Anti-tumor Immune Function Research Articles

Valproic Acid-Like Compounds Enhance and Prolong the Radiotherapy Effect on Breast Cancer by Activating and Maintaining Anti-Tumor Immune Function.

Inadequate sustained immune activation and tumor recurrence are major limitations of radiotherapy (RT), sustained and targeted activation of the tumor microenvironment can overcome this obstacle. Here, by two models of a primary rat breast cancer and cell co-culture, we demonstrated that valproic acid (VPA) and its derivative (HPTA) are effective immune activators for RT to inhibit tumor growth by inducing myeloid-derived macrophages and polarizing them toward the M1 phenotype, thus elevate the expression of cytokines such as IL-12, IL-6, IFN-γ and TNF-α during the early stage of the combination treatment. Meanwhile, activated CD8+ T cells increased, angiogenesis of tumors is inhibited, and the vasculature becomes sparse. Furthermore, it was suggested that VPA/HPTA can enhance the effects of RT via macrophage-mediated and macrophage-CD8+ T cell-mediated anti-tumor immunity. The combination of VPA/HPTA and RT treatment slowed the growth of tumors and prolong the anti-tumor effect by continuously maintaining the activated immune response. These are promising findings for the development of new effective, low-cost concurrent cancer therapy.

A genome-wide CRISPR screen reveals a role for the BRD9-containing non-canonical BAF complex in Foxp3 expression and regulatory T cell function

Abstract Regulatory T cells (Tregs) play a pivotal role in suppressing auto-reactive T cells and maintaining immune homeostasis. Treg development and function are dependent on the transcription factor Foxp3. Although a large body of work has been devoted to dissecting the molecular mechanisms regulating Foxp3 expression, a systematic, genome-wide approach has not been reported. Here we performed a genome-wide CRISPR/Cas9 knockout screen to identify the regulators of Foxp3 in mouse primary natural Tregs. The screen results not only confirmed a number of known Foxp3 regulators, but also revealed many novel factors that control Foxp3 expression. Gene ontology analysis showed that Foxp3 regulators are highly enriched in genes encoding SWI/SNF and SAGA complex subunits. Among the three SWI/SNF-related complexes, the non-canonical or ncBAF (also called GBAF or BRD9-containing BAF) complex promoted the expression of Foxp3, whereas the PBAF complex repressed its expression. Gene ablation of BRD9 led to reduced Foxp3 expression and compromised Treg function in inflammatory disease and anti-tumor immunity. BRD9 co-localized with Foxp3 in genome-wide binding studies, including at the CNS0 and CNS2 enhancers at the Foxp3 locus. Functional genomics revealed that BRD9 is required for Foxp3 binding and expression of a subset of Foxp3 target genes. Thus, we provide an unbiased analysis of genes and networks regulating Foxp3, and reveal the ncBAF complex as a novel target that could be exploited to manipulate Treg function.

025 ALA-PDT inhibits skin squamous cell carcinoma (cSCC) via regulating formation of tertiary lymphoid structures

Topical ALA-mediated PDT, ALA-PDT, is a novel therapeutic modality widely used to treat actinic keratosis, Bowen’s Disease, superficial skin SCC, and other cancerous and precancerous skin diseases. Several studies have proved that ALA-PDT can inhibit SCC growth. Subsequent research suggested that ALA-PDT not only directly induced tumor cells apoptosis, but also improve tumor microenvironment through regulation of immune cells. However, the anti-tumor immune function of ALA-PDT is still need to be elucidated. Here, we found that tertiary lymphoid structures (TLSs), which are ectopic lymphoid organs that develop in non-lymphoid tissues at sites of chronic inflammation including tumors, play a pivotal role in anti-tumor immune function of ALA-PDT. We analyzed 77 samples of cSCC patient in our hospital. TLS was observed in 79% of the patient samples, and the density of TLS is negatively correlation with the Brodes classification of cSCC. Intriguingly, immunohistochemistry showed that ALA-PDT could promote the formation of TLS in vivo. We also found that promotion effect of ALA-PDT on TLS is mediated by M1 macrophage, which function as lymphoid tissue inducer cells in TLS formation and recruited by ALA-PDT. Further investigation substantiated that PDT-secreted exosomes were involved in M1 macrophage polarization. Our study elucidated a mechanism that ALA-PDT influence M1 macrophage polarization via PDT-secreted exosomes, which could facilitate the formation of TLS. Thus, our research may contribute to in-depth molecular understanding of the ALA-PDT on anti-tumor immune function.

