bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Tumor-Associated Macrophages Can Contribute to Antitumor Activity through FcγR-Mediated Processing of Antibody–Drug Conjugates
Authors: Li Fu , Ulrich Michelle , Jonas Mechthild , Stone Ivan J. , Linares Germein , Zhang Xinqun , Westendorf Lori , Benjamin Dennis R. , Law Che-Leung
Published: April 2023

Abstract
The primary mechanism of antibody–drug conjugates (ADC) is targeted delivery of a cytotoxic payload to tumor cells via cancer-associated membrane receptors. However, the tumor microenvironment likely plays a role in ADC penetration, distribution, and processing and thus impacts the overall antitumor activity. Here, we report on the potential contribution of Fc–FcγR interactions between ADCs and tumor-associated macrophages (TAM) to the preclinical antitumor activities of ADCs. In the CD30+ L-428 Hodgkin lymphoma model, anti-CD30-vcMMAE and a non-binding control (hIgG-vcMMAE) demonstrated similar antitumor activity as well as similar payload release in the tumors. IHC analysis revealed L-428 tumors contained highly abundant TAMs, which were confirmed to bind ADCs by IHC and flow cytometry. The infiltration of TAMs was further found to correlate with the antitumor activity of the non-binding hIgG-vcMMAE in five additional xenograft models. hIgG1V1-vcMMAE, bearing a mutation in the Fc region which ablates Fc gamma receptor (FcγR) binding, lost antitumor activity in three TAM-high xenograft models, suggesting Fc–FcγR interactions modulate the TAM-ADC interaction. Our results suggest that TAMs can contribute to ADC processing through FcγR interaction in preclinical tumor models and may represent an important additional mechanism for drug release from ADCs. Correlative studies in clinical trials will further shed light on whether TAMs play a role in patients' response to ADC therapies. Mol Cancer Ther; 16(7); 1347–54. ©2017 AACR.
Data from Anti-PSMA/CD3 Bispecific Antibody Delivery and Antitumor Activity Using a Polymeric Depot Formulation
Authors: Leconet Wilhem , Liu He , Guo Ming , Le Lamer-Déchamps Sophie , Molinier Charlotte , Kim Sae , Vrlinic Tjasa , Oster Murielle , Liu Fang , Navarro Vicente , Batra Jaspreet S. , Noriega Adolfo Lopez , Grizot Sylvestre , Bander Neil H.
Published: April 2023

Abstract
Small therapeutic proteins represent a promising novel approach to treat cancer. Nevertheless, their clinical application is often adversely impacted by their short plasma half-life. Controlled long-term delivery of small biologicals has become a challenge because of their hydrophilic properties and in some cases their limited stability. Here, an in situ forming depot-injectable polymeric system was used to deliver BiJ591, a bispecific T-cell engager (BiTE) targeting both prostate-specific membrane antigen (PSMA) and the CD3 T-cell receptor in prostate cancer. BiJ591 induced T-cell activation, prostate cancer–directed cell lysis, and tumor growth inhibition. The use of diblock (DB) and triblock (TB) biodegradable polyethylene glycol–poly(lactic acid; PEG-PLA) copolymers solubilized in tripropionin, a small-chain triglyceride, allowed maintenance of BiJ591 stability and functionality in the formed depot and controlled its release. In mice, after a single subcutaneous injection, one of the polymeric candidates, TB1/DB4, provided the most sustained release of BiJ591 for up to 21 days. Moreover, the use of BiJ591-TB1/DB4 formulation in prostate cancer xenograft models showed significant therapeutic activity in both low and high PSMA–expressing tumors, whereas daily intravenous administration of BiJ591 was less efficient. Collectively, these data provide new insights into the development of controlled delivery of small therapeutic proteins in cancer. Mol Cancer Ther; 17(9); 1927–40. ©2018 AACR.
Data from Mechanisms of NT5C2-Mediated Thiopurine Resistance in Acute Lymphoblastic Leukemia
Authors: Moriyama Takaya , Liu Shuguang , Li Jing , Meyer Julia , Zhao Xujie , Yang Wentao , Shao Youming , Heath Richard , Hnízda Aleš , Carroll William L. , Yang Jun J.
Published: April 2023

Abstract
Relapse remains a formidable challenge for acute lymphoblastic leukemia (ALL). Recently, recurrent mutations in NT5C2 were identified as a common genomic lesion unique in relapsed ALL and were linked to acquired thiopurine resistance. However, molecular mechanisms by which NT5C2 regulates thiopurine cytotoxicity were incompletely understood. To this end, we sought to comprehensively characterize the biochemical and cellular effects of NT5C2 mutations. Compared with wild-type NT5C2, mutant proteins showed elevated 5′-nucleotidase activity with a stark preference of thiopurine metabolites over endogenous purine nucleotides, suggesting neomorphic effects specific to thiopurine metabolism. Expression of mutant NT5C2 mutations also significantly reduced thiopurine uptake in vitro with concomitant increase in efflux of 6-mercaptopurine (MP) metabolites, plausibly via indirect effects on drug transporter pathways. Finally, intracellular metabolomic profiling revealed significant shifts in nucleotide homeostasis induced by mutant NT5C2 at baseline; MP treatment also resulted in global changes in metabolomic profiles with completely divergent effects in cells with mutant versus wild-type NT5C2. Collectively, our data indicated that NT5C2 mutations alter thiopurine metabolism and cellular disposition, but also influence endogenous nucleotide homeostasis and thiopurine-induced metabolomic response. These complex mechanisms contributed to NT5C2-mediated drug resistance in ALL and pointed to potential opportunities for therapeutic targeting in relapsed ALL.
Data from Glibenclamide Targets Sulfonylurea Receptor 1 to Inhibit p70S6K Activity and Upregulate KLF4 Expression to Suppress Non-Small Cell Lung Carcinoma
Authors: Xu Kexin , Sun Geng , Li Min , Chen Hongling , Zhang Zuhao , Qian Xixi , Li Ping , Xu Lin , Huang Wenbin , Wang Xuerong
Published: April 2023

