bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5106: The addition of radiation therapy to simultaneous targeting of PD-1 and IL2Rβγ enhances antigen-specific CD8 T cell anti-tumor immunity in pancreatic cancer models
Authors: Hoen Maureen , Piper Miles , Darragh Laurel B. , Nugyen Diemmy , Gadwa Jacob , Amann Maria , Durini Greta , Knitz Michael , Van Court Benjamin , Pham Tiffany , Kedl Ross , Umana Pablo , Codarri-Deak Laura , Klein Christian , D’Alessandro Angelo , Karam Sana D.
Published: April 2023

Although IL-2 has been shown to critically mediate NK and T cell effector functions, when used in clinical trials this cytokine has failed due to toxicity and severe side effects of the systemic treatment. Our lab has investigated the use of a next-generation IL-2 compound, murine PD1-IL2v, which consists of an anti-PD-1 antibody fused to a IL2-variant with abolished binding to IL2Rα. Previous work shows this molecule results in tumor-antigen specific T cell expansion and differentiation towards immune effectors. Here, we report for the first time that radiation therapy amplifies the anti-tumor effect observed with PD1-IL2v treatment of murine orthotopic models of local tumor growth and metastasis. This anti-tumor effect is correlated with increases in proliferative and polyfunctional CD8 T cells across tumor, lymph node, and blood compartments. Furthermore, using an OVA-antigen specific orthotopic model, we found that radiation in combination with PD1-IL2v treatment enhances infiltration and activation of antigen-specific CD8 T cells, which have been shown to be critical for immunotherapy clinical success. A portion of tumor-bearing mice treated with radiation and PD1-IL2v fully eradicated their primary tumors and tumor rechallenges, which we show correlates with increased markers of memory in CD8 T cells of these mice. To demonstrate the clinical relevance of these preclinical findings, we performed RNA sequencing on paired pancreatic adenocarcinoma (PDAC) patient tumor tissue samples prior to and following radiation. We observed enhanced IL2Rβ and IL2Rγ expression as well as IL2R signaling in PDAC-patient responders to radiation therapy compared to non-responders. In conclusion, the antigen-specific, lasting memory response to preclinical PDAC models demonstrated here provides rationale for the treatment of PDAC with a radiation and PD1-IL2v combination.

Citation Format: Maureen Hoen, Miles Piper, Laurel B. Darragh, Diemmy Nugyen, Jacob Gadwa, Maria Amann, Greta Durini, Michael Knitz, Benjamin Van Court, Tiffany Pham, Ross Kedl, Pablo Umana, Laura Codarri-Deak, Christian Klein, Angelo D’Alessandro, Sana D. Karam. The addition of radiation therapy to simultaneous targeting of PD-1 and IL2Rβγ enhances antigen-specific CD8 T cell anti-tumor immunity in pancreatic cancer models. [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 5106.
Abstract 5130: Multi-plex cytokine profiling for mice on Bleomycin induced systemic sclerosis model
Authors: Yuan Yanping , Qian Cheng , Yangmeng Chuanzhi , Fang Li , Yan Ruiqing
Published: April 2023

Systemic sclerosis (SSc) is recognized as an autoimmune disorder characterized by immune dysregulation, obliterative vasculopathy, excessive accumulation of extracellular matrix, and fibrosis of the skin and other organs. It also ranks on the highest position of disease-related mortality and morbidity with an impaired quality of life among the rheumatologic illnesses. However, the immunological mechanism related to SSc remains to be elucidated systemically, which may lay for the novel therapeutic methodology. In the present study, we combined the MSD methodology and flow cytometry to landscape the chemokine or cytokine network, immunology response and signaling axis in mice on Blomycin induced SSc model. Our results provide the reference and path for the future direction of novel therapeutic reagents discovery.

Citation Format: Yanping Yuan, Cheng Qian, Chuanzhi Yangmeng, Li Fang, Ruiqing Yan. Multi-plex cytokine profiling for mice on Bleomycin induced systemic sclerosis model. [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 5130.
Abstract 5161: Molecular mechanisms of early-stage lung adenocarcinoma initiation
Authors: Rodriguez-Fuguet Alice , Silva Luis , Asara John , Li Xiaoman , Kobayashi Susumu , Zhang Wen Cai
Published: April 2023

Lung cancer initiation and progression driven by epidermal growth factor receptor (EGFR) mutation has been extensively studied in the past decade. Though many leaps have been made in the field in terms of treatment, early diagnosis screening is not widely available or well established. Meanwhile, EGFR mutation is a significant driver of tumorigenesis and drug resistance. This receptor is related to many downstream pathways that influence cancer metabolism and the ability of tumor cells to adapt to changes in the tumor microenvironment. Our study finds that cancer metabolism plays a prominent role in the progression of tumorigenesis in early-stage lung cancer. Kynurenine, a metabolite from the tryptophan pathway, was highly upregulated in early-stage, EGFR-mutation driven lung cancer by metabolomic analysis. The connection between EGFR mutation and the upregulation of kynurenine remains largely unknown. We hypothesize that kynurenine, based on our preliminary data and as a ligand of the aryl hydrocarbon receptor, may have a dual role in early tumorigenesis. We propose that kynurenine may be used by tumor cells to modulate their response to stress by triggering anti-apoptotic pathways. Additionally, kynurenine may be used as a signaling molecule in the tumor microenvironment to allow the early tumor cells to evade immune detection through the transformation of macrophages from M1 to M2. Currently, our data show that indoleamine 2,3- dioxygenase 1 (IDO1), the rate-limiting enzyme of the tryptophan-kynurenine pathway, is significantly upregulated in early-stage, EGFR-driven lung cancer murine models by RNA-seq analysis. This supports our idea that IDO1 could help to modulate the response to cellular stress and to regulate the immune response to favor tumor development. We hope that IDO1 could be a potential clinical target and a potential biomarker for the dysregulation of this pathway.