CARM1 Inhibition Enables Immunotherapy of Resistant Tumors by Dual Action on Tumor Cells and T Cells.

A number of cancer drugs activate innate immune pathways in tumor cells but unfortunately also compromise antitumor immune function. We discovered that inhibition of CARM1, an epigenetic enzyme and cotranscriptional activator, elicited beneficial antitumor activity in both cytotoxic T cells and tumor cells. In T cells, Carm1 inactivation substantially enhanced their antitumor function and preserved memory-like populations required for sustained antitumor immunity. In tumor cells, Carm1 inactivation induced a potent type 1 interferon response that sensitized resistant tumors to cytotoxic T cells. Substantially increased numbers of dendritic cells, CD8 T cells, and natural killer cells were present in Carm1-deficient tumors, and infiltrating CD8 T cells expressed low levels of exhaustion markers. Targeting of CARM1 with a small molecule elicited potent antitumor immunity and sensitized resistant tumors to checkpoint blockade. Targeting of this cotranscriptional regulator thus offers an opportunity to enhance immune function while simultaneously sensitizing resistant tumor cells to immune attack. SIGNIFICANCE: Resistance to cancer immunotherapy remains a major challenge. Targeting of CARM1 enables immunotherapy of resistant tumors by enhancing T-cell functionality and preserving memory-like T-cell populations within tumors. CARM1 inhibition also sensitizes resistant tumor cells to immune attack by inducing a tumor cell-intrinsic type 1 interferon response.This article is highlighted in the In This Issue feature, p. 1861.

P80.01 A Multicenter, Randomized, Phase 3 Trial of Penpulimab in Combination With Anlotinib or Chemotherapy as First-Line Treatment in Advanced NSCLC

Penpulimab, a human immunoglobulin G1 (IgG1) monoclonal antibody (mAb) directed against human programmed cell death-1 (PD-1), was engineered to optimize receptor occupancy by improving duration of drug binding, and to eliminate Fc-mediated effector function that compromises anti-tumor immune cell function. Anlotinib, an antiangiogenic multi-target tyrosine kinase inhibitor. Chemotherapy is the standard treatment for advanced non-small cell lung cancer (NSCLC). A combination of the PD-1 checkpoint inhibitor and the antiangiogenic drug may improve clinical response in NSCLC by blocking the accumulation of immunosuppressive cells and promoting influx of effector T cells into tumors.

Neem leaf glycoprotein salvages T cell functions from Myeloid-derived suppressor cells-suppression by altering IL-10/STAT3 axis in melanoma tumor microenvironment.

Myeloid-derived suppressor cells (MDSCs) suppress antitumor immune functions. We have observed that an immunomodulator, neem leaf glycoprotein (NLGP), inhibits tumor-resident MDSCs and enhances antitumor CD8+ T cell immunity. NLGP inhibits the number as well as functions of tumor-resident MDSCs (Gr1±CD11b±) and enhances antitumor CD8± T cell immunity by downregulating arginase 1 and inducible nitric oxide synthase production in MDSCs. Accordingly, decreased T cell anergy and helper to regulatory T cell conversion have been observed in the presence of NLGP, which ultimately augments T cell functions. Mechanistically, NLGP-mediated rectification of T cell suppressive functions of MDSCs was primarily associated with downregulation of the interleukin (IL)-10/signal transducer and activator of transcription 3 (STAT3) signaling axis within the tumor microenvironment, as confirmed by knockdown of STAT3 (by STAT3-siRNA) and using IL-10-/- mice. Thus, NLGP-mediated suppression of MDSC functions in tumor hosts is appeared to be another associated effective mechanism for the eradication of murine melanoma by NLGP.

