Establishment of a visualized mouse orthotopic xenograft model of nasopharyngeal carcinoma.

Mouse orthotopic xenograft tumor models are commonly employed in studies investigating the mechanisms underlying the development and progression of tumors and their preclinical treatment. However, the unavailability of mature and visualized orthotopic xenograft models of nasopharyngeal carcinoma limits the development of treatment strategies for this cancer. The aim of this study was to provide a simple and reliable method for building an orthotopic xenograft model of nasopharyngeal carcinoma. Human nasopharyngeal carcinoma (C666-1-luc) cells, stably expressing the firefly luciferase gene, were injected subcutaneously into the right axilla of BALB/C nude mice. Four weeks later, the resulting subcutaneous tumors were cut into small blocks and grafted into the nasopharynx of immunodeficient BALB/C nude mice to induce tumor formation. Tumor growth was monitored by bioluminescence imaging and small animal magnetic resonance imaging (MRI). The expression of histological and immunological antigens associated with orthotopic xenograft nasopharyngeal carcinoma was analyzed by tissue section analysis and immunohistochemistry (IHC). A visualized orthotopic xenograft nasopharyngeal carcinoma model was successfully developed in this study. Luminescence signal detection, micro-MRI, and hematoxylin and eosin staining revealed the successful growth of tumors in the nasopharynx of the nude mice. Moreover, IHC analysis detected cytokeratin (CK), CK5/6, P40, and P63 expression in the orthotopic tumors, consistent with the reported expression of these antigens in human nasopharyngeal tumors. This study established a reproducible, visual, and less lethal orthotopic xenograft model of nasopharyngeal carcinoma, providing a platform for preclinical research.

The Agonistic Activity of the Human Epidermal Growth Factor is Reduced by the D46G Substitution.

BACKGROUND: Resistance to anti-tumor agents targeting the epidermal growth factor receptor (EGFR) reduces treatment response and requires the development of novel EGFR antagonists. Mutant epidermal growth factor (EGF) forms with reduced agonistic activity could be promising agents in cancer treatment. METHODS: EGF D46G affinity to EGFR domain III was assessed with affinity chromatography. EGF D46G acute toxicity in Af albino mice at 320 and 3200 µg/kg subcutaneous doses was evaluated. EGF D46G activity in human epidermoid carcinoma cells at 10 ng/mL concentration in serum-free medium and in subcutaneous Ehrlich ascites carcinoma mice model at 320 µg/kg dose was studied. RESULTS: The D46G substitution decreases the thermal stability of EGF complexes with EGFR domain III by decreasing the ability of the C-terminus to be released from the intermolecular ß- sheet. However, with remaining binding sites for EGFR domain I, EGF D46G effectively competes with other EGF-like growth factors for binding to EGFR and does not demonstrate toxic effects in mice. EGF D46G inhibits the proliferation of human epidermoid carcinoma cells compared to native EGF. A single subcutaneous administration of EGF D46G along with Ehrlich carcinoma cells injection inhibits the proliferation of these cells and delays tumor formation for up to seven days. CONCLUSION: EGF D46G can be defined as a partial EGFR agonist as this mutant form demonstrates reduced agonistic activity compared to native EGF. The study emphasizes the role of the EGF C-terminus in establishing interactions with EGFR domain III, which are necessary for EGFR activation and subsequent proliferation of cells.

Defucosylated Monoclonal Antibody (H2Mab-139-mG2a-f) Exerted Antitumor Activities in Mouse Xenograft Models of Breast Cancers against Human Epidermal Growth Factor Receptor 2.