Citation Format: Alice Rodriguez-Fuguet, Luis Silva, John Asara, Xiaoman Li, Susumu Kobayashi, Wen Cai Zhang. Molecular mechanisms of early-stage lung adenocarcinoma initiation. [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 5161.
Abstract 5165: In vivo therapeutic validation of NRT-YHD_001, a novel macrophage checkpoint inhibitor, in liver cancer
Authors: Nam Suk Woo , Kim Sangyean , Na Min Jeong , Shin Eunbi , Ha Jin Woong , Jeon Soyoung , Yoon Jeong Hwan
Published: April 2023

Previously, we demonstrated that recovery of Histone deacetylase 6 (HDAC6) elicited let-7i-5p suppression to de-repress the thrombospondin-1 (TSP1) expression, and thereby it occupied CD47 receptor to block CD47-SIRPα mediated anti-phagocytosis of macrophage in liver cancer. NRT-YHD_001 is a modified antisense miRNA of let-7i-5p for liver cancer treatment. NRT-YHD_001 was substituted with O-methoxyethyl and phosphothioate for stability, and N-acetylgalactosamine was attached for specific delivery to the liver. In in vitro phagocytosis assay, NRT-YHD_001 showed macrophage phagocytosis activity with similar or better results compared to non-modified antisense let-7i-5p (AS_let-7i-5p). In addition, it was confirmed that NRT-YHD_001 inhibited endogenous let-7i-5p of liver cancer cells and activated macrophage phagocytosis at 4°C or 25°C for up to 2 weeks. When NRT-YHD_001 activity was investigated in vivo after 1 week, 2 weeks, and 4 weeks after administration of single dose NRT-YHD_001 to the tail vein of a mouse, NRT-YHD_001 was not degraded, and let-7i-5p was inhibited while maintaining activity in liver tissue for 2 weeks after injection. To investigate the in vivo therapeutic effect, a mouse spontaneous (Ras-transgenic) liver cancer model was used and compared with the sorafenib-administered group. After administration of NRT-YHD_001 and sorafenib each week at the time when liver mass was detected, the tumor size and growth rate were compared. As a result, the NRT-YHD_001 administration group showed significant therapeutic effect compared to sorafenib. These results showed that NRT-YHD_001 had a strong anticancer effect by converting the macrophage of don't eat me into eat me signal in the tumor microenvironment of liver cancer, thereby removing or inhibiting liver cancer cells.

Keywords: let-7i-5p, macrophage, phagocytosis, liver cancer, NRT-YHD_001

Citation Format: Suk Woo Nam, Sangyean Kim, Min Jeong Na, Eunbi Shin, Jin Woong Ha, Soyoung Jeon, Jeong Hwan Yoon. In vivo therapeutic validation of NRT-YHD_001, a novel macrophage checkpoint inhibitor, in liver 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 5165.
Abstract 5181: The role of Neuropilin-1 signaling in CD4 conventional T cell immune function in non-small cell lung cancer
Authors: Kahangi Bitta P. , Lim Raymond J. , Abascal Jensen , Crosson William P. , Reyes Edgar Perez , Dumitras Camelia , Tran Linh M. , Krysan Kostyantyn , Salehi-Rad Ramin , Dubinett Steven M. , Liu Bin
Published: April 2023

Despite advances in therapy, lung cancer remains the leading cause of cancer related deaths. In non-small cell lung cancer (NSCLC), a subset of tumor-infiltrating CD4 T cells express Neuropilin-1 (NRP1), a transmembrane protein that binds to multiple ligands with diverse functions in tumor and immune cells. Recent studies reveal that NRP1 can be induced upon T cell receptor stimulation and that NRP1 signaling promotes increased stabilization and function of CD4+ FoxP3+ regulatory T cells (Tregs), while signaling on CD8 T cells results in inhibition of CD8 T cell migration and cytotoxicity; therefore, NRP1 is implicated as a newly defined immune checkpoint protein. The functional role of NRP1 in CD4+ FoxP3- T conventional cells (CD4 Tconv) and the ligands involved are largely unexplored. We have found that human lung cancer malignant pleural effusions (MPE) from a subset of metastatic lung cancer patients contain both NRP1+ and NRP1- CD4 Tconv cells, providing a relevant clinical setting to study the function of NRP1 in human T cells. Our preliminary studies reveal that MPE-derived NRP1+ CD4 Tconv cells have altered expression of CD25, suggestive of a more activated state as compared to that of NRP1- CD4 Tconv cells. Additionally, we have observed disproportionate cell death of NRP1+ CD4 Tconv cells that can be partially rescued with anti-NRP1 monoclonal antibodies (mAbs). Surviving NRP1+ cells are more likely to express PD-1 and secrete effector cytokines. In our murine NSCLC model carrying key driver mutations KrasG12D P53−/− Lkb1−/− and higher tumor mutational burden (KPL-3M), analysis of tumor-infiltrating CD4 Tconv cells confirm that NRP1+ CD4 T cells are preferentially found within tumors and with higher PD-1 expression as compared to NRP1- cells. Furthermore, treatment with anti-NRP1 mAbs confers anti-tumor efficacy in this model, indicating that NRP1 is a functionally important inhibitor of host anti-tumor immune responses. These data suggest that CD4 Tconv cells expressing NRP1 are activated but prone to cell death, warranting further investigation into its potential as a therapeutic target for NSCLC.

