Oncolytic adenoviruses expressing checkpoint inhibitors for cancer therapy.

Despite the remarkable success of immune checkpoint inhibitors (ICIs), primary resistance to ICIs causes only subsets of patients to achieve durable responses due to the complex tumor microenvironment (TME). Oncolytic viruses (OVs) can overcome the immunosuppressive TME and promote systemic antitumor immunity in hosts. Engineered OVs armed with ICIs would likely have improved effectiveness as a cancer therapy. According to the diverse immune cell landscapes among different types of tumors, we rationally and precisely generated three recombinant oncolytic adenoviruses (OAds): OAd-SIRPα-Fc, OAd-Siglec10-Fc and OAd-TIGIT-Fc. These viruses were designed to locally deliver SIRPα-Fc, Siglec10-Fc or TIGIT-Fc fusion proteins recognizing CD47, CD24 or CD155, respectively, in the TME to achieve enhanced antitumor effects. Our results suggested that OAd-SIRPα-Fc and OAd-Siglec10-Fc both showed outstanding efficacy in tumor suppression of macrophage-dominated tumors, while OAd-TIGIT-Fc showed the best antitumor immunity in CD8+ T-cell-dominated tumors. Importantly, the recombinant OAds activated an inflammatory immune response and generated long-term antitumor memory. In addition, the combination of OAd-Siglec10-Fc with anti-PD-1 significantly enhanced the antitumor effect in a 4T1 tumor model by remodeling the TME. In summary, rationally designed OAds expressing ICIs tailored to the immune cell landscape in the TME can precisely achieve tumor-specific immunotherapy of cancer.

In Situ Cocktail Nanovaccine for Cancer Immunotherapy.

In situ vaccination is a desirable strategy for cancer immunotherapy due to its convenience and capacity to target tumor antigens. Here, an in situ nanovaccine based on a cationic peptide with cholesterol-modified, DP7-C, for cancer immunotherapy is rationally designed, and developed a cancer nanovaccine that is easy to preparate. The nanovaccine includes cocktail small interfering RNAs (siRNAs) and immunologic adjuvant CpG ODNs, has synergistic effect in the cancer treatment. This nanovaccine can induce tumor cell death, promote antigen presentation and relieve immune suppression in the tumor microenvironment (TME). Moreover, this nanovaccine is administered to CT26 (hot) and B16F10 (cold) tumor model mice, in which it targeted the primary tumors and induced systemic antitumor immunity to inhibit metastasis. It is validated that the nanovaccine can convert cold tumors into hot tumors. Furthermore, the nanovaccine increased the immune response to anti-PD-1 therapy by modulating the TME in both CT26- and B16F10-tumor-bearing mice. The siRNA cocktail/CpG ODN/self-assembling peptide nanovaccine is a simple and universal tool that can effectively generate specific tumor cell antigens and can be combined with immuno-oncology agents to enhance antitumor immune activity. The versatile methodology provides an alternative approach for developing cancer nanovaccines.

A Dendrimer Peptide (KK2DP7) Delivery System with Dual Functions of Lymph Node Targeting and Immune Adjuvants as a General Strategy for Cancer Immunotherapy.

The clinical efficacy of personalized cancer vaccines still needs to be improved due to their insufficient immune effect. The development of innovative adjuvants and lymph node-targeted delivery systems is the key to improving the clinical efficacy of personalized vaccines. However, there is still a lack of an adjuvant delivery system that is simple in preparation and capable of mass production and integrates adjuvant and lymph node targeted delivery functions. Here, this work reports that a simple dendrimer polypeptide (KK2DP7) nanoparticle enhances the immune efficacy of an OVA/neoantigen-based vaccine. Due to its multiple functions as a delivery vehicle, immune adjuvant, and facilitator of dendritic cell migration, KK2DP7 efficiently increases the efficiency of antigen uptake and cross-presentation by antigen-presenting cells (APCs) and delivers antigens to lymph nodes via APCs. Strikingly, the antitumor effect of KK2DP7/OVA is superior to that of commonly used adjuvants such as poly(I:C), CpG, and aluminum adjuvant combined with OVA. Furthermore, KK2DP7/OVA combined with anti-PD-1 antibody is able to prevent tumor recurrence in a postoperative recurrent tumor model. Thus, KK2DP7-based cancer vaccines alone or in combination with immune checkpoint blockade therapies to treat tumors or postoperative tumor recurrence are a powerful strategy to enhance antitumor immunity.