Abstract
Sulfonylurea receptor 1 (SUR1) is the regulatory subunit of ATP-sensitive potassium channels (KATP channels) and the receptor of antidiabetic drugs, such as glibenclamide, which induce insulin secretion in pancreatic β cells. However, the expression and role of SUR1 in cancer are unknown. In this study, we found that SUR1 expression was elevated in human non–small cell lung carcinoma (NSCLC) tissues and cell lines. SUR1 silencing suppressed the growth of NSCLC cells, while SUR1 overexpression promoted cell growth. Targeting SUR1 with glibenclamide suppressed cell growth, cell-cycle progression, epithelial–mesenchymal transition (EMT), and cell migration. Moreover, SUR1 directly interacted with p70S6K and upregulated p70S6K phosphorylation and activity. In addition, glibenclamide inhibited p70S6K, and overexpression of p70S6K partially reversed the growth-inhibiting effect of glibenclamide. Furthermore, glibenclamide upregulated the expression of the tumor suppressor Krüppel-like factor 4 (KLF4), and silencing KLF4 partially reversed the inhibitory effect of glibenclamide on cell growth, EMT, and migration. We found that SUR1 targeted p70S6K to downregulate KLF4 expression by enhancing DNA-methyltransferase 1–mediated methylation of the KLF4 promoter. Finally, in xenograft mouse models, SUR1 expression silencing or glibenclamide treatment inhibited the growth of A549 tumors, downregulated p70S6K activity, and upregulated KLF4 expression. These findings suggested that SUR1 expression was elevated in some NSCLC tissues and functioned as a tumor enhancer. Targeting SUR1 with glibenclamide inhibited NSCLC through downregulation of p70S6K activity and subsequent upregulation of the expression of the tumor suppressor gene KLF4. SUR1 can be developed as a new target for cancer therapy and glibenclamide has potential anticancer effects.
Data from MET Copy Number Gain Is Associated with Gefitinib Resistance in Leptomeningeal Carcinomatosis of EGFR-mutant Lung Cancer
Authors: Nanjo Shigeki , Arai Sachiko , Wang Wei , Takeuchi Shinji , Yamada Tadaaki , Hata Akito , Katakami Nobuyuki , Okada Yasunori , Yano Seiji
Published: April 2023

Abstract
Leptomeningeal carcinomatosis occurs frequently in EGFR-mutant lung cancer, and develops acquired resistance to EGFR tyrosine kinase inhibitors (EGFR-TKIs). This study aimed to clarify the mechanism of EGFR-TKI resistance in leptomeningeal carcinomatosis and seek for a novel therapeutic strategy. We examined EGFR mutations, including the T790M gatekeeper mutation, in 32 re-biopsy specimens from 12 leptomeningeal carcinomatosis and 20 extracranial lesions of EGFR-mutant lung cancer patients who became refractory to EGFR-TKI treatment. All the 32 specimens had the same baseline EGFR mutations, but the T790M mutation was less frequent in leptomeningeal carcinomatosis specimens than in extracranial specimens (8% vs. 55%, P < 0.01). To study molecular mechanisms of acquired EGFR-TKI resistance in leptomeningeal carcinomatosis, we utilized our previously developed mouse model of leptomeningeal carcinomatosis with the EGFR-mutant lung cancer cell line PC-9/ffluc cells, in which acquired resistance to gefitinib was induced by continuous oral treatment. Compared with subcutaneously inoculated gefitinib-resistant tumors, the T790M mutation was less frequent in leptomeningeal carcinomatosis that acquired resistance to gefitinib. PC-9/LMC-GR cells were established from the gefitinib-resistant leptomeningeal carcinomatosis model, and they were found to be intermediately resistant to gefitinib and osimertinib (third-generation EGFR-TKI). Although EGFR-T790M was negative, gefitinib resistance of PC-9/LMC-GR cells was related to MET copy number gain with MET activation. Moreover, combined use of EGFR-TKI and crizotinib, a MET inhibitor, dramatically regressed leptomeningeal carcinomatosis with acquired resistance to gefitinib or osimertinib. These findings suggest that combination therapy with MET inhibitors may be promising for controlling leptomeningeal carcinomatosis that acquires resistance to EGFR-TKIs. Mol Cancer Ther; 16(3); 506–15. ©2017 AACR.
Abstract 1065: Cancer-specific molecular subtypes of cholangiocarcinoma reveal unique molecular dependency with clinical relevance
Authors: Kwon Woo Young , LEE Sung Hwan
Published: April 2023

Introduction: Cholangiocarcinoma is a heterogeneous tumor entity harboring distinct tumor biology and clinical phenotype according to their unique tumor microenvironment (TME), especially anatomic location in terms of intrahepatic, peri-hilar, and distal extrahepatic portion. Therefore, there is a clear unmet need for the precision strategy based on cancer-specific genomic features rather than previous molecular subtypes from bulk RNA sequencing.

