Frequent Cancer Research Articles

The epidemiology of the most frequent cancers in Poland in 2015-2021 and the impact of the COVID-19 pandemic on cancer incidence.

The late diagnosis, despite the improving availability and accessibility of diagnostic procedures during the last decade in Poland and cooperation between specialist cancer centres, remains an unsolved problem. Moreover, the accessibility to healthcare resources and diagnostic procedures has been drastically reduced because of the COVID-19 pandemic in 2019-2020. The study aimed to present the epidemiology of the most frequent cancers diagnosed in Poland as well as the impact of the COVID-19 pandemic on cancers' incidence. Depersonalized, epidemiological data was obtained from the National Health Fund of Poland. In this retrospective study, the epidemiological analysis was performed and divided into subregions, according to patients' domicile. For each of the subregions, we have calculated the incidence rate per 100,000 standardized to the European Standard Population 2013. The time points of providing the first healthcare service were considered as the time of cancer diagnosis. In the 2015-2019 period, before the COVID-19 pandemic occurred, the nationwide incidence of analysed cancers remained stable or slightly decreased (as the lung cancer). Simultaneusly, during the same period the prevalence of the prostate cancer has increased and the large differences between subregions with the least and the highest incidence were observed. Subsequently, the incidences of all analyzed cancers decreased in 2020, compared to the period before the COVID-19 pandemic occurred. Then, in 2021 a disproportionate increase in cancers' incidence rates was noted. Our results show a significant decrease in the incidence rate of the most frequent cancers diagnosed in Poland in 2020 compared to 2019. Subsequently, in 2021 the increase of the incidence ratios was noted, most likely due to the gradual reduction of epidemic restrictions.

Antitumoral Activity of the Universal Methyl Donor S-Adenosylmethionine in Glioblastoma Cells

Glioblastoma (GBM), the most frequent and lethal brain cancer in adults, is characterized by short survival times and high mortality rates. Due to the resistance of GBM cells to conventional therapeutic treatments, scientific interest is focusing on the search for alternative and efficient adjuvant treatments. S-Adenosylmethionine (AdoMet), the well-studied physiological methyl donor, has emerged as a promising anticancer compound and a modulator of multiple cancer-related signaling pathways. We report here for the first time that AdoMet selectively inhibited the viability and proliferation of U87MG, U343MG, and U251MG GBM cells. In these cell lines, AdoMet induced S and G2/M cell cycle arrest and apoptosis and downregulated the expression and activation of proteins involved in homologous recombination DNA repair, including RAD51, BRCA1, and Chk1. Furthermore, AdoMet was able to maintain DNA in a damaged state, as indicated by the increased γH2AX/H2AX ratio. AdoMet promoted mitotic catastrophe through inhibiting Aurora B kinase expression, phosphorylation, and localization causing GBM cells to undergo mitotic catastrophe-induced death. Finally, AdoMet inhibited DNA repair and induced cell cycle arrest, apoptosis, and mitotic catastrophe in patient-derived GBM cells. In light of these results, AdoMet could be considered a potential adjuvant in GBM therapy.

The intraoperative application of indocyanine green (ICG), in breast reconstructive surgery using a latissimus dorsi flap, as a preventive factor for tissue ischemia and postoperative complications

Introduction: Breast cancer is the most frequent cancer among women. Nowadays more and more women decide to undergo breast reconstruction, both using implants as well as autologous flaps. The most important factor in avoiding bad wound healing is the preservation of blood supply and good tissue perfusion. Latissimus dorsi flap is a commonly used method, mainly reserved for irradiated patients, delayed reconstruction, or salvage procedures. With damaged tissues, objectively assessing the conditions is much more difficult than in primary surgery.Aim: The aim of the study is to present the intraoperative application of ICG, in breast reconstructive surgery with a latissimus dorsi flap, as a preventive factor for tissue ischemia and postoperative complications. Materials and methods: A single-center retrospective review of the outcomes of 12 patients who underwent breast reconstruction using an LD flap, with an intraoperative assessment of flap blood supply using ICG. Patient characteristics, comorbidities, and smoking were taken into account. Results: In two cases, after an initial assessment of tissue perfusion, it was decided to resection the distal part of the flap with less blood supply. All reconstructions were successful and the wounds healed properly. None of the patients required reoperation, there were no postoperative complications or surgical site infections.Conclusion: The use of ICG in breast reconstruction with an LD flap is an important predictor of postoperative complications, especially flap necrosis and loss of reconstruction.