Factors Associated with Lymph Node Count in Mucosal Squamous Cell Carcinoma Neck Dissection

Decreased lymph node count (LNC) from neck dissection (ND) for mucosal head and neck squamous cell carcinoma (HNSCC) patients is correlated with decreased survival. Advanced age and low BMI due to undernutrition from dysphagia from advanced T-stage tumors are common in patients with HNSCC. We studied the relationship between these two well-described causes for immune dysfunction and LNC in patients undergoing neck dissection. We conducted a retrospective review at a single tertiary care institution of patients with HNSCC that underwent neck dissection from 2006 to 2017. Stepwise linear and logistic regression analyses were performed on 247 subjects to identify independent significant factors associated with 1) the LNC per neck level dissected; 2) advanced T-stage. One-way ANOVA was utilized to demonstrate differences between the p16 positive and negative subgroups. Low BMI (<23 vs. ≥23) (P = .03), extra nodal extension (ENE) (P = .0178), and advanced age (P = .005) were associated with decreased LNC per neck level dissected on multivariable analysis. Higher T-stage (P = .0005) was correlated with low BMI (<23) after controlling for the effects of tobacco, smoking, sex, ECE, and p16 status. p16+ patients, on average had higher BMI, were younger and produced a higher nodal yield (P < .0001, .007, and .035). Patient intrinsic factors known to correlate with decreased immune function and worse outcomes, including p16 negative status, advanced age, and low BMI from undernutrition and ENE are associated with low nodal yield in neck dissections. LNC may be a metric for anti-tumor immune function that correlates with prognosis and T-stage. 3 Laryngoscope, 2021.

Stromal infiltrating mast cells identify immunoevasive subtype high-grade serous ovarian cancer with poor prognosis and inferior immunotherapeutic response

ABSTRACT Tumor infiltrating mast cells (TIMs), with pro- or anti-tumorigenic role in different types of malignancies, have been implicated in resistance to anti-PD1 therapy. Here, we aimed to identify the relevance of TIMs with the prognosis, immune contexture, and immunotherapy in high-grade serous ovarian cancer (HGSOC). Tissue microarrays containing 197 HGSOC patients were assessed by immunohistochemistry (IHC) for detecting the expression of mast cell tryptase and other immune markers. Kaplan-Meier curve, log-rank test, and Cox regression model were applied to perform survival analysis. Single-cell RNA-seq analysis and flow cytometric analysis were selected to characterize TIMs. Furthermore, short-term HGSOC organoids were employed to validate the effect of TIMs on anti-PD1 therapy. Abundance of stromal TIMs (sTIMs) predicted dismal prognosis and linked to immunoevasive subtype of HGSOC, characterized by increased infiltration of pro-tumor cells (Treg cells, M2-polarized macrophages, and neutrophils) and impaired anti-tumor immune functions. Intensive inter-cell interactions between TIMs and other immune cells were identified, suggesting potential cross-talks to foster an immunosuppressive microenvironment. Organoids derived from sTIMs-low patients were associated with increased response to anti-PD-1 treatment other than the presence of high sTIMs infiltration. A nomogram, constructed by combining FIGO stage, sTIMs, and PD-L1, with an area under the curve (AUC) for predicting 5-year overall survival of 0.771 was better than that of FIGO staging system of 0.619. sTIMs/PD-L1-based classifier has potential clinical application in predicting prognosis of patients with HGSOC. sTIMs-high tumors correlate with immunosuppressive tumor microenvironment (TME) and possess potential insensitivity to immunotherapy.

Regulatory B cells in cancer.