Citation Format: Bitta P. Kahangi, Raymond J. Lim, Jensen Abascal, William P. Crosson, Edgar Perez Reyes, Camelia Dumitras, Linh M. Tran, Kostyantyn Krysan, Ramin Salehi-Rad, Steven M. Dubinett, Bin Liu. The role of Neuropilin-1 signaling in CD4 conventional T cell immune function in non-small cell lung 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 5181.
Abstract 5328: Novel KRAS-specific in-well lysis ELISA for high throughput screening
Authors: Park Hyun Min , Poole Jason , Jelinek Mary Anne
Published: April 2023

RAS is one of the most frequently mutated genes in cancer. Gain–of–function missense mutations promoting oncogenesis cluster at codons 12, 13, and 61 of KRAS, NRAS and HRAS. These result in enhanced GTP binding due to fast exchange of nucleotide and/or impairment of GAP (GTPase Activating Protein) binding. Although these mutants are all activating, they are not equal in their oncogenic potential and differences in patient survival are associated with different RAS mutants and which RAS allele is affected, with KRAS being the most prevalent. Current RAS therapeutic approaches have migrated to be isoform specific. With this in mind, we have developed an activation state specific high throughput screening compatible (HTS) assay that is specific for KRAS. A novel rabbit anti–KRAS recombinant antibody was isolated via cloning of immunoglobulin variable regions from antigen positive B cells isolated from rabbits immunized with a peptide immunogen targeting a region of low homology between the RAS isoforms. Antibody specificity was validated with recombinant HRAS, KRAS and NRAS proteins over a wide concentration range by both immunoblotting and by a bead–based multiplex ELISA where all three RAS isoforms were present. The new assay allows for direct in–well lysis of cells cultured in 96–well plate format, either directly in media or following a PBS wash step. Activated KRAS in the lysate is captured on the assay plate and detected via chemiluminescence using the KRAS specific antibody. Preliminary assay validation has included EGF treatment of HeLa cells to show activation in cells expressing wildtype KRAS and inhibition with the selective non–covalent KRASG12D inhibitor MRTX1133 in Aspc1 cells known to express KRASG12D over a wide range of cell seeding densities. We anticipate that this assay will further the discovery of novel KRAS specific therapeutics.

Citation Format: Hyun Min Park, Jason Poole, Mary Anne Jelinek. Novel KRAS-specific in-well lysis ELISA for high throughput screening. [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 5328.
Abstract 5475: Improved prediction of immunotherapy responses by constitutive neoantigen load and LINE-1 methylation
Authors: Bang Hyoeun , Kim Jeong Yeon , Joe Cheolyong , Kim Kyeonghui , Bang Younghak , Kim Hyemin , Cha Honghi , Oh Eunjoo , Lee Naeun , Shin Inkyung , Noh Seung-Jae , Cho Dae-Yeon , Lee Se-Hoon , Choi Jung Kyoon
Published: April 2023

Previous biomarkers such as PD-L1 expression, tumor mutation burden (TMB), and neoantigen load have limited accuracy in predicting responses to immune checkpoint inhibitor (ICI) therapy. In this work, we propose an improved predictor of ICI responses on the basis of our previous findings on the role of neoantigen functionality and genomic hypomethylation in tumor immunity. Specifically, we developed a metric of constitutive neoantigen load given the association of constitutive neoantigens derived from genes indispensable for cancer growth with favorable clinical responses to ICI (Clin. Transl. Med. 12:e714). A panel that captures somatic mutations on the 300 most essential genes identified by genome-wide screening in ~1,800 pan-cancer cell lines was used. Our DeepNeo algorithm (Nat. Commun. 11:951; Nat. Genet. 2023) was employed to identify T cell-reactive neoantigens from somatic mutations binding patient-matched HLA proteins. Also, considering the role of genomic hypomethylation in inducing resistance of tumors to ICI therapy (Nat. Commun. 10:4278), we developed an assay that estimates genomic hypomethylation by targeted sequencing of LINE-1 elements. Constitutive neoantigen load and LINE-1 methylation status were evaluated in our cohort consisting of 335 lung cancer patients treated with ICIs. As a result, constitutive neoantigen load (HR=0.671; p-value=3.13E-03) and LINE-1 methylation (HR=0.597; p-value=0.012) outperformed PD-L1 (HR=0.74; p-value=0.199), TMB (HR=0.917; p-value=0.511), and the combination of PD-L1 and TMB (HR=0.707; p-value=0.218). The combination of constitutive neoantigen load and LINE-1 methylation (HR=0.245; p-value=1.11E-05) showed even better predictive power. In conclusion, our approach can be used as new companion diagnostics for ICI-based immune-oncology.

