Tumor Induction Research Articles

Enhanced Anti-tumor Effect of Flavopiridol in Combination With Gemcitabine in Pancreatic Cancer.

The efficacy of current chemotherapies for pancreatic ductal adenocarcinoma (PDAC) is still unsatisfactory. Flavopiridol inhibits multiple cyclin-dependent kinases, causing cell cycle arrest and inducing cancer cell apoptosis. This study aimed to evaluate the anti-tumor effect of flavopiridol and gemcitabine in PDAC in vitro and in vivo. PANC-1 and MIA PaCa-2 cell lines were treated with gemcitabine and flavopiridol alone, in combination, and sequentially, and cell proliferation, apoptosis, and the cell cycle were evaluated. Proteins related to cell cycle progression (cyclin A, CDK2, E2F-1, and p53) were quantified using western blotting. A xenograft mouse model was generated, and the effects of gemcitabine and flavopiridol, administered alone or in combination, were evaluated by measuring tumor volume and apoptosis degree using the TUNEL assay. Sequential administration of gemcitabine and flavopiridol suppressed PDAC cell proliferation and induced apoptosis. Flavopiridol treatment led to an increase in the number of cells in the S and a decrease in those in the G0/G1 phases. Gemcitabine increased and decreased the number of S- and G2/M-phase cells, respectively. Furthermore, flavopiridol treatment decreased cyclin A and CDK2 expression and increased E2F-1 expression. In a xenograft mouse model, the combined administration of gemcitabine and flavopiridol demonstrated the most significant reduction in tumor volume and induction of apoptosis. Flavopiridol potentiates the anti-tumor activity of gemcitabine by inducing cell cycle arrest and apoptosis. Its synergistic inhibition of PDAC cell proliferation, when combined with gemcitabine, positions flavopiridol as a promising candidate for cancer treatment.

A4 DNA HYPOMETHYLATION INHIBITS COLITIS-ASSOCIATED CANCER

Abstract Background Colorectal cancer is the second leading cause of cancer death in Canada. A major risk factor for the development of colorectal cancer is chronic inflammation leading to colitis-associated cancer (CAC). We previously described a CAC mouse model in which tumors arise from DCLK1+ cells following loss of the tumor suppressor adenomatous polyposis coli (APC) and induction of colitis. Interestingly, both colitis and CAC are associated with DNA methylation changes that lead to altered gene expression. Moreover, inhibition of DNA methylation has recently been shown to lead to a viral mimicry response. The effects of DNA methylation on colonic tumorigenesis, however, is not known. Aims Thus, we aim to investigate whether inhibition of DNA methylation leads to a viral mimicry response that inhibits colitis-associated tumorigenesis. Methods We first examined the effects of DNA hypomethylation on CAC, by crossing Dclk1-CreERT2/Apcf/f mice to Dnmt1f/f mice which allowed for knock-out of the DNA methyltransferase enzyme DNMT1 in DCLK1+ cells. Dclk1/Apcf/f/Dnmt1f/f and Dclk1/Apcf/f (control group) mice were administered three doses of tamoxifen followed by 2.5% dextran sodium sulfate (DSS) for five days to induce colitis. Fourteen weeks later, we assessed colonic tumor number and size. In a separate cohort of Dclk1/Apcf/f mice treated with DSS, we compared colonic tumor number in mice treated with six doses of 5-AZA-2’-deoxycytidine (5-AZA) versus vehicle. The effects of 5-AZA and DSS on DNA methylation were assessed using the Infinium Mouse Methylation BeadChip Array on colonic epithelial cells isolated from mice treated with vehicle versus 5-AZA in the presence or absence of colitis. RNA expression of viral transposable elements and type I interferon response related genes were measured as a readout of a viral mimicry response. Lastly, we investigated the mechanism by which DNA hypomethylation affects tumorigenesis by crossing Dclk1/Apcf/f/Dnmt1f/f mice to MAVS knockout mice and examining tumor number following tamoxifen and DSS administration as above. Results Deletion of DNMT1 in DCLK1+ cells or treatment with 5-AZA significantly decreased colonic tumor number and size. Treatment with 5-AZA or DNMT1 loss led to DNA hypomethylation and subsequent upregulation of endogenous retroviral expression. Both 5-AZA treatment and DNMT1 knockout in DCLK1+ cells led to increased expression of type I interferon response genes. Knockout of the type I interferon response related gene, MAVS, reversed the anti-tumor effect observed with DNMT1 loss. Conclusions Our findings demonstrate that DNA hypomethylation by 5-AZA or loss of DNMT1 reduces colitis-associated tumorigenesis through activation of a viral mimicry response. Funding Agencies CAG, CIHRCancer Research Society, Canada Foundation for Innovation, Lawson Health Research Institute

