bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2384: Quantitative phase imaging provides new insights into diverse morphologies in cell lines and their behavior in response to alternative cancer treatments
Authors: Khan Rida , McDonald Meaghan , Mikhaylova Albina , Rehman Fatima K.
Published: April 2023

Quantitative phase imaging (QPI) through holographic time-lapse microscopy allows for non-invasive measurements of cell dry mass and the ability to track cells for long periods of time with minimal damage, making it most beneficial for research on cancer treatment. Research with this enhanced method of observation can help decipher various morphologies present in specific cell lines and show how each cell can respond differently to provided signals. In this assay, we studied N2a murine neuroblast differentiation in response to Retinoic acid, as well as the anticancerous activity of Saffron and UV in MDA-MB-468 breast carcinoma and HCT116 colon carcinoma cell lines. Cells were observed using Telight’s Q-Phase, a multimodal microscope that utilizes QPI holographic time-lapse technology with label-free monitoring. The cells were able to survive in optimum conditions for up to 96 hours without any appreciative loss of viability.  With the emergence of phytochemicals being used as alternative cancer treatments, specific pigments in saffron have shown therapeutic effects against cancer tissues such as apoptosis and decreased carcinogen presence. A microanalysis of the effects of saffron on reducing the invasion potential, viability, and tumorigenicity of breast cancer cell lines was performed. QPI microscopy was performed on MDA-MB-468 human breast adenocarcinoma and HCT116 colon carcinoma cell lines treated with varying doses of saffron and ultraviolet radiation. Cell lines were observed under the microscope for 58-hour periods for changes in mass, movement, and replication. The quantitative data showed a reduction of projections from saffron treatment but did not show any significant reduction in cell movement over the 58-hours. These results were later confirmed through scratch wound and Boyden chamber assays. Similarly, when examining the process of N2a neuroblast differentiation, it was observed that cells do not move towards differentiation in a linear manner, but rather changed their direction and path several times before committing to their final phenotype. These types of observations are critical in understanding the biological processes behind differentiation and drug response and could not have been done without using the various quantitative and analysis tools provided by the QPI technology.

Citation Format: Rida Khan, Meaghan McDonald, Albina Mikhaylova, Fatima K. Rehman. Quantitative phase imaging provides new insights into diverse morphologies in cell lines and their behavior in response to alternative cancer treatments [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2384.
Abstract 2507: Chronic stress promotes tumor progression and metastasis through the tumor microenvironment in triple-negative breast cancer
Authors: Zhang Chao , Wang Lizhong , Liu Runhua
Published: April 2023

Background: Breast cancer is the most common cancer diagnosed in American women. Triple-negative breast cancer (TNBC) is three times more likely to end in death than other breast cancer subtypes and disproportionately affects young African American women. Chronic stress as a risk factor promotes tumor progression, but its role in TNBC progression and metastasis remains elusive.

Methods: In this study, mice were implanted with 4T1-luciferase cells in the tibia or the fourth mammary pad of BALB/c mice and NSG mice. After four weeks of restraining inducing chronic stress and (or) β-adrenergic receptor inhibitor treatment, the impact of chronic stress on the progression and metastasis of tumor, tumor microenvironment was assessed.

Results: Our results showed that chronic stress facilitated tumor progression and metastasis in both BALB/c mice and NSG TNBC mice models. In contrast, chronic stress reduced the number and function of tumor-infiltrating TCD8+ cells but increased the expression of the β2-adrenergic receptor of TCD8+ cells in the TNBC tumor microenvironment (TME) of BALB/c mice. Interestingly, chronic stress increased the density of neuron fiber in the TME of both BALB/c and NSG TNBC mice. Notably, the treatment of Propranolol (Prop, a β1- and β2-adrenergic receptor inhibitor) significantly decreased the tumor burden and metastasis of both BALB/c mice and NSG TNBC mice model. Accompany by recovering the function and number of TCD8+ cells in the TME of BALB/c mice.

Conclusion: Our study indicates that chronic stress promotes TNBC progression and metastasis by regulating the tumor-immune network in the TME.

Citation Format: Chao Zhang, Lizhong Wang, Runhua Liu. Chronic stress promotes tumor progression and metastasis through the tumor microenvironment in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2507.
Abstract 2534: Targeting defective necroptosis in colorectal cancer to overcome therapeutic resistance
Authors: Ermine Kaylee , Chen Dongshi , Lu Xinyan , Yu Jian , Zhang Lin
Published: April 2023

