Immune-stimulating Agents Research Articles

Recent Advances in Bacterium-Based Therapeutic Modalities for Melanoma Treatment.

Melanoma is one of the most severe skin cancer indications with rapid progression and a high risk of metastasis. However, despite the accumulated advances in melanoma treatment including adjuvant radiation, chemotherapy, and immunotherapy, the overall melanoma treatment efficacy in the clinics is still not satisfactory. Interestingly, bacterial therapeutics have demonstrated unique properties for tumor-related therapeutic applications, such as tumor-targeted motility, tailorable cytotoxicity, and immunomodulatory capacity of the tumor microenvironment, which have emerged as a promising platform for melanoma therapy. Indeed, the recent advances in genetic engineering and nanotechnologies have boosted the application potential of bacterium-based therapeutics for treating melanoma by further enhancing their tumor-homing, cell-killing, drug delivery, and immunostimulatory capacities. This review provides a comprehensive summary of the state-of-the-art bacterium-based anti-melanoma modalities, which are categorized according to their unique functional merits, including tumor-specific cytotoxins, tumor-targeted drug delivery platforms, and immune-stimulatory agents. Furthermore, a perspective is provided discussing the potential challenges and breakthroughs in this area. The insights in this review may facilitate the development of more advanced bacterium-based therapeutic modalities for improved melanoma treatment efficacy.

Abstract 7240: Intratumor delivery of plasmid DNA encoding immune stimulating agents induces both local and systemic responses

Abstract Immune checkpoint inhibitors (ICIs) have significant clinical benefits, but are limited by both primary and acquired resistance. The absence of T cells within the tumor microenvironment (TME) is a significant hurdle, reducing the efficacy of ICIs against solid tumors. Multiple studies have been initiated to modify the TME to enhance the effectiveness of ICIs. We previously demonstrated that the intratumor delivery of a plasmid encoding interleukin-12 (pIL-12) using gene electrotransfer (GET) resulted in a significant increase in T effector cells and a reduction in T regulatory cells and myeloid derived suppressor cells within the TME, inducing a robust immune response. Combining pIL-12 GET with ICIs resulted in local and systemic responses in both preclinical and clinical studies. While encouraging results were obtained, improvements were possible. For example, the established GET technology requires high applied voltage for plasmid delivery. In addition, satisfactory delivery cannot be immediately detected. Our current research has focused on developing the next generation GET technology to overcome these issues. This new system includes a pulse generator, a heat source and a tissue impedance measuring device. An array that incorporates independently addressable electrodes was also developed. A moderate elevation of the tissue temperature coupled with the electrode array enabled a 75% reduction in the applied voltage. We tested the approach in the B16.F10 mouse melanoma model. Delivery of pIL-12 with the new system resulted in 100% long-term complete regression and 100% protection from challenge. In addition, monitoring impedance within each independent section of the electrode array revealed different pulse numbers were needed to achieve delivery in each section. We next evaluated a combination therapy, pIL-12 plasmid combined with a plasmid encoding a PD1 peptide, delivered with the next-gen device. In a multi-tumor model, the plasmids were delivered to the subcutaneous tumor; tumors generated with an intraperitoneal injection were untreated. This new therapy induced complete regression of the subcutaneous tumor and blockage of peritoneal tumor growth as assessed via in vivo imaging. Peritoneal tumor growth was not affected by single plasmid delivery or when both plasmids were delivered with the established GET technology. Work is ongoing to translate this approach to clinical evaluation. Citation Format: Richard Heller, Loree C. Heller, Guilan Shi, Jody Synowiec, Julie Singh, Alex Otten, Mark J. Jaroszeski. Intratumor delivery of plasmid DNA encoding immune stimulating agents induces both local and systemic responses [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7240.

Natural Immunomodulatory Agents as a Complementary Therapy for Poxviruses.