Tumorigenesis proceeds through discrete steps where acquisition of genetic lesions and changes in the surrounding microenvironment combine to drive unrestricted neoplastic proliferation and metastasis. The ability of tumor-infiltrating immune cells to promote tumor growth via the provision of signals that enable tumor cell survival and proliferation as well as contribute to immune suppression is an active area of research. Recent efforts have provided us with mechanistic insights into how B cells can positively and negatively regulate immune responses. Negative regulation of immune responses in cancer can be mediated by regulatory B cells and is often a result of increased production of cytokines that can directly and indirectly affect anti-tumor immune function and cancer cell growth. Signals that lead to the expansion of regulatory B cells and the spectrum of their functional roles are not well understood and are the subject of active research by many groups. Here, we elaborate broadly on the history of regulatory B cells in cancer and summarize recent studies that have established genetic models for the study of regulatory B cell function and their potential for therapeutic intervention in the setting of solid cancers.

Psychological stress and cancer: new evidence of an increasingly strong link

To date stress, a highly complex process that disrupts homeostasis and involves environmental and psychosocial factors, is considered as one of the most crucial factor that affects our daily life, especially urban dweller’s life.Clinical and experimental studies widely support the notion that adrenergic stimulation due to chronic stress affects inflammation and metabolism.In this work, supported by several recent scientific evidences, we show how stress plays a positive role in cancer initiation, progression and cancer metastasis, a negative role for anti-tumor immune function and therapy response.Understanding the intricacies of this interaction could provide an additional help on how act in cancer prevention and therapy.

Oncolytic Viruses and Immune Checkpoint Inhibitors: Preclinical Developments to Clinical Trials.

Immuno-oncology (IO) has been an active area of oncology research. Following US FDA approval of the first immune checkpoint inhibitor (ICI), ipilimumab (human IgG1 k anti-CTLA-4 monoclonal antibody), in 2011, and of the first oncolytic virus, Imlygic (talimogene laherparepvec), in 2015, there has been renewed interest in IO. In the past decade, ICIs have changed the treatment paradigm for many cancers by enabling better therapeutic control, resuming immune surveillance, suppressing tumor immunosuppression, and restoring antitumor immune function. However, ICI therapies are effective only in a small subset of patients and show limited therapeutic potential due to their inability to demonstrate efficacy in ‘cold’ or unresponsive tumor microenvironments (TMEs). Relatedly, oncolytic viruses (OVs) have been shown to induce antitumor immune responses, augment the efficacy of existing cancer treatments, and reform unresponsive TME to turn ‘cold’ tumors ‘hot,’ increasing their susceptibility to checkpoint blockade immunotherapies. For this reason, OVs serve as ideal complements to ICIs, and multiple preclinical studies and clinical trials are demonstrating their combined therapeutic efficacy. This review will discuss the merits and limitations of OVs and ICIs as monotherapy then progress onto the preclinical rationale and the results of clinical trials of key combination therapies.

The Multifaceted Output of c-Jun Biological Activity: Focus at the Junction of CD8 T Cell Activation and Exhaustion.

c-Jun is a major component of the dimeric transcription factor activator protein-1 (AP-1), a paradigm for transcriptional response to extracellular signaling, whose components are basic-Leucine Zipper (bZIP) transcription factors of the Jun, Fos, activating transcription factor (ATF), ATF-like (BATF) and Jun dimerization protein 2 (JDP2) gene families. Extracellular signals regulate c-Jun/AP-1 activity at multiple levels, including transcriptional and posttranscriptional regulation of c-Jun expression and transactivity, in turn, establishing the magnitude and the duration of c-Jun/AP-1 activation. Another important level of c-Jun/AP-1 regulation is due to the capability of Jun family members to bind DNA as a heterodimer with every other member of the AP-1 family, and to interact with other classes of transcription factors, thereby acquiring the potential to integrate diverse extrinsic and intrinsic signals into combinatorial regulation of gene expression. Here, we review how these features of c-Jun/AP-1 regulation underlie the multifaceted output of c-Jun biological activity, eliciting quite distinct cellular responses, such as neoplastic transformation, differentiation and apoptosis, in different cell types. In particular, we focus on the current understanding of the role of c-Jun/AP-1 in the response of CD8 T cells to acute infection and cancer. We highlight the transcriptional and epigenetic regulatory mechanisms through which c-Jun/AP-1 participates in the productive immune response of CD8 T cells, and how its downregulation may contribute to the dysfunctional state of tumor infiltrating CD8 T cells. Additionally, we discuss recent insights pointing at c-Jun as a suitable target for immunotherapy-based combination approaches to reinvigorate anti-tumor immune functions.