Engineering strategies to enhance oncolytic viruses in cancer immunotherapy.

Oncolytic viruses (OVs) are emerging as potentially useful platforms in treatment methods for patients with tumors. They preferentially target and kill tumor cells, leaving healthy cells unharmed. In addition to direct oncolysis, the essential and attractive aspect of oncolytic virotherapy is based on the intrinsic induction of both innate and adaptive immune responses. To further augment this efficacious response, OVs have been genetically engineered to express immune regulators that enhance or restore antitumor immunity. Recently, combinations of OVs with other immunotherapies, such as immune checkpoint inhibitors (ICIs), chimeric antigen receptors (CARs), antigen-specific T-cell receptors (TCRs) and autologous tumor-infiltrating lymphocytes (TILs), have led to promising progress in cancer treatment. This review summarizes the intrinsic mechanisms of OVs, describes the optimization strategies for using armed OVs to enhance the effects of antitumor immunity and highlights rational combinations of OVs with other immunotherapies in recent preclinical and clinical studies.

Lactobacillus reuteri Alleviates Gastrointestinal Toxicity of Rituximab by Regulating the Proinflammatory T Cells in vivo.

Rituximab (RTX) is a widely used anticancer drug with gastrointestinal side effects, such as nausea, vomiting, and diarrhea. The reason for these side effects is still poorly understood. Previous studies have reported that the intestinal microbiota is associated with the occurrence of disease and the therapeutic effect of drugs. In this study, we observed mucosal damage, inflammatory cell infiltration and increased intestinal inflammatory factor expression in RTX-treated mice. RTX also changed the diversity of the intestinal microbiota in mice, and decreased abundance of Lactobacillus reuteri was observed in RTX-treated mice. Further experiments revealed that intragastric administration of L. reuteri in RTX-treated mice attenuated the intestinal inflammatory response induced by RTX and regulated the proportion of helper T (Th) cells. In conclusion, our data characterize the effect of the intestinal microbiota on RTX-induced intestinal inflammation, suggesting that modifying the gut microbiota may represent a positive strategy for managing adverse reactions.

Rational design of innate defense regulator peptides as tumor vaccine adjuvants.

The development of adjuvants has been an empirical process. Efforts to develop a new design and evaluation system for novel adjuvants are not only desirable but also necessary. Moreover, composite adjuvants that contain two or more types of adjuvants to synergistically enhance the immune response are important for adjuvant and vaccine design. Innate defense regulator peptides (IDRs) are promising adjuvants for clinical immunotherapy because they exhibit multifaceted immunomodulatory capabilities. However, the rational design and discovery of IDRs that have improved immunomodulatory activities have been hampered by the lack of screening techniques and the great challenges in the identification of their interaction partners. Here, we describe a screening and evaluation system for IDR design. On the basis of in vitro screening, the optimized IDR DP7 recruited neutrophils, monocytes and macrophages to the site of infection. The adjuvant, comprising the DP7 and CpG oligonucleotide (CpG), induced chemokine/cytokine expression, enhanced the antigen uptake by dendritic cells and upregulated surface marker expression in dendritic cells. Vaccination with the NY-ESO-1 or OVA antigens combined with the adjuvant alum/CpG/DP7 strongly suppressed tumor growth in mice which was due to the improvement of antigen-specific humoral and cellular immunity. Regarding the mechanism of action, GPR35 may be the potential interaction partner of DP7. Our study revealed the potential application of the screening and evaluation system as a strategy for rationally designing effective IDRs or composite adjuvants and identifying their mechanism of action.

Sustained and targeted delivery of siRNA/DP7-C nanoparticles from injectable thermosensitive hydrogel for hepatocellular carcinoma therapy.