The HN protein of Newcastle disease virus induces cell apoptosis through the induction of lysosomal membrane permeabilization.

Lysosomes are acidic organelles that mediate the degradation and recycling of cellular waste materials. Damage to lysosomes can cause lysosomal membrane permeabilization (LMP) and trigger different types of cell death, including apoptosis. Newcastle disease virus (NDV) can naturally infect most birds. Additionally, it serves as a promising oncolytic virus known for its effective infection of tumor cells and induction of intensive apoptotic responses. However, the involvement of lysosomes in NDV-induced apoptosis remains poorly understood. Here, we demonstrate that NDV infection profoundly triggers LMP, leading to the translocation of cathepsin B and D and subsequent mitochondria-dependent apoptosis in various tumor and avian cells. Notably, the released cathepsin B and D exacerbate NDV-induced LMP by inducing the generation of reactive oxygen species. Additionally, we uncover that the viral Hemagglutinin neuraminidase (HN) protein induces the deglycosylation and degradation of lysosome-associated membrane protein 1 (LAMP1) and LAMP2 dependent on its sialidase activity, which finally contributes to NDV-induced LMP and cellular apoptosis. Overall, our findings elucidate the role of LMP in NDV-induced cell apoptosis and provide novel insights into the function of HN during NDV-induced LMP, which provide innovative approaches for the development of NDV-based oncolytic agents.

Sex differences in colonic gene expression and fecal microbiota composition in a mouse model of obesity-associated colorectal cancer

This study investigated the sex-specific correlation between obesity and colorectal cancer emphasizing a more pronounced association in males. Estrogen, chromosomal genes, and gut bacteria were assessed in C57BL6/J male, female and ovariectomized (OVX) female mice, subjected to either a low-fat diet (LFD) or high-fat diet (HFD) for 14 weeks. Induction of colon tumor involved azoxymethane (10 mg/kg) administration, followed by three cycles of dextran sulfate sodium. Male mice on HFD exhibited higher final body weight and increased colon tumors compared to females. Colonic mucin 2 expression was significantly higher in females. HFD-modulated differentially expressed genes numbered 290 for males, 64 for females, and 137 for OVX females. Only one up-regulated gene (Gfra3) overlapped between females and OVX females, while two down-regulated genes (Thrsp and Gbp11) overlapped between males and OVX females. Genes up-regulated by HFD in males were linked to cytokine-cytokine interaction, HIF-1 signaling pathway, central carbon metabolism in cancer. Sex-specific changes in gut microbial composition in response to HFD were observed. These findings suggest a male-specific vulnerability to HFD-induced colon tumor formation, implicating key genes and colonic bacteria in colon tumorigenesis.

PBRM1 loss is associated with increased sensitivity to MCL1 and CDK9 inhibition in clear cell renal cancer.