Methods: Comprehensive analyses were performed using the dataset from the cancer dependency map (DepMap) project, including cancer-specific molecular characterization with multi-omics data, genome-wide loss-of-function screening with CRISPR-Cas9 system, and drug sensitivity with compound screening to uncover cancer-specific molecular subtypes showing clinical relevance. The cancer-specific molecular signatures were validated in independent translational cohorts (TCGA-CHOL; n=45, ICGC-BTC; n=181, GSE132305; n=220).

Results: Integrative profiling of transcriptome from bile duct cancer cell lines (n=39) from the Cancer Cell Line Encyclopedia (CCLE) revealed a total of three cancer-specific molecular subtypes showing distinct tumor biology through all omics layer as well as clinical relevance with prognostic significance. Major molecular features of each subtype were reproducible in the validation cohorts. Subtype-specific molecular biomarkers, including mutational signature and metabolites, were identified. Finally, the target drug with subtype-specific genetic dependency was analyzed to provide precision strategy according to distinct subtypes’ molecular characterization.

Conclusions: Comprehensive analysis using a multi-omics dataset revealed precision strategies based on cancer-specific molecular subtypes of cholangiocarcinoma in terms of tumor classification and discriminative therapeutic chances. Prospective translational studies companion with clinical trials based on cancer-specific molecular subtypes is mandatory to establish the precision strategy for managing cholangiocarcinoma.

Citation Format: Woo Young Kwon, Sung Hwan LEE. Cancer-specific molecular subtypes of cholangiocarcinoma reveal unique molecular dependency with clinical relevance [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 1065.
Abstract 1076: Targeting EGFR and PI3K pathways with KRASG12D inhibitor overcomes oncogenic dependence in pancreatic cancer
Authors: Bhavar Prashant Kashinath , Surampudi Uday Kumar , Gandham Adilakshmi , Sarma Partha Pratim
Published: April 2023

In the U.S., pancreatic cancer is 9th most commonly diagnosed cancer (depending on gender), but the fourth leading cause of cancer death in men and women. FOLFIRINOX or gemcitabine plus nab-paclitaxel have been the standard of care for pancreatic cancer, but late stage prognosis often leads to disease progression of disease, with an overall 5 year survival of 11.5%, in the US, and 9% in Japan. KRASG12D mutant isoform is found in 36% of pancreatic adenocarcinoma patients and hence targeting this major mutation that appear early in the disease may help overcome cancer progression. Several signaling pathways are known to converge on the MAPK and PI3K pathways as effectors of cellular response within the cell. Pancreatic cancer is driven by activating mutations in KRAS, which in turn is expected to activate PI3K signalling through the p110α subunit, indicating that a large proportion of patients could benefit from effective targeting of this pathway. Role of EGFR on the other hand in KRAS-dependent pancreatic cancer progression is not well understood, and cetuximab as single agent has not provided benefit in PDAC patients. Recent studies of KRASG12C inhibitors in combination with EGFR inhibitors have shown clinical benefit in colorectal cancer possibly due to enhanced inhibition of KRAS-dependent signaling or overcoming adaptive feedback mechanisms. This opens an opportunity to investigate combinations of KRASG12D inhibitors with other inhibitors of MAPK and PI3K pathways. Here we report a data for 40-point combination dose response matrix combining VRTX153, a novel and highly selective inhibitor of KRASG12D with Alpelisib (a p110α inhibitor) and an EGFR inhibitor using MTT assay. Effect on downstream markers such as pERK, pEGFR and pAKT is being investigated. In vivo pharmacological effect of these agents will be investigated in the due course of development.

Citation Format: Prashant Kashinath Bhavar, Uday Kumar Surampudi, Adilakshmi Gandham, Partha Pratim Sarma. Targeting EGFR and PI3K pathways with KRASG12D inhibitor overcomes oncogenic dependence in pancreatic 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 1076.
Abstract 1132: Exploiting MGMT loss using a new class of DNA modifiers which selectively target tumor DNA and overcome therapy resistance mechanisms across multiple tumor types
Authors: Ruggeri Bruce , Tarantino Kyle , Sundaram Ranjini , Lin Kingson , Herzon Seth , Park Joseph , Johnson Spenser , Gueble Susan , Bindra Ranjit S.
Published: April 2023

DNA damage response (DDR) defects are common in cancer, and it is well established that these tumor-associated DNA repair vulnerabilities can be exploited for a therapeutic gain. Small molecule inhibitors of DDR proteins have been developed to target these cancers, which are now FDA-approved and/or currently being tested in clinical trials. Our team recently reported on an entirely new approach to exploit DDR defects in cancer, via DNA modification, eliminating the need to target proteins (Lin et al., Science 2022). In this recent study, we demonstrated that our new class of “DNA modifiers (DMs)” can be engineered to exploit loss of O6-methylguanine methyltransferase (MGMT), which leads to robust and selective killing of MGMT-deficient glioma tumors, both in vivo and in vitro. We also found that our approach could overcome acquired and intrinsic defects in another DDR pathway, mismatch repair (MMR), which is a known resistance mechanism to standard-of-care cytotoxic therapies in glioma, including TMZ.