Poxviruses target innate immunity mediators such as tumor necrosis factors, interleukins, interferons, complement, and chemokines. It also targets adaptive immunity such as CD4+ T cells, CD4+ T cells, and B cells. Emerging of the recent epidemic of monkeypox virus (MPXV), a zoonotic disease native to Central and Western Africa, besides the lack of permitted treatments for poxviruses infections, encouraged researchers to identify effective inhibitors to help in preventing and treating poxviruses infections. Natural bioactive components, particularly polyphenolics, are promising for creating powerful antioxidants, anti-inflammatory, immune-stimulating, and antiviral agents. As a result, they are potentially effective therapies for preventing and treating viral diseases, such as infections caused by poxviruses including the recent pandemic MPXV. Polyphenolics: rosmarinic acid, caffeic acid, resveratrol, quercitrin, myricitrin, gingerol, gallotannin, and propolis-benzofuran A, as well as isoquinoline alkaloids: galanthamine and thalimonine represent prospective antiviral agents against MPXV, they can inhibit MPXV and other poxviruses via targeting different viral elements including DNA Topoisomerase I (TOP1), Thymidine Kinase (TK), serine/threonine protein kinase (Ser/Thr kinase), and protein A48R. The bioactive extracts of different traditional plants including Guiera senegalensis, Larrea tridentata, Sarracenia purpurea, Kalanchoe pinnata (Lam.) Pers., Zingiber officinale Roscoe, Quercus infectoria, Rhus chinensis, Prunella vulgaris L., Salvia rosmarinus, and Origanum vulgare also can inhibit the growth of different poxviruses including MPXV, vaccinia virus (VACV), variola virus, buffalopox virus, fowlpox virus, and cowpox virus. There is an urgent need for additional molecular studies to identify and confirm the anti-poxviruses properties of various natural bioactive components, especially those that showed potent antiviral activity against other viruses.

Rise of Antibody-Drug Conjugates: The Present and Future.

Antibody-drug conjugates (ADCs) embody a simple, but elegant, vision for cancer therapy-the delivery of a potent cytotoxic agent to tumor cells with minimal damage to normal cells-so-called smart chemo. Although there were significant challenges in achieving this milestone culminating in the first Food and Drug Administration approval in 2000, subsequent advancements in technology have led to rapid drug development with regulatory approvals for ADCs targeting a variety of tumor types. The most successful application for solid tumors has been in breast cancer, with ADCs becoming the standard of care across traditional human epidermal growth factor receptor 2 (HER2)+, hormone receptor+ (HR+) and triple-negative disease subtypes. Moreover, the improved features and gains in potency with the development of ADCs have expanded the treatment-eligible population to those with low/heterogeneous expression of the target antigen on the tumor with trastuzumab deruxtecan or in the case of sacituzumab govitecan, agnostic to target expression. Despite their antibody-directed homing, these novel agents come with their share of toxicities obligating appropriate patient selection and vigilant monitoring while on treatment. As more ADCs are included in the treatment armamentarium, mechanisms of resistance need to be studied and understood for optimal sequencing. Modifying the payload to use immune-stimulating agents or combination therapies with immunotherapy and other effective targeted therapies may further extend the utility of these agents in the treatment of solid tumors.

The role of viruses in cancer development versus cancer therapy: An oncological perspective.

Despite great medical advances, oncological research is still looking for novel therapeutic approaches due to the limitation of conventional therapeutic agents. Virotherapy is one of these new emerging therapeutic approaches that attract attention with their widespread applications. Virotherapy use lives oncolytic viruses or genetically engineered viruses that selectively infect the tumor cells, replicate, and disrupt the cancerous cells that also induce their anticancer activity by stimulating the host antitumor immune response. Moreover, viruses are widely used as target delivery vectors for specifically delivering different genes, therapeutic agents, and immune-stimulating agents. In addition to having antitumor activity by themselves in combination with conventional therapeutic agents like immune therapy and chemotherapy, Virotherapy agents also elicit promising outcomes. Therefore, in addition to their promising result in monotherapy use, virotherapy agents can also be used in combination with conventional cancer therapy, epigenetic modulators, and even microRNAs without any cross-resistance, which allows the patient not to be deprived of her routine medicine. Still, this combination therapy reduces the adverse effect of the conventional therapies. All together suggest that virotherapy agents as novel potential agents in the field of cancer therapy.

Detecting monocyte trafficking in an animal model of glioblastoma using R2* and quantitative susceptibility mapping.

The role of tumor-associated macrophages (TAMs) in glioblastoma (GBM) disease progression has received increasing attention. Recent advances have shown that TAMs can be re-programmed to exert a pro-inflammatory, anti-tumor effect to control GBMs. However, imaging methods capable of differentiating tumor progression from immunotherapy treatment effects have been lacking, making timely assessment of treatment response difficult. We showed that tracking monocytes using iron oxide nanoparticle (USPIO) with MRI can be a sensitive imaging method to detect therapy response directed at the innate immune system. We implanted syngeneic mouse glioma stem cells into C57/BL6 mice and treated the animals with either niacin (a stimulator of innate immunity) or vehicle. Animals were imaged using an anatomical MRI sequence, R2* mapping, and quantitative susceptibility mapping (QSM) before and after USPIO injection. Compared to vehicles, niacin-treated animals showed significantly higher susceptibility and R2*, representing USPIO and monocyte infiltration into the tumor. We observed a significant reduction in tumor size in the niacin-treated group 7days later. We validated our MRI results with flow cytometry and immunofluoresence, which showed that niacin decreased pro-inflammatory Ly6C high monocytes in the blood but increased CD16/32 pro-inflammatory macrophages within the tumor, consistent with migration of these pro-inflammatory innate immune cells from the blood to the tumor. MRI with USPIO injection can detect therapeutic responses of innate immune stimulating agents before changes in tumor size have occurred, providing a potential complementary imaging technique to monitor cancer immunotherapies. We show that iron oxide nanoparticles (USPIOs) can be used to label innate immune cells and detect the trafficking of pro-inflammatory monocytes into the glioblastoma. This preceded changes in tumor size, making it a more sensitive imaging technique.

