Xu, Z., M. Elaish, C. P. Wong, B. B. Hassan, J. Lopez-Orozco, A. Felix-Lopez, N. S. Ogando, L. Nagata, L. K. Mahal, and A. Kumar, The Wnt/β-catenin pathway is important for replication of SARS-CoV-2 and other pathogenic RNA viruses, , vol. 2, issue 1: Nature Publishing Group UK London, pp. 6, 2024. Abstract
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Abd El-Hameed, R. H., M. S. Mohamed, S. M. Awad, B. B. Hassan, M. A. E. - F. Khodair, and Y. E. Mansour, Novel benzo chromene derivatives: Design, synthesis, molecular docking, cell cycle arrest, and apoptosis induction in human acute myeloid leukemia HL-60 cells, , vol. 38, issue 1: Taylor & Francis, pp. 405 - 422, 2023. Abstract
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Hassan, B. B., A. K. Al-Mokaddem, H. A. Abdelrahman, A. Samir, and M. R. Mousa, Cutaneous tumors in dogs: a retrospective epidemiological and histological study of 112 cases, , vol. 10, issue 1, pp. 170 - 182, 2022. Abstract
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Elshafae, S. M., B. B. Hassan, W. Supsavhad, W. P. Dirksen, R. Y. Camiener, H. Ding, M. F. Tweedle, and T. J. Rosol, "Gastrin-releasing peptide receptor (GRPr) promotes EMT, growth, and invasion in canine prostate cancer", The Prostate, vol. 76, no. 9, pp. 796–809, 2016. Abstract
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Simmons, J. K., B. E. Hildreth III, W. Supsavhad, S. M. Elshafae, B. B. Hassan, W. P. Dirksen, R. E. Toribio, and T. J. Rosol, "Animal models of bone metastasis", Veterinary pathology, vol. 52, no. 5: SAGE Publications Sage CA: Los Angeles, CA, pp. 827–841, 2015. Abstract
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Supsavhad, W., B. B. Hassan, J. K. Simmons, W. P. Dirksen, S. M. Elshafae, N. A. Kohart, A. A. Demirer, and T. J. Rosol, "Effect of Dickkopf-1 (Dkk-1) and SP600125, a JNK Inhibitor, on Wnt Signaling in Canine Prostate Cancer Growth and Bone Metastases.", Veterinary sciences, vol. 8, issue 8, 2021. Abstract

Human Dickkopf-1 (Dkk-1) upregulates a noncanonical Wnt/JNK pathway, resulting in osteoclast stimulation, cell proliferation, and epithelial-to-mesenchymal transition (EMT) of cancer cells. Ace-1-Dkk-1, a canine prostate cancer (PCa) cell line overexpressing Dkk-1, was used to investigate Wnt signaling pathways in PCa tumor growth. SP600125, a JNK inhibitor, was used to examine whether it would decrease tumor growth and bone tumor phenotype in canine PCa cells in vitro and in vivo. Ace-1-Vector and Ace-1-Dkk-1 cells were transplanted subcutaneously, while Ace-1-Dkk-1 was transplanted intratibially into nude mice. The effects of Dkk-1 and SP600125 on cell proliferation, in vivo tumor growth, and bone tumor phenotype were investigated. The mRNA expression levels of Wnt/JNK-related genes were measured using RT-qPCR. Dkk-1 significantly increased the mRNA expression of Wnt/JNK-signaling-related genes. SP600125 significantly upregulated the mRNA expression of osteoblast differentiation genes and downregulated osteoclastic-bone-lysis-related genes in vitro. SP600125 significantly decreased tumor volume and induced spindle-shaped tumor cells in vivo. Mice bearing intratibial tumors had increased radiographic density of the intramedullary new bone, large foci of osteolysis, and increased cortical lysis with abundant periosteal new bone formation. Finally, SP600125 has the potential to serve as an alternative adjuvant therapy in some early-stage PCa patients, especially those with high Dkk-1 expression.