Emerging Role of Immunotherapy for Colorectal Cancer with Liver Metastasis.

Colorectal cancer (CRC) is the third most common malignant tumor in the world and the second leading cause of cancer-related deaths, with the liver as the most common site of distant metastasis. The prognosis of CRC with liver metastasis is poor, and most patients cannot undergo surgery. In addition, conventional antitumor approaches such as chemotherapy, radiotherapy, targeted therapy, and surgery result in unsatisfactory outcomes. In recent years, immunotherapy has shown good prospects in the treatment of assorted tumors by enhancing the host’s antitumor immune function, and it may become a new effective treatment for liver metastasis of CRC. However, challenges remain in applying immunotherapy to CRC with liver metastasis. This review examines how the microenvironment and immunosuppressive landscape of the liver favor tumor progression. It also highlights the latest research advances in immunotherapy for colorectal liver metastasis and identifies immunotherapy as a treatment regimen with a promising future in clinical applications.

Immune Activation in Patients with Locally Advanced Cervical Cancer Treated with Ipilimumab Following Definitive Chemoradiation (GOG-9929).

A phase I clinical trial (GOG-9929) examined the safety and efficacy of adjuvant immune-modulation therapy with the checkpoint inhibitor ipilimumab [anti-CTL antigen-4 (anti-CTLA-4)] following chemoradiation therapy (CRT) for newly diagnosed node-positive human papillomavirus (HPV)-related cervical cancer. To better understand the mechanism of action and to identify predictive biomarkers, immunologic and viral correlates were assessed before, during, and after treatment. Twenty-one patients who received CRT and ≥2 doses of ipilimumab and 5 patients who received CRT only were evaluable for translational endpoints. Circulating T-cell subsets were evaluated by multiparameter flow cytometry. Cytokines were evaluated by multiplex ELISA. HPV-specific T cells were evaluated in a subset of patients by IFNγ ELISpot. Expression of the activation markers ICOS and PD-1 significantly increased on T-cell subsets following CRT and were sustained or increased following ipilimumab treatment. Combined CRT/ipilimumab treatment resulted in a significant expansion of both central and effector memory T-cell populations. Genotype-specific E6/E7-specific T-cell responses increased post-CRT in 1 of 8 HPV16+ patients and in 2 of 3 HPV18+ patients. Elevation in levels of tumor-promoting circulating cytokines (TNFα, IL6, IL8) post-CRT was significantly associated with worse progression-free survival. Our data indicate that CRT alone and combined with ipilimumab immunotherapy show immune-modulating activity in women with locally advanced cervical cancer and may be a promising therapeutic option for the enhancement of antitumor immune cell function after primary CRT for this population at high risk for recurrence and metastasis. Several key immune biomarkers were identified that were associated with clinical response.

Immune infiltration-associated serum amyloid A1 predicts favorable prognosis for hepatocellular carcinoma.