The clinically approved human epidermal growth factor receptor 2 (HER2)-targeting monoclonal antibodies (mAbs), trastuzumab, and pertuzumab, target domains IV and II, respectively. Trastuzumab is now the standard treatment for HER2-overexpressed breast and gastric cancers, and trastuzumab in combination with pertuzumab showed clinical benefit. However, there still exist patients who do not respond to the therapy. Furthermore, HER2 mutants that cannot be recognized by pertuzumab were found in tumors. Therefore, novel anti-HER2 mAbs and modalities have been desired. In our previous study, we developed a novel anti-HER2 domain I mAb, H2Mab-139 (mouse IgG1, kappa). We herein produced a defucosylated mouse IgG2a type of mAb against HER2 (H2Mab-139-mG2a-f) to enhance antibody-dependent cellular cytotoxicity (ADCC)-mediated antitumor activity. H2Mab-139-mG2a-f exhibits a high binding affinity in flow cytometry with the dissociation constant (KD) determined to be 3.9 × 10-9 M and 7.7 × 10-9 M against HER2-overexpressed Chinese hamster ovary (CHO)-K1 (CHO/HER2) and HER2-positive BT-474 cells, respectively. Moreover, we showed that H2Mab-139-mG2a-f exerted ADCC and complement-dependent cytotoxicity against CHO/HER2 and BT-474 in vitro and exhibited potent antitumor activities in mouse xenograft models. These results indicated that H2Mab-139-mG2a-f exerts antitumor effects against HER2-positive human breast cancers and is useful as an antibody treatment for HER2-positive human cancers.

A Coumarin-Imidazothiadiazole Derivative, SP11 Abrogates Tumor Growth by Targeting HSP90 and Its Client Proteins.

Despite several treatment options for blood cancer, mortality remains high due to relapse and the disease's aggressive nature. Elevated levels of HSP90, a molecular chaperone essential for protein folding, are associated with poor prognosis in leukemia and lymphoma. HSP90 as a target for chemotherapy has been met with limited success due to toxicity and induction of heat shock. This study tested the activity of an HSP90 inhibitor, SP11, against leukemic cells, mouse lymphoma allograft, and xenograft models. SP11 induced cytotoxicity in vitro in leukemic cell lines and induced cell death via apoptosis, with minimal effect on normal cells. SP11 induced cell death by altering the status of HSP90 client proteins both in vitro and in vivo. SP11 reduced the tumor burden in allograft and xenograft mouse models without apparent toxicity. The half-life of SP11 in the plasma was approximately 2 h. SP11 binding was observed at both the N-terminal and C-terminal domains of HSP90. C-terminal binding was more potent than N-terminal binding of HSP90 in silico and in vitro using isothermal calorimetry. SP11 bioavailability and minimal toxicity in vivo make it a potential candidate to be developed as a novel anticancer agent.

Combination of PD-1 Checkpoint Blockade and Botulinum Toxin Type A1 Improves Antitumor Responses in Mouse Tumor Models of Melanoma and Colon Carcinoma.

BACKGROUND: Tumor innervation has been shown to be utilized by some solid cancers to support tumor initiation, growth, progression, and metastasis, as well as confer resistance to immune checkpoint blockade through suppression of antitumor immunologic responses. Since botulinum neurotoxin type A1 (BoNT/A1) blocks neuronal cholinergic signaling, its potential use as an anticancer drug in combination with anti-PD-1 therapy was investigated in four different syngeneic mouse tumor models. METHODS: Mice implanted with breast (4T1), lung (LLC1), colon (MC38), and melanoma (B16-F10) tumors were administered a single intratumoral injection of 15 U/kg BoNT/A1, repeated intraperitoneal injections of 5 mg/kg anti-PD-1 (RMP1-14), or both. RESULTS: Compared to the single-agent treatments, anti-PD-1 and BoNT/A1 combination treatment elicited significant reduction in tumor growth among B16-F10 and MC38 tumor-bearing mice. The combination treatment also lowered serum exosome levels in these mice compared to the placebo control group. In the B16-F10 syngeneic mouse tumor model, anti-PD-1 + BoNT/A1 combination treatment lowered the proportion of MDSCs, negated the increased proportion of Treg cells, and elicited a higher number of tumor-infiltrating CD4+ and CD8+ T lymphocytes into the tumor microenvironment compared to anti-PD-1 treatment alone. CONCLUSION: Our findings demonstrate the synergistic antitumor effects of BoNT/A1 and PD-1 checkpoint blockade in mouse tumor models of melanoma and colon carcinoma. These findings provide some evidence on the potential application of BoNT/A1 as an anticancer drug in combination with immune checkpoint blockade and should be further explored.

In vivo Assessment of the Impact of Molecular Weight on Constructs of 68Ga-DOTA-Manocept in a Syngeneic Mouse Tumor Model.