Li, J., M. Haque, C. Shang, B. Hassan, D. Liu, W. Chen, and R. Lai, "Identification and Characterization of Cancer Stem-Like Cells in ALK-Positive Anaplastic Large Cell Lymphoma Using the SORE6 Reporter.", Current issues in molecular biology, vol. 43, issue 2, pp. 543-557, 2021. Abstract

Transcription factors Sox2 and Oct4 are essential in maintaining the pluripotency of embryonic stem cells and conferring stemness in cancer stem-like (CSL) cells. SORE6, an in-vitro reporter system, was designed to quantify the transcription activity of Sox2/Oct4 and identify CSL cells in non-hematologic cancers. Using SORE6, we identified and enriched CSL cells in ALK-positive anaplastic large cell lymphoma (ALK + ALCL). Two ALK + ALCL cell lines, SupM2 and UCONN-L2, contained approximately 20% of SORE6+ cells, which were purified based on their expression of green fluorescent protein. We then performed functional studies using single-cell clones derived from SORE6- and SORE6+ cells. Compared to SORE6- cells, SORE6+ cells were significantly more chemoresistant and clonogenic in colony-formation assays. Sox2/Oct4 are directly involved in conferring these CSL properties, since the shRNA knockdown of Sox2 in SORE6+ significantly lowered their chemoresistance, while enforced expression of Sox2/Oct4 in SORE6- cells produced opposite effects. Using Western blots, we found that the expression and subcellular localization of Sox2/Oct4 were similar between SORE6- and SORE6+ cells. However, in SORE6+ but not SORE6- cells, Sox2 and Oct4 abundantly bound to a probe containing the SORE6 consensus sequence. c-Myc, previously shown to regulate cancer stemness in ALK + ALCL, regulated the SORE6 activity. In conclusion, SORE6 is useful in identifying/enriching CSL cells in ALK + ALCL.

Shang, C., B. Hassan, M. Haque, Y. Song, J. Li, D. Liu, E. Lipke, W. Chen, S. Giuriato, and R. Lai, "Crizotinib Resistance Mediated by Autophagy Is Higher in the Stem-Like Cell Subset in ALK-Positive Anaplastic Large Cell Lymphoma, and This Effect Is MYC-Dependent.", Cancers, vol. 13, issue 2, 2021. Abstract

Previously it was shown that autophagy contributes to crizotinib resistance in ALK-positive anaplastic large cell lymphoma (ALK + ALCL). We asked if autophagy is equally important in two distinct subsets of ALK + ALCL, namely eporter nresponsive () and eporter esponsive (), of which RR cells display stem-like properties. Autophagic flux was assessed with a fluorescence tagged LC3 reporter and immunoblots to detect endogenous LC3 alongside chloroquine, an autophagy inhibitor. The stem-like RR cells displayed significantly higher autophagic response upon crizotinib treatment. Their exaggerated autophagic response is cytoprotective against crizotinib, as inhibition of autophagy using chloroquine or shRNA against or led to a decrease in their viability. In contrast, autophagy inhibition in RU resulted in minimal changes. Since the differential protein expression of MYC is a regulator of the RU/RR dichotomy and is higher in RR cells, we asked if MYC regulates the autophagy-mediated cytoprotective effect. Inhibition of MYC in RR cells using shRNA significantly blunted crizotinib-induced autophagic response and effectively suppressed this cytoprotective effect. In conclusion, stem-like RR cells respond with rapid and intense autophagic flux which manifests with crizotinib resistance. For the first time, we have highlighted the direct role of MYC in regulating autophagy and its associated chemoresistance phenotype in ALK + ALCL stem-like cells.

