Baofa Yu

Objectives: Studying the sentinel lymph nodes (SLNs) to define the metastasis and the mechanisms of abscopal effect. Method: Using scRNA-seq to investigate the cellular and genomic changes of SLNs before or after treatment of the primar pancreatic carcinoma site by intratumoral injection of cytotoxic chemotherapy drugs plus hapten. Result: A very significant decrease of SLN carcinoma cells, but increased macrophages, monocytes, CD8 T-effector cells, and fibroblasts. At the genomic and functional levels, the above treatment increased the ability of angiogenesis, epithelial mesenchymal transition (EMT), and cancer-associated fibroblast (CAF) signaling. In addition, matricellular proteins were upregulated. Alternative conclusion: Our study demonstrated a marked reduction in carcinoma cells within SLNs following treatment, alongside a notable increase in inflammatory and stromal cells, which suggested a dynamic reorganization of the lymph node microenvironment rather than straightforward tumor growth. The absence of significant enlargement of SLNs, despite the increase in cellular density, may be attributed to the remodeling effects induced by the treatment. The observed upregulation of angiogenic factors, EMT pathways, and CAF signaling highlighted a complex interaction between tumor and immune responses. These findings provided new insights into the abscopal effect, revealing how targeted therapies can modulate lymph node microenvironments to enhance local immune responses and potentially improve systemic antitumor efficacy.

Background: Pancreatic cancer is a highly lethal cancer with no treatment options when diagnosed at an advanced stage. Hapten combined with chemotherapy drugs have been successfully injected into the tumor for local treatment of pancreatic cancer, but due to the heterogeneity of the tumor, the survival time of patients with similar stages is different. Methods: SCNA-SEQ and Olink proteomics methods were used to observe the cell and genome changes in the survival group treated with hyrazine combined with HEIC for more than 10 months and less than 6 months. scRNA-Seq shows T cell activity of cDCs and polygene-expressed pDCs. Results: scRNA-Seq confirmed the initiation of immune response: NaiveT and CD8Teff increased, proliferation increased, NK decreased, and ribosomal protein gene upregulated after treatment. The Neutrophils_4 gene set score of precursor cell neutrophilS_4 was higher. There was high expression of interferon-stimulating gene in neutrophils s_3, and high expression of s_3 characteristic gene set in interferon-associated neutrophils. Due to immune action, proteomic results showed that GZMB, GZMZ, IL18, CASP-8, CD8A, HO-1 and AOA genes were up-regulated by 8%. DCN, MCP-1, CX3cl1, CD40, CD27, IL33, TIE2, Gal-9, PGE, MCP-3, CD28, PD-L1, CD38, CCL3, MCP-2, MMP7, laptgf-β-1 genes were down-regulated by 18%. Conclusions: Overall, the study showed that patients with similar stages of pancreatic cancer had an enhanced immune response after the same regimen of HEIC treatment, and found that gene expression increased by 8% and decreased by 18% in the longer survival group compared with the shorter survival group due to tumor heterogeneity.

Immunotherapy has emerged as a novel treatment strategy for many types of cancers, among them, liver cancer. The major advances and achievements in recent years is the use of programmed death-1 (PD-1) blockers for cancer treatment. Since patients’ immune systems are already weak following concurrent surgery plus chemo or radiotherapy, it is necessary to restore and awake their immune cells to maximize the effect for immune therapy. Methods: Liver cancers were treated twice with hapten enhanced local chemotherapy (HELC) like tumor lysates vaccine and following PD-1. Single-cell RNA sequencing (scRNA-seq) was used to analyze the changes of immune responses prior and after treatments. Results: We observed upregulation of cytotoxicity-related genes of CD8 effector T cells and NK cells in untreated tumor; Both Bmem and Naive B cells in untreated tumor showed a significant increase in MHC II signaling pathway-related genes, while MHC I-related genes was upregulated in plasma cells. Significant tumor size shrinkage was observed in both treated and untreated tumors following the HELC+PD-1 therapy. Conclusions: This study provides new biological insights into the abscopal effect at the single-cell level related to the composition of T and B cells in untreated liver cancers before and after major primary tumors treated by HELC. Our data showed that intra-tumor HELC can kill tumor cells and induced immune activity, which is likely vital in modifying tumor associate antigens (TAAs) into neo TAAs. It induces immune response just like vaccine can wake up immune cells and therefore increasing the efficacy of PD1 therapy.