MCL1 is a member of the BCL2 family of apoptosis regulators, which play a critical role in promoting cancer survival and drug resistance. We previously described PRT1419, a potent, MCL1 inhibitor with anti-tumor efficacy in various solid and hematologic malignancies. To identify novel biomarkers that predict sensitivity to MCL1 inhibition, we conducted a gene essentiality analysis using gene dependency data generated from CRISPR/Cas9 cell viability screens. We observed that clear cell renal cancer (ccRCC) cell lines with damaging PBRM1 mutations displayed a strong dependency on MCL1. PBRM1 (BAF180), is a chromatin-targeting subunit of mammalian pBAF complexes. PBRM1 is frequently altered in various cancers particularly ccRCC with ~40% of tumors harboring damaging PBRM1 alterations. We observed potent inhibition of tumor growth and induction of apoptosis by PRT1419 in various preclinical models of PBRM1-mutant ccRCC but not PBRM1-WT. Depletion of PBRM1 in PBRM1-WT ccRCC cell lines induced sensitivity to PRT1419. Mechanistically, PBRM1 depletion coincided with increased expression of pro-apoptotic factors, priming cells for caspase-mediated apoptosis following MCL1 inhibition. Increased MCL1 activity has been described as a resistance mechanism to Sunitinib and Everolimus, two approved agents for ccRCC. PRT1419 synergized with both agents to potently inhibit tumor growth in PBRM1-loss ccRCC. PRT2527, a potent CDK9 inhibitor which depletes MCL1, was similarly efficacious in monotherapy and in combination with Sunitinib in PBRM1-loss cells. Taken together, these findings suggest PBRM1 loss is associated with MCL1i sensitivity in ccRCC and provide rationale for the evaluation of PRT1419 and PRT2527 for the treatment for PBRM1-deficient ccRCC.

Multiple Bowen's disease due to long-term narrow-band ultraviolet B phototherapy: A case report and literature review.

By presenting a case study on multiple instances of Bowen's disease and the consistent use of narrow-band ultraviolet B (NB-UVB) phototherapy over a three-year period, our aim is to enhance the comprehension of domestic clinicians regarding the disease. Additionally, we seek to review existing literature, encouraging dermatologists to consider clinical secondary primary lesion diagnoses. Our approach involves analyzing a diagnosed case of multiple Bowen's disease, examining clinical manifestations, histopathology, imaging results, and treatment methods related to NB-UVB phototherapy. We aim to facilitate discussion and understanding through a comprehensive literature analysis. An elderly male with a 30-year history of psoriasis vulgaris initiated continuous NB-UVB therapy three years ago. A year later, he developed red patches and plaques with distinct borders and scaly surfaces on his face, trunk, lower extremities, and scrotum. Histopathological examination confirmed Bowen's disease. Treatment involved liquid nitrogen cryotherapy, with no recurrence observed during the one-year follow-up. This case highlights that Bowen's disease, typically solitary, can manifest as multiple instances, especially in individuals with a history of psoriasis vulgaris. While NB-UVB stands as the primary treatment for psoriasis vulgaris, caution is warranted due to the potential risk of skin tumor induction with prolonged high-dose usage. Clinicians should be vigilant in monitoring and assessing the long-term implications of such therapies.

Biodistribution and pharmacokinetics of [89Zr]-anti-VEGF mAbs using PET in glioblastoma rat models

IntroductionGlioblastomas present intensive angiogenesis, thus anti–Vascular Endothelial Growth Factor (VEGF) antibodies (mAbs) have been proposed as promising therapies. However, the results of clinical trials reported moderate toxicity and limited effectiveness. This study evaluates the in vivo pharmacokinetics and biodistribution of these mAbs in a growing model of glioblastoma in rats using Positron Emission Tomography (PET). Material&Methods:mAbs were radiolabeled with zirconium-89. Four days after the model induction, animals were injected with 2.33 ± 1.3 MBq of [89Zr]-DFO-bevacizumab (n = 8) or 2.35 ± 0.26 MBq of [89Zr]-DFO-aflibercept (n = 6). PETs were performed at 0H, 48H, 168H, 240H, and 336H post-injection. Tumor induction was confirmed using [18F]-Fluorodeoxyglucose-PET and immunohistochemistry. Radiotracer uptake was estimated in all pre-defined Volumes-of-Interest. ResultsAnti-VEGF mAbs showed 100 % Radiochemical-Purity. [89Zr]-DFO-bevacizumab showed a significantly higher bioavailability in whole-blood. A significant increase in the tumor uptake was detectable at 168H PET with [89Zr]-DFO-bevacizumab meanwhile with [89Zr]-DFO-aflibercept it was only detectable at 336H. [89Zr]-DFO-bevacizumab tumor uptake was significantly higher than that of [89Zr]-DFO-aflibercept in all the scans. Tumor induction was confirmed in all animal models. ConclusionMAbs detect VEGF-expression in glioblastoma models. Tumors were earlier targeted by Bevacizumab. The lower blood availability of aflibercept resulted in a lower tumor uptake than bevacizumab in all the scans.