Here, we sought to determine the extent to which our approach can be expanded to selectively target a wide range of cancers beyond glioma in which loss of MGMT, as well as acquired and intrinsic MMR defect, are commonly seen at various frequencies. To this end, we profiled over 900 cell line models, as well as several collections of patient-derived cultures derived from several tumor types for sensitivity to our DMs based on MGMT and MMR status in vitro. We then validated our findings in cell line xenograft and patient-derived xenograft models, in vivo. In parallel, we compared the activity of DMs to commonly used, standard-of-care cytotoxic drugs, in vitro and in vivo. Our data indicate that DMs can effectively target MGMT loss, as well as intrinsic and acquired MMR defects, in multiple non-CNS tumor types. Taken together, the findings reported here support an entirely new approach to selectively exploit DDR defects via direct DNA modification. This new approach has the potential to overcome known resistance mechanisms to currently used small molecule inhibitors of DDR proteins, across a broad range of cancers.

Citation Format: Bruce Ruggeri, Kyle Tarantino, Ranjini Sundaram, Kingson Lin, Seth Herzon, Joseph Park, Spenser Johnson, Susan Gueble, Ranjit S. Bindra. Exploiting MGMT loss using a new class of DNA modifiers which selectively target tumor DNA and overcome therapy resistance mechanisms across multiple tumor types [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 1132.
Abstract 1170: Scalable co-sequencing of RNA and DNA from individual nuclei
Authors: Olsen Timothy R. , Talla Pranay , Furnari Julia , Zha Shan , Canoll Peter , Bruce Jeffrey , Sims Peter
Published: April 2023

A scalable co-assay for single cell mRNA and gDNA sequencing could potentially harmonize the identification of rare cell populations and cell states with corresponding cell-lineage relationships, drug sensitivities, and genetic mutations. Such an assay would provide new insights into the relationship between genetic alterations and tumor cell phenotype. However, simultaneously measuring the genomic and transcriptomic content of individual cells at high throughput remains challenging. Here we present single-cell lithium assisted nucleosome depleted sequencing (scLAND-seq), a scalable method for co-sequencing RNA and DNA from single nuclei that uses the widely available 10x Genomics microfluidics platform. In scLAND-seq, nuclei are treated with a mild detergent, lithium diiodosalicylate, which denatures protein and renders gDNA free of nucleosome packing while preserving the nuclear membrane. The nuclei are then tagmented with Tn5 transposase, which randomly fragments gDNA and integrates universal sequencing adapters. Tagmented nuclei are loaded into a microfluidic droplet generator which co-encapsulates nuclei with beads containing barcoded primers for reverse transcription of mRNA and replication of gDNA. Ultimately two libraries are created: one for nuclear mRNA and one for gDNA. Each library contains barcodes linking it to its nuclei of origin, thus allowing simultaneous analysis of the transcriptome and genome of each nucleus. Once nuclei have been depleted of nucleosomes, all steps can be performed using a 10x Genomics Chromium Controller and 10x Multiome Kit without further experimental modification. We characterize LAND-seq with BJ fibroblasts, a human diploid cell line, and demonstrate the detection of rare subpopulations with copy number variants (CNVs) associated with specific transcriptional clusters. Analysis of transcription start site enrichment indicates near-complete disruption of chromatin structure, resulting in high-complexity libraries that could not be sequenced to saturation even with >1M reads/cell. The corresponding gene expression library is of high-quality, containing ~20,000 gene counts per nuclei. Finally, to demonstrate scLAND-seq’s compatibility with primary cells, we applied scLAND-seq to cryopreserved, human glioblastoma (GBM) surgical specimens, enabling direct investigation into the relationship between phenotype and genotype in thousands of individual cells.

Character limit: 2,600

Citation Format: Timothy R. Olsen, Pranay Talla, Julia Furnari, Shan Zha, Peter Canoll, Jeffrey Bruce, Peter Sims. Scalable co-sequencing of RNA and DNA from individual nuclei [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 1170.
Abstract 1190: Mimicking CAFs, pial cells support leptomeningeal metastasis
Authors: Snyder Jenna , Remsik Jan , Prescott Nicholas , Boire Adrienne
Published: April 2023

Leptomeningeal metastasis (LM), the spread of cancer into cerebrospinal fluid (CSF)-filled coverings of the brain and spinal cord, is a dismal complication of cancer leading to rapid neurologic disability and death. While a diverse array of primary tumors may result in LM, recent work from our laboratory (Chi Y et al 2020 Science) demonstrates a conserved transcriptional signature, independent of primary tumor, suggesting that tumor-microenvironmental interactions play a dominant role in this pathology. We have proteomically interrogated CSF collected from breast (n = 45), lung (n = 30), and melanoma (n = 27) patients with and without LM. We detect robust expression of inflammatory cytokines in all LM+ samples, 15 of these cytokines including IL8, IL6, and CXCL1 are conserved across tumor types in the presence of LM. Because cancer cells in the CSF do not express mRNA associated with these cytokines, we hypothesized that these inflammatory cytokines may originate from the microenvironment. Within the leptomeninges, LM cells adhere to and interact with the innermost layer of the meninges, the pia mater, which contains fibroblast-like cells. Co-culture of cancer cells with pial cells induced changes in the pial cell transcriptome and secretome, leading to increased IL8, IL6, and CXCL1 production. This communication between cancer cells and pial cells is preserved in the absence of direct cell-cell contact. To identify cancer cell signal(s) responsible for these pial cell changes, we subjected cancer cell conditioned media to gel filtration, and uncovered TGFBI as a key secreted factor inducing pial cell transcriptional changes. These pial cells support cancer cell growth in vivo, as measured by co-implantation experiments. In autopsy samples, we detect IL8, IL6, and CXCL1 expression in the pia of patients harboring LM. Taken together, these observations support a cancer-associated-fibroblast like role for pial cells in the setting of LM. We are currently carrying out single cell RNA sequencing of both human and mouse LM samples to capture heterogeneity in both cancer and pial cell responses, and to place these cells in the larger context of the CAF knowledgebase. Our work extends the CAF phenotype beyond classical fibroblasts to include pial cells, suggesting novel therapeutic opportunities within CNS malignancies.