Colorectal cancer (CRC) is a leading cause of cancer related deaths in the United States. CRC patients are often treated with the chemotherapeutic drug 5-fluorouracil (5-FU), but resistance often occurs. It is well-established that 5-FU induces apoptosis in cancer cells, but apoptosis inhibition does not block the anticancer effect of 5-FU, suggesting a role of non-apoptotic cell death. Necroptosis is a regulated form of necrosis controlled by Receptor-Interacting Protein kinase 1 (RIP1), RIP3, and Mixed Lineage Kinase domain-Like (MLKL), and its role in anticancer therapy remains unclear. TCGA RNAseq data show that RIP3 is significantly downregulated in primary CRC tumors, which is correlated with worse outcomes. RIP3 downregulation also occurs in established CRC cell lines and is associated with decreased 5-FU sensitivity. Therefore, we hypothesize that necroptosis may play a critical role in anticancer therapy and that restoring defective necroptosis may enhance therapeutic sensitivity. Our studies show that RIP3-expressing (RIP3+) CRC cells undergo both apoptosis and necroptosis in response to 5-FU. As expected, knock-out of RIP3 in RIP3+ CRC cells resulted in decreased 5-FU sensitivity, as well as abrogated necroptosis induction. Additionally, stable restoration of RIP3 in RIP3-silenced (RIP3-) CRC cells appeared to enhance 5-FU response, under conditions where apoptosis was inhibited. Furthermore, we explored alternative ways to induce necroptosis in RIP3-silenced cells. Together, our results suggest that necroptosis plays an important role in 5-FU-induced cell death in CRC. These results provide novel insights into the role of necroptosis in anticancer therapy in CRC, which could be used for development of improved therapies and personalized medicine based on RIP3 status.

Citation Format: Kaylee Ermine, Dongshi Chen, Xinyan Lu, Jian Yu, Lin Zhang. Targeting defective necroptosis in colorectal cancer to overcome therapeutic resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2534.
Abstract 2546: Targeting of AKT/β-catenin signaling pathways driven hepatocellular carcinoma cell lines by combined treatment of HDAC and multi-kinase inhibitors
Authors: Elberawi Mohamed Mahmoud , Abdulqadir Mohamed , Sultan Ahmed Samir
Published: April 2023

Hepatocellular carcinoma (HCC) is the dominant type of liver cancer exhibiting high mortality and poor prognosis with the fastest rate of death due to its aggressiveness and lack of effective therapies. Beta-catenin is a key player transcriptional factor, which orchestrates gene expression in HCC progression. Activation of β-catenin is promoted by its phosphorylation by AKT. Although the studies of metastatic behavior of β-catenin over-expressing HCC cells are not clear, the present study is focusing on the relationship between chromatin remodelling through deacetylation and phosphorylation of β-catenin by AKT. The current study investigated the role of a combination of Lenvatinib, a multi-kinase inhibitor, and Belinostat, a histone deacetylase inhibitor (HDACI), on two HCC cell lines, Hep G2 and Hep 3B, respectively. In-depth, our data showed that combined treatment significantly induced morphological changes in the tested cell lines with activation of caspase-3 and pre-apoptotic signals of Bcl-2, Bcl-XL, and Bax, indicating apoptotic induction after 48 hours of treatment compared to the control or each drug separately. The calculated IC50 was 1µmol/L for Belinostat and 25 n mole/L for Lenvatinib. Western blotting analysis reveals that the combined treatment for 48 hours inhibited the oncogenic β-catenin, AKT, and TGFβ expression in a dose-dependent manner.In addition, the combined treatment inhibited the localization of beta-catenin in the tested cell nuclei after 48 hours as confirmed by IHC staining. Statistical analysis proved that the combination of both drugs significantly decreased the proliferation and invasion of both cell lines (P<0.01, ANOVA) compared to each drug alone. In addition, the percentage of apoptotic cells was significantly increased and showed a cell cycle arrest in the G0/G1 phase of Hep-G2 cells after 48 hours of combined treatment as confirmed by flow cytometry. All in all, our data show that targeting AKT/β-catenin driven HCC progression can be targeted by Lenvatinib and Belinostat combined treatment.

Citation Format: Mohamed Mahmoud Elberawi, Mohamed Abdulqadir, Ahmed Samir Sultan. Targeting of AKT/β-catenin signaling pathways driven hepatocellular carcinoma cell lines by combined treatment of HDAC and multi-kinase inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2546.
Abstract 2587: AHR-WLS complex correlates with poor prognosis in B-cell precursor acute lymphoblastic leukemia
Authors: Wang Li-Ting , Hsu Shih-Hsien
Published: April 2023

B cell precursor acute lymphoblastic leukemia (BCP ALL), the most common childhood cancer, is a malignant disorder that originates from the lymphoid precursor cells committed to B-cell lineage in the bone marrow. Although genetic abnormalities disturb the normal maturation process of these lymphoid precursor cells resulting in the formation of leukemia cells and suppression of normal hematopoiesis, environmental factors seem to modulate disease progression. The underline mechanism is not clear yet. In this study, we hypothesize environmental pollutants-AHR and small molecule chaperone-WLS are involved in promoting the oncogenic effect and disease relapse of BCP-ALL leukemia cells and inhibiting normal hematopoietic function. Here, we show that Indeno [1,2,3-cd] pyrene (IP) promotes oncogenic activities in vitro and in vivo. Mechanistically, IP treatment activates AHR-IDOs axis to enhance tryptophan metabolism and bioactive metabolite- kynurenine (kyn) level, further promoting kyn-AHR signal enhanced loop. In orthotopic xenograft mouse model, IP treatment decreases disease relapse time and increases cell population of BCP ALL. These results show PAH (IP) exposure promotes the disease progression of BCP ALL.