Immune suppression is associated with enhanced systemic inflammatory, endothelial and procoagulant responses in critically ill patients

Patients admitted to the Intensive Care Unit (ICU) oftentimes show immunological signs of immune suppression. Consequently, immune stimulatory agents have been proposed as an adjunctive therapy approach in the ICU. The objective of this study was to determine the relationship between the degree of immune suppression and systemic inflammation in patients shortly after admission to the ICU. Design: An observational study in two ICUs in the Netherlands. The capacity of blood leukocytes to produce cytokines upon stimulation with lipopolysaccharide (LPS) was measured in 77 patients on the first morning after ICU admission. Patients were divided in four groups based on quartiles of LPS stimulated tumor necrosis factor (TNF)-α release, reflecting increasing extents of immune suppression. 15 host response biomarkers indicative of aberrations in inflammatory pathways implicated in sepsis pathogenesis were measured in plasma. A diminished capacity of blood leukocytes to produce TNF-α upon stimulation with LPS was accompanied by a correspondingly reduced ability to release of IL-1β and IL-6. Concurrently measured plasma concentrations of host response biomarkers demonstrated that the degree of reduction in TNF-α release by blood leukocytes was associated with increasing systemic inflammation, stronger endothelial cell activation, loss of endothelial barrier integrity and enhanced procoagulant responses. In patients admitted to the ICU the strongest immune suppression occurs in those who simultaneously display signs of stronger systemic inflammation. These findings may have relevance for the selection of patients eligible for administration of immune enhancing agents. ClinicalTrials.gov identifier NCT01905033.

Abstract 2869: ADG138, a novel HER2×CD3 POWERbody™ integrating bispecific TCE with precision masking to control cytokine release syndrome and on-target off-tumor toxicity for single agent and combination therapies in HER2-expressing solid tumors

Abstract Bispecific T-cell engagers (TCEs) have shown therapeutic promise in treating hematologic cancers. However, their utility in treating solid tumors has yet to be effectively demonstrated due to on-target off-tumor toxicity and systemic cytokine release syndrome (CRS). To explore the full potential of TCEs as a powerful modality of immunotherapy for solid tumors, we developed ADG138, a novel HER2×CD3 POWERbody™ TCE by applying our SAFEbody® precision masking technology to double mask the binding interfaces in both ADG138 arms using covalently linked designer peptides in order to limit on-target off-tumor toxicities in normal tissues and to avert systemic CRS. In the tumor microenvironment (TME), the double masked ADG138 prodrug can be activated for concomitant in situ binding to the targeted cancer cells by the HER2 arm and to T cells by the CD3 arm to selectively enhance T cell responses against tumor cells specifically. Nonclinical studies demonstrated that when activated, ADG138 was highly potent in inducing T cell mediated cytotoxicity of both high and low HER2-expressing tumor cells in vitro, whereas in its double masked prodrug form ADG138 exhibited high masking efficiency with greatly reduced HER2 binding, T cell activation, and T cell mediated cytotoxicity. In in vivo human PBMC-engrafted tumor xenograft models, ADG138 induced robust tumor regression of both high and low HER2-expressing tumors in mice. Furthermore, combination of ADG138 with our species cross-reactive immuno-oncology therapeutics mediated strong synergistic antitumor activities in HER2 positive syngeneic solid tumor models in human CD3e knock-in mice. The double masked ADG138 prodrug also exhibited excellent safety profiles and normal pharmacokinetics in exploratory toxicology studies in cynomolgus monkeys, achieving over a 300-fold higher tolerated dose in striking contrast with its parental TCE that lacks masking. These nonclinical studies demonstrated that ADG138, a novel double masked HER2×CD3 POWERbody TCE, exhibited an impressively high therapeutic index relative to its parental non-masked TCE in HER2 high and low expressing solid tumors. This provides strong support for advancing the ADG138 prodrug into clinical development for treating HER2-expressing solid tumors as a single agent and in combination with other immune stimulatory agents. Citation Format: Guizhong Liu, Yan Li, Zhengxi Dai, Jiagui Qu, Jianfeng Shi, Qi Zhao, Linhai Qu, Aaron N. Nguyen, Songmao Zheng, Jiangchun Xu, Felix Du, Peter Luo. ADG138, a novel HER2×CD3 POWERbody™ integrating bispecific TCE with precision masking to control cytokine release syndrome and on-target off-tumor toxicity for single agent and combination therapies in HER2-expressing solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2869.