Effects of non-contact electric fields on kidney and liver histology in tumour-induced rats

Background A novel modality of cancer treatment based on exposure to non-contact electric fields has been developed to reduce cancer incidence. However, the safety of the electric field exposure was not fully investigated. Therefore, This study aimed to observe the safety of electric field exposure on kidney and liver structures. Methods Female Sprague-Dawley rats were divided into one control group and three treatment groups. Animals were treated with 7,12-dimethylbenz[a]anthracene for mammary tumour induction and exposed to non-contact electric fields individually for 10 hours a day for three weeks. Fresh samples of the kidney and liver were collected for observation of structural damage in both organs. Both organs were prepared for histopathological cross-sectioning using the paraffin method and Hematoxylin and Eosin staining followed by histological scoring using the post-examination masking method. Results Damages found in the kidney were as follows: thickening of the Bowman capsule, karyolysis, karyorrhexis, pyknosis, cloudy swelling, epithelial sloughing, inflammation, haemorrhage, and congestion. In addition, the number of inflammation and haemorrhage in the kidney structure of healthy rats that were exposed to electric fields was significantly lower than that in the control group. All damages to the kidneys were also found in the liver, but each showed a different degree of damage. Exposure to this electric field can cause haemorrhagic damage to the livers of healthy rats, but not to rats with breast cancer. Conclusions Exposure to non-contact electric fields can cause haemorrhagic damage to the livers of healthy rats, however, in other liver tissues and the kidneys, exposure to this electric field was safe. It can even decrease the number of inflammations and haemorrhages in the kidneys.

Self-assembled fructo-oligosaccharide conjugated ferulic acid microparticle: Anticancer, anti-inflammatory and immunomodulatory effects

Fructo-oligosaccharide ferulic acid conjugate was synthesized, which self-assembles into disc shaped amorphous microparticles (FA FOS I), that could be used for targeted delivery to the colon. It is accessed for its ability to ameliorate colorectal cancer and inflammation. The microparticle exhibited selective cytotoxicity and apoptosis induction in colon cancer cells. The microparticle arrested the cell cycle in the G0-G1 phase. In-vivo anti-colorectal cancer activity were evaluated in colitis associated colon cancer mice model. There was a reduction in 60.83% of tumour lesions and 48.27% reduction in the malignant cells. The immunohistochemistry (IHC) revealed the suppression of inflammation and related inflammatory cytokines in the colon. The inhibition of cell proliferation, up-regulation of tumour suppressor protein and induction of apoptosis in the malignant cells were observed. The evaluation of immune modulation by FA FOS I microparticle revealed a 9% increase in WBC count and an increase in the infiltration of T-helper cells and cytotoxic T-cells into the tumour microenvironment. There was a considerable decrease in the expression of immune suppressing PD-L1 by the tumour cells on treatment with FA FOS I microparticle. All these data implicate better efficacy of the FA FOS I microparticle delivery to colon and amelioration of colitis associated colorectal cancer through multimodal route of inflammation reduction, apoptosis induction and positive modulation of immune system in the tumour microenvironment. FA FOS I was proposed to be a candidate drug which is physiologically and metabolically stable under gastro-intestinal condition and potentiating its effect in the target site large intestine; without being absorbed or transported to other tissue nor causing any adverse cytotoxicity to the neighbouring normal tissue.