Citation Format: Hyoeun Bang, Jeong Yeon Kim, Cheolyong Joe, Kyeonghui Kim, Younghak Bang, Hyemin Kim, Honghi Cha, Eunjoo Oh, Naeun Lee, Inkyung Shin, Seung-Jae Noh, Dae-Yeon Cho, Se-Hoon Lee, Jung Kyoon Choi. Improved prediction of immunotherapy responses by constitutive neoantigen load and LINE-1 methylation. [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 5475.
Abstract 5527: Undergraduate cancer research scholars’ community field experiences encourage community engagement to curb future disparities in the cancer workforce
Authors: Kovacic Melinda Butsch , Nolan Rachael D. , Loyd Melissa , Sauter Sharon , Ramos Karen , Kues John R.
Published: April 2023

Lack of access to cancer prevention, early screening, and treatment, particularly in low socioeconomic, underserved communities, are cited as substantial barriers to improving survivorship. Overcoming the complexity of this problem will require a larger, diverse, and dedicated workforce. Community engagement can lead to greater awareness of and both access and accrual to research. Encouraging trainees understanding of and participation in community engagement early on is one approach to curb cancer research related disparities. The current pipeline of cancer-focused healthcare professionals and researchers that understand, appropriately engage, and connect with their target communities and patient populations is dangerously inadequate. To expand our future culturally competent, cancer-focused workforce, the University of Cincinnati Cancer Center has established the Cancer Research Scholars Program which is supported in part by a National Cancer Institute R25. The program offers to 20-23 undergraduate scholars each year, unique and inspiring paid research opportunities across the cancer continuum and educational experiences that included cancer topics, professional development, team science, and both cultural humility and microaggression training. Importantly, all scholars prepare for and participate in one of three community-engaged, cancer-focused summer field experiences offered with our community partners to encourage the supporting of and//or active participation in community-engaged research, education, and outreach in their future cancer careers. Twelve of the 23 summer 2022 Scholars were also invited to continue their research into the academic year either in-person or virtually. After year 1, Scholars and mentors completed surveys and on-camera interviews about the program highlighting the program’s successes as well as areas for improvement. Nearly all Scholars appreciated the community engagement opportunities offered and understood their value to their potential careers. Annual tracking of these Scholars will determine the program’s longer-term impact on the cancer workforce.

Citation Format: Melinda Butsch Kovacic, Rachael D. Nolan, Melissa Loyd, Sharon Sauter, Karen Ramos, John R. Kues. Undergraduate cancer research scholars’ community field experiences encourage community engagement to curb future disparities in the cancer workforce. [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 5527.
Abstract 5540: Spouse caregivers of patients with glioblastoma report unmet needs throughout the disease trajectory
Authors: Coman Diana L. , Chard Megan , Desautels Lisa , Lutz Barbara J. , Minns Laurie
Published: April 2023

Glioblastoma (GBM) is a devastating and terminal cancer that results in memory loss, cognitive declines, personality changes, neuropsychiatric symptoms, and physical impairments that greatly affect quality of life of both the patient with GBM and their primary caregiver, who is often a spouse or partner. These caregivers describe feeling unsupported and unprepared to assume the responsibilities associated with addressing the progressive physical, emotional, and psychological support needs of their loved ones with GBM. In this qualitative study, female spouse primary caregivers were recruited through a private Facebook support group to write letters describing their experiences in caring for their loved ones with GBM. Using a reflexive thematic analysis, 101 letters from caregivers were analyzed for unmet needs, challenges, and opportunities for better care and support. Caregivers expressed feeling unsupported and poorly prepared, and described different levels and types of needs and challenges throughout the GBM disease trajectory. The 4 phases of the trajectory included the acute phase (first 2-4 months post diagnosis), living with disease progression, end of life stage, and the bereavement stage. Challenges included difficulties adapting to sudden life changes, the need for more education on what to expect, feelings of sudden loss, relationship strain, and not having enough knowledge about GBM. Associated emotional responses included feelings of fear, lack of control, agony, disappointment, and exhaustion. These challenges and emotional impact were greatest during the acute and living with disease progression phases. Caregivers also described relying heavily on friends, community supports (such as faith groups), peer-support through the Facebook group during this living with disease progression phase, but still experienced high levels of relationship strain and other unmet needs. These findings on the primary caregivers’ experiences provide opportunities to guide more timely and tailored interventions to provide support and improve care for patient/caregiver dyads to help mitigate the burden of this progressive disease and improve quality of life for caregivers.

Citation Format: Diana L. Coman, Megan Chard, Lisa Desautels, Barbara J. Lutz, Laurie Minns. Spouse caregivers of patients with glioblastoma report unmet needs throughout the disease trajectory. [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 5540.
Abstract 5567: Circulating bacterial DNA is a non-invasive biomarker for early recurrence of NSCLC patients
Authors: Chen Haiming , Lu Yiming , Qiu Mantang
Published: April 2023

Background: Alteration of human microbiota has been implicated in non-small cell lung cancer (NSCLC). However, most metagenomics studies utilized fecal samples, circulating bacteria DNA in patients with respectable NSCLC remain unexplored. This study aimed to characterize bacteria DNA in plasma of respectable NSCLC patients and build a machine-learning model to detect early recurrence of NSCLC.

