achieve a clinically safe, simply made, and environment-friendly vaccine with enhanced antitumor immune response.…”

“…Therefore, different strategies for UPs enrichment have been developed to achieve a clinically safe, simply made, and environment-friendly vaccine with enhanced antitumor immune response. [19][20][21] In our previous studies, we enriched UPs from tumor cells after proteasome inhibition by Ni-NTA agarose beads conjugated with ubiquitin-binding protein Vx3. We found that the UPs have the ability to be an effective cancer vaccine.…”

“…Nevertheless, those UPs are lack of highly immunogenic and the approach is time-consuming. 19,20 To optimize the therapeutic vaccine based on UPs, a CONH linker was applied in this study to couple Vx3 protein to α-Al 2 O 3 nanoparticles to generate Vx3-conjugated α-Al 2 O 3 nanoparticles (denoted as α-Al 2 O 3 -CONH-Vx3), which enriched UPs from 4T1 cancer cell lysate (α-Al 2 O 3 -CONH-Vx3-UPs, denoted as α-Al 2 O 3 -UPs) by a single centrifugation step. By using α-Al 2 O 3 -CONH-Vx3, Vx3 pulls UPs out of tumor cell lysate like a fishhook, and α-Al 2 O 3 functions as an immune adjuvant to enhance the immunogenicity of UPs.…”