Citation Format: Li-Ting Wang, Shih-Hsien Hsu. AHR-WLS complex correlates with poor prognosis in B-cell precursor acute lymphoblastic leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2587.
Abstract 2606: Requirement of MDM2 E3 ligase activity for regulating p53 during normal development, cell cycle regulation and genome integrity
Authors: Chinnam Meenalakshmi , Lama Rati , Xu Chao , Zhang Xiaojing , Cedeno Carlos , Wang Yanqing , Stablewski Aimee B. , Goodrich David W. , Wang Xinjiang
Published: April 2023

p53 tumor suppressor, the most mutated gene in human cancer, is negatively regulated by MDM2 and MDM4 proteins. Two distinct mechanisms of MDM2/MDM4 regulation of p53 include, one, suppression of p53 transactivation activity by direct interaction of MDM2/MDM4 with p53 protein, thus blocking its transactivation interface and two, targeting p53 to proteasomal degradation by MDM2 E3 ligase activity. However, the importance of these mechanisms in normal p53 regulation in vivo has remained controversial. To genetically separate these two mechanisms of p53 regulation we generated a novel Mdm2 mutant mouse (Mdm2L466A) that lacks E3 ligase activity but retains the ability of Mdm2 to heterodimerize with Mdm4 and inhibit p53 transactivation. Homozygous Mdm2L466A mice are embryonic lethal due to dysregulated p53 activity, thus demonstrating Mdm2 E3 ligase mediated regulation of p53 is essential during embryonic development. Additionally, we uncovered novel p53-independent functions of MDM2 E3 ligase in cell cycle regulation and genome integrity in cells. Cells lacking MDM2 E3 ligase activity have defective G2-M transition during cell cycle and developed elevated levels of aneuploidy regardless of p53 status. This study unequivocally demonstrates the requirement of Mdm2 E3 ligase activity for normal regulation of p53 and uncovers previously unknown p53-independent role of Mdm2 in cell cycle regulation and maintaining genome integrity. A current therapeutic approach involves blocking MDM2-p53 interaction to promote p53 tumor suppression function in cancer cells. Our findings suggest that this approach alone is insufficient because MDM2 has other potential oncogenic mechanisms independent of its role as negative regulator of p53. Hence, targeting MDM2 E3 ligase activity is likely to be more effective cancer therapy as it addresses both p53-dependent and p53-independent oncogenic mechanisms of MDM2.

Citation Format: Meenalakshmi Chinnam, Rati Lama, Chao Xu, Xiaojing Zhang, Carlos Cedeno, Yanqing Wang, Aimee B. Stablewski, David W. Goodrich, Xinjiang Wang. Requirement of MDM2 E3 ligase activity for regulating p53 during normal development, cell cycle regulation and genome integrity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2606.
Abstract 2649: Discovery of a potent, selective, and tumor-suppressing antibody antagonist of DKK1
Authors: Wang Linya , Peterson Sean M. , Lujan Ana , Chan Kara Y. , Hu Cameron F. , Villalta Mouna , Yuan Tom Z. , Han Zhen , Yang Marisa , Prabhu Vishwas , Htoy Tammy , VanDyke Paul , Holliday Carson , Franco Hector , Wadhwa Hansika , Giang Hoa , Stafford Ryan , Axelrod Fumiko , Sato Aaron
Published: April 2023

Wnt signaling plays an important role in embryonic development and tumorigenesis. These biological effects are exerted by activation of the β-catenin/TCF transcription complex and consequent regulation of a set of downstream genes. Dickkopf WNT Signaling Pathway Inhibitor 1 (DKK1) has been shown to be a potent inhibitor of Wnt signaling via competing Wnt binding to LRP5/6. DKK1 is tumorigenic in multiple cancer types and also immunosuppressive via MDSCs and NK cells. Emerging evidence indicates that DKK1 has been involved in T cell differentiation and induction of cancer evasion of immune surveillance by accumulating MDSCs. Consequently, DKK1 has become a promising target for cancer immunotherapy, and the mechanisms of DKK1 affecting cancers and immune cells have received great attention. With Twist’s precision DNA writing technologies, we have created phage display libraries with diversity greater than 1 × 1010, and utilized machine learning model for optimal discovery. In this study, we discovered anti-DKK1 antibodies blocking the binding of DKK1 to the receptor. The in vitro functional assays showed that the interaction of Wnt to its receptor was restored in the presence of anti-DKK1 antibodies, resulting in TCF/LEF signaling upregulation. Moreover, anti-DKK1 antibodies induced immune cell activation, and led to tumor cell cytotoxicity. In addition, anti-DKK1 antibodies binding to different domains of DKK1 led to different functional effect. In vivo study indicated anti-DKK1 antibodies is potent in tumor regression. Our anti-DKK1 leads shows promising efficacy in cancer immunotherapy.