Methods: We performed whole genome sequencing of plasma from 101 NSCLC patients with (n = 36, R) or without (n = 65, NR) tumor recurrence within three years after surgery. Microbial DNA was obtained by removing the host genome and relative abundance was measured by mapping reads into microbial genomes. Significant microbial features were identified in the discovery set and built into a random forest model, which was tested in the validation set.

Results: We enrolled 22 R and 39 NR patients with matching demographic features in the discovery set. The remaining 14 R and 26 NR patients were included in the validation set. PCoA analysis using Bray-Curtis distance showed significant differences between R and NR patients at genus level and species level in the discovery set. Notably, in the discovery set, 4 genera and 13 species showed significantly lower abundance in R patients, whereas 11 genera and 21 species were conversely enriched. Based on these significant microbial features, the random-forest model obtained the maximal AUC value of 0.9044 (95% CI, 0.8203-0.9886) when 11 features were selected. In the validation set, this model distinguished R from NR patients with an AUC of 0. 8434 (95% CI, 0.7189-0.9679).

Conclusions: This pioneer study characterized alteration of circulating bacterial DNA in NSCLC patients with early recurrence, and illustrated significant difference in microbial composition between R and NR patients. The predictive model constructed with selected circulating microbial features was an accurate and non-invasive tool for prediction in early NSCLC recurrence.

Citation Format: Haiming Chen, Yiming Lu, Mantang Qiu. Circulating bacterial DNA is a non-invasive biomarker for early recurrence of NSCLC patients. [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 5567.
Abstract 5662: Early unwanted immunogenicity assessment of immuno oncology drugs
Authors: Pattyn Sofie , Vlaming Martijn , Pieters Alanah , Schockaert Jana
Published: April 2023

Bispecifics targeting multiple antigens or epitopes have been showing great promise for treatment of cancer. However, managing unwanted immunogenicity has become a challenge in the development cycle of these promising therapeutics as there is a trend towards higher unwanted immune responses compared with classical monoclonal antibodies. In vitro assays using human primary immune cells can be used to assess the risk of induction of a cytokine release storm or induction of unwanted immunogenicity. For the latter, T cell activation and proliferation assays can be used to assess and predict an unwanted immune response and avoid induction of anti-drug antibodies later on. In order to achieve reliable and consistent results, high quality primary immune cells should be used in combination with sensitive fit-for-purpose in vitro assays.

Citation Format: Sofie Pattyn, Martijn Vlaming, Alanah Pieters, Jana Schockaert. Early unwanted immunogenicity assessment of immuno oncology drugs. [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 5662.
Abstract 5677: MB101, a novel PD-L1 x CD3 targeted bispecific antibody, exhibits potent tumor-killing efficacy with minimal off-tumor toxicity
Authors: Kim Suna , Bang Hyojoo , Jung Yongjun , Cho Sunjung , Kim Youngsam , Park Sang-hyun , Kang Seok Chan , Park Youngjin , Jung Sungyoub
Published: April 2023

Bispecific antibody (bsAb) which targets tumor-associated antigens and simultaneously induces CD3-mediated T cell activation, is one of the promising therapeutic approaches for cancer treatment. However, despite numerous CD3-based bsAb have been developed, the clinical application faces pronounced hurdles, such as increased on-target off-tumor toxicity and cytokine release syndrome (CRS) by using high-affinity anti-CD3 moieties. Accordingly, there are still needs for the development of CD3-based bsAb with more favorable properties, which are tumor-specific targeting and potent tumor killing activity without excessive cytokine release. Thus, we developed MB101, a novel PD-L1 x CD3 bsAb with low affinity to CD3, which is uncoupled potent tumor cytotoxicity from severe cytokine release. MB101 showed a strong binding affinity to PD-L1 (KD = 0.4 nM) but a weak binding affinity to CD3 (KD = 1.0 μM), indicating preferentially targeting to PD-L1+ tumor cells. MB101 showed an effective tumor cell killing activity with higher Emax compared to avelumab mono- or combination treatment with CD3 mAb in PD-L1+ MDA-MB-231 cells. T cell-mediated cytotoxicity stimulated by MB101 in co-culture with target cells was accompanied by lower secretion of pro-inflammatory cytokines than CD3 mAb and bsAb comparator with high-affinity anti-CD3 arm. Additionally, in the absence of target cells, MB101 treatment to PBMC induced a very low level of pro-inflammatory cytokines, suggesting the possibility of reduced CRS risk. Furthermore, we proved that T cell-mediated cytotoxicity by MB101 was increased in a PD-L1 expression-dependent manner of tumor cells, but not normal cells, indicating minimal on-target off-tumor toxicity. Consistently with in vitro results, MB101 effectively regressed tumor volume (TGI=83%) without induction of excessive cytokine release in human CD3 knock-in mouse model. In addition, MB101 showed superior tumor growth inhibition (TGI=70%) than that of avelumab (TGI=44%) in humanized mouse xenograft model. In summary, MB101 selectively triggers robust PD-L1-dependent tumor cytotoxicity while inducing very low cytokine release, suggesting that MB101 might be a promising therapeutic option as CD3-based bsAb for cancer treatment.