Exploring the SARS-Cov-2 Main Protease (Mpro) and RdRp Targets by Updating Current Structure-based Drug Design Utilizing Co-crystals to Combat COVID-19.

The unprecedented pandemic of COVID-19 caused by the novel strain of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) engulfs millions of death worldwide. It has directly hit the socio-economic status of the affected countries. There are more than 219 countries badly affected by the COVID-19. There are no particular small molecule inhibitors to combat the dreadful virus. Many antivirals, antimalarials, antiparasitic, antibacterials, immunosuppressive antiinflammatory, and immune stimulatory agents have been repurposed for the treatment of COVID-19. But the exact mechanism of action of these drugs towards COVID-19 targets has not been experimented with yet. Under the effect of chemotherapeutics, the virus may change its genetic material and produces various strains, which are the main reasons behind the dreadful attack of COVID-19. The nuclear genetic components are composed of main protease and RNA-dependent RNA polymerase (RdRp) which are responsible for producing nascent virion and viral replication in the host cells. To explore the biochemical mechanisms of various small molecule inhibitors, structure-based drug design can be attempted utilizing NMR crystallography. The process identifies and validates the target protein involved in the disease pathogenesis by the binding of a chemical ligand at a well-defined pocket on the protein surface. In this way, the mode of binding of the ligands inside the target cavity can be predicted for the design of potent SARS-CoV-2 inhibitors.

Immunomodulatory Activity of a Polyherbal Immune Blend in Wistar Rats

Abstract Objectives The immune system plays an important role in protection against foreign organisms and its modulation is implicated in infection, inflammatory and autoimmune diseases. Several plant extracts have been explored as immune modulators which work either as immunosuppressive or immune-stimulating agents. he present study explores an in vivo immunomodulatory activity of an in-house formulated combination of plant extracts, called as Immune blend consisting of extracts of Boswellia, Ashwagandha, Neem, Ultrasol Curcumin, and Star anise. Methods The study was performed using 32 male wistar rats divided into 4 groups (vehicle control, dexamethasone, immune blend, dexamethasone + immune blend). Immunosuppression was induced in rats using dexamethasone (5 mg/kg body weight) intraperitoneally. The in-vivo immunomodulatory activity of Immune blend (at a dose of 50 mg/kg body weight) was evaluated by measurement of hemagglutination titer, delayed-type hypersensitivity (DTH) response against a foreign antigen and carbon clearance test. Additionally, synergy of the combination of plant extracts was measured in lipopolysaccharide (LPS) -induced THP1 cells for their ability to inhibit inflammatory cytokines through fluorescence-activated cell-sorting (FACS). Results Immune blend demonstrated significant (P < 0.05) increase in hemagglutination titer against sheep red blood cell (SRBC) as compared to dexamethasone control animals. Further immune blend also led to significant increase (P < 0.05) in cell mediated immune response against SRBC as measured by foot paw edema as compared to dexamethasone control animals. Pretreatment with the immune blend also showed a significant rise (P < 0.01) in phagocytic index indicated by phagocytic activity. Immune blend significantly inhibited expression of TNF-α and IL-2 in LPS-stimulated THP1 cells with synergistic response with the blend as compared to individual ingredients. Conclusions The study revealed that the immune blend stimulates both the cellular and the humoral immune responses under normal and immunosuppressed conditions in wistar rat model suggesting its therapeutic usefulness. Funding Sources OmniActive Health Technologies Limited.

