Autophagy in cancer therapy

Foreword 

Autophagy is a regulatory mechanism that removes unnecessary or dysfunctional cellular components and recycles metabolic substrates. In response to stress signals in the tumor microenvironment, the autophagy pathway is altered in tumor cells and immune cells, resulting in different effects on tumor progression, immunity, and treatment.

Recent studies have shown that the autophagy pathway is involved in the survival and apoptosis, differentiation, activation, effector function, and trafficking of immune cell subsets to tumors. At the same time, tumor autonomous autophagy can alter tumor growth by regulating immune responses. Furthermore, by combining immune checkpoint therapy with autophagy inhibitors, the tumor-promoting effects of autophagy can be eliminated. Therefore, autophagy is a complex but promising target in cancer therapy.

Autophagy of tumor cells 

The tumor microenvironment (TME) plays a key role in cancer progression, metastasis and treatment resistance. In the TME, autophagy in tumor cells can be induced by intracellular and extracellular stress signals, including metabolic stress, hypoxia, redox stress, and immune signals.

Metabolic stress

Insufficient nutrients absorbed by the TME will affect the metabolic mechanism and lead to intracellular metabolic stress. In response to metabolic stress, tumor cells restructure their metabolic pathways by upregulating nutrient transporters and activating autophagy.

Mechanistically, 5′-AMP-activated protein kinase (AMPK) and mTOR complex 1 (mTORC1) are two opposite regulatory kinases that alter autophagy induction under conditions of nutrient deprivation. As an AMP sensor, AMPK is activated by an increase in the AMP to ATP ratio, whereas mTORC1 activity is reduced by amino acid deficiency. This results in phosphorylation of targets in the pre-autophagy initiation complex, thereby initiating autophagy.

Hypoxic stress

Hypoxia is a characteristic of solid tumors. Hypoxia causes the inhibition of mitochondrial oxidative phosphorylation, increases the ratio of AMP to ATP, activates AMPK, and hypoxia also Inhibits mTOR signaling. In addition, hypoxia leads to the activation of activating transcription factor 4 (ATF4), which upregulates the expression of LC3B and autophagy-related protein 5 (ATG5) and maintains high levels of autophagy.

Oxidative stress

Oxidative stress reflects an imbalance between free radicals and antioxidants. Reactive oxygen species (ROS) are generated within cells through oxygen metabolism and other processes, and excess ROS may increase the risk of DNA damage and promote tumorigenesis. In response to elevated ROS, ataxia telangiectasia mutated (ATM) activates the TSC2 tumor suppressor through the liver kinase B1 (LKB1) and AMPK metabolic pathways in the cytoplasm to inhibit mTORC1 and induce autophagy. Oxidative stress can also promote autophagy through NF-κB-mediated upregulation of p62/SQSTM1.

Immune signaling

Immune signaling can regulate the autophagy pathway in the TME. Damage-associated molecular patterns (DAMPs) and cytokines are the main mediators that regulate autophagic responses. Extracellular DAMP signals are sensed by extracellular or intracellular pattern recognition receptors, such as Toll-like receptors (TLRs), and autophagy induction occurs when various TLRs recognize DAMPs and activate downstream signals.

In addition, in Drosophila, cytokines such as TNF and IL-6-like signaling can activate autophagy, thereby promoting early tumor growth and invasion. TGF-β increases the transcription levels of BECN1, ATG5 and ATG7 through SMAD-dependent and SMAD-independent pathways and activates autophagy, which can delay the apoptosis of human hepatocellular carcinoma and breast cancer cells in vitro.

 Autophagy-mediated immune evasion 

Evasion of anti-tumor immune responses is an important survival strategy for various tumors. Recent evidence suggests that autophagy plays an important role in tumor immune evasion. Studies have found that the downregulation of MHC class I molecules in pancreatic ductal adenocarcinoma (PDAC) is mediated through selective autophagic degradation, and inhibition of autophagy releases a strong anti-tumor immune response.

On the other hand, MDSCs play an immunosuppressive role in the TME, and studies have proven that autophagy in MDSC is a key mechanism for inhibiting the anti-tumor immune activity of melanoma. Autophagy in MDSC immune cells is central to the degradation of MHC class II molecules, preventing the initiation and activation of anti-tumor T cells.

 Autophagy and drug resistance

As a mechanism for (cancer) cells to respond to threatening stressors, autophagy is considered an important mechanism of treatment resistance in cancer treatment. There is evidence that tumor cell resistance to cisplatin is mediated, at least in part, by increased autophagy in ovarian cancer cell lines. Similar evidence suggests that cisplatin, doxorubicin, and methotrexate overcome chemotherapy resistance by inhibiting autophagy in osteosarcoma.

Interestingly, for example, the interaction between cisplatin and autophagy is a continuum, and even in cells that are not resistant to certain chemotherapeutic drugs themselves, both in vitro and in vivo, the addition of autophagy Phagocytosis inhibitors such as chloroquine (CQ) can also improve treatment effectiveness. This has been demonstrated in mouse models of adrenocortical carcinoma, colon cancer cell lines, and 5-fluorouracil- and temozolomide-induced cytotoxicity in glioma cells.

Additionally, similar results have been reported with antibody-based treatments, such that in trastuzumab-resistant breast cancer cells, autophagy inhibition using CQ resulted in almost complete tumor progression. eliminate. Similarly, inhibition of autophagy using CQ can also effectively antagonize bevacizumab-induced autophagy in colorectal cancer cells and reduce tumor growth in in vivo mouse tumor models.