Citation Format: Jenna Snyder, Jan Remsik, Nicholas Prescott, Adrienne Boire. Mimicking CAFs, pial cells support leptomeningeal metastasis [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 1190.
Abstract 1218: Mercury(II)-induced abnormalities in processing of topoisomerase I-mediated DNA damage
Authors: Ursu Giorgiana Madalina , Cyran Anna M. , Zhitkovich Anatoly
Published: April 2023

Mercury (Hg) is a persistent environmental pollutant that has long been associated with neurotoxic and other noncancer health effects in exposed populations. Recent large epidemiological studies using different internal indices of Hg exposure have found significantly elevated risks for skin cancers in both men and women. To understand potential carcinogenic mechanisms for Hg(II), we examined its mutagenicity and impact on processing of different types of DNA damage. In agreement with its lack of DNA reactivity, we found that Hg(II) was not mutagenic at the mammalian Hprt locus and did not change DNA incorporation of rNMPs which are the most abundant endogenous nucleotide abnormalities in the genome. We next tested the impact of Hg(II) at nontoxic or only mildly toxic concentrations on sensitivity of cells to several genotoxicants producing specific classes of DNA damage. We found that even nontoxic concentrations of Hg(II) impaired cellular resistance to topoisomerase I-mediated DNA damage in human epidermal keratinocytes. Hg(II) had no significant effect on cytotoxicity of topoisomerase II-mediated DNA lesions and examination of numerous components of DNA damage response activated by topoisomerase II-targeting drugs showed no major differences between control and Hg(II)-exposed cells. In contrast, Hg(II)-treated keratinocytes displayed a distorted DNA damage response to topoisomerase I-produced DNA damage, manifested in a misbalanced activation of ATM and ATR targets. Consistent with their lower survival, Hg(II)-treated cells accumulated higher levels of toxic DNA double-strand breaks (DSBs). Further analyses indicated that Hg(II) caused impartment of DNA repair of replication-associated DSBs. Thus, although Hg(II) is not directly genotoxic, it can increase genomic instability by interfering with accurate processing of endogenous DNA damage produced by topoisomerase I.

Citation Format: Giorgiana Madalina Ursu, Anna M. Cyran, Anatoly Zhitkovich. Mercury(II)-induced abnormalities in processing of topoisomerase I-mediated DNA damage [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 1218.
Abstract 1227: Esophageal cancer shows spontaneous plasticity between epithelial and mesenchymal status
Authors: Tsuchihashi Kenji , Hirata Yuki , Yamasaki Juntaro , Suina Kentaro , Tanoue Kenro , Yae Toshifumi , Masuda Kenta , Baba Eishi , Akashi Koichi , Kitagawa Yuko , Saya Hideyuki , Nagano Osamu
Published: April 2023

Epithelial-mesenchymal transition (EMT) is one of essential processes in cancer progression. Plasticity between epithelial and mesenchymal status is referred as epithelial-mesenchymal plasticity (EMP). The significance of spontaneous change of EMP is under investigation in the field of EMT. To date, spontaneous EMP is reported in breast and prostate cancer, which leads to the acquisition of stem-cell properties and chemoresistance. On the other hand, the presence of spontaneous EMP is still not examined in other cancer including esophageal cancer. In the present study, we tackled the question of whether esophageal cancer have the potency of spontaneous EMP. We screened eleven esophageal squamous cancer cell (ESCC) cell lines with CD44 isoform expression, which is known to reflect epithelial and mesenchymal status. Interestingly, KYSE520 was found to comprise heterogenous populations consisting of CD44v+ and CD44v- subpopulations. CD44v+ and CD44v- cells showed the expression of epithelial and mesenchymal markers, respectively. Single-cell sorting of CD44v+ and CD44v- cells revealed both cells gave rise to cell populations consisting of CD44v+ and CD44v- cells, indicating CD44v+ epithelial-like and CD44v- mesenchymal-like cells can generate counterparts, respectively. This EMP was maintained in next generation. Further, we found the generation of counterparts between CD44v+ and CD44v- cells in vivo. Epithelial splicing regulatory protein 1 (ESRP1) is reported as the regulator of CD44 mRNA splicing. The role of ESRP1 in EMP is not well understood in esophageal cancer. The inhibition of ESRP1 shifted CD44v+ to CD44v- cells in KYSE520. However, the expression of epithelial-mesenchymal transition (EMT)-related markers or transcriptional factors were almost not affected, suggesting ESRP1 functions downstream of EMP. Our results firstly revealed the presence of spontaneous EMP in esophageal cancer and give a new insight in the field of EMT.