Ilixadencel, a Cell-based Immune Primer, plus Sunitinib Versus Sunitinib Alone in Metastatic Renal Cell Carcinoma: A Randomized Phase 2 Study

BackgroundThe prognosis of patients with synchronous metastatic renal cell carcinoma (mRCC) is poor. Whereas single-agent tyrosine kinase inhibition (TKI) is clearly insufficient, the effects can be enhanced by combinations with immune checkpoint inhibitors. Innovative treatment options combining TKI and other immune-stimulating agents could prove beneficial. ObjectiveTo evaluate the clinical effects on metastatic disease when two doses of allogeneic monocyte-derived dendritic cells (ilixadencel) are administrated intratumorally followed by nephrectomy and treatment with sunitinib compared with nephrectomy and sunitinib monotherapy, in patients with synchronous mRCC. Design, setting, and participantsA randomized (2:1) phase 2 multicenter trial enrolled 88 patients with newly diagnosed mRCC to treatment with the combination ilixadencel/sunitinib (ILIXA/SUN; 58 patients) or sunitinib alone (SUN; 30 patients). Outcome measurements and statistical analysisThe primary endpoints were 18-mo survival rate and overall survival (OS). A secondary endpoint was objective response rate (ORR) assessed up to 18 mo after enrollment. Statistic evaluations included Kaplan-Meier estimates, log-rank tests, Cox regression, and stratified Cochran-Mantel-Haenszel tests. Results and limitationsThe median OS was 35.6 mo in the ILIXA/SUN arm versus 25.3 mo in the SUN arm (hazard ratio 0.73, 95% confidence interval 0.42–1.27; p = 0.25), while the 18-mo OS rates were 63% and 66% in the ILIXA/SUN and SUN arms, respectively. The confirmed ORR in the ILIXA/SUN arm were 42.2% (19/45), including three patients with complete response, versus 24.0% (six/25) in the SUN arm (p = 0.13) without complete responses. The study was not adequately powered to detect modest differences in survival. ConclusionsThe study failed to meet its primary endpoints. However, ilixadencel in combination with sunitinib was associated with a numerically higher, nonsignificant, confirmed response rate, including complete responses, compared with sunitinib monotherapy. Patient summaryWe studied the effects of intratumoral vaccination with ilixadencel followed by sunitinib versus sunitinib only in a randomized phase 2 study. The combination treatment showed numerically higher numbers of confirmed responses, suggesting an immunologic effect.

Collagen-targeted tumor-specific transepithelial penetration enhancer mediated intravesical chemoimmunotherapy for non-muscle-invasive bladder cancer

Intravesical instillation of chemotherapeutics or immune-stimulating agents could reduce the recurrence rate of non-muscle-invasive bladder cancer (NMIBC) after transurethral resection of the bladder tumors. Its efficacy, however, remains to be improved due to the bladder epithelial barrier. Although certain transmucosal delivery carriers are able to enhance the transepithelial penetration of intravesical agents, they could hardly differentiate carcinoma and adjacent normal tissues of the bladder wall. Here, we reported polyethylene glycol (PEG) & glutaraldehyde co-modified fluorinated chitosan (PGFCS) as a collagen-targeted transepithelial penetration enhancer, which could create a tumor-targeted adhesive interface by the aldehyde-selective reaction with collagen amines enriched in the tumor, thus opening the transepithelial-delivery barrier at the tumor site though the fluorinated-chitosan-mediated tight junction regulation. Interestingly, with the help of PGFCS pre-treatment, intravesical instillation of chemotherapeutics pirabucin (THP) combined with immune stimulating agent interleukin-12 could trigger potent antitumor chemoimmunotherapeutic responses in destructing orthotopic bladder tumors and inhibiting cancer recurrence. Our work presents a unique type of tumor-specific transepithelial penetration enhancer, which shows great potential for safe and effective intravesical instillation of NMIBC.

A Second-Generation Nanoluc-IL27 Fusion Cytokine for Targeted-Gene-Therapy Applications.

An emerging approach in treating skeletal malignancies utilizes osteoimmunology to investigate new multifunctional immune-stimulatory agents that can simultaneously combat tumor growth and promote bone repair. We have hypothesized that cytokine Interleukin-27 (IL-27) is an excellent candidate biologic to help rebalance the prostate tumor cells and bone cell environment. In this work, we examined the proof of principle for a short, secreted luciferase (Nanoluc or Nluc) fusion with IL-27 to produce a novel cytokine-based biologic (Nluc-27), whereby we examined its efficacy in vitro in reducing prostate tumor growth and rebalancing bone cell proliferation and differentiation. This work demonstrates the targeting and anti-tumor efficacy of the Nluc-27 fusion cytokine in cancer and bone cell models. The fusion cytokine is detectable in conditioned media, and bioactive in different cell systems. This novel Nluc-27 cytokine will allow flexible incorporation of other targeting domains and may serve as flexible tool to augment IL-27′s bioactivity and reengineer its efficacy against prostate tumor or bone cells, and may prove applicable to several other cell types for targeted gene therapy applications.