Citation Format: Suna Kim, Hyojoo Bang, Yongjun Jung, Sunjung Cho, Youngsam Kim, Sang-hyun Park, Seok Chan Kang, Youngjin Park, Sungyoub Jung. MB101, a novel PD-L1 x CD3 targeted bispecific antibody, exhibits potent tumor-killing efficacy with minimal off-tumor toxicity. [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 5677.
Abstract 5792: The novel therapeutic diiminoquinone exhibits anticancer effects on human prostate cancer cells in 2D and 3D in vitro models
Authors: Yehya Amani , Monzer Alissar , Wakimian Kevork , Ghamlouche Fatima , Saheb Nour , Tawil Ayman , Gali-Muhtasib Hala , Mukherji Deborah , El-Hajj Albert , Abou-Kheir Wassim G.
Published: April 2023

Aim: To investigate the anticancer potential of the novel quinone DIQ on prostate cancer (PCa) cells in two-dimensional (2D) and three-dimensional (3D) in vitro.

Methods: The anti-proliferative effect of DIQ on PC3 and DU145 PCa cell lines was determined in vitro by using 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and trypan blue exclusion assays. In addition, the potency of DIQ was tested on the formation and size of PCa cell-derived prostatospheres and organoids grown in a specialized 3D-culture system. Also, 3D organoids were established from fresh tissue samples representing different stages of PCa obtained from consented patients who underwent radical prostatectomy at the American University of Beirut Medical Center (AUBMC) according to appropriate Institutional Review Board (IRB) approval guidelines. This model was used to assess DIQ

individualized tumor response on organoids growth and the effect was evaluated by quantifying the number of organoids formed (OFC) and calculating the average size (diameters).

Results: Our results showed that DIQ significantly attenuated cell proliferation and viability in PC3 and DU145 PCa cell lines. DIQ also reduced the formation ability and size of PC3 and DU145 prostatospheres and organoids at sub-toxic doses of 1 µM. Potently, DIQ displayed a significant decrease in both the count and the size of the different patient-derived organoids as compared to their respective control.

Conclusion: This study provides promising clues for the use of the novel anticancer therapeutic DIQ in targeting CSCs, findings that may have promising therapeutic implications for PCa patients.

Citation Format: Amani Yehya, Alissar Monzer, Kevork Wakimian, Fatima Ghamlouche, Nour Saheb, Ayman Tawil, Hala Gali-Muhtasib, Deborah Mukherji, Albert El-Hajj, Wassim G. Abou-Kheir. The novel therapeutic diiminoquinone exhibits anticancer effects on human prostate cancer cells in 2D and 3D in vitro models [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 5792.
Abstract 5804: Targeting methyl donor synthesis inhibits platinum-induced enrichment of ovarian cancer stem cells
Authors: Zhang Shu , Metcalfe Tara X. , Hassel Christiane A. , O'Hagan Heather M. , Nephew Kenneth P.
Published: April 2023

Ovarian cancer (OC) is one of the deadliest gynecologic cancers in US. Platinum (Pt)-based chemotherapy can reprogram cancer cells to ovarian cancer stem cells (OCSCs) and contribute to incurable recurrence. We have previously shown that Pt-containing drugs, e.g., cisplatin, enrich for OCSCs and Pt-induced alterations in DNA methylation enhanced OCSC malignant properties. CSCs have been shown to be dependent on methionine (Met) in many cancers, and in the current study, we investigated the role of Met and the interplay between methylation and Met metabolism in OCSC enrichment. In high grade serous ovarian cancer cell lines (OVCAR5, OVCAR3), cisplatin treatment (IC50, 16hr) increased (p<0.01) the percentage of cells that were positive for aldehyde dehydrogenase (ALDH+ cells), a known OCSCs marker. Cisplatin treatment also increased (p<0.05) the whole cell level of 5-methylcytosine, indicating Pt-induced methylation changes. In addition, Met metabolism was altered by cisplatin, demonstrated by decreased (p<0.01) level of Met derivative, S-adenosyl-methionine (SAM), and increased (p<0.01) expression of Met utilization enzyme, methionine adenosyltransferase 2A (MAT2A). To investigate a possible role of Met metabolism in Pt-induced enrichment of OCSCs, OVCAR5 cells were cultured in complete or Met-depleted (Met-) media, treated with cisplatin or vehicle and examined for CSC properties, including ALDH+ cells and the ability to form spheroids under low-attachment conditions. Met depletion blocked (p<0.01) both the Pt-induced increase of ALDH+ cells and spheroid formation. Both Met- only and Met- in combination with cisplatin activated p38, determined by increased phosphorylated p38, which may have responded to Met depletion and promoted cell cycle arrest. In addition, MAT2A expression was higher (p<0.01) in ALDH+ compared to ALDH- OC cells. MAT2A inhibitor FIDAS-5 combined with cisplatin blocked the Pt-induced increase in ALDH+ cells (p<0.01), demonstrating a requirement for Met utilization in Pt-induced OCSC enrichment. As MAT2A is a therapeutic target in cancer, MAT2A inhibition could represent a therapeutic approach for inhibiting CSC in ovarian cancer. We are further investigating the mechanism of MAT2A-mediated inhibition on Pt-induced OCSCs enrichment by examining the OC methylome and cell cycle arrest.