Background: the abscopal is a hypothesis of effect on non-irradiated tumors after localized radiation therapy which associated with the products of tumor-associated gene as autoantibodies (aTAAs) in reaction to the tumor-associated antigens (TAAs). When TAAs interact with hapten within tumor after hapten plus chemotherapy drugs intratumoral injection to pancreatic cancer like tumor lysates vaccine, TAAs-hapten stimulates the immune system as a neu antigen and produces autologous tumor antibodies, it is called induced tumor-associated autoantibodies (iTAAs) which is lightly difference to aTAAs since epitope of antigen linked with hapten. Method: immunofluorescence (IF) was applied for detect the binding of the iTAAs in tumor cells at sentinel lymph node. Results: aTAAS naturally could not target and bind the TAAs in tumor since due to immune tolerance. The iTAAs can target and bind the TAAS, also the iTAAs targeted in the tumors of sentinel lymph node with complements C help, it was found Cmyc (P = 0.0017 at one week; P = 0.0001 at two weeks), p53 (P = 0.0373; 29.13 ± 6.91, and P = 0.0254), and Zeta (P = 0.1513, P = 0.0044) increased significantly in the tumor cells or perinuclear tumor cells in lymph node at one and two weeks after treatment. Conversely, IMP1 (P = 0.6154; P = 0.0138), Koc (P = 0.5684, P = 0.0103), Survivn (P = 0.1020; P = 0.0147) and Rala (P = 0.3226; P = 0.0249) increased significantly in the cell or perinuclear of tumor in lymph node at two weeks later following. Conclusions: it indicated all of iTAAs plays a function in the binding of original cellular tumor genes at sentinel lymph node while the aTAAs could not bind the cellular tumor genes at lymph node due to immune tolerance. We could make a hypothesis: TAAs inside the cancer cells’ nuclei can be targeted by the iTAAs which produced by hapten enhanced intratumoral chemotherapy. The iTAA may play a distinctive role in controlling tumor cell growth and resulting in an abscopal effect, which is one of hapten associated with TAA induced immune response.

Objective: Nanotechnology offers many advantages in various fields of cancer therapy. This study describes a new method of self-forming nanoparticles (self-nano) using ferric chloride (FeCl₃) and hydrogen peroxide (H₂O₂) to form the self-nanoparticles in an in vivo tumor. The treatment effect is evaluated.Method: A solution of 3% FeCl₃ (0.5 mL) and 1.8% H₂O₂ (1.0 mL) was injected into the tumor. At various time points post-injection, tumors were collected, and sections were prepared for electron microscopy to evaluate the size of the self-nano particles. Single-cell RNA sequencing (scRNA-seq) was used to analyze the immune changes and their effect on tumor growth.Result: The formation of self-nano in vitro was observed and confirmed, with particles averaging 421 nm in size for the FeCl₃ + H₂O₂ solution. Over time points ranging from 1 to 14 days, the formed self-nano remained stable at a regular size of 421 ± 8 nm. The self-nano, primarily consisting of iron, induced ferroptosis under the influence of an external magnetic field, leading to tumor growth control through iron-induced cell death and immune reactions. These self-nanoparticles also showed stronger enrichment of pathways related to CD8⁺ T effect cells (Teff), T cell activation, and regulation of T cell proliferation.Conclusion: The FeCl₃ + H₂O₂ solution can form Fe₂O₃-based self-nanoparticles within tumors through H₂O₂-incubated oxidation of FeCl₃. The self-nano remains effective for over 14 days, inducing ferroptosis and upregulating immune cells under magnetic field treatment. This method offers a novel approach for cancer treatment that can be combined with other modalities.

Ascites and pleural effusion are recognized complications of pancreatic cancer. These diseases are accompanied by ascites and pleural effusion, and drug treatment is limited by high costs, long hospital stays, and failure rates. Immunotherapy may offer new option, but in most patients with late stages of cancer, immune cells may lose the ability to recognize tumor cells, how to activate their immune cells is a major problem, sodium glucosidate (SSG) is injected into ascites as a protein tyrosine phosphatase inhibitor to wake up immune cells and prepare for immunotherapy. We used single-cell RNA sequencing (scRNA-seq) to investigate whether and how SSG injected into ascites of pancreatic cancer elicits an immune response. Our study showed that the process of SSG fusion treatment of ascites and pleural effusion, the interaction between TandNK cells, MPs cells, monocytes and neutrophils was induced, and large numbers of genes were expressed, resulting in upregulation of immune response, which also approved that SSG is not only used as a protein tyrosine phosphatase inhibitor, but also it works as a protein tyrosine phosphatase inhibitor. It can also be used to regulate immune cell function, recruiting immune cells to the right place with the help of PD-1 or PD-L1 to fight cancer cells in ascites and pleural effusions in cancer patients.

Background: Cervical cancer is one of the most common and deadly cancers in women, for who with persistent infection of high-risk human papillomavirus (HPV). Current treatment of cervical cancer involves radical hysterectomy, radiotherapy and chemotherapy or a combination. Objective: We investigated if hapten enhanced intratumoral chemotherapy (HEIC) was effective in boosting immunity for effective treatment of precancerous cervical lesions and HPV infection Study Design: We used single-cell RNA sequencing (scRNA-Seq) to obtain transcriptome profiles of 40239 cells from biopsy of precancerous cervical lesions at the cervix directly from one patient before the start of HEIC and about 1 week after HEIC. The blood samples were taken at same time as biopsies. We compared the expression characteristics of malignant epithelial cells and immune cells, including Epithelial Cells, Ecs, Fibroblasts, Mural Cells, Tcells, Bcells, TandNK Neutrophils, Mast Cells, MPs, and Platelets, as well as the dynamic changes in cell percentage and cell subtype heterogeneity. Results: Intratumoral injection of chemotherapy drug plus hapten induces acute immune response in precancerous cervical lesions with HPV, and further awakens immune cells to prevent abnormal proliferation of the precancerous cells. Conclusion: HEIC provides a potential treatment methodfor cervical cancer and HPV infection tailored to each patient's condition.