COVID-19 Response: The case for Phytomedicines in Africa with particular focus on Cameroon

Despite enormous efforts deployed and considerable positive results obtained in the global fight against the Coronavirus disease 2019 (COVID-19) the scourge remains a major international public health hazard. The main control measures at the onset consisted in the application of barrier and hygiene measures to stop the spread of the virus and case identification and clinical management of symptoms in the absence of widely available anti-COVOD-19 drugs. Vaccination as a major control measure became widely available in the advanced countries of the global north, but not in Africa where less than 5-10% 0f the population are vaccinated against COVID-19. However, African countries, possibly excluding South Africa, have been less impacted by COVID-19 pandemic as they registered fewer cases, hospitalizations and deaths. Herein it is postulated that the wide use of African traditional Phytomedicines (herbal medicines) has contributed, at least in part, to the better control of the COVID-19 pandemic in Africa. Abundant evidence in the literature suggests the availability of anti-viral, anti-oxidant and immune-stimulatory agents in the proposed COVID-19 herbal remedies., these activities being similar to those the standard drugs used in the standard treatment/ management of COVID-19. The review also examines a number of COVID-19 herbal medicines including COVID Organics CVO (Madagascar) ADSAR, ELISIR COVID, COROCUR (Cameroon) IHP Detox Tea (Nigeria) and COVIDEX (Uganda) and notes that though approved by the competent authorities in the respective African countries, these phytomedicines have not been approved by the WHO. It is proposed that additional studies be carried out to validate the Africa herbal remedies for possible use as stand-alone or complementary treatment of COVID-19 in addition to vaccination and barrier and hygiene control measures.

Reeducating Tumor-Associated Macrophages Using CpG@Au Nanocomposites to Modulate Immunosuppressive Microenvironment for Improved Radio-Immunotherapy.

With the recent success of immune checkpoint blockade (ICB) in cancer immunotherapy, there has been renewed interest in evaluating the combination of ICB inhibitors with radiotherapy (RT) in clinical trials in view of the localized RT-initiated vaccination effect, which can be augmented further by systemic immune-stimulating agents. Unfortunately, traditional RT/ICB accompanies severe toxicity from high-dose ionizing irradiation and low response rate from RT-aggravated immunosuppression, among which M2-type tumor-associated macrophages (TAMs) play an important role. Herein, CpG-decorated gold (Au) nanoparticles (CpG@Au NPs) were fabricated to improve the RT/ICB efficacy by immune modulation under low-dose X-ray exposure, where Au NPs served as radioenhancers to minimize the radiotoxicity, and yet acted as nanocarriers to deliver CpG, a toll-like receptor 9 agonist, to re-educate immunosuppressive M2 TAMs to immunostimulatory M1 counterparts, thus arousing innate immunity and meanwhile priming T cell activation. When combined with an anti-programmed death 1 antibody, irradiated CpG@Au led to consistent abscopal responses that efficiently suppressed distant tumors in a bilateral GL261 tumor-bearing model. This work thus demonstrates that CpG@Au-mediated macrophage reeducation could efficiently modulate the tumor-immune microenvironment for synergistic RT/ICB.

Conditional PD-1/PD-L1 Probody Therapeutics Induce Comparable Antitumor Immunity but Reduced Systemic Toxicity Compared with Traditional Anti-PD-1/PD-L1 Agents.

Immune-checkpoint blockade has revolutionized cancer treatment. However, most patients do not respond to single-agent therapy. Combining checkpoint inhibitors with other immune-stimulating agents increases both efficacy and toxicity due to systemic T-cell activation. Protease-activatable antibody prodrugs, known as Probody therapeutics (Pb-Tx), localize antibody activity by attenuating capacity to bind antigen until protease activation in the tumor microenvironment. Herein, we show that systemic administration of anti-programmed cell death ligand 1 (anti-PD-L1) and anti-programmed cell death protein 1 (anti-PD-1) Pb-Tx to tumor-bearing mice elicited antitumor activity similar to that of traditional PD-1/PD-L1-targeted antibodies. Pb-Tx exhibited reduced systemic activity and an improved nonclinical safety profile, with markedly reduced target occupancy on peripheral T cells and reduced incidence of early-onset autoimmune diabetes in nonobese diabetic mice. Our results confirm that localized PD-1/PD-L1 inhibition by Pb-Tx can elicit robust antitumor immunity and minimize systemic immune-mediated toxicity. These data provide further preclinical rationale to support the ongoing development of the anti-PD-L1 Pb-Tx CX-072, which is currently in clinical trials.