Citation Format: Linya Wang, Sean M. Peterson, Ana Lujan, Kara Y. Chan, Cameron F. Hu, Mouna Villalta, Tom Z. Yuan, Zhen Han, Marisa Yang, Vishwas Prabhu, Tammy Htoy, Paul VanDyke, Carson Holliday, Hector Franco, Hansika Wadhwa, Hoa Giang, Ryan Stafford, Fumiko Axelrod, Aaron Sato. Discovery of a potent, selective, and tumor-suppressing antibody antagonist of DKK1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2649.
Abstract 2668: Long non-cording RNA TUG1 promotes cisplatin resistance in ovarian cancer via DNA polymerase eta
Authors: Sonobe Ryosuke
Published: April 2023

Ovarian cancer (OVCA) is the most malignant gynecologic cancer. Despite the use of surgery and platinum-based chemotherapy, the 5-year survival rate is approximately 30% at the time of diagnosis. Cisplatin inhibits DNA synthesis by cross-linking DNA and then induces cell apoptosis. However, cisplatin resistance often causes treatment failure and tumor relapse in OVCA patients. To overcome the cisplatin resistance in OVCAs, we studied the mechanism of chemoresistance by a long non-coding RNA (lncRNA), TUG1 (taurine upregulated gene 1), which is associated with carcinogenesis and drug sensitivity in OVCAs. The expression level of TUG1 in cisplatin-resistant OVCA clinical samples is significantly higher than that of cisplatin-sensitive samples by public database. A depletion of TUG1 increased the sensitivity of OVCA cells to cisplatin together with upregulation of a set of miRNAs in vitro. We examined the targets of miRNAs and found that TUG1 depletion downregulates DNA polymerase eta (Polη) by upregulating the miRNAs. Polη is a major translesion DNA synthesis (TLS) DNA polymerase, which bypasses DNA lesions produced by DNA damaging agents and helps DNA replication. The expression level of Polη showed a positive correlation with that of TUG1 in OVCA clinical samples. Overexpression of Polη recovered cisplatin resistance in TUG1-depleted cells. Finally, cisplatin treatment combined with TUG1 depletion effectively inhibited tumor growth in xenograft mouse models, indicating the efficacy of TUG1 depletion in vivo. Our data provide a strong rationale for targeting TUG1 as a novel therapeutic approach to overcome cisplatin resistance in OVCA.

Citation Format: Ryosuke Sonobe. Long non-cording RNA TUG1 promotes cisplatin resistance in ovarian cancer via DNA polymerase eta [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2668.
Abstract 2689: Targeting the proteasome pathway in dactinomycin resistant Wilms tumors
Authors: Tiburcio Patricia Denise Ballesteros , Chen Kenneth S.
Published: April 2023

Nephroblastomas or Wilms tumors are the most common pediatric renal cancer. Across all cases, the 5-year overall survival is over 90%. This is largely in part due to brute-force, chemotherapy regimens such as dactinomycin (actD) that have been routinely used to treat Wilms tumors. However, those that do respond to treatment still face a 15% chance of relapse. Patients with anaplastic tumor histology require more toxic therapies, and yet they still face a worse prognosis of only 66% 4-year survival and relapse rate of 43%. Furthermore, recurrence cuts survival rate down to 50%. With the widespread use of actD in Wilms tumors and other childhood malignancies, understanding the mechanism to recalcitrance against this chemotherapy can potentially reveal important vulnerabilities that can be exploited in the clinic. While transcriptomic and genomic analyses have been done in Wilms tumors, no targetable pathways have been identified in actD chemotherapy-resistant and anaplastic tumor cases. To that end, we performed ribosome sequencing of actD-treated anaplastic Wilms tumor cell line WiT49 and uncovered that the proteasome pathway is preferentially translated. Moreover, loss of proteasome subunits in a CRISPR library dropout screen sensitized the cells to actD. Consistent with these findings we found upregulation of proteasome subunit levels and proteasome enzymatic activity in Wilms tumor lines following actD treatment. To interrogate the effect of cotreating bortezomib with dactinomycin in vivo, we subcutaneously implanted anaplastic Wilms tumor patient-derived xenograft (PDX) lines in NOD/SCID Gamma (NSG) mice. Combinatorial treatment was found to be retard tumor growth compared to vehicle and single drug conditions. With further investigation, our findings can provide a foundation for exploring proteasome inhibitors with dactinomycin in clinical trials. Our study has important implications to improving the outcomes for not just for recurrent and anaplastic Wilms tumors, but to other pediatric cancers that use actD as well.

Citation Format: Patricia Denise Ballesteros Tiburcio, Kenneth S. Chen. Targeting the proteasome pathway in dactinomycin resistant Wilms tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2689.
Abstract 2701: pH-responsive targeted-nanoparticle releases encapsulated ERK-inhibitor effectively in the hypoxic microenvironment and sensitizes gemcitabine in pancreatic cancer cells
Authors: Dutta Debasmita , De Archana , Hazra Raj Shankar , Ghosh Pratyusha , Quadir Mohiuddin , Banerjee Snigdha , Banerjee Sushanta K.
Published: April 2023

