Reduction Of Metastatic Lesions Research Articles

Abstract B004: ATR-CHK1-WEE1 pathway is a critical dependency in the context of DNA damage and replicative stress in osteosarcoma

Abstract Osteosarcoma (OS) is characterized by widespread somatic copy number alterations (SCNAs) and structural variations (SVs) with few recurrent point mutations. We previously demonstrated proof-of-principle for a genome-informed strategy for treatment of OS based on rank-ordering SCNAs within specific oncogenes identified by WGS and RNAseq. Although we identified several potentially effective therapeutic strategies using this approach, it is likely that effective therapy for OS will require use of combination therapies. To identify new combination therapy approaches for OS, we used a panel of PDX-derived cell lines (PDXC) and performed a combination drug screen. We assessed 5 drug backbones against 15 targeted agents in 8 PDXC and 2 established OS cell lines. When we combined Gemcitabine with agents that target the ATR-CHK1-WEE1 pathway, we observed strong synergy across all PDXC tested. These results were validated in a secondary screen using a combination matrix across 10 PDXC. Use of targeted agents inhibiting ATR, CHK1 or WEE1 in combination with Gemcitabine led to decreased proliferation and a marked increase in apoptosis in vitro. In subcutaneous tumor models, we observed that decreased tumor growth with either ATRi or Gemcitabine alone, whereas tumors shrank when treated with the combination. However, when we treated OS774 in vivo, there was no effect on tumor growth for either single agent alone or when in combination. This effect was dependent on the presence of ATR as a PDXC with no ATR detectable by western blotting showed not effect in vivo. In an orthotopic model in which PDXC are implanted along the tibia, this combination therapy effectively decreased tumor growth. In a lung metastasis model, ATRi and Gemcitabine resulted a durable reduction in metastatic lesions over time. In summary, we have identified a susceptibility to the ATR-CHK1-WEE1 pathway when combined with gemcitabine. These studies suggest that further investigation ATR-CHK1-WEE1 and gemcitabine are warranted to address the unmet need for new therapeutic approaches for relapsed and recurrent OS patients. Citation Format: Leanne C. Sayles, Henry Martell, Amanda Koehne, Kean-Hooi Ang, Chris Wilson, Michelle Arkin, E. Alejandro Sweet-Cordero. ATR-CHK1-WEE1 pathway is a critical dependency in the context of DNA damage and replicative stress in osteosarcoma [abstract]. In: Proceedings of the AACR Special Conference: Sarcomas; 2022 May 9-12; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(18_Suppl):Abstract nr B004.

Generation of cancer vaccine immunogens derived from major histocompatibility complex (MHC) class I molecules using variable epitope libraries

Although various immune checkpoint inhibitors (ICIs), used for the treatment of advanced cancer, showed remarkably durable tumor regression in a subset of patients, there are important limitations in a large group of non-responders, and the generation of novel immunogens capable of inducing protective cellular immune responses is a priority in cancer immunotherapy field. During the last decades, several types of vaccine immunogens have been used in numerous preclinical studies and clinical trials. However, although immunity to tumor Ags can be elicited by most vaccines tested, their clinical efficacy remains modest. Recently, we have developed an innovative vaccine concept, called Variable Epitope Libraries (VELs), with the purpose to exploit the high antigenic variability of many important pathogens and tumor cells as starting points for the construction of a new class of vaccine immunogens capable of inducing the largest possible repertoire of both B and T cells. In the present study, we decided to generate VEL immunogens derived from both classical and non-classical major histocompatibility complex (MHC) class I molecules. The MHC molecules, responsible for antigen presentation and subsequent activation of T lymphocytes, undergo multiple modifications that directly affect their proper function, resulting in immune escape of tumor cells. Two large VELs derived from multi-epitope region of H2-Kd and Qa-2 sequences (46 and 34 amino acids long, respectively), along with their wild type counterparts have been generated as synthetic peptides and tested in an aggressive 4T1 mouse model of breast cancer. Significant inhibition of tumor growth and the reduction of metastatic lesions in the lungs of immunized mice were observed. This study demonstrated for the first time the successful application of VELs carrying combinatorial libraries of epitope variants derived from MHC class I molecules as novel vaccine immunogens.

In vitro and in vivo anti-tumor effects of selected platinum(IV) and dinuclear platinum(II) complexes against lung cancer cells.