Applications of Magnetite Nanoparticles in Cancer Immunotherapies: Present Hallmarks and Future Perspectives.

Current immuno-oncotherapeutic protocols that inhibit tumor immune evasion have demonstrated great clinical success. However, the therapeutic response is limited only to a percentage of patients, and the immune-related adverse events can compromise the therapeutic benefits. Therefore, improving cancer immunotherapeutic approaches that pursue high tumor suppression efficiency and low side effects turn out to be a clinical priority. Novel magnetite nanoparticles (MNPs) exhibit great potential for therapeutic and imaging applications by utilizing their properties of superparamagnetism, good biocompatibility, as well as the easy synthesis and modulation/functionalization. In particular, the MNPs can exert magnetic hyperthermia to induce immunogenic cell death of tumor cells for effective antigen release and presentation, and meanwhile polarize tumor-associated macrophages (TAMs) to M1 phenotype for improved tumor killing capability, thus enhancing the anti-tumor immune effects. Furthermore, immune checkpoint antibodies, immune-stimulating agents, or tumor-targeting agents can be decorated on MNPs, thereby improving their selectivity for the tumor or immune cells by the unique magnetic navigation capability of MNPs to promote the tumor killing immune therapeutics with fewer side effects. This mini-review summarizes the recent progress in MNP-based immuno-oncotherapies, including activation of macrophage, promotion of cytotoxic T lymphocyte (CTL) infiltration within tumors and modulation of immune checkpoint blockade, thus further supporting the applications of MNPs in clinical therapeutic protocols.

Immunological tumor heterogeneity and diagnostic profiling for advanced and immune therapies

Immunotherapies have changed the way how we treat cancer at all stages. The understanding of the immune system in individual tumor specimens guides the selection of immune-modulating agents such as immune checkpoint inhibitors alone or in combination with other therapeutic agents that target, modulate or unleash the patient's immune system. Despite the similar histopathological diagnosis, each tumor is unique at its primary site and site of metastasis, also depending on previous treatment regimens or genetic alterations, such as chromosomal instability or acquired mutations. The clinically well-established use of PD-1/PD-L1 inhibitors already requires the assessment of its target molecules in different cells (viable tumor cells alone or in combination with immune cells or immune cells alone) with different thresholds in various indications. Anyhow, checkpoint inhibitors show the best overall response rate when immune effector cells like tumor-infiltrating lymphocytes are in close spatial proximity without being suppressed by other humoral or cellular regulatory mechanisms. Therefore, immune cell-rich tumors (“hot tumors”) are usually quite reactive to immune-modulating agents, whereas other immune-depleted or immune-excluded tumor areas are less responsive and require alternative treatment regimens such as modified immune effectors cells or immune-stimulating agents, for example, oncolytic viruses. Here, we summarize the relevance to understand the entire tumor heterogeneity and its environment, the contextual relationship and spatial quantification of all immune and tumor cells along with the genetic background of the individual cancer through the application of multiplex in-situ technologies and the application of machine learning tools.

Thermal Ablation of Colorectal Cancer Metastases Leads to Activation of Individual Cancer-Specific T Cell Immunity