PURPOSE: Manocept™ constructs are mannosylated amine dextrans (MADs) that bind with high affinity to the mannose receptor, CD206. Tumor-associated macrophages (TAMs) are the most numerous immune cells in the tumor microenvironment and a recognized target for tumor imaging and cancer immunotherapies. Most TAMs express CD206, suggesting utility of MADs to deliver imaging moieties or therapeutics to TAMs. The liver Kupffer cells also express CD206, making them an off-target localization site when targeting CD206 on TAMs. We evaluated TAM targeting strategies using two novel MADs differing in molecular weight in a syngeneic mouse tumor model to determine how varying MAD molecular weights would impact tumor localization. Increased mass dose of the non-labeled construct or a higher molecular weight (HMW) construct were also used to block liver localization and enhance tumor to liver ratios. PROCEDURES: Two MADs, 8.7 kDa and 22.6 kDa modified with DOTA chelators, were synthesized and radiolabeled with 68Ga. A HMW MAD (300 kDa) was also synthesized as a competitive blocking agent for Kupffer cell localization. Balb/c mice, with and without CT26 tumors, underwent dynamic PET imaging for 90 min followed by biodistribution analyses in selected tissues. RESULTS: The new constructs were readily synthesized and labeled with 68Ga with ≥ 95% radiochemical purity in 15 min at 65 °C. When injected at doses of 0.57 nmol, the 8.7 kDa MAD provided 7-fold higher 68Ga tumor uptake compared to the 22.6 kDa MAD (2.87 ± 0.73%ID/g vs. 0.41 ± 0.02%ID/g). Studies with increased mass of unlabeled competitors showed reduced liver localization of the [68Ga]MAD-8.7 to varying degrees without significant reductions in tumor localization, resulting in enhanced tumor to liver signal ratios. CONCLUSION: Novel [68Ga]Manocept constructs were synthesized and studied in in vivo applications, showing that the smaller MAD localized to CT26 tumors more effectively than the larger MAD and that the unlabeled HMW construct could selectively block liver binding of [68Ga]MAD-8.7 without diminishing the localization to tumors. Promising results using the [68Ga]MAD-8.7 show a potential path to clinical applications.

In vivo Preclinical Tumor-Specific Imaging of Superparamagnetic Iron Oxide Nanoparticles Using Magnetic Particle Imaging for Cancer Diagnosis.

Purpose: Magnetic particle imaging (MPI) is an emerging radiation-free, non-invasive three-dimensional tomographic technology that can visualize the concentrations of superparamagnetic iron oxide nanoparticles (SPIONs). To verify the applicability of the previously proposed point-of-care testing MPI (PoCT-MPI) in medical diagnosis and therapeutics, we imaged SPIONs in animal tumor models. Methods: CT26 or MC38 mouse colon carcinoma cells (2 × 106 cells) were subcutaneously injected into the right flank of BALB/c mice. SPIONs were either injected directly into the tumor lesions in the intratumoral group or through tail veins in the intravenous group. CT26 and MC38 tumor models were examined both intratumorally and intravenously to confirm the biological availability of SPIONs using PoCT-MPI. Results: Signals were observed in the tumor lesions from day 1 to day 7. This is the first study to successfully image the pathological region and show the biodistribution of SPIONs in CT26 tumor models using the recently developed PoCT-MPI technology. Furthermore, MC38 tumor models were examined, resulting in similar images to those of the CT26 tumor model in both intratumoral and intravenous groups. Conclusion: The present study demonstrates the biological applicability of PoCT-MPI, which promises to be a powerful diagnostic and therapeutic technique in biomedical imaging.

Gene Gun Her2/neu DNA Vaccination: Evaluation of Vaccine Efficacy in a Syngeneic Her2/neu Mouse Tumor Model.

Genetic vaccination using naked plasmid DNA is an immunization strategy both against infectious diseases and cancer.In order to improve efficacy of DNA vaccines, particularly in large animals and humans, different strategies have been pursued. These vaccination strategies are based on different application routes, schedules and coexpression of immunomodulatory molecules as adjuvants. Our mouse tumor model offers the possibility to investigate Her2/neu DNA vaccines in different settings, that is, intramuscular or intradermal application with or without coexpression of adjuvants. The immunogenicity of predicted peptides for Her2/neu specific memory T cells were screened and confirmed after intramuscular and intradermal application. Protection from tumor growth in tumor challenge experiments and both T cell and humoral immune responses against Her2/neu peptides are used as surrogate parameters for vaccine efficacy.

Comparison of Whiskbroom and Pushbroom darkfield elastic light scattering spectroscopic imaging for head and neck cancer identification in a mouse model.