BACKGROUNDSerum amyloid A1 (SAA1) is an acute-phase protein involved in acute or chronic hepatitis. Its function is still controversial. In addition, the effect of the expression of SAA1 and its molecular function on the progression in hepatocellular carcinoma (HCC) is still unclear.AIMTo demonstrate the expression of SAA1 and its effect on the prognosis in HCC and explain further the correlation of SAA1 and immunity pathways.METHODSSAA1 expression in HCC was conducted with The Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) in GEPIA tool, and the survival analysis based on the SAA1 expression level was achieved in the Kaplan-Meier portal. The high or low expression group was then drawn based on the median level of SAA1 expression. The correlation of SAA1 and the clinical features were conducted in the UALCAN web-based portal with TCGA-LIHC, including tumor grade, patient disease stage, and the TP53 mutation. The correlation analysis between SAA1 expression and TP53 mutation was subjected to the TCGA portal. The tumor purity score and the immune score were analyzed with CIBERSORT. The correlation of SAA1 expression and tumor-infiltrating lymphocytes was achieved in TISIDB web-based integrated repository portal for tumor-immune system interactions. GSE125336 dataset was used to test the SAA1 expression in the responsive or resistant group with anti-PD1 therapy. Gene set enrichment analysis was applied to evaluate the gene enrichment signaling pathway in HCC. The similar genes of SAA1 in HCC were identified in GEPIA, and the protein-protein interaction of SAA1 was conducted in the Metascape tool. The expression of C-X-C motif chemokine ligand 2, C-C motif chemokine ligand 23, and complement C5a receptor 1 was studied and overall survival analysis in HCC was conducted in GEPIA and Kaplan-Meier portal, respectively.RESULTSSAA1 expression was decreased in HCC, and lower SAA1 expression predicted poorer overall survival, progression-free survival, and disease-specific survival. Furthermore, SAA1 expression was further decreased with increased tumor grade and patient disease stage. Also, SAA1 expression was further downregulated in patients with TP53 mutation compared with patients with wild type TP53. SAA1 expression was negatively correlated with the TP53 mutation. Lower SAA1 predicted poorer survival rate, especially in the patients with no hepatitis virus infection, other than those with hepatitis virus infection. Moreover, the SAA1 expression was negatively correlated with tumor purity. In contrast, SAA1 expression was positively correlated with the immune score in HCC, and the correlation analysis between SAA1 expression and tumor-infiltrating lymphocytes also showed a positive correlation in HCC. Decreased SAA1 was closely associated with the immune tolerance of HCC. C-X-C motif chemokine ligand 2 and C-C motif chemokine ligand 23 genes were identified as the hub genes associated with SAA1, which could also serve as favorable prognosis markers for HCC.CONCLUSIONSAA1 is downregulated in the liver tumor, and it is closely involved in the progression of HCC. Lower SAA1 expression indicates lower survival rate, especially for those patients without hepatitis virus infection. Lower SAA1 expression also suggests lower immune infiltrating cells, especially for those with immune cells exerting anti-tumor immune function. SAA1 expression is closely associated with the anti-tumor immune pathways.

Abstract 4507: TransConTM IL-2 β/γ: a novel long-acting prodrug of receptor-biased IL-2 designed for improved pharmocokinetics and optimal activation of T cells for the treatment of cancer