Therapeutic strategies for Pancreatic ductal adenocarcinoma (PDAC), one of the deadliest types of aggressive malignancies, remain challenging with poor prognosis, high mutational rate, and lowest patient survival among other cancers. The treatment options for managing PDAC are limited to surgery, radiation, and chemotherapy. However, surgery is the only treatment option that can cure the disease in about twenty percent of cases. Thus, Gemcitabine (GEM) remains the first-line drug in the clinic for PDAC treatment despite suboptimal clinical effects. These are primarily due to weak drug delivery and the development of chemoresistance resulting from mutant-K-RAS/AKT/ERK signaling-mediated desmoplastic cages. Several new therapeutic approaches, including nanoparticle-based drug delivery procedures, are currently being investigated. In this study, we designed a pH-responsive PEG-b-poly (carbonate) block copolymer encapsulated ERK inhibitor (SCH772984) nanoparticles surface functionalized with tumor-penetrating peptide, iRGD to specifically target PDAC tumor tissue. By utilizing the ERKi-encapsulated pH-responsive targeted nanoparticles, we performed a combinational drug treatment of GEM and ERKi. We found that the pH-responsive targeted nanocarrier efficiently released ERKi in the hypoxic environment. The free GEM, combined with nanoencapsulated ERKi, demonstrated significant synergistic outcomes in vitro colony forming ability, migration, and motility in pancreatic cancer cell line MiaPaCa2. Moreover, combination therapy significantly enhanced the GEM effect in reducing PDAC growth in KPC mice and increasing the survival of the mice. Thus, our studies indicate that pH-responsive targeted nanocarrier-based delivery of ERKi in the hypoxic area of PDAC enhances the efficacy of GEM in PDAC.

Citation Format: Debasmita Dutta, Archana De, Raj Shankar Hazra, Pratyusha Ghosh, Mohiuddin Quadir, Snigdha Banerjee, Sushanta K. Banerjee. pH-responsive targeted-nanoparticle releases encapsulated ERK-inhibitor effectively in the hypoxic microenvironment and sensitizes gemcitabine in pancreatic cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2701.
Abstract 2715: Utilizing a pH sensitive peptide (pHLIP) for tumor targeted delivery of an immunogenic peptide motif
Authors: Yurkevicz Annali M. , Liu Yanfeng , Glazer Peter M.
Published: April 2023

Chimeric antigen receptor (CAR) T cell and immune-cell therapies are revolutionizing how patients are treated in the clinic by targeting tumor-specific antigens. Specifically, advances in antigen targeting of CD20 and CD19 with CAR T cell therapy has increased overall survival of this patient population and has influenced to expand T cell repertoire to novel antigen targets. However, limitations to these cell-based therapies are the lack of development against solid tumor antigens, partly due to the paucity of antigen candidates that are sufficiently specific to epithelial cancers. Therefore, technologies that can mark tumor cells for immune recognition could prove beneficial in the treatment of these solid cancers. This project aims to artificially deliver an antigen to tumor cells by harnessing a universal property of tumors: acidic microenvironment. The pH low insertion peptide (pHLIP) is a technology that targets solid tumors based on the low pH produced by the Warburg effect. pHLIP-conjugates have undergone extensive development for drug delivery, with one notable candidate CBX12 entering phase I clinical trials in 2021. Our system uses a pHLIP-conjugated SIINFEKL peptide (derived from ovalbumin) to serve as our model antigen system, with SIINFEKL-targeting T cells (OT1) that can be used as a tool to investigate antigen delivery, recognition, and functionality. We have observed through immunofluorescence that B16 F10 mouse melanoma cells treated in vitro with the 1uM pHLIP-SIINFEKL construct under acidic conditions can recruit OT1 cells and induce a T cell synapse. Preliminary in vivo experiments in C57BL/6-Tg OT1 mice bearing B16 F10 flank tumors showed that treatment with pHLIP-SIINFEKL by intravenous administration led to a suppression of tumor growth compared to controls, with approximately 50% smaller fold change in tumor volume. Cumulatively, these initial results suggest that pHLIP-SIINFEKL can be delivered to tumors to recruit OT1 cells to engage in antigen recognition and lead to efficacy in tumor growth delay in vivo. This technology has the capability to provide a novel approach to decorate solid tumors with engineered antigens for enhanced immune recognition and anti-tumor efficacy.

Citation Format: Annali M. Yurkevicz, Yanfeng Liu, Peter M. Glazer. Utilizing a pH sensitive peptide (pHLIP) for tumor targeted delivery of an immunogenic peptide motif [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2715.
Abstract 2742: Small interfering RNA targeting CD24 inhibits proliferation and migration of triple-negative breast cancer cells
Authors: Liu Yanli , Cheng Kun
Published: April 2023

Triple-negative breast cancer (TNBC) is defined as the lack of human epidermal growth factor receptor 2 (HER2), progesterone receptor (PR), and estrogen receptor (ER) expression. TNBC accounts for 10-20% of all cancer cases and is more aggressive than other subtypes of breast cancer. Although chemotherapy is the mainstay of treatment for TNBC, its therapeutic efficacy is limited. Therefore, developing new therapies to address the unmet need of TNBC is urgently needed.

CD24 protein is a heavily glycosylated glycosylphosphatidylinositol (GPI)-anchored surface protein. The high expression level of CD24 is observed in a variety of cancers, such as TNBC and ovarian cancer. CD24 plays an important role in tumor growth, invasion, and metastasis of different types of cancers. Besides, overexpression of CD24 in tumors is associated with resistance to chemotherapy agents and targeted therapies.