The early detection of head and neck cancer is a prolonged challenging task. It requires a precise and accurate identification of tissue alterations as well as a distinct discrimination of cancerous from healthy tissue areas. A novel approach for this purpose uses microspectroscopic techniques with special focus on hyperspectral imaging (HSI) methods. Our proof-of-principle study presents the implementation and application of darkfield elastic light scattering spectroscopy (DF ELSS) as a non-destructive, high-resolution, and fast imaging modality to distinguish lingual healthy from altered tissue regions in a mouse model. The main aspect of our study deals with the comparison of two varying HSI detection principles, which are a point-by-point and line scanning imaging, and whether one might be more appropriate in differentiating several tissue types. Statistical models are formed by deploying a principal component analysis (PCA) with the Bayesian discriminant analysis (DA) on the elastic light scattering (ELS) spectra. Overall accuracy, sensitivity, and precision values of 98% are achieved for both models whereas the overall specificity results in 99%. An additional classification of model-unknown ELS spectra is performed. The predictions are verified with histopathological evaluations of identical HE-stained tissue areas to prove the model's capability of tissue distinction. In the context of our proof-of-principle study, we assess the Pushbroom PCA-DA model to be more suitable for tissue type differentiations and thus tissue classification. In addition to the HE-examination in head and neck cancer diagnosis, the usage of HSI-based statistical models might be conceivable in a daily clinical routine.

Sensitivity and Resistance of Oncogenic RAS-Driven Tumors to Dual MEK and ERK Inhibition.

Oncogenic mutations in RAS family genes arise frequently in metastatic human cancers. Here we developed new mouse and cellular models of oncogenic HrasG12V-driven undifferentiated pleomorphic sarcoma metastasis and of KrasG12D-driven pancreatic ductal adenocarcinoma metastasis. Through analyses of these cells and of human oncogenic KRAS-, NRAS- and BRAF-driven cancer cell lines we identified that resistance to single MEK inhibitor and ERK inhibitor treatments arise rapidly but combination therapy completely blocks the emergence of resistance. The prior evolution of resistance to either single agent frequently leads to resistance to dual treatment. Dual MEK inhibitor plus ERK inhibitor therapy shows anti-tumor efficacy in an HrasG12V-driven autochthonous sarcoma model but features of drug resistance in vivo were also evident. Array-based kinome activity profiling revealed an absence of common patterns of signaling rewiring in single or double MEK and ERK inhibitor resistant cells, showing that the development of resistance to downstream signaling inhibition in oncogenic RAS-driven tumors represents a heterogeneous process. Nonetheless, in some single and double MEK and ERK inhibitor resistant cell lines we identified newly acquired drug sensitivities. These may represent additional therapeutic targets in oncogenic RAS-driven tumors and provide general proof-of-principle that therapeutic vulnerabilities of drug resistant cells can be identified.

Induction of Therapeutic Protection in an HPV16-Associated Mouse Tumor Model Through Targeting the Human Papillomavirus-16 E5 Protein to Dendritic Cells.

HPV E5 is an oncoprotein mainly expressed in premalignant lesions, which makes it an important target for a vaccine to prevent or cure cervical cancer (CC). In this study, we evaluated whether E5 targeted to DEC-205, present in dendritic cells (DCs), could induce a therapeutic protection against HPV16-induced tumor cells in a mouse model. The HPV-16 E5 (16E5) protein was cross-linked to a monoclonal antibody (mAb) specific to mouse DEC-205 (anti-DEC-205:16E5) or to an isotype control mAb (isotype:16E5). Rotavirus VP6 was cross-linked to the mouse anti-DEC-205 mAb (anti-DEC-205:VP6) as a non-specific antigen control. BALB/c mice were inoculated subcutaneously (s.c.) with the 16E5-expressing BMK-16/myc tumor cells, and 7 and 14 days later the mice were immunized s.c. with the conjugates, free 16E5 or PBS in the presence of adjuvant. Tumor growth was monitored to evaluate protection. A strong protective immune response against the tumor cells was induced when the mice were inoculated with the anti-DEC-205:16E5 conjugate, since 70% of the mice controlled the tumor growth and survived, whereas the remaining 30% developed tumors and died by day 72. In contrast, 100% of the mice in the control groups died by day 30. The anti-DEC-205:16E5 conjugate was found to induce 16E5-specific memory T cells, with a Th1/Th17 profile. Both CD4+ and CD8+ T cells contributed to the observed protection. Finally, treating mice that had developed tumors with an anti-PD-1 mAb, delayed the tumor growth for more than 20 days. These results show that targeting 16E5 to DEC-205, alone or combined with an immune checkpoint blockade, could be a promising protocol for the treatment of the early stages of HPV-associated cancer.