Hepatocellular carcinoma (HCC) is one of the most lethal cancers in humans. The inhibition of peptidyl-prolyl cis/trans isomerase (Pin1) gene expression may have great potential in the treatment of HCC. N-Acetylgalactosamine (GalNAc) was used to target the liver. Cholesterol-modified antimicrobial peptide DP7 (DP7-C) acts as a carrier, the GalNAc-siRNA/DP7-C complex increases the uptake of GalNAc-siRNA and the escape of endosomes in hepatocytes. In addition, DP7-C nanoparticles and hydrogel-assisted GalNAc-Pin1 siRNA delivery can effectively enhance the stability and prolong the silencing effects of Pin1 siRNA. In an orthotopic liver cancer model, the GalNAc-Pin1 siRNA/DP7-C/hydrogel complex can potentially regulate Pin1 expression in hepatocellular carcinoma cells and effectively inhibit tumor progression. Our study proves that Pin1 siRNA is an efficient method for the treatment of HCC and provides a sustainable and effective drug delivery system for the suppression of liver cancer.

TLR2 mediates autophagy through ERK signaling pathway in Mycoplasma gallisepticum-infected RAW264.7 cells.

Toll-like receptor 2 (TLR2) plays a crucial role in early innate immune response of host to various microorganisms. Mycoplasma gallisepticum (MG) is one of the major pathogen that can cause chronic respiratory diseases in chickens, but the molecular mechanism of MG infection still remained unclear. In this study, we examined the typical hallmarks of autophagy and multiple signaling pathways by western blot, immunofluorescence microscopy and electron microscopy. The results indicated that infection of mouse macrophage cell line RAW264.7 with MG activated autophagy and mitogen-activated protein kinases (MAPKs). Silencing of TLR2 by siRNA substantially down-regulated MG-triggered autophagy in macrophages, and markedly reduced MG-induced extracellular regulated protein kinase (ERK) in macrophages but did not down-regulate c-Jun N-terminal kinase (JNK) and p38. Importantly, in macrophages, inhibition of ERK by PD98059 (ERK inhibitor) also significantly attenuated the level of autophagy upon MG infection, and the simultaneous treatment of TLR2 siRNA and PD98059 showed a similar effect on MG-induced autophagy as compared with TLR2 siRNA treatment alone. These findings thus demonstrate that TLR2 may mediate MG-induced autophagy through ERK signaling pathway in macrophage.

p53 Mediates Colistin-Induced Autophagy and Apoptosis in PC-12 Cells.

The mechanism of colistin-induced neurotoxicity is still unknown. Our recent study (L. Zhang, Y. H. Zhao, W. J. Ding, G. Z. Jiang, Z. Y. Lu, L. Li, J. L. Wang, J. Li, and J. C. Li, Antimicrob Agents Chemother 59:2189-2197, 2015, http://dx.doi.org/10.1128/AAC.04092-14; H. Jiang, J. C. Li, T. Zhou, C. H. Wang, H. Zhang, and H. Wang, Int J Mol Med 33:1298-1304, 2014, http://dx.doi.org/10.3892/ijmm.2014.1684) indicates that colistin induces autophagy and apoptosis in rat adrenal medulla PC-12 cells, and there is interplay between both cellular events. As an important cellular stress sensor, phosphoprotein p53 can trigger cell cycle arrest and apoptosis and regulate autophagy. The aim of the present study was to investigate the involvement of the p53 pathway in colistin-induced neurotoxicity in PC-12 cells. Specifically, cells were treated with colistin (125 µg/ml) in the absence and presence of a p53 inhibitor, pifithrin-α (PFT-α; 20 nM), for 12 h and 24 h, and the typical hallmarks of autophagy and apoptosis were examined by fluorescence/immunofluorescence microscopy and electron microscopy, real-time PCR, and Western blotting. The results indicate that colistin had a stimulatory effect on the expression levels of the target genes and proteins involved in autophagy and apoptosis, including LC3-II/I, p53, DRAM (damage-regulated autophagy modulator), PUMA (p53 upregulated modulator of apoptosis), Bax, p-AMPK (activated form of AMP-activated protein kinase), and caspase-3. In contrast, colistin appeared to have an inhibitory effect on the expression of p-mTOR (activated form of mammalian target of rapamycin), which is another target protein in autophagy. Importantly, analysis of the levels of p53 in the cells treated with colistin revealed an increase in nuclear p53 at 12 h and cytoplasmic p53 at 24 h. Pretreatment of colistin-treated cells with PFT-α inhibited autophagy and promoted colistin-induced apoptosis. This is the first study to demonstrate that colistin-induced autophagy and apoptosis are associated with the p53-mediated pathway.