Orthotopic Model of Hepatocellular Carcinoma in Mice.

Orthotopic models of hepatocellular carcinoma (HCC) consist in the implantation of tumor cells into the liver by direct intrahepatic injection. In this model, tumorigenesis is triggered within the hepatic microenvironment, thus mimicking the metastatic behavior of HCC. Herein, we detail a surgically mediated methodology that allows the reproducible and effective induction of liver-sessile tumors in mice. We enumerate the steps to be followed before and after the surgical procedure, including HCC cell preparation, the quantity of cancer cells to be injected, presurgical preparation of the mice, and finally, postoperative care. The surgical procedure involves laparotomy to expose the liver, injection of cells into the left-lateral hepatic lobe, and closure of the incision with sutures followed by wound clips. We also provide information concerning the subsequent tumor growth follow-up, as well as the application of bioluminescence imaging to monitor tumor development.

Establishing and Characterizing Chemotherapy-Induced Immunological Dormant Tumor Cell Lines.

Chemotherapy, together with radiotherapy, targeted therapies, and immunotherapy, is the main option to treat cancer patients in neoadjuvant/adjuvant setting to reduce the risk of disease progression and metastasis formation from disseminated tumor cells. Cancer cells that survived chemotherapy treatment may emerge with novel characteristics, one of which is the ability to stimulate the native and adaptive immune systems. Models allowing the characterization of chemotherapy-induced tumor cell plasticity and induction of immune response or adaptation are needed to identify novel mechanisms and devise novel therapeutic strategies to prevent relapses. Here we describe a protocol for selecting chemotherapy-resistant cancer cells and testing the in vivo effects on the local and systemic immune responses. While originally developed to characterize the effects of methotrexate and doxorubicin on murine 4T1 breast cancer cells and the relative immune response, the method can be broadened to other chemotherapies and syngeneic cancer models.

MRI monitoring of combined therapy with transcatheter arterial delivery of NK cells and systemic administration of sorafenib for the treatment of HCC.

This preclinical study explored the synergistic potential of sorafenib and NK cell chemoimmunotherapy to combat hepatocellular carcinoma (HCC) in a rat model. We aimed to enhance NK cell cytotoxicity through IL-12/18 cytokines supplementation and elucidate the underlying molecular mechanisms driving this collaborative antitumor action. Twenty-four Sprague-Dawley rats were divided into distinct treatment groups, receiving sorafenib via gavage and NK cells via catheterization of the proper hepatic artery. Tumor growth and treatment response were monitored through weekly MRI scans, including T1w, T2w, DCE, and DWI sequences. Histological examinations assessed tumor cell viability, apoptosis fraction, and microvessel density. The combined therapy demonstrated significant inhibition of tumor growth, angiogenesis, and induction of durable antitumor immunity compared to either modality alone. DCE-MRI and DWI revealed distinct alterations in tumor microvasculature, highlighting the effectiveness of the combination. Our findings highlight the promise of sorafenib-augmented NK cell chemoimmunotherapy as a potential therapeutic strategy for HCC management. The targeted delivery of IL-12/18 cytokines supplemented NK cells effectively enhanced cytotoxicity within the tumor microenvironment, leading to improved antitumor responses. Further investigation in clinical trials is warranted to validate these findings in human patients and explore the translational potential of this approach.

EBP50 Depletion and Nuclear β-Catenin Accumulation Engender Aggressive Behavior of Colorectal Carcinoma through Induction of Tumor Budding.