Lipid bubbles combined with low-intensity ultrasound enhance the intratumoral accumulation and antitumor effect of pegylated liposomal doxorubicin in vivo.

Pegylated liposomal doxorubicin (PLD) is a representative nanomedicine that has improved tumor selectivity and safety profile. However, the therapeutic superiority of PLD over conventional doxorubicin has been reported to be insignificant in clinical medicine. Combination treatment with microbubbles and ultrasound (US) is a promising strategy for enhancing the antitumor effects of chemotherapeutics by improving drug delivery. Recently, several preclinical studies have shown the drug delivery potential of lipid bubbles (LBs), newly developed monolayer microbubbles, in combination with low-intensity US (LIUS). This study aimed to elucidate whether the combined use of LBs and LIUS enhanced the intratumoral accumulation and antitumor effect of PLD in syngeneic mouse tumor models. Contrast-enhanced US imaging using LBs showed a significant decrease in contrast enhancement after LIUS, indicating that LIUS exposure induced the destruction of LBs in the tumor tissue. A quantitative evaluation revealed that the combined use of LBs and LIUS improved the intratumoral accumulation of PLD. Furthermore, tumor growth was inhibited by combined treatment with PLD, LBs, and LIUS. Therefore, the combined use of LBs and LIUS enhanced the antitumor effect of PLD by increasing its accumulation in the tumor tissue. In conclusion, the present study provides important evidence that the combination of LBs and LIUS is an effective method for enhancing the intratumoral delivery and antitumor effect of PLD in vivo.

A study of the possible role of Fab-glycosylated IgG in tumor immunity.

Previously we reported that administration of IgG could inhibit tumor progression in mouse models. At the same time, we also found that some IgGs have glycosylation modifications on their Fab fragments, which may have different biological functions than non-glycosylated IgG. In this study, we employed mouse tumor models to explore the roles of two different forms of IgG, i.e. Fab-glycosylated and Fab-non-glycosylated IgG, in tumor progression. The two types of IgGs were separated with ConA absorption which could react with glycan on the Fab arm but could not access glycan on the Fc fragment. In addition, we performed cytokine array, ELISA, western blotting, immunocytochemistry and other techniques to investigate the possible mechanisms of the actions of Fab-glycosylated IgG in the models. We found that Fab-glycosylated IgG, unlike Fab-non-glycosylated IgG, did not inhibit tumor growth and metastasis in the model. On the contrary, Fab-glycosylated IgG may bind to antigen-bound IgG molecules and macrophages through the glycosidic chain on the Fab fragment to affect antigen-antibody binding and macrophage polarization, which are likely to help tumor cells to evade the immune surveillance. A new mechanism of immune evasion with Fab-glycosylated IgG playing a significant role was proposed.

ST09, A Novel Curcumin Derivative, Blocks Cell Migration by Inhibiting Matrix Metalloproteases in Breast Cancer Cells and Inhibits Tumor Progression in EAC Mouse Tumor Models.