Citation Format: Kenji Tsuchihashi, Yuki Hirata, Juntaro Yamasaki, Kentaro Suina, Kenro Tanoue, Toshifumi Yae, Kenta Masuda, Eishi Baba, Koichi Akashi, Yuko Kitagawa, Hideyuki Saya, Osamu Nagano. Esophageal cancer shows spontaneous plasticity between epithelial and mesenchymal status [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 1227.
Abstract 126: Extreme selection constrains ability of EGFR-driven lung adenocarcinoma to diversify in response to erlotinib therapy
Authors: Fisk J Nicholas , Gaffney Stephen , Politi Katerina , Gettinger Scott , de Miguel Fernando , Townsend Jeffrey
Published: April 2023

The acquisition of resistance by tumors to targeted molecular therapies remains one of the greatest challenges in precision medicine. Evolutionary techniques, such as phylogenetic analysis and the thoughtful calculation of selection intensities, are well suited to offer guiding insights in describing and, ultimately, overcoming therapeutic resistance.

Here, we quantify the relative importance of mutations occurring during EGFR targeted therapy on a cohort of Lung Adenocarcinoma (LUAD) patients using clinical records to calibrate phylogenetic chronograms to identify variants occurring on branches of the tumor phylogeny coinciding with targeted EGFR therapy and calculate the evolutionarily-derived cancer effect size for those variants. We then compare these results to other cancer contexts to reveal the strikingly high effect size of the EGFR T790M mutation. To probe and quantify the constraints therapeutically driven selection pressure places on the cohort, we predict the neoantigen composition for each patient for both extant and ancestral tumor states, using a general allele frequency selection model to calculate selection on cancer variants constituting putative neoantigens. We then compare the proportion of negatively and positively selected neoantigens to non-neoantigen variants within the tumor to deduce if neoantigens are subject to additional immune scrutiny in a background of therapeutics. Lastly, we perform ancestral state reconstruction and mutational signature analysis to provide further insight on mutational processes on both individual patient and cohort levels, detailing how these endogenous and exogenous processes shift between etiologically pertinent time-points. Taken together, our results detail the degree of selection on cancer undergoing targeted therapy and highlights unique shifts in mutational processes in response to such therapies, ultimately offering quantitative insights that can inform clinical decision making regarding therapeutic strategies in Lung Adenocarcinoma and provide a framework for analyzing similar cancer systems.

Citation Format: J Nicholas Fisk, Stephen Gaffney, Katerina Politi, Scott Gettinger, Fernando de Miguel, Jeffrey Townsend. Extreme selection constrains ability of EGFR-driven lung adenocarcinoma to diversify in response to erlotinib therapy [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 126.
Abstract 1293: GPRC5A drives an implantation-like phenotype in early-stage lung disseminated cancer cells of esophageal squamous carcinoma
Authors: Zhou Hongyu , Liu Beilei , Wong Ching Ngar , Guan Xinyuan
Published: April 2023

Esophageal squamous carcinoma (ESCC) is one of the leading malignancies worldwide, especially in Asian countries, largely threatening human life. The 5-year survival rate of ESCC is less than 20%, and patients with distant metastases have dismal survival with <5% of five-year survival rate. Traditional therapies e.g., surgery, chemotherapy, radiotherapy demonstrate poor clinical outcomes for ESCC patients. Metastases are commonly seen in ESCC patients, and lung metastases approximately account for 5% of all ESCC metastases. The underlying mechanism of ESCC lung metastasis remains unsettled. In our study, we established ESCC lung metastasis mice model via intravenous injection of ESCC cells in multiple timepoints. Lung disseminated cancer cell (DCCs) of different timepoints were isolated and thrown into single cell RNA sequencing (scRNA-seq). Via bioinformatic analysis, we found the upregulated genes in early-stage lung DCCs were highly enriched in embryo development, placenta development, pre-implantation embryos processes, neither in late-stage lung DCCs. It highly suggested that ESCC lung metastasis process might be similar to embryo implantation. Based on above findings, 5 intersected genes (GPRC5A, FSTL3, WNT7A, TM4SF1, NT5E) of early-stage lung DCCs and embryo implantation were figured out to construct an implantation-like signature. Enrichment analysis results implied that ESCCs subgroups with implantation-like phenotype showed higher ability to migration, focal adhesion and response to stress. Preliminary data indicated that the representative gene GPRC5A was upregulated in ESCC tumors compared with normal esophageal epithelium in clinical samples and public databases. GPRC5A overexpression promoted tumor growth, invasion, migration and sphere formation both in vivo and in vitro, as the same as other two signature genes, WNT7A and FSTL3. Moreover, GPRC5A-overexpresssing ESCC cells displayed high ability to develop resistance to stress and generated less reactive oxygen species (ROS) in vitro. In vivo, GPRC5A overexpression increased the quantalities of early-stage lung DCCs in ESCC lung metastasis mice model and disruption of GPRC5A expression abolished its pro-metastatic effect. Together, our preliminary findings identify an implantation-like signature in early-stage ESCC lung metastasis and disrupting this phenotype helps to prevent ESCC tumorigenesis and metastasis.

Citation Format: Hongyu Zhou, Beilei Liu, Ching Ngar Wong, Xinyuan Guan. GPRC5A drives an implantation-like phenotype in early-stage lung disseminated cancer cells of esophageal squamous carcinoma [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 1293.
Abstract 1319: Saffron (Crocus sativus) extract inhibits migration and invasion of cancer cells through modulation of MMP-3 and MMP-9 expression
Authors: King Tyler , Moscardini Stefana , Martin McKenzie , Rehman Fatima K.
Published: April 2023

Saffron is a spice that has been used throughout history for various therapeutic properties. Recently, it has also been documented to have a significant anticancer effect on primary tumors. However, limited studies are found studying its effect on the metastatic potential of cancers. The goal of our study was to explore the effect of saffron on the migration and metastatic potential of breast cancer cells.