 Research status of autophagy in tumor treatment

Autophagy inhibitors are divided into early inhibitors targeting ULK1/ULK2 or VPS34, such as SBI-0206965, 3MA and wortmannin , as well as late inhibitors targeting lysosomes, such as CQ, hydroxychloroquine (HCQ), bafilomycin A1 and monensin. CQ and HCQ inhibit autophagosome degradation by interfering with lysosomal acidification. However, in clinical trials, HCQ monotherapy failed to control tumor growth in patients with advanced pancreatic cancer. Currently, autophagy inhibition is combined with other cancer treatments to improve the therapeutic effect.

Chemotherapy

High autophagy flux in cancer is associated with reduced response to chemotherapy and associated with poor survival in cancer patients. Preclinical studies have shown that inhibiting autophagy can overcome chemotherapy resistance in NSCLC, bladder cancer, thyroid cancer, and pancreatic cancer. In addition, results from some studies suggest that autophagy inhibition may synergize with inhibition of MEK-ERK signaling.

An early phase II study in 2014 used HCQ monotherapy to treat patients with metastatic pancreatic cancer who had been previously treated with other methods. The primary endpoint was two-month progression-free survival. As a result, autophagy levels were reduced to varying degrees in different patients, but the primary endpoint was not significantly improved. Another study combining HCQ, gemcitabine, and nab-paclitaxel in patients with advanced or metastatic pancreatic cancer also failed to demonstrate a 12-month extension in overall survival. Importantly, however, patients taking HCQ showed a significantly better response rate (38.2% vs. 21.1%).

Radiotherapy

Autophagy plays a key role in protecting tumor cells from cell death caused by radiotherapy. In breast cancer cells, radiation induces the expression of autophagy-related genes, accompanied by the accumulation of autophagosomes. Short-term inhibition of autophagy during radiotherapy can enhance the cytotoxicity of radiotherapy against drug-resistant cancer cells. Similarly, hypoxia enhances the radioresistance of A549 lung cancer cells by inducing autophagy.

In glioblastoma, radiation therapy induces autophagy by increasing the expression of mammalian STE20-like protein kinase 4 (MST4), which stimulates autophagy through ATG4B phosphorylation. The small molecule inhibitor NSC185058, which targets ATG4B, impairs intracranial xenograft growth of glioblastoma and prolongs survival in treated mice when used in combination with radiation therapy. Therefore, targeting tumor autophagy may enhance the efficacy of radiotherapy. In fact, autophagy inhibitors have been used in combination with radiation therapy in clinical trials in cancer patients.

Immunotherapy

Taking advantage of the immune system is an important way to fight cancer. Inhibiting autophagy may impair systemic immunity because autophagy is involved in immune system development and the survival and function of effector T cells. However, short-term systemic inhibition of autophagy by CQ did not impair T cell function in preclinical models of melanoma and breast cancer. Data suggest that the immune system may be resistant to some degree of autophagy inhibition. However, given that autophagy can regulate tumor immune responses, targeting autophagy can improve the efficacy of immunotherapy and overcome immunotherapy resistance.

For example, inhibition of VPS34 kinase activity using inhibitors SB02024 or SAR405 leads to increased levels of CCL5, CXCL10, and IFN-γ in the TME, thereby inhibiting NK and T cell tumors in melanoma and colorectal cancer models. Increased levels of infiltration. In these models, VPS34 inhibition also reversed resistance to anti-PD1 or anti-PD-L1 therapy. Furthermore, CQ treatment blocks autophagy-mediated MHC class I degradation, synergizing with dual ICB treatment (anti-PD1 and anti-CTLA4 antibodies) to produce enhanced anti-tumor immune responses in pancreatic cancer mouse models.

Thus, targeting autophagy may enhance immunotherapy. Clinical trials of HCQ combined with immunotherapy to treat patients with different types of cancer are currently underway.

In addition, CAR-T cell therapy has achieved clinical success in the treatment of hematological tumors, but its efficacy in the treatment of solid cancers is still limited. Modulation of autophagy may provide some benefits to cancer patients treated with CAR-T cells. It is well known that the TME is a barrier to CAR-T cell infiltration and function in solid tumors. Given that autophagy inhibition can reshape the TME and promote the production of TH1-type chemokines, autophagy inhibition can promote the transport of CAR-T cells to the tumor. CAR -Enhanced autophagy of T cells may support T cell adaptation and survival in the TME. In addition, inhibition of tumor autophagy may lead to increased antigen expression, thereby enhancing CAR-T cell-mediated tumor killing. Finally, autophagy inhibition may improve cytokine release syndrome and provide clinical benefit to patients. Overall, the potential of inhibiting autophagy to enhance the efficacy of immunotherapy is a promising area of ongoing exploration.

Summary

Autophagy is an important mechanism for research in many fields such as tumor biology and immunology. Given that autophagy can be induced by different factors in different cells in the TME, its induction and activation can promote or inhibit tumor progression. Autophagy in T cell subsets may play an active role in anti-tumor immune responses, while functional autophagy in tumor cells may support tumor antigen presentation and recognition, in which case inhibition of autophagy may be detrimental to anti-tumor immunity. On the other hand, the autophagy pathway may be related to tumor cell survival, tumor antigen degradation, reduction of TH1-type chemokines, and enhancement of Treg cells and MDSC. Therefore, systemic targeting of autophagy for cancer treatment is challenging.

Therefore, in order to target the autophagy pathway in tumor immunotherapy, it is crucial to explore the biological activity of autophagy in major immune cell subsets. Recent studies have begun to evaluate the autophagy pathway in T cells, macrophages, and dendritic cells. Future work may expand to B cells, NK cells, and NKT cells. In addition, as cancer progresses, the autophagy pathway will dynamically change in response to different stimuli in the TME. It is necessary to understand how autophagy simultaneously participates in the function and survival of immune cells, tumor cells, and stromal cells in the TME, so as to can ultimately provide clinical benefit to cancer patients.