Abstract Background: Recombinant Interleukin-2 (IL-2, aldesleukin) is an approved immunotherapy for metastatic melanoma and renal cell carcinoma. However, treatment with IL-2 can induce severe side effects, such as vascular leak syndrome (VLS) and eosinophilic infiltration of cardiac and pulmonary tissues. IL-2 promotes the survival, proliferation and anti-tumor functions of CD8+ cytotoxic T cells, CD4+ effector T cells and natural killer cells, which mostly express the dimeric IL-2Rβγ receptor with low IL-2Rα levels. However, IL-2 also potently activates IL-2Rα+ immunosuppressive regulatory T cells as well as IL-2Rα+ eosinophils, type 2 innate lymphoid cells and endothelial cells, which may promote VLS. Aldesleukin is rapidly cleared with elimination T1/2 of 85 minutes requiring thrice daily dosing and resulting in large fluctuations in serum exposure, likely contributing to its toxicity. Thus, IL-2 has two critical drawbacks for cancer immunotherapy: 1) potent activation of undesired cells types expressing IL-2Rα and 2) high Cmax with rapid clearance. Methods: We developed TransCon (transient conjugation) IL-2 β/γ, a novel sustained release prodrug of a receptor-biased IL-2 (IL-2 β/γ) to optimally address each of these drawbacks. First, to block IL-2Rα binding yet retain IL-2Rβγ activity, we created IL2 β/γ by permanently attaching a small PEG moiety to an engineered cysteine placed at the IL-2Rα binding site. Second, to improve PK properties, we attached the receptor-biased IL-2 β/γ to a TransCon PEG carrier via a TransCon linker, shielding bioactivity and creating a prodrug. Under physiological conditions, TransCon IL-2 β/γ was designed for sustained release of the bioactive IL-2 β/γ from the PEG carrier, aiming for a much lower Cmax and longer effective half-life of released IL-2 β/γ compared to aldesleukin. Results: In binding and cell-based functional assays, free IL-2 β/γ demonstrated desirable IL-2 receptor selectivity maintaining IL-2Rβγ potency while losing IL2Rα potency. In vitro, TransCon IL-2 β/γ showed expected slow-release kinetics, supporting a half-life in humans of 2-3 days. In mouse tumor models, TransCon IL2 β/γ promoted CD8 and NK cell proliferation and activation. In cynomolgus monkeys, a single dose of TransCon IL-2 β/γ was well tolerated and induced a more robust CD8+ T cell and NK cell expansion relative to CD4+ T cells or eosinophils as compared to daily aldesleukin treatment. Consistent with these observations, TransCon IL-2 β/γ induced lower levels of systemic inflammatory cytokines and endothelial activity biomarkers when compared to aldesleukin. Summary: The data presented here provide evidence that TransCon IL-2 β/γ may promote anti-tumor immune effector function with improved tolerability. TransCon IL-2 β/γ was designed as a novel long-acting, receptor-biased IL-2 prodrug that has the potential to overcome the challenges of existing IL-2 treatments. Citation Format: David Brian Rosen, Burkhardt Laufer, Thomas Knappe, Meike Schnabel, Diana Begaliew, Joachim Zettler, Mathias Krusch, Ana Bernhard, Stefan Heinig, Valentino Konjik, Thomas Wegge, Vibeke Breinholt, Mohammad Tabrizi, Kennett Sprogoe, Juha Punnonen. TransConTM IL-2 β/γ: a novel long-acting prodrug of receptor-biased IL-2 designed for improved pharmocokinetics and optimal activation of T cells for the treatment of cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4507.

Abstract 6621: A combined VHH and small molecule approach to modulate the antitumor immunity function of ERAP1

Abstract ERAP1 is a multifunctional enzyme trimming N-terminal amino acids of peptides. Its most characterised function is its role in generating peptide ligands in the endoplasmic reticulum for loading onto MHC class I (MHCI) molecules for presentation at the cell surface. Here it acts as an antigenic peptide editor influencing the peptide repertoire displayed at the cell surface to circulating CD8+ T cells and NK cells. ERAP1 modulates the immunopeptidome and could play a key role in tumor antigenicity. ERAP1 is a highly dynamic enzyme and its conformation is crucial for intracellular immuno-signals and also various extracellular functions. The precise molecular mechanisms behind the secretion and conformational status of ERAP1 remain unclear. To study ERAP1, we combined a VHH and a small molecule approach. After Lama immunization, the selected ERAP1 binders were filtered for functional activity. To measure ERAP1 activity with relevant substrates, we developed a mass spectrometry assay and a fluorogenic substrate mimicking ERAP1 peptidic substrates. Nanobodies modulators of ERAP1 activity are currently used for structural studies. In addition to the mechanistic studies, the team also developed ERAP1 small molecule modulators to induce or restore tumor immunogenicity. Herein, we report a chemical family of potent ERAP1 inhibitors that demonstrate structure activity relationship. The nanobodies and small molecule modulators we developed provide useful tools to better understand the biology of ERAP1 in immune responses. Citation Format: Nicolas Basse. A combined VHH and small molecule approach to modulate the antitumor immunity function of ERAP1 [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6621.

Antagonistic anti-LILRB1 monoclonal antibody regulates antitumor functions of natural killer cells.