Ezin-radixin-moesin-binding phosphoprotein 50 (EBP50) is a scaffold protein that interacts with several partner molecules including β-catenin. Here, we examined the crosstalk between EBP50 and nuclear catenin during colorectal carcinoma (CRC) progression. In clinical samples, there were no correlations between the subcellular location of EBP50 and any clinicopathological factors. However, EBP50 expression was significantly lower specifically in the outer areas of tumor lesions, in regions where tumor budding (BD) was observed. Low EBP50 expression was also significantly associated with several unfavorable prognostic factors, suggesting that EBP50 depletion rather than its overexpression or subcellular distribution plays an important role in CRC progression. In CRC cell lines, knockout of EBP50 induced epithelial-mesenchymal transition (EMT)-like features, decreased proliferation, accelerated migration capability, and stabilized nuclear β-catenin due to disruption of the interaction between EBP50 and β-catenin at the plasma membrane. In addition, Slug expression was significantly higher in outer lesions, particularly in BD areas, and was positively correlated with nuclear β-catenin status, consistent with β-catenin-driven transactivation of the Slug promoter. Together, our data suggest that EBP50 depletion releases β-catenin from the plasma membrane in outer tumor lesions, allowing β-catenin to accumulate and translocate to the nucleus, where it transactivates the Slug gene to promote EMT. This in turn triggers tumor budding and contributes to the progression of CRC to a more aggressive phase.

HPV16 E6/E7-mediated regulation of PiwiL1 expression induces tumorigenesis in cervical cancer cells.

PiwiL1 has been reported to be over-expressed in many cancers. However, the molecular mechanism by which these proteins contribute to tumorigenesis and their regulation in cancer cells is still unclear. We intend to understand the role of PiwiL1 in tumorigenesis and also its regulation in cervical cells. We studied the effect of loss of PiwiL1 function on tumor properties of cervical cancer cells in vitro and in vivo. Also we have looked into the effect of PiwiL1 overexpression in the malignant transformation of normal cells both in vitro and in vivo. Further RNA-seq and RIP-seq analyses were done to get insight of the direct and indirect targets of PiwiL1 in the cervical cancer cells. Here, we report that PiwiL1 is not only over-expressed, but also play a major role in tumor induction and progression. Abolition of PiwiL1 in CaSki cells led to a decrease in the tumor-associated properties, whereas, its upregulation conferred malignant transformation of normal HaCaT cells. Our study delineates a new link between HPV oncogenes, E6 and E7 with PiwiL1. p53 and E2F1 directly bind and differentially regulate PiwiL1 promoter in a context-dependant manner. Further, RNA-seq together with RIP-RNA-seq suggested a strong and direct role for PiwiL1 in promoting metastasis in cervical cancer cells. Our study demonstrates that PiwiL1 act as an oncogene in cervical cancer by inducing tumor-associated properties and EMT pathway. The finding that HPV oncogenes, E6/E7 can positively regulate PiwiL1 suggests a possible mechanism behind HPV-mediated tumorigenesis in cervical cancer.

Anti-tumor Effect of High Doses of Carbon Ions and X-Rays during Irradiation of Ehrlich Ascites Carcinoma Cells Ex Vivo.

The effect of carbon ions (12C) with the energy of 400 MeV/nucleon on the dynamics of induction and growth rate of solid tumors in mice under irradiation of Ehrlich ascites carcinoma cells (EAC) ex vivo at doses of 5-30 Gy relative to the action of equally effective doses of X-ray radiation was studied. The dynamics of tumor induction under the action of 12C and X-rays had a similar character and depended on the dose during 3 months of observation. The value of the latent period, both when irradiating cells with 12C and X-ray, increased with increasing dose, and the interval for tumor induction decreased. The rate of tumor growth after ex vivo irradiation of EAC cells was independent of either dose or type of radiation. The dose at which EAC tumors are not induced within 90 days was 30 Gy for carbon ions and 60 Gy for X-rays. The value of the relative biological effectiveness of carbon ions, calculated from an equally effective dose of 50% probability of tumors, was 2.59.