PURPOSE: Curcumin is known for its anticancer and migrastatic activity in various cancers, including breast cancer. Newer curcumin derivatives are being explored to overcome limitations of curcumin like low bioavailability, stability, and side effects due to its higher dose. In this study, the synthesis of ST09, a novel curcumin derivative, and its antiproliferative, cytotoxic, and migrastatic properties have been explored both in vitro and in vivo. METHODS: After ST09 synthesis, anticancer activity was studied by performing standard cytotoxicity assays namely, lactate dehydrogenase (LDH) release assay, 3-(4, 5-dimethylthiazol-2-yl)-2-5-diphenyletrazolium bromide (MTT), and trypan blue exclusion assay. Annexin-FITC, cell cycle analysis using flow cytometry, and Western blotting were performed to elucidate cell death mechanisms. The effect on the inhibition of cell migration was studied by transwell migration assay. An EAC (Ehrlich Ascites carcinoma) induced mouse tumor model was used to study the effect of ST09 on tumor regression. Drug toxicity was measured using aspartate aminotransferase (AST), alanine aminotransferase (ALT), blood urea nitrogen (BUN), and flow-cytometry based lymphocyte count. Histological analysis was performed for assessment of any tissue injury post ST09 treatment. RESULTS: ST09 shows an approximate 100-fold higher potency than curcumin, its parent compound, on breast tumor cell lines MCF-7 and MDA-MB231. ST09 arrests the cell cycle in a cell type-specific manner and induces an intrinsic apoptotic pathway both in vitro and in vivo. ST09 inhibits migration by downregulating matrix metalloprotease 1,2 (MMP1,2) and Vimentin. In vivo, ST09 administration led to decreased tumor volume in a mouse allograft model by boosting immunity with no significant drug toxicity. CONCLUSION: ST09 exhibits antiproliferative and cytotoxic activity at nanomolar concentrations. It induces cell death by activation of the intrinsic pathway of apoptosis both in vitro and in vivo. It also inhibits migration and invasion. This study provides evidence that ST09 can potentially be developed as a novel antitumor drug candidate for highly metastatic and aggressive breast cancer.

A Transposon-Based Mouse Model of Hepatocellular Carcinoma via Hydrodynamic Tail Vein Injection.

Transgenic mouse are reliable, convenient models for studying human hepatocellular carcinoma (HCC). The development of a synthetically engineered Sleeping Beauty (SB) transposon system further enables the viral-free, efficient delivery of desired oncogenes to mouse tissues. Here, we describe an SB transposon-based approach to induce HCC in mice by expressing a hyperactive form of N-RAS, N-RASG12V, while silencing the endogenous Trp53 gene via hydrodynamic tail vein injection, a method to rapidly deliver naked plasmids to mouse liver.

Synthesis of dual functional procaine-derived carbon dots for bioimaging and anticancer therapy.

Aim: Procaine-derived carbon dots, termed P-dots, expectedly offer both fluorescent biomarker function and anticancer activity. Materials & methods: P-dots were synthesized by condensing procaine, citric acid and ethylenediamine via hydrothermal synthesis and characterized by transmission electron microscopy, Fourier transform infrared spectroscopy and x-ray photoelectron spectroscopy. The cellular uptake behavior and the bioimaging performance of P-dots were assessed by confocal laser scanning microscopy. Their antitumor activity was evaluated using the CCK-8 assays and in vivo antitumor experiments, and the underlying mechanism was evaluated by flow cytometry and western blotting. Results: P-dots exhibited excellent luminescence properties suitable for bioimaging and considerable anticancer activity associated with caspase-3-related cell apoptosis. Conclusion: The synthesized procaine-derived carbon dots presented a dual function consisting of bioimaging and anticancer activity, which may enable their implementation as safe and effective clinical nanotherapeutics.

Live-attenuated lymphocytic choriomeningitis virus-based vaccines for active immunotherapy of HPV16-positive cancer.

Infection with human papillomavirus (HPV) is associated with a variety of cancer types and limited therapy options. Therapeutic cancer vaccines targeting the HPV16 oncoproteins E6 and E7 have recently been extensively explored as a promising immunotherapy approach to drive durable antitumor T cell immunity and induce effective tumor control. With the goal to achieve potent and lasting antitumor T cell responses, we generated a novel lymphocytic choriomeningitis virus (LCMV)-based vaccine, TT1-E7E6, targeting HPV16 E6 and E7. This replication-competent vector was stably attenuated using a three-segmented viral genome packaging strategy. Compared to wild-type LCMV, TT1-E7E6 demonstrated significantly reduced viremia and CNS immunopathology. Intravenous vaccination of mice with TT1-E7E6 induced robust expansion of HPV16-specific CD8+ T cells producing IFN-γ, TNF-α and IL-2. In the HPV16 E6 and E7-expressing TC-1 tumor model, mice immunized with TT1-E7E6 showed significantly delayed tumor growth or complete tumor clearance accompanied with prolonged survival. Tumor control by TT1-E7E6 was also achieved in established large-sized tumors in this model. Furthermore, a combination of TT1-E7E6 with anti-PD-1 therapy led to enhanced antitumor efficacy with complete tumor regression in the majority of tumor-bearing mice that were resistant to anti-PD-1 treatment alone. TT1-E7E6 vector itself did not exhibit oncolytic properties in TC-1 cells, while the antitumor effect was associated with the accumulation of HPV16-specific CD8+ T cells with reduced PD-1 expression in the tumor tissues. Together, our results suggest that TT1-E7E6 is a promising therapeutic vaccine for HPV-positive cancers.