In the present study, cytotoxic effects of cisplatin, the most usually used chemotherapeutic agent, were compared with new designed platinum(IV) ([PtCl4(en)] (en=ethylenediamine) and [PtCl4(dach)]) (dach=(±)-trans-1,2-diaminocyclohexane) and platinum(II) complexes ([{trans-Pt(NH3)2Cl}2(μ-pyrazine)](ClO4)2 (Pt1), [{trans-Pt(NH3)2Cl}2(μ-4,4'-bipyridyl)](ClO4)2DMF(Pt2),[{trans-Pt(NH3)2Cl}2(μ-1,2-bis(4pyridyl)ethane)](ClO4)2 (Pt3)), in vitro and in vivo against human and murine lung cancer cells, to determine anti-tumor potential of newly synthesized platinum-based drugs in the therapy of lung cancer. Results obtained by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide], Lactate dehydrogenase and Annexin V/Propidium Iodide assays showed that, among all tested complexes, [PtCl4(en)] had the highest cytotoxicity against human and murine lung carcinoma cells in vitro. [PtCl4(en)] showed significantly higher cytotoxicity then cisplatin in all tested concentrations, mainly by inducing apoptosis in lung cancer cells. [PtCl4(en)] was well tolerated in vivo. Clinical signs of [PtCl4(en)]-induced toxicity, such as changes in food, water consumption or body weight, nephrotoxicity or hepatotoxicity was not observed in [PtCl4(en)]-treated mice. [PtCl4(en)] managed to increase presence of CD45+leukocytes, including F4/80+macrophages, CD11c+dendritic cells, CD4+helper and CD8+cytotoxic T cells (CTLs) in the lungs, cytotoxic NK, NKT and CTLs in the spleens of tumor bearing mice, resulting with reduction of metastatic lesions in the lungs, indicating its potential to stimulate anti-tumor immune response in vivo. Due to its anti-tumor cytotoxicity, biocompatibility, and potential for stimulation of anti-tumor immune response, [PtCl4(en)] may be a good candidate for further testing in the field of medicinal chemistry.

Promotion of hepatic metastases by liver resection in the rat.

In the early period following radical hepatectomy for hepatoma, recurrences in the remaining liver are frequently found. In regenerating liver, implantation and growth of tumour cells released into the portal system during surgical treatment might be promoted. We examined the relationship between liver regeneration and the formation of metastases following hepatic resection. Intraportal injections of rat ascites containing hepatoma AH130 cells at a concentration of 1 x 10(5) cells 0.2 ml-1 were made at various periods following two thirds liver resection in rats. Tumour cell injections immediately at 24 h after surgery resulted in an increased number of hepatic metastases compared with control animals. Tumour cell injections 2 weeks after hepatectomy, however, had no significant difference in effect compared with control rats. In contrast, tumour cells injected immediately after removal of half of the caudate lobe resulted in the same number of metastases as control animals. These results demonstrate that the number of artificially induced hepatic metastases was increased during an initial period of active liver regeneration and was proportional to the volume of hepatectomy. The effect of 5-fluorouracil (5FU) or mitomycin C (MMC) as inhibitors of hepatic regeneration on liver metastasis after hepatectomy was studied. The administration of 5FU (20 mg kg-1) or MMC (0.2 mg kg-1) immediately, 24 and 48 h after hepatectomy resulted in a marked reduction in metastatic lesions. The administration of 5FU caused delays in weight gain and decreases in the wet weight of remaining liver, while MMC had no effect on either. Accordingly, results of 5FU administration may be due to inhibitory effects on liver regeneration whilst that of MMC administration may be due to cytocidal antitumour effect. The effect of OK-432 as an immunoactivator on the implantation and growth of tumour cells in regenerating liver was also studied. Pretreatment with OK-432, 0.5 mg intraperitoneally on 7 consecutive days, had no effect on hepatic metastases. The pathophysiology of liver regeneration may enhance hematogenous hepatic metastasis and release of tumour cells during surgical manipulation may represent an important cause of recurrence following hepatic resection.

Interference With Tumor Cell-Induced Degradation of Endothelial Matrix on the Antimetastatic Action of Nafazatrom2

The antithrombotic compound nafazatrom was evaluated in several in vivo and in vitro assays to elucidate the mechanism of its antimetastatic activity. C57BL/6 mice bearing B16 amelanotic subcutaneous tumors treated with 100 mg nafazatrom/kg/day exhibited a sixfold reduction in metastatic pulmonary lesions compared to lesion numbers in controls. The reduction in metastatic lesions was not accompanied by changes in primary tumor growth, and up to 1 microgram nafazatrom/ml did not inhibit tumor cell proliferation in vitro. Treatment of C57BL/6 mice with nafazatrom prior to iv inoculation of tumor cells failed to inhibit lung colony formation. In vitro exposure of exponentially growing B16 amelanotic cells to nafazatrom (1 microgram/ml for 72 hr) in culture did not change their ability to adhere to endothelial cell monolayers. B16 amelanotic cells degraded the matrix material of bovine endothelial cell monolayers; a heparin sulfate proteoglycan appeared to be the predominant matrix component released by these tumor cells, as judged by resistance to chondroitin ABC lyase and sensitivity to heparitinase and pronase degradation. Nafazatrom (1 microgram/ml for 72 hr) inhibited the solubilization of matrix components by approximately 60%. Tumor cell degradation of matrix components is an important event in the pathogenesis of metastasis. Thus the interference with this process appears to provide an explanation for the inhibition of malignant cell dissemination in vivo by nafazatrom.