Methods: MDA468 breast carcinoma cells were treated with varying concentrations of Saffron (100-1000 ug/mL) and examined using Q-phase holographic time-lapse microscopy to measure the movement of cells over time. The anticancer activity of Saffron was confirmed using cell viability assays. Invasion and metastatic potential were studied using Boyden-chamber and scratch wound assays. Finally, the tumorigenicity of the cell line was examined through colony formation assays in the presence and absence of Saffron.

Results: Q-phase microscopy revealed a reduction in projections and significantly reduced movements in cells after Saffron treatment. The inhibition of migration, cell mobility and invasion by Saffron was confirmed through scratch wound and Boyden chamber assays (p<.01). Saffron extract also resulted in inhibition of viability in a dose-dependent manner and colony formation assays found markedly reduced size and number of colonies in the presence of the extract. Western blot results showed marked reduction of MMP-3 and MMP-9 expression in Saffron-treated cells.

Conclusions: Our results found saffron to be effective in decreasing breast cancer carcinoma cells MDA468’s viability, migration, and tumorigenicity. Initial results indicate that Saffron may reduce migratory ability of cancer cells through the modulation of metalloproteinases.

Citation Format: Tyler King, Stefana Moscardini, McKenzie Martin, Fatima K. Rehman. Saffron (Crocus sativus) extract inhibits migration and invasion of cancer cells through modulation of MMP-3 and MMP-9 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 1319.
Abstract 1352: SOX17 plays a critical role in immune evasion of colorectal cancer
Authors: Goto Norihiro , Goto Saori , Westcott Peter , Imada Shinya , Agudo Judith , Yilmaz Omer
Published: April 2023

Colorectal cancers (CRCs) are a leading cause of cancer-related death. Most CRCs are immune cold, and do not respond to checkpoint blockade therapy except for microsatellite-high CRCs harboring high mutational burdens. Deciphering the mechanism by which CRC cells evade immune surveillance has the potential to dramatically improve the prognosis of CRC patients. During tumor evolution of CRCs, epigenetic changes play critical roles in addition to accumulation of genetic mutations. However, distinct mutational patterns and patient backgrounds render it challenging to distinguish the driver epigenetic alterations from passenger epigenetic alterations induced by gene mutations or other environmental factors. Identification of the driver epigenetic alterations may provide novel mechanistic insights into colon cancer biology including how they evade immune surveillance. Here, we utilized the colon cancer organoid orthotopic transplantation approach to establish colon cancer organoids from different stages of the tumors, and performed comprehensive epigenomic and transcriptomic analyses to understand the epigenetic alterations during tumor evolution. We found that in vivo environment induces epigenetic alterations that converge on SOX17, a transcription factor that is required for endoderm development. SOX17 is re-expressed in colon cancers in vivo, but not in the in vitro organoid culture, and reprograms tumor cell fate with fetal intestinal gene expressions. Importantly, SOX17 knockout leads to tumor rejection in immunocompetent mice, but not in immunodeficient mice, by turning immune cold tumors into hot tumors with robust intratumoral infiltration of activated CD8+ T cells. Mechanistically, SOX17 directly downregulates Ifngr1 expression and mitigates MHC-I expression to evade CD8+ T cell-mediated tumor cell killing. Together, our result reveals that SOX17 is a master transcription factor that induces the in vivo epigenetic reprograming of tumors, which contributes to the immune evasion of colon cancers.

Citation Format: Norihiro Goto, Saori Goto, Peter Westcott, Shinya Imada, Judith Agudo, Omer Yilmaz. SOX17 plays a critical role in immune evasion of colorectal 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 1352.
Abstract 1499: HOXC6 gene functions as an oncogenic driver in HPV+ head and neck squamous cell carcinoma (HNSCC)
Authors: Choudhury Subhendu Roy , Mukhina Vera , Vyshkvorkina Iuliia , Gore Jessica , Anderson Alexa , Gaykalova Daria A.
Published: April 2023

Homeobox C6 (HOXC6) belongs to the member of the homeobox family, a highly conserved transcription factor that plays an essential role in morphogenesis and cellular differentiation during embryonic development. Previous studies reported that the dysregulation of HOXC6 gene expression plays a crucial role in carcinogenesis. HOXC6 is reportedly over-expressed in several types of human cancer, including breast, leukemia, and lung cancers. Moreover, higher expression of HOXC6 leads to lymph node metastasis. In addition, it has been also demonstrated that in prostate cancer cells, HOXC6 downregulation reduced cell proliferation by inducing apoptosis. Although the precise mechanisms of HOXC6 in carcinogenesis still need to be investigated. In a secondary data analysis from our previous study, we identified HOXC6 is upregulated and enriched with tumor type-specific enhancer region in HPV+HNSCC samples. Moreover, we used publicly available TCGA data consisting of 520 HNSCC samples and a JHU cohort with 47 HPV+ HNSCC samples to illustrate that HOXC6 is more highly expressed in HPV+ HNSCC than in adjacent normal samples. We further validated the HOXC6 overexpression through an in vitro study using HPV+HNSCC cell lines and patient tissue samples. Clinicopathological data from TCGA indicated that higher HOXC6 expression correlated with poor overall survival. To explore the function of HOXC6 in HNSCC, we knocked down HOXC6 in HPV+ HNSCC cells via siRNA and performed the cell proliferation assay. We observed that HOXC6 downregulation decreased the cell proliferation rate compared to control samples. In conclusion, our results suggest HOXC6 acts as an oncogenic driver in HPV+ HNSCC progression, which is potentially regulated by tumor type-specific enhancers and may serve as a potential biomarker.