Efficacy of High Doses of Protons on the Induction of Solid Tumors in Mice by <i>Ex vivo</i> Irradiation of Ehrlich Ascitic Carcinoma Cells

We study the induction patterns of the solid form of Ehrlich ascitic carcinoma (EAC) in mice in the short(1 month) and long-term (18 months) periods after inoculation of ascitic cells irradiated ex vivo with a proton beam at doses of 30, 60, 90, 120, and 150 Gr. The growth rate of solid tumors after inoculation of irradiated cells ex vivo coincided with the growth of tumors in the control group. The frequency of tumor induction in mice transplanted with EAC cells was dose-dependent and decreased from 80% to 0% along with an increase in radiation dose. Thus, using the ex vivo model of tumor cell irradiation followed by in vivo grafting allowed us to determine the parameters of the antitumor effect of new proton therapy regimens.

EXTH-43. TARGETING THE DNA DAMAGE RESPONSE THROUGH COMBINATION MDM2 AND AKT INHIBITOR THERAPY IMPROVES TEMOZOLOMIDE EFFECTIVENESS IN CHEMO-RESISTANT GLIOBLASTOMA

Abstract Temozolomide remains the lone pharmacotherapeutic option for glioblastoma (GBM), yet the development of resistance to temozolomide has been a major challenge contributing to the persistent median < 2-year survival for patients after diagnosis. Tumor heterogeneity and induction of treatment response networks, such as the DNA damage response (DDR), are major contributors to temozolomide resistance in GBM. Targeting DDR treatment response networks, such as the MDM2/p53/p73 and PI3K/AKT/mTOR networks, with small-molecule inhibitors (SMIs) presents an opportunity to disrupt resistance mechanisms and enhance temozolomide efficacy. We utilized a triple drug combination of clinically relevant concentrations of the blood-brain-barrier penetrant SMIs of AKT (ipatasertib; GDC-0068) and MDM2 (idasanutlin; RG7388) with temozolomide to evaluate this targeted strategy using the recurrent, temozolomide-resistant, p53wt GBM10 xenoline. Proliferation studies demonstrated dose-related additive to synergistic inhibition of proliferation at clinically relevant concentrations of ipatasertib and idasanutlin. Further, IncuCyte live-cell imaging demonstrated dose-and time-related growth inhibition of these GBM cells and apoptosis marked by increased cleaved caspase 3 expression following the temozolomide+idasanutlin+ipatasertib triple combination treatment. Cells treated with temozolomide+idasanutlin+ipatasertib also displayed senescence phenotypes, with increased cell cycle arrest and elevated expression of SPiDER β-Gal expression and cell-cycle inhibitors such as p53 and p21. Experiments are in progress to determine the extent to which the effects of temozolomide+idasanutlin+ipatasertib combination therapy are dependent on p53 using siRNA knockdown of p53. In the present study, targeting the temzolomide-induced DNA damage response with idasanutlin+ipatasertib increased the effectiveness of temozolomide. These results indicate that this triple combination may be a promising approach to improving patient outcomes in temozolomide-resistant GBM.

EXTH-52. LONG-ACTING RECOMBINANT INTERLEUKIN-7, NT-I7, IMPROVES SURVIVAL FOLLOWING ONCOLYTIC ZIKA VIRUS TREATMENT IN THE SB28 IMMUNOSUPPRESSIVE AND TREATMENT-RESISTANT MURINE GLIOMA MODEL