Introduction: The underlying cellular mechanisms of successful immunotherapy are still insufficiently characterized. Especially, priming of tumor-specific T cells against relevant antigens and in situ tumor-recognition is crucial for therapy success in the development of active immunotherapeutic approaches, but remain a serious challenge. Thermal ablation of cancer, e.g. percutaneous radiofrequency ablation (RFA) is a well-established treatment option, particularly for patients with contraindications for surgical resection or in non-resectable primary or secondary liver cancers. RFA induces local tumor destruction by coagulation necrosis, leading to the release of danger signals (e.g. heat shock proteins) and cellular content that can induce and stimulate adaptive and innate immune responses, including tumor-antigen specific T cells. By breaking local immune tolerance, RFA may support the development of antigen-specific anti-tumor responses. Here, we analyzed patient-individual immune responses against autologous HLA-presented tumor antigens after RFA treatment.Materials and Methods: Patients (n=6) with liver metastases of colorectal carcinoma (CRC) in different liver segments were included. First, non-resectable manifestations were ablated using RFA. Subsequently, patients underwent metachronous liver surgery for resectable residual tumors. Using surgical specimens, immunopeptidome analyses of tumor and corresponding non-malignant liver tissues were performed by uHPLC-coupled tandem mass spectrometry (MS) after immunoprecipitation of HLA-ligands. Ligandome data were combined with results from whole transcriptome sequencing (WTS) in order to select for patient-specific HLA-ligands potentially relevant for the patient's individual tumor. Furthermore, identified peptides were compared with an extensive in house database of HLA ligands, derived from different non-transformed tissues including non-malignant colon (NMC) specimens. Peripheral blood mononuclear cells (PBMC) obtained at pre-defined time points before and after RFA and surgery were stimulated in vitro with these T cell epitope candidates. Functional T cell responses were characterized by intracellular cytokine staining (ICS).Results: Combining WTS data and analyses of autologous tumor-exclusive HLA-ligandomes, we selected 32 individual T cell epitope candidates for the respective patients. Antigen-specific T cells could be detected against approx. one third of the candidate epitopes. Both boosting of pre-existing as well as newly induced T cell responses against these epitopes could be detected after RFA. With regard to pre-existing immune responses, CD8+ and CD4+ T cell reactivities could be demonstrated against both HLA class I and class II epitopes. In 4/6 patients (66%) analyzed, de novo induced antigen-specific T cells could be detected after RFA. These reactivities comprised CD8+ as well as CD4+ T cells. Of particular interest, in one patient (diagnosed with mCRC) antigen-specific CD4+ T cells against a mutated HLA-class II peptide were detectable, which were induced by RFA. Of note, this epitope could only be detected on the transcriptome level but not directly as an HLA-ligand by MS.Conclusions: Employing this individualized approach, tumor-antigen specific T cells were more frequently detected as compared to our previous observations employing broadly expressed tumor-associated antigens. These findings suggest that RFA leads to induction of antigen-specific T cells more frequently than previously appreciated, and interestingly also encompasses responses against neo-antigens. The detected immune responses induced by RFA themselves may not be solely sufficient for effectively rejecting established tumors. However, these findings justify the clinical investigation of combinatorial therapies, using immune stimulatory agents or immune checkpoint inhibitors together with thermal ablation. DisclosuresNo relevant conflicts of interest to declare.

Eradication of Systemic Lymphoma By Local Immunotherapy

Background: It has recently become apparent that the immune system can cure cancer. There are variety of methods for generating these immune responses. In some of these strategies, such as CAR-T cells and the somatic mutation vaccines, the antigen targets are pre-identified and therapies are custom-made against these targets for each patient. In other approaches, such as checkpoint antibodies, “off the shelf” antibodies are used to remove the breaks on the immune system, allowing pre-existing T cells to attack unknown targets on cancer cells.We have developed another non-customized approach called in situ vaccination. Here, immune enhancing agents are injected locally into one site of tumor, that triggers a T cell immune response locally that then travels to attack cancer throughout the body.In order to screen for the best local immune stimulatory agents, we have employed a preclinical strategy whereby the same syngeneic tumor is implanted at two separate sites in the body. One tumor is then injected with the test agents and the resulting immune response is detected by the regression of the distant, untreated tumor.Using this assay for abscopal therapeutic effects we have screened a variety of ligands for Toll-Like Receptors (TLRs), antibodies against immune checkpoints and T cell co-stimulatory targets in multiple combinations of these agents.Among these, the combination of unmethylated CG-enriched oligodeoxynucleotide (CpG) -a TLR9 ligand and anti-OX40 antibody has provided the most impressive results. TLRs are components of the innate immune system that recognize molecular patterns on bacterial, fungal, or viral pathogens. Intratumoral (IT) injection of CpG, results in tumor eradication at the injected site but no in systemic anti-tumor immune response. However, it induced the enhanced expression of OX40 on the local tumor-infiltrating T cells. The addition of antibody against OX40, a potent costimulatory activator of T cells, resulted in eradication of both local and distant disease.This treatment resulted in a T cell immune response that could be measured both in vitro and in vivo. The systemic anti-tumor response required the presence of both CD4+ and CD8+ T cells as mice treated with the corresponding depleting antibodies were unable to control tumor growth.In addition, the cured animals were protected from re-challenge with the same A20 tumor but not unrelated tumors.To examine the potential of in situ vaccination to treat spontaneous cancers, we chose MMTV-PyVT a mouse tumor model that allowed for local injected at a predicable site of disease. These animals develop invasive breast cancer in all of their mammary glands by 12 weeks of age and widespread lethal metastases by 20 weeks of age. Injections of CpG and anti-OX40 antibody into the first arising tumor not only prevented its growth but prevented the outgrowth of subsequent tumors at un-injected mammary glands, reduced lung metastases and significantly improved overall survival.Impact: Anti-OX40 and CpG are both currently in phase-I trials as single agents. Each has proved safe with slight evidence of anti tumor effects. Our results provide strong rationale for using these two agents in combination as an in situ therapeutic vaccine for lymphoma tumors. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.