Here, we design four siRNAs targeting different regions of the CD24 mRNA. The silencing activity of the predesigned siRNAs is evaluated by quantitative real-time RT-PCR and western blot. The biological effects of designed siRNAs on human breast cancer cells are investigated by cell proliferation, migration, invasion, cell cycle arrest, and apoptosis. The data demonstrate that the CD24 siRNAs significantly reduce CD24 expressions at both mRNA level and protein level in human TNBC cells. Moreover, silencing of CD24 suppresses breast cancer cell proliferation. Cell cycle assay shows that the anti-proliferation effect of CD24 siRNA is mediated by arresting the cells in the S phase. Furthermore, we reveal that silencing of CD24 inhibits migration and invasion of human breast cancer cells. Significant apoptosis is observed in CD24 siRNA transfected cells compared with the scrambled siRNA group. These data suggest that silencing CD24 using siRNA can be a potential therapeutic strategy for triple-negative breast cancer.

Citation Format: Yanli Liu, Kun Cheng. Small interfering RNA targeting CD24 inhibits proliferation and migration of triple-negative breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2742.
Abstract 2755: Identification of novel GPX4 inhibitors using global transcriptional reporters
Authors: Read Timothy , Winigrad Zoe , Goliaei Ardeshir , Liechty Cole , Grochala Carter , Damon Leah , Dickson John , Harris Jason , Pham Casey , Rimel Jenna , Rhine Christy , Simpson Dave , Martin Eric , Azofeifa Joey
Published: April 2023

Induction of Ferroptosis is a promising strategy for treating therapy-resistant tumors including mesenchymal tumor types. We performed a phenotypic screen to identify small molecules that induce ferroptosis in mesenchymal cancer cells by screening for compounds whose toxicity could be rescued by the lipophilic antioxidant Ferrostatin-1. To identify compounds that induce GPX4-mediated ferroptosis, we identified several pharmacodynamic biomarkers specific to GPX4 inhibition using our nascent RNA sequencing platform, which allows for time-resolved snapshots of global transcription. We measured the transcriptional changes that occur in the hours following inhibition of GPX4 and identified HMOX1 as a robust biomarker of GPX4i-induced ferroptosis in mesenchymal cancer cell lines. Using HMOX1-induction as a guide, we uncovered a small number of compounds as potential GPX4 inhibitors. One such compound was validated as a bonafide GPX4 inhibitor through a variety of biochemical assays and selected for a hit-to-lead campaign. In addition to standard medicinal chemistry strategies, we are currently employing a newly-developed global transcriptional reporter system to stratify several novel series of GPX4 inhibitors by their transcriptional signatures. This approach represents a unique strategy for determining on- and off- target effects of a compound and for defining structure activity relationships within a chemical series.

Citation Format: Timothy Read, Zoe Winigrad, Ardeshir Goliaei, Cole Liechty, Carter Grochala, Leah Damon, John Dickson, Jason Harris, Casey Pham, Jenna Rimel, Christy Rhine, Dave Simpson, Eric Martin, Joey Azofeifa. Identification of novel GPX4 inhibitors using global transcriptional reporters [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2755.
Abstract 2846: The mitochondrial dynamics of breast cancer regulate P-glycoprotein expression
Authors: Terrero David , Malla Saloni , Raman Dayanidhi , Babu Jayachandra , Tiwari Amit
Published: April 2023

Multidrug efflux transporter ABCB1 (MDR1) encodes P-glycoprotein (P-gp), a well-known transporter that is believed to contribute to the development of multidrug resistance (MDR). Furthermore, P-gp plays an important role in drug disposition, pharmacokinetics, and drug development. Thus, P-gp transporter expression, function, and modulation of MDR in human cancers have been extensively studied. In spite of significant progress in understanding P-gp, ABCB1 regulation and its impact on clinical trials remain unclear. Through sequential exposure of SUM159PT to paclitaxel, we created a paclitaxel resistant triple negative breast cancer cell line SUM159PT-PAC200. We observed a significant increase in the phosphorylation of the mitochondrial fission protein dynamin related protein kinase 1 (Drp1) at Ser616 and its upstream kinase ERK1/2a of Drp1. Through the use of CRISPR-Cas9 technology, we knocked out the gene encoding Drp1 in PAC200. When Drp1 is genetically abated (PAC200 Drp1-/-), paclitaxel resistance is significantly reversed. It was also observed that clonogenicity under paclitaxel pressure was significantly decreased in both the PAC200/Drp1-/- line and the partial KO Drp1-/+ line, compared to the PAC200 line. Parallelly, a significant increase in the expression of P-gp transporters was also observed in the PAC200 cell line. As a surprise, genetically targeting Drp1 in PAC200 cells (i.e., PAC200 Drp1/KO) reduced P-gp expression to virtually undetectable levels. Until now, there has been no description of how Drp1 and P-gp are related. Furthermore, PAC200 Drp1/KO cells showed reduced proliferation, migration, and invasion potential, while their soft agar clonogenic capacities were almost nonexistent, suggesting that Drp1 plays a key role in breast cancer proliferation and dissemination. It appears that modulating or targeting Drp1 might be a useful strategy for modulating P-gp in the development and maintenance of the cancer's MDR phenotype, proliferation, and invasion. Understanding how mitochondrial dynamics determine the transactivation and coordination of drug-resistant P-gp phenotypes would make Drp1 an attractive molecular target for cancer treatment.

