Based on the growth characteristics, tumors can be divided into "malignant" and "benign". The difference between "hot" and "cold" tumors will be explained in the following paragraphs.

According to the spatial distribution of immune cells in TME, tumors can be classified into three basic immunophenotypes: immune-inflamed, immune-excluded, and immune-desert phenotypes.

"Hot tumors", also known as Immune-inflamed tumors, are characterized by high T-cell infiltration, increased interferon-γ (IFN-γ) signaling, expression of PD-L1, and high tumor mutational burden (TMB) . (In general, tumors with a higher TMB carry a higher neoantigen load that can be recognized by T cells, making them more likely to prime the immune system). CD8⁺ T lymphocytes play roles in the prolonged survival of cancer patients and increased efficacy of immunotherapy. Infiltration of CD8⁺ T lymphocytes is critical for cancer immunotherapies. Immune-excluded and immune-desert tumors can be described as "cold tumors". In immune-excluded tumors, CD8⁺ T lymphocytes localize only at invasion margins and do not efficiently infiltrate the tumor. In immune-desert tumors, CD8⁺ T lymphocytes are absent from the tumor and its periphery.

Immunosuppressive mechanisms of the TME in cold tumors: Besides poor T cell infiltration, the T cell priming may be inhibited in cold tumor TME. Furthermore, the deposition of extracellular matrix (ECM) and stiff stroma-induced hypoxia in cold tumor lesions build a physical and chemical barrier to obstruct the T cell infiltration. More notably, numerous immunosuppressive cells are widely settled in the TME of cold tumors and suppress the function of CD8⁺ T cells via T cell exhaustion^[2](Figure 2). In summary, TME in cold tumors lacks innate immunity, or "cold tumors" possess innate anti-tumor immunity characteristics.

Immune checkpoints are the inhibitory pathways on the surface of T cells that inhibit immune activation. Immune checkpoint Inhibitors (ICIs) have revolutionized cancer treatment by activating T-cell-based antitumor immunity. The TME in hot tumors is infiltrated by numerous T cells, and the immune cells in "hot" TME tumors are more active than in cold tumors. When ICIs remove the inhibitory effect of the immune checkpoint, immune response of T cells can be activated again, then the effector T cells kill the cancer cell. "Hot tumors" with inflammatory phenotypes tend to be more sensitive to ICIs therapy; however, ICIs are not effective against cold tumors due to their "stiff" and immunosuppressive microenvironment.

Therefore, conversion of "cold tumors" to "hot tumors" has become an attractive strategy for cancer treatment, which could increase the sensitivity ofthese tumors to ICIs and improveclinical use ofimmunotherapy

Driven T cells into the cold tumor TME improve the efficacy of ICI

STING agonists, oncolytic viruses, chemotherapy, radiotherapy, etc.can induce immunogenic cell death (ICD) to promote T-cell priming and activation^[1][3].

For example, activation of the STING signaling pathway mediates the expression of proinflammatory cytokines (e.g., type I IFNs) and activates the innate immune responses mediated by T cells.The activated immune system changes the immunophenotype of the tumor.It switches the "cold tumor" with low T cell reactivity to the "hot tumor" with high T cell reactivity.

As illustrated in figure 3, In the article entitled Development of Potent Immune Modulators Targeting Stimulator of Interferon Genes Receptor, STING agonist 4c significantly decreased tumor volume in a CT26 murine colorectal carcinoma model. Additionally, In a 4C-treated mouse model, immunological memory-derived cancer inhibition was observed. The results confirm the therapeutic potential of compound 4c for cancer immunotherapy via STING-mediated immune activation^[3]. In line with the above observatione, an article published in Science also reported that MSA-2 (a STING agonist) stimulates the secretion of IFN-β and induces tumor regression. In addition, MSA-2 increases CD8⁺ T lymphocyte infiltration in low PD-1 blocking response mouse tumor models.

Increase the abundance of immune checkpoints and promote effective T cells infiltration,"assisting" cold tumors turn to hot

In March 2022, Dr. Jinfang Zhang's team at Wuhan University published their study entitled "USP8 inhibition reshapes an inflamed tumor microenvironment that potentiates the latest achievement in the field of tumor immunotherapy". The study reveals that inhibition of the deubiquitination enzyme USP8 can reshape the tumor immune microenvironment (TME), and turn "cold" tumors to "hot", thereby improving an inflamed TME where immunotherapy can be more effective.

The article shows that in vitro inhibition of USP8 can enhance the ubiquitination modification of PD-L1 K63 junction and up-regulate PD-L1 protein level. Additionally, a combination of USP8 inhibitor with anti-PD-L1 antibody significantly suppressed tumor growth and improved the overall survival rates of MC38 tumor-bearing C57BL/6 mice(Fig 4a). A combination of DUB-IN-2 and anti-PD-L1 antibodies could significantly increase the percentage of CD8⁺ T cells (Fig. 4b). Meanwhile, USP8 inhibitor combined with anti-PD-L1 treatment also significantly elevated the expression of the T cell activation marker GzmB and reduced the expression of the exhausted T cell marker TIM3 on infiltrated CD8⁺ T cells (Fig.4c-d). This study provides evidence to support the enhanced anti-tumor efficacy of the combined therapies of a USP8 inhibitor and PD-1/PDL-1 blockade.

The nanoparticle-based treatment for turning "cold" tumors into "hot"

Nanotechnologies are rapidly increasing their role in immunotherapy in line with the need for novel therapeutic strategies. Nanoparticle (NP)-based drugs for turning “cold tumors” into “hot tumors”.Four potential pathways in Fig 5 show the mechanisms of NP-based drugs turning cold tumors hot^[5]:

1.The difference between "hot" and "cold" tumors is mainly due to the spatial distribution of immune cells, and "cold" tumors are often "insensitive" to immunotherapy.

2.Promoting T cell initiation could produce by increasing tumor antigen expression and restoring antigen processing and presentation mechanisms (e.g., STING agonist^[3]). Furthermore, reprogramming the tumor microenvironment, promoting T-cell transportation and allowing T-cells to infiltrate effectively^[4]. These methods potentially could turn "cold" tumors to "hot" and therefore improve immunotherapy efficiency.