Mycobacterium indicus pranii therapy induces tumor regression in MyD88- and TLR2-dependent manner.

OBJECTIVES: Mycobacterium indicus pranii (MIP) is an atypical mycobacterium species with potent antitumor efficacy. Macrophages and dendritic cells (DCs) are antigen-presenting cells, playing key roles in the activation of antitumor immunity. We have previously shown the potent activation of macrophages and DCs by MIP, which is mediated by MyD88-TLR2 signaling axis. In the present study, we further examined the role of MyD88 and TLR2 in MIP-mediated tumor regression. RESULTS: Wild-type and MyD88-/- mice were implanted with B16F10 tumor cells, treated with MIP or phosphate-buffered saline (PBS) and monitored for tumor growth. As expected, MIP therapy led to significant tumor regression in wild-type mice. However, antitumor efficacy of MIP was lost in MyD88-/- animals. Both PBS-treated (control) and MIP-treated MyD88-/- mice developed tumors with comparable volume. Since MyD88 relays TLR engagement signals, we analyzed the antitumor efficacy of MIP in TLR2-/- and TLR4-/- mice. It was observed that MIP therapy reduced tumor burden in wild-type and TLR4-/- mice but not in TLR2-/- mice. Tumor volume in MIP-treated TLR2-/- mice were comparable with those in PBS-treated wild-type animals. These results implicated the MyD88-TLR2 signaling axis in the antitumor efficacy of MIP.

Upregulation of DLC-1 inhibits pancreatic cancer progression: Studies with clinical samples and a pancreatic cancer model.

Deleted in liver cancer 1 (DLC-1) serves a vital role in the progression of multiple cancers, including those of the pancreas. Numerous studies have aimed to reveal the anti-cancer mechanisms of the DLC-1 gene, though few have focused on its impact on the development of pancreatic cancer. Using clinical pancreatic cancer samples and pancreatic cancer cell lines, the present study aimed to reveal the role of DLC-1 in this disease. The expression levels of DLC-1 were determined in pancreatic cancer and adjacent normal tissues from patients with pancreatic cancer, indicating a decreased expression level of DLC-1 in cancerous tissues. Using the pancreatic cancer cell line SW1990, the effect of DLC overexpression on cell proliferation, invasive capacity and the cell cycle and were assessed. Using a mouse tumor model, the tumor-progression capacity of transfected and untransfected SW1990 cells was investigated, indicating that DLC-1 transfection reduced the capacity for tumor progression. Thus, the present study indicated that the overexpression of DLC-1 inhibited the proliferation and reduced the invasive capacity of SW1990 cells both in vitro and in vivo, and that it may have significant inhibitory effects on the development of pancreatic cancer.

CD49b, CD87, and CD95 Are Markers for Activated Cancer-Associated Fibroblasts Whereas CD39 Marks Quiescent Normal Fibroblasts in Murine Tumor Models.

Fibroblasts are thought to be key players in the tumor microenvironment. Means to identify and isolate fibroblasts as well as an understanding of their cancer-specific features are essential to dissect their role in tumor biology. To date, the identification of cancer-associated fibroblasts is widely based on generic markers for activated fibroblasts in combination with their origin in tumor tissue. This study was focused on a deep characterization of the cell surface marker profile of cancer-associated fibroblasts in widely used mouse tumor models and defining aberrant expression profiles by comparing them to their healthy counterparts. We established a generic workflow to isolate healthy and cancer-associated fibroblasts from solid tissues, thereby reducing bias, and background noise introduced by non-target cells. We identified CD87, CD44, CD49b, CD95, and Ly-6C as cancer-associated fibroblast cell surface markers, while CD39 was identified to mark normal fibroblasts from healthy tissues. In addition, we found a functional association of most cancer-related fibroblast markers to proliferation and a systemic upregulation of CD87, and CD49b in tumor-bearing mice, even in non-affected tissues. These novel markers will facilitate the characterization of fibroblasts and shed further light in their functions and implication in cancer progression.