Citation Format: David Terrero, Saloni Malla, Dayanidhi Raman, Jayachandra Babu, Amit Tiwari. The mitochondrial dynamics of breast cancer regulate P-glycoprotein expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2846.
Abstract 2978: YH013, a novel EGFR x MET bispecific antibody-drug conjugate, exhibits potent antitumor efficacy
Authors: Han Yanfei , Shang Chengzhang , li Zhuon , Han Zhenyan , Li Jun , Cui Zixu , An Gao , Hao Wanjun , Liu Yujie , Li Hao , Liu Baihong , Zhang Ellen , Lin Qingcong , Yang Yi , Shen Yuelei
Published: April 2023

Bispecific antibodies (BsAb) that target dual tumor-associated antigens have the advantages of invoking synergistic effects between two signaling pathways, increasing target tissue specificity, and reducing systemic toxicity. By combining the antibody-mediated specific targeting with potent killing from a cytotoxic payload, antibody-drug conjugates (ADC), and especially bispecific ADCs (BsADC), have become powerful therapeutic strategies. EGFR and MET are oncogenic proteins that are co-expressed in a wide range of tumors. Moreover, MET amplification is largely associated with drug resistance of EGFR tyrosine kinase inhibitors in non-small cell lung cancer (NSCLC) patients. Biocytogen developed a fully human EGFR x MET BsADC using our proprietary common light chain RenLiteTM mouse platform and knobs-into-holes technology. The BsAb showed enhanced internalization and binding affinity compared to parental monoclonal and monovalent antibodies in the EGFR/MET co-expressing NCI-H1975 cell line. After conjugating the BsAb with monomethyl auristatin E (MMAE) via a protease-cleavable linker, the resulting BsADC, YH013, exhibited a remarkable and dose-dependent anti-tumor efficacy in NCI-H1975 and NCI-H292 cell line-derived xenograft models. Moreover, in multiple patient-derived xenografts of NSCLC and pancreatic ductal adenocarcinoma (PDAC), which co-express EGFR and MET, YH013 demonstrated superior and durable efficacy that outperformed benchmark antibodies at a lower dose (3 mg/kg). Collectively, these results suggest that YH013 can be an effective treatment option for EGFR and MET co-expressing tumors and overcome MET-driven EGFR-TKI resistance to improve patient outcomes.

Citation Format: Yanfei Han, Chengzhang Shang, Zhuon li, Zhenyan Han, Jun Li, Zixu Cui, Gao An, Wanjun Hao, Yujie Liu, Hao Li, Baihong Liu, Ellen Zhang, Qingcong Lin, Yi Yang, Yuelei Shen. YH013, a novel EGFR x MET bispecific antibody-drug conjugate, exhibits potent antitumor efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2978.
Abstract 3023: Identifying colorectal cancer biomarkers using a targeted proteomics platform with extensive coverage
Authors: Harbs Justin , Rentoft Matilda , Liu Xijia , Melin Beatrice S. , Harlid Sophia , Van Guelpen Bethany
Published: April 2023

Despite advances in treatment, colorectal cancer remains one of the leading causes of cancer-specific death. To decrease colorectal cancer mortality, better and more cost-efficient tools for risk prediction and early detection are needed. One area that has shown promise is using protein biomarkers, measured in blood, to detect individuals with early-stage disease or at increased risk. The primary aim of this study was therefore to identify novel protein biomarkers associated with colorectal cancer risk and the secondary aim was to validate protein biomarkers that had been described in previous studies. We included data and samples from 185 incident colorectal cancer cases and 185 controls (matched on age, sex, sampling date, and the number of freeze-thaw cycles) from the Northern Sweden Health and Disease Study (NSHDS). Samples were collected between six years and three months prior to the diagnosis date of each case and concentrations of 1536 individual proteins were measured in plasma samples using the Olink Explore 1536 platform (Olink Proteomics). Prior to statistical analyses, 249 proteins were excluded, either due to not passing quality control or having more than 50% of the concentrations below the limit of detection. In addition, we also excluded extreme outliers. Conditional logistic regression was used to estimate Odds Ratios (ORs) and 95% confidence intervals (95% CI) for the association between protein concentrations and colorectal cancer risk. Multivariable models were adjusted for predetermined colorectal cancer risk factors, including body mass index, alcohol consumption, physical activity, smoking status, and level of education. When considering p<0.005 as a cut-off, we identified 20 proteins that were associated with colorectal cancer risk. Our top hit was TFF3 (Trefoil Factor 3) which was positively associated with colorectal cancer at a high significance level (multivariable OR per SD: 1.84 95% CI: 1.38 - 2.45). TFF3 was significant in men (FDR<0.1), but not in women (pheterogeneity <0.05). In addition, several of the top 20 proteins such as AREG, MMP7, and CXCL16 have been linked to the diagnosis or prognosis of colorectal cancer in earlier studies. To conclude, we identified one novel protein biomarker, TFF3, strongly associated with colorectal cancer risk and validated several proteins previously linked to the diagnosis or prognosis of colorectal cancer.