Hassan, B. B., L. A. Altstadt, W. P. Dirksen, S. M. Elshafae, and T. J. Rosol, "Canine Thyroid Cancer: Molecular Characterization and Cell Line Growth in Nude Mice.", Veterinary pathology, vol. 57, issue 2, pp. 227-240, 2020. Abstract

Thyroid cancer is the most common endocrine malignancy in dogs. Dogs and humans are similar in the spontaneous development of thyroid cancer and metastasis to lungs; however, thyroid cancer has a higher incidence of metastasis in dogs. This study developed a preclinical nude mouse model of canine thyroid cancer using a canine thyroid adenocarcinoma cell line (CTAC) and measured the expression of important invasion and metastasis genes in spontaneous canine thyroid carcinomas and CTAC cells. CTAC cells were examined by electron microscopy. Short tandem repeat analysis was performed for both the original neoplasm and CTAC cells. CTAC cells were transduced with luciferase and injected subcutaneously and into the tail vein. Tumors and metastases were monitored using bioluminescent imaging and confirmed with gross necropsy and histopathology. Invasion and metastasis genes were characterized in 8 follicular thyroid carcinomas (FTCs), 4 C-cell thyroid carcinomas, 3 normal thyroids, and CTAC cells. CTAC cells grew well as xenografts in the subcutis, and they resembled the primary neoplasm. Metastasis to the kidney and lung occurred infrequently following subcutaneous and tail vein injection of CTAC cells. STR analysis confirmed that CTAC cells were derived from the original neoplasm and were of canine origin. Finally, 24 genes were differentially expressed in spontaneous canine thyroid carcinomas, CTAC, and normal thyroids. This study demonstrated the usefulness of a nude mouse model of experimental canine thyroid carcinoma and identified potential molecular targets of canine follicular and C-cell thyroid carcinoma.

Elshafae, S. M., W. P. Dirksen, A. Alasonyalilar-Demirer, J. Breitbach, S. Yuan, N. Kantake, W. Supsavhad, B. B. Hassan, Z. Attia, L. B. Alstadt, et al., "Canine prostatic cancer cell line (LuMa) with osteoblastic bone metastasis.", The Prostate, vol. 80, issue 9, pp. 698-714, 2020. Abstract

BACKGROUND: Osteoblastic bone metastasis represents the most common complication in men with prostate cancer (PCa). During progression and bone metastasis, PCa cells acquire properties similar to bone cells in a phenomenon called osteomimicry, which promotes their ability to metastasize, proliferate, and survive in the bone microenvironment. The mechanism of osteomimicry resulting in osteoblastic bone metastasis is unclear.

METHODS: We developed and characterized a novel canine prostatic cancer cell line (LuMa) that will be useful to investigate the relationship between osteoblastic bone metastasis and osteomimicry in PCa. The LuMa cell line was established from a primary prostate carcinoma of a 13-year old mixed breed castrated male dog. Cell proliferation and gene expression of LuMa were measured and compared to three other canine prostatic cancer cell lines (Probasco, Ace-1, and Leo) in vitro. The effect of LuMa cells on calvaria and murine preosteoblastic (MC3T3-E1) cells was measured by quantitative reverse-transcription polymerase chain reaction and alkaline phosphatase assay. LuMa cells were transduced with luciferase for monitoring in vivo tumor growth and metastasis using different inoculation routes (subcutaneous, intratibial [IT], and intracardiac [IC]). Xenograft tumors and metastases were evaluated using radiography and histopathology.

RESULTS: After left ventricular injection, LuMa cells metastasized to bone, brain, and adrenal glands. IT injections induced tumors with intramedullary new bone formation. LuMa cells had the highest messenger RNA levels of osteomimicry genes (RUNX2, RANKL, and Osteopontin [OPN]), CD44, E-cadherin, and MYOF compared to Ace-1, Probasco, and Leo cells. LuMa cells induced growth in calvaria defects and modulated gene expression in MC3T3-E1 cells.

CONCLUSIONS: LuMa is a novel canine PCa cell line with osteomimicry and stemness properties. LuMa cells induced osteoblastic bone formation in vitro and in vivo. LuMa PCa cells will serve as an excellent model for studying the mechanisms of osteomimicry and osteoblastic bone and brain metastasis in prostate cancer.