Citation Format: Subhendu Roy Choudhury, Vera Mukhina, Iuliia Vyshkvorkina, Jessica Gore, Alexa Anderson, Daria A. Gaykalova. HOXC6 gene functions as an oncogenic driver in HPV+ head and neck squamous cell carcinoma (HNSCC) [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 1499.
Abstract 1548: CO-1008, an ADC program targeting leukemic progenitor and blast AML cells, but not normal hematopoetic progenitor cells
Authors: Kaushik Garima , Vishwakarma Bandana , Bell Marina , Komurov Kakajan
Published: April 2023

The overall survival of patients diagnosed with Acute Myeloid Leukemia (AML) remains low. While initial responses to therapy are favorable, the duration of response is short, and overcoming therapeutic resistance has proven difficult. The presence of chemoresistant leukemic stem cells (LSCs), among other mechanisms, has been implicated as a driver in the observed relapse. Antibody-drug conjugates (ADC) are targeted cancer therapeutics that leverage antigen specificity to deliver a potent cytotoxic payload. Recently, ADCs have shown promise in the clinic, but can also present safety and efficacy concerns. Mylotarg, an FDA approved CD33-directed ADC used to treat Acute Myeloid Leukemia, was the first ADC to be approved. The therapeutic is active against most AML tumors but can lead to severe myelotoxicity given the expression of CD33 on normal myeloid progenitor cells. We leveraged a pharmaco-pheno-multiomic analysis of primary AML tumors to uncover a previously unexplored target expressed on both LSCs and AML blasts, but not normal hematopoetic progenitor cells or other normal tissue. The target also showed capacity to rapidly internalize and has activity of well-established oncogenic kinases associated with expression. A novel monoclonal antibody (IgG1) with favorable binding and biophysical characteristics was established against this novel target and conjugated to a highly potent cytotoxic agent, creating a novel ADC. This ADC shows strong binding across AML tumors with effective internalization and highly potent killing of AML tumors.

Citation Format: Garima Kaushik, Bandana Vishwakarma, Marina Bell, Kakajan Komurov. CO-1008, an ADC program targeting leukemic progenitor and blast AML cells, but not normal hematopoetic progenitor 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 1548.
Abstract 1563: Concomitant inhibition of Aurora kinase A and WEE1 kinases results in synergistic tumor control and heightens DNA replication stress in head and neck and lung carcinomas
Authors: Lee Jong Woo , Kim Sundong , Cruz-Gomez Sebastian , Shi Jackie , Yang Cindy , Burtness Barbara
Published: April 2023

Human papillomavirus (HPV)-negative head and neck squamous cell carcinoma (HNSCC) and lung cancer mainly harbor disruptive mutations in TP53 and/or CDKN2A tumor suppressor genes, which are associated with refractoriness to treatment as well as poor prognosis and survival. Mutational loss of TP53 function fosters elevated expression of Aurora kinase A (AURKA) that plays a crucial role in mitotic progression at G2/M and stabilizes DNA replication forks at S phase. We have previously demonstrated that combined inhibition of AURKA and WEE1, a G2/M checkpoint kinase induced by replication stress, results in synergistic anti-tumor effects in HNSCC in vitro and in vivo. We have now further confirmed synergy using the highly selective AURKA inhibitor VIC1911 in HNSCC and lung cancers harboring TP53 mutations and explored mechanisms of cell death. VIC1911 and adavosertib combination synergistically suppressed cell growth and survival in both 2D- and 3D-culture systems relative to vehicle or single-agent treatment in TP53-mutated HNSCC FaDu and CAL27, and lung cancer A549 and NCI-H358 cells, with no observable toxicity in normal cells, predicting a favorable therapeutic index. Furthermore, combination treatment accelerated mitotic entry and resulted in accumulation of mitotic catastrophe, as demonstrated by time-lapse imaging, and apoptotic cell death as evidenced by cleaved PARP assays. Notably, we found that combination VIC1911 and adavosertib in HNSCC cells drastically slowed and stalled progression of replication forks in DNA fiber analysis (Mean: DMSO=3.117; adavosertib=1.495; VIC1911=0.9757; combination=0.5548 kb/min; P <0.0001) and amplified DNA damage as measured by γH2AX, indicating induction of severe replication stress. In vivo, this combination resulted in significant tumor regression in both HNSCC and lung adenocarcinoma patient-derived and cancer cell-derived xenografted mice compared with either vehicle or single-agent treatment. Taken together, these results suggest that AURKA inhibition increases dependency on WEE1 by enhancing replication stress and mitotic catastrophe, and support clinical evaluation of combined AURKA and WEE1 inhibition as a novel and effective treatment for HNSCC and lung cancer patients with elevated AURKA expression.

Citation Format: Jong Woo Lee, Sundong Kim, Sebastian Cruz-Gomez, Jackie Shi, Cindy Yang, Barbara Burtness. Concomitant inhibition of Aurora kinase A and WEE1 kinases results in synergistic tumor control and heightens DNA replication stress in head and neck and lung carcinomas [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 1563.