BackgroundCurrent immune checkpoint blockade strategies have been successful in treating certain types of solid cancer. However, checkpoint blockade monotherapies have not been successful against most hematological malignancies including multiple myeloma and leukemia. There is an urgent need to identify new targets for development of cancer immunotherapy. LILRB1, an immunoreceptor tyrosine-based inhibitory motif-containing receptor, is widely expressed on human immune cells, including B cells, monocytes and macrophages, dendritic cells and subsets of natural killer (NK) cells and T cells. The ligands of LILRB1, such as major histocompatibility complex (MHC) class I molecules, activate LILRB1 and transduce a suppressive signal, which inhibits the immune responses. However, it is not clear whether LILRB1 blockade can be effectively used for cancer treatment.MethodsFirst, we measured the LILRB1 expression on NK cells from cancer patients to determine whether LILRB1 upregulated on NK cells from patients with cancer, compared with NK cells from healthy donors. Then, we developed specific antagonistic anti-LILRB1 monoclonal antibodies and studied the effects of LILRB1 blockade on the antitumor immune function of NK cells, especially in multiple myeloma models, in vitro and in vivo xenograft model using non-obese diabetic (NOD)-SCID interleukin-2Rγ-null mice.ResultsWe demonstrate that percentage of LILRB1+ NK cells is significantly higher in patients with persistent multiple myeloma after treatment than that in healthy donors. Further, the percentage of LILRB1+ NK cells is also significantly higher in patients with late-stage prostate cancer than that in healthy donors. Significantly, we showed that LILRB1 blockade by our antagonistic LILRB1 antibody increased the tumoricidal activity of NK cells against several types of cancer cells, including multiple myeloma, leukemia, lymphoma and solid tumors, in vitro and in vivo.ConclusionsOur results indicate that blocking LILRB1 signaling on immune effector cells such as NK cells may represent a novel strategy for the development of anticancer immunotherapy.

617 Skin squamous cell carcinoma (SCC)-derived exosomes after ALA-PDT treatment deliver microRNA-3473 to induce M1 macrophage polarization via NF-κB pathway

Topical ALA-mediated PDT, ALA-PDT, is a novel therapeutic modality widely used to treat actinic keratosis, Bowen’s Disease, superficial skin SCC, and other cancerous and precancerous skin diseases. Several studies have proved that ALA-PDT can inhibit SCC growth and reduce tumor volume. Subsequent research suggested that ALA-PDT not only directly induced tumor cells apoptosis, but also improve tumor microenvironment through regulation of immune cells. However, the anti-tumor immune function of ALA-PDT is still need to be elucidated.

Thermosensitive gel of polysaccharide from Ganoderma applanatum combined with paclitaxel for mice with 4T1 breast cancer

Polysaccharide from Ganoderma applanatum has the activities of anti-tumor and enhancing immune function. There were no reports on antitumor effect of its intratumoral injection. In this study, the polysaccharide was extracted from G. applanatum by water extraction and alcohol precipitation, and purified by ceramic membrane after removing protein by Sevage method. The total polysaccharide content from G. applanatum(PGA)was about 63%. The combination of PGA and paclitaxel showed synergistic effect on cytotoxicity of 4 T1 cells at lower concentrations in vitro. In addition, the growth curve of 4 T1 cells showed that PGA could retard the growth of 4 T1 cells gradually. The PGA thermosensitive gel(PGA-TG)was prepared by using poloxamer 188 and 407. The gel temperature was 36 ℃, and the PGA-TG could effectively slow down the release rate of PGA in vitro. 4 T1 breast cancer-bearing mice were used as a model to evaluate the therapeutic effect of intratumoral injection of PGA combined with tail vein injection of nanoparticle albumin-bound paclitaxel(nab-PTX). In high and low dose PGA groups, each mice was given with 2.25, 1.125 mg PGA respectively, twice in total, and the dosage of paclitaxel was 15 mg·kg~(-1), once every 3 days, for a total of five times. The tumor inhibition rate was 29.65% in the high dose PGA-TG group, 58.58% in the nab-PTX group, 63.37% in low dose PGA-TG combined with nab-PTX group, and 68.10% in high dose PGA-TG combined with nab-PTX group respectively. The inhibitory effect in high dose PGA-TG group combined with nab-PTX on tumors was significantly higher than that in nab-PTX group(P&lt;0.05). The results showed that paclitaxel therapy combined with intratumoral injection of PGA-TG could improve the therapeutic effect for 4 T1 mice and reduce the side effects of chemotherapy.