Citation Format: Shu Zhang, Tara X. Metcalfe, Christiane A. Hassel, Heather M. O'Hagan, Kenneth P. Nephew. Targeting methyl donor synthesis inhibits platinum-induced enrichment of ovarian cancer stem 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 5804.
Abstract 5806: Kinase library screening identifies IGF-1R/PI3K/AKT axis as a crucial pathway for survival of intrahepatic cholangiocarcinoma cells with stemness characteristics
Authors: Rattanasinchai Chotirat , Chornkrathok Chidchanok , Sriphan Donlaporn , Navasumrit Panida , Ruchirawat Mathuros
Published: April 2023

Intrahepatic cholangiocarcinoma (iCCA) is a subtype of biliary malignancy that arises along the peripheral bile duct epithelium. Given its limited and inefficacious treatment options, the mortality rate of iCCA remains high and thus a novel therapeutic strategy is urgently needed. The concept of cancer stem cells (CSCs) explains a subpopulation of cancer cells that possesses self-renewal and differentiation capacities. These CSCs become the culprit for cancer metastasis, cancer recurrence and chemotherapeutic resistance, making them promising targets for cancer treatment. The propose of this study is to identify the signaling pathways crucial for the survival of iCCA with stemness characteristics. Gene expression analysis on a dataset of Thai iCCA patients (n=91) showed that iCCA tumors are enriched in the Adult Stem Cell (ASC) signature which are the shared characteristics among the epithelial-derived cancer with stemness characteristics. Although iCCA cells exhibit some degree of enrichment in stem cells markers when compared with cholangiocyte cells, further upregulation of stem cell-related genes, e.g. Nanog, Oct3/4, Sox2, and Sox9, were found in iCCA cells grown as 3D tumorspheres. A kinase inhibitor screening library was then performed on iCCA tumorspheres. Our initial screening demonstrated that 11 out of the top 50 kinase inhibitors that effectively decrease viability of iCCA tumorspheres belongs to the insulin-like growth factor-1 receptor (IGF-1R)/phosphoinositide-3-kinase (PI3K)/protein kinase B (PKB/AKT) axis. Additionally, these inhibitors increase cell apoptosis, judged by the caspase 3/7 activity, within iCCA tumorspheres. Conversely, IGF-1 treatment which is sufficient to induce activation of IGF-1R and AKT also enhances cell viability and decrease cell apoptosis in iCCA. Taken together, our data demonstrate that IGF-1R/PI3K/AKT axis is crucial for survival of iCCA cells with stemness characteristics and this pathway could be a potential therapeutic target in iCCA.

Citation Format: Chotirat Rattanasinchai, Chidchanok Chornkrathok, Donlaporn Sriphan, Panida Navasumrit, Mathuros Ruchirawat. Kinase library screening identifies IGF-1R/PI3K/AKT axis as a crucial pathway for survival of intrahepatic cholangiocarcinoma cells with stemness characteristics. [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 5806.
Abstract 5808: Integrin-linked kinase regulates Wnt transcriptional activity in platinum resistant ovarian cancer stem cells
Authors: Atwani Rula , Lazar Virginie , Sandusky George Earl , Condello Salvatore
Published: April 2023

Ovarian cancer (OC) is the most lethal gynecological cancer, characterized by chemo-resistance and fatal tumor recurrence after primary treatment. The metastatic progression is mediated by exfoliation of tumor cells into the peritoneal cavity, the formation of compact OC spheroids, and their implantation onto the overlying peritoneal surface. Spheroid formation is promoted by a change of cell adhesion properties, enhanced secretion of extracellular matrix (ECM) components, such as fibronectin (FN), that stiffen the tumor stroma, promote the aberrant activation of integrins, and recruit the adaptor protein integrin-linked kinase (ILK). ILK drives cytoskeletal assembly and modulates key processes, including survival, invasion, and stemness. However, the functional role of ILK in chemoresistance of ovarian cancer stem cells (OCSCs) remains incompletely understood. We tested the hypothesis that the formation of FN-integrin complexes at the cell membrane promotes survival of OCSCs and supports OC spheroid formation by activating ILK and downstream oncogenic signaling. In OCSCs compared with non-CSC, ILK, FN, and integrin β1 mRNA expression levels were increased (P<0.001) and further enriched in OC spheroids compared with monolayers (P<0.01). ILK blockade with the specific inhibitor cpd-22 suppressed spheroid proliferation and tumor initiating capacity in xenograft mouse model. Furthermore, key oncogenic signaling pathways, in particular decreased β-catenin signaling essential to sustaining cancer cell stemness, were altered in OC spheroids treated with cpd-22. Of the genes examined, expression of Wnt receptor Frizzled 7 (Fzd7) was the mostly highly downregulated in OC spheroids treated with cpd-22, indicating a direct correlation between ILK activation and Wnt signaling. By using co-immunoprecipitation and proximity ligation assay, we demonstrated that ILK co-localizes with Fzd7 in OC spheroids. Mechanistically, treatment of OC spheroids with the combination of carboplatin and ILK-Fzd7 blockade decreased β-catenin signaling, inhibited phospho-Akt at Ser473 and increased levels of cleaved-caspase-3 compared to single agent alone, resulting in apoptosis. By regulating β-catenin signaling, Fzd7 expression and ILK activation are essential for OCSCs survival. Targeting Fzd7-ILK clusters may represent a new therapeutic approach to eradicate OCSC and ultimately improve patient outcomes.