Abstract Glioblastoma (GBM) does not respond to immunotherapy for several reasons, including limited cancer-specific neoantigens and poor T-cell infiltration. The SB28 murine glioma model recapitulates these aspects of human GBM. Zika virus (ZIKV) treatment of GL261 and CT2A-bearing mice resulted in significant survival improvement through multiple mechanisms, including direct tumor killing and induction of a local CD8+ T-cell anti-tumor response, but ZIKV is less effective against SB28 tumors. Previously, we published that a long-acting human IL-7, rhIL-7-hyFc (NT-I7, efineptakin alfa; NeoImmuneTech Inc.), increases T-cell abundance and improves tumor infiltration. Therefore, we hypothesize that combining NT-I7 with oncolytic ZIKV therapy will enhance the anti-tumor response and improve survival in mice bearing SB28 syngeneic tumors. 1×103 SB28 tumor cells were intracranially implanted in C57BL/6J mice. Mice were treated with NT-I7 on post tumor implantation day 7 and 10, followed by intratumoral ZIKV on day 14. Tumor burden was assessed using bioluminescence imaging; mice were monitored for survival and bled weekly to assess the systemic T-cell response to NT-I7. Subcutaneous injection of NT-I7 significantly boosted systemic T-cell abundance compared to control (5,623 cells/uL vs. 136 cells/uL, p < 0.05), particularly cytotoxic CD8+ T-cells (4,776 cells/uL vs. 47 cells/uL, p < 0.05). This increased abundance of CD8+ T-cells peaked at 7 days after initial treatment and persisted for 21 days. NT-I7 monotherapy did not confer any survival benefit. However, the combination of NT-I7 and ZIKV treatment resulted in significant improvement in survival compared to NT-I7 or ZIKV treatment alone (p < 0.05). Treatment with NT-I7 primes the immune system by expanding the circulating CD8+ T-cell population by 100-fold. Timing the oncolytic ZIKV injection with the peak in peripheral CD8+ T-cells greatly improved survival in the immunotherapy resistant SB28 glioma model. These data suggest a role for NT-I7 as an adjunct to various T-cell mediated treatment strategies for GBM.

Heme Oxygenase-1 and Its Role in Colorectal Cancer.

Heme oxygenase-1 (HO-1) is an enzyme located at the endoplasmic reticulum, which is responsible for the degradation of cellular heme into ferrous iron, carbon monoxide and biliverdin-IXa. In addition to this main function, the enzyme is involved in many other homeostatic, toxic and cancer-related mechanisms. In this review, we first summarize the importance of HO-1 in physiology and pathophysiology with a focus on the digestive system. We then detail its structure and function, followed by a section on the regulatory mechanisms that control HO-1 expression and activity. Moreover, HO-2 as important further HO isoform is discussed, highlighting the similarities and differences with regard to HO-1. Subsequently, we describe the direct and indirect cytoprotective functions of HO-1 and its breakdown products carbon monoxide and biliverdin-IXa, but also highlight possible pro-inflammatory effects. Finally, we address the role of HO-1 in cancer with a particular focus on colorectal cancer. Here, relevant pathways and mechanisms are presented, through which HO-1 impacts tumor induction and tumor progression. These include oxidative stress and DNA damage, ferroptosis, cell cycle progression and apoptosis as well as migration, proliferation, and epithelial-mesenchymal transition.

Antitumor activity of pegylated human interferon β as monotherapy or in combination with immune checkpoint inhibitors via tumor growth inhibition and dendritic cell activation

Type I interferons (IFN), especially human IFN alpha (IFNα), have been utilized for antitumor therapy for decades. Human interferon beta (IFNβ) is rarely used for cancer treatment, despite advantages over IFNα in biological activities such as tumor growth inhibition and dendritic cell (DC) activation. The utilization of pegylated human IFNβ (PEG-IFNβ), as monotherapy or in combination with immune checkpoint inhibitors (ICIs) was evaluated in this study through in vivo efficacy studies in syngeneic mouse melanoma, non-small cell lung cancer (NSCLC), and colon adenocarcinoma (COAD) models resistant to immune checkpoint inhibitors (ICIs). In vitro comparative study of PEG-IFNβ and pegylated IFNα-2b was performed in terms of tumor growth inhibition against human melanoma, NSCLC and COAD cell lines and activation of human monocyte-derived DCs (MoDCs). Our data demonstrate that the in vivo antitumor effects of PEG-IFNβ are partially attributable to tumor growth-inhibitory effects and DC-activating activities, superior to pegylated IFNα-2b. Our findings suggest that utilizing PEG-IFNβ as an antitumor therapy can enhance the therapeutic effect of ICIs in ICI-resistant tumors by directly inhibiting tumor growth and induction of DC maturation.