Citation Format: Justin Harbs, Matilda Rentoft, Xijia Liu, Beatrice S. Melin, Sophia Harlid, Bethany Van Guelpen. Identifying colorectal cancer biomarkers using a targeted proteomics platform with extensive coverage [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3023.
Abstract 3072: Estrogen receptor β4 regulates chemotherapy resistance and induces cancer stem cells in triple negative breast cancer
Authors: Bano Ayesha , Stevens Jessica , Modi Paulomi S. , Strom Anders , Gustafsson Jan-Ake
Published: April 2023

TNBC has the worst prognosis among all breast cancers, and survival in patients with recurrence is rarely beyond 12 months due to acquired resistance to chemotherapy, which is the standard of care for these patients. Our hypothesis is that Estrogen Receptor Beta 1 (ERB1) increases response to chemotherapy but is opposed by ERB4, which it preferentially dimerizes with. The role of ERB1 and ERB4 in influencing chemotherapy sensitivity has never been studied before. CAS9/CRSPR was used to truncate ERB1 Ligand Binding Domain (LBD) and knock down the exon unique to ERB4. We show that the truncated ERB1 LBD in a variety of mutant p53 TNBC cell lines, where ERB1 ligand dependent function was inactivated, had increased resistance to Paclitaxel, whereas the ERB4 knockdown cell line was sensitized to Paclitaxel. We further show that ERB1 LBD truncation as well as treatment with ERB1 antagonist PHTPP leads to increase in the drug efflux transporters. Hypoxia Inducible Factors (HIFs) activate factors involved in pluripotency and regulate the stem cell phenotype, both in normal and cancer cells. Here we show that the ERβ1 and ERB4 regulate these stem cell markers like SOX2, OCT4, and Nanog in an opposing manner; and we further show that this regulation is mediated by HIF. We show the upregulation of cancer cell stemness due to ERB1 LBD truncation is attenuated when HIF1/2a is knocked down by siRNA. Finally we show, an increase in the frequency of breast cancer stem cell population due to ERB1 antagonist using both ALDEFLUOR and SORE6 reporter in SUM159 and MDA-MB-231 cell lines. Since most TNBC cancers are ERβ4 positive, while only a small proportion of TNBC patients are ERβ1 positive, we believe that simultaneous activation of ERß1 with agonists and inactivation of ERB4, in combination with paclitaxel, can be more efficacious and yield better outcome for chemo resistant TNBC patients.

Citation Format: Ayesha Bano, Jessica Stevens, Paulomi S. Modi, Anders Strom, Jan-Ake Gustafsson. Estrogen receptor β4 regulates chemotherapy resistance and induces cancer stem cells in triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3072.
Abstract 3184: Single-cell analysis of CAR T-cells helps uncover distinct functional signatures following CAR T-cell manufacturing
Authors: Colina Alfredo Sebastian , Shah Ravi , Kozlik Tanya , Dwinell Michael B. , Johnson Bryon , Shah Nirav , Zamora Anthony
Published: April 2023

Chimeric antigen receptor T cell (CAR-T) therapy has proven a major success for patients with B cell malignancies. Despite recent advancements, some patients never mount an effective antitumor response and ultimately succumb to their disease. While modifications to specific CAR construct domains have resulted in greater CAR-T expansion and functional responses in some patients, it has become increasingly evident that more studies are needed to evaluate the impact of intra- and inter-patient functional heterogeneity on clinical efficacy. To begin deciphering the factors governing these disparate functionalities, we performed single-cell polyfunctional assessments of bispecific CD20/CD19 CAR-T cells and accounted for how differences in their manufacturing could further alter functional capacity. Based on our preliminary data, we hypothesized that differences in CAR-T’s underlying biology, such as their phenotypic, transcriptional, and functional status, mediate their persistence, cytotoxicity, and clinical efficacy. By taking advantage of BLI’s Lightning platform, which interrogates and links CAR-T’s cytotoxic function, secretory profile, and transcriptional state at single-cell resolution, we uncovered key manufacturing processes that alter CAR-T cell antitumor function. We’ve also identified distinct immune signatures associated with clinical response. Future studies will utilize in vivo mouse tumor models to validate the immunological signatures driving CAR-T cell functionality and persistence after infusion. Our studies suggest that differences in CAR T-cell manufacturing protocols alter the immunological signatures of T cells, which can be exploited to guide future therapeutic interventions and the design of future clinical trials.

Citation Format: Alfredo Sebastian Colina, Ravi Shah, Tanya Kozlik, Michael B. Dwinell, Bryon Johnson, Nirav Shah, Anthony Zamora. Single-cell analysis of CAR T-cells helps uncover distinct functional signatures following CAR T-cell manufacturing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3184.