Citation Format: Rula Atwani, Virginie Lazar, George Earl Sandusky, Salvatore Condello. Integrin-linked kinase regulates Wnt transcriptional activity in platinum resistant ovarian cancer stem 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 5808.
Abstract 5817: LKB1 overcomes RAS pathway-mediated loss of epithelial integrity
Authors: Tervonen Topi A. , Pant Shishir M. , Englund Johanna , Klefström Juha
Published: April 2023

Oncogenic RAS pathway disassembles cell-cell and cell-basement membrane junctions, thus promoting epithelial cell motility and invasion. To find tumor proteomic changes responsible for the disrupting effects of RAS on epithelial integrity, we explored cBioportal cell line proteomics data across all cancer types for correlations between oncogenic KRAS or HRAS mutations and possible regulators of epithelial integrity. We found inverse correlation between these RAS mutations and tumor suppressor LKB1 protein levels in a cohort where LKB1 was not inactivated by a mutation. Furthermore, LKB1 protein levels were also positively correlated with NF2 protein (a negative regulator of RAS) in human breast tumors and inversely correlated with c-RAF (RAS effector) activating phosphorylation at serine 338 in breast cancer cell lines. We sought to functionally clarify the relationship between RAS pathway and LKB1, finding that an ectopic constitutive or inducible expression of HRASV12 downregulates LKB1 protein in mammary epithelial MCF10A cells and HMEC-hTERT cells. Furthermore, RAS-mutant MCF10A cells regained LKB1 protein levels upon treatment with MEK/ERK inhibitors. We establish that the RAS-mediated downregulation of LKB1 protein occurs through proteasomal activity. LKB1 reconstituted MCF10A-HRASV12 (RAS-LKB1) cells displayed restored desmoplakin, γ-catenin and E-cadherin pericellular immunostaining pattern in monolayer cultures, suggesting restoration of desmosomes and adherence junctions. We also cultured RAS-LKB1 cells in 3D egg-white and observe restoration of E-cadherin, γ-catenin, α6-integrin, and laminin-332, indicating a rescue of epithelial integrity in 3D context. Our findings suggest that LKB1 rescues mutant RAS-mediated disruption of cell-cell and cell-basement membrane junctions and general epithelial integrity. Our current aim is to define how LKB1 suppresses RAS mediated disruption of breast epithelial integrity, that is expected to shed new light into early stages of tumor invasion and uncover novel therapeutic opportunities via restoration of LKB1.

Citation Format: Topi A. Tervonen, Shishir M. Pant, Johanna Englund, Juha Klefström. LKB1 overcomes RAS pathway-mediated loss of epithelial 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 5817.
Abstract 5906: Gut microbiota signatures are associated with immunotherapy induced cognitive decline and neurotoxicity
Authors: Banerjee Anik , Roland Christina , Johnson Sarah , Ajami Nadim , Wargo Jennifer
Published: April 2023

Studies have demonstrated promising outcomes of combined immune checkpoint blockade (CICB) modulating tumor cytotoxicity via targeting cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and programmed cell death 1 (PD-1). The emergence of long-term patient management has been transformed among health care professionals as patient survival rates are increasing significantly. Despite promising advances, patient outcomes are heterogenic and are correlative to host intrinsic and extrinsic factors. Multiple reports have highlighted the negative effects of cancer treatments towards inducing cognitive impairment and neurotoxicity often referred to as “chemofog”. More recently, the gut microbiome has been increasingly recognized to regulate antitumor immunity and further distinct microbiome signatures can confer favorable outcomes in the context of CICB. Interestingly, specific microbes within the gut microbiome have been shown to regulate social behavior and cognition via the bidirectional communication along the gut-brain axis though changes in microbiota-derived metabolites and vagal afferent fibers. Insights into mechanisms of immunotherapy induced cognitive decline are warranted. Thus, we hypothesize CICB exacerbates cognitive impairment and neurotoxicity via the detrimental alterations to the gut microbiome. To address this, we profiled the gut microbiome, through 16S rRNA gene and metagenomic sequencing of fecal samples from patients (n=146) with advanced melanoma treated with CICB stratified based on severity of cognitive dissonance. Beta-diversity, incorporating weighted UniFrac distances by principal coordinate analysis (PCoA), revealed a cluster of samples from patients with cognitive impairment as compared to samples from those with minimal to absent cognitive decline. Further, taxonomical analysis showed significant alterations in relative abundances of microbial candidates (e.g., L. intestinalis, A. muciniphila, F. prausnitizii, and R. hominis). These results suggests that CICB patients with cognitive impairments and neurotoxicity have altered microbiome signatures which may potentially exacerbate downstream neuroinflammatory signaling. Further studies investigating specific host-microbe pathways and profiling microbiota-derived metabolites which are implicated in these detrimental effects are essential for biomarker discovery and mechanistic insight.

Citation Format: Anik Banerjee, Christina Roland, Sarah Johnson, Nadim Ajami, Jennifer Wargo. Gut microbiota signatures are associated with immunotherapy induced cognitive decline and neurotoxicity. [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 5906.