Abstract LB060: Engineering KRAS mutation-specific affinity binders: A new frontier in cancer research

Abstract Cancer is a complex disease involving genetic and proteomic dysregulation. Better understanding of this disease requires precise tools to analyze how cells change from a normal to a diseased state. Among the genes implicated in cancer, the checkpoint inhibitor KRAS is notable for its frequent cancer causing mutations. These mutations, especially in glycine codon 12, are a driving force in several cancers, including those of the pancreas, lung, and colon. Recently, researchers have made significant strides in treating one variant, targeting G12C, using small molecules. However, other mutations, including the G12D and G12V variants, which are more common in disease, still lack targeted treatments. To overcome this challenge, we are using a unique immunization and screening strategy to identify a series of KRAS mutant-specific binders. These affinity binders, targeting the wild-type glycine and variants like G12D, G12S, G12C, G12A, G12R, and G12V, are being designed for research purposes to dissect cancer pathways and understand the spatial organization of cancer cells. They may also hold potential for clinical use as tools for prognosis and therapy. Deep characterization of these and other potentially widely used tools will be critical for advancing their uptake in the laboratory and clinic. Citation Format: Ramon Serrano, Ryan Wallace, Sergei Bibikov, Michael Weiner, Kevin J. Harvey. Engineering KRAS mutation-specific affinity binders: A new frontier in cancer research [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB060.

“Accepting Unexpected Health Realities”: Exploring the Life Experiences of Ovarian Cancer Patients – A Qualitative Study

Introduction: Among women, ovarian cancer is the eighth most frequent cancer globally and the fifth leading cause of cancer-related deaths in women. Ovarian cancer is termed the "silent killer" because of its slow onset and unclear symptoms. Most of the advanced cases of ovarian cancer were found due to poor screening. The main objective of this study was to understand the emotional experiences, coping and resilience of patients undergoing ovarian cancer treatment. Methods: This qualitative study employs a phenomenological approach to explore the life experiences of ovarian cancer patients. An in-depth interview was conducted among patients undergoing cancer treatment. A total of 11 participants were included in this study. Thematic analysis was done. Results: The analysis of the data revealed eight main themes and 32 sub-themes that were characterized based on the experiences of cancer patients: 1) Misconceptions about symptoms, 2) Barriers in accessing healthcare, 3) Physical struggles, 4) Emotional turmoil, 5) Social issues, 6) Hope giving factors, 7) Sacrifice for family well-being, and 8) Emotional resilience. Conclusion: One of the key findings of this study is the importance of accepting unexpected health realities. This study shows the emotional journey of accepting difficult realities and mainly trusting medical professionals to guide one through treatment. Develop targeted educational campaigns to raise awareness about the symptoms of ovarian cancer and dispel common misconceptions. 

Global cancer statistics 2022: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries.

This article presents global cancer statistics by world region for the year 2022 based on updated estimates from the International Agency for Research on Cancer (IARC). There were close to 20 million new cases of cancer in the year 2022 (including nonmelanoma skin cancers [NMSCs]) alongside 9.7 million deaths from cancer (including NMSC). The estimates suggest that approximately one in five men or women develop cancer in a lifetime, whereas around one in nine men and one in 12 women die from it. Lung cancer was the most frequently diagnosed cancer in 2022, responsible for almost 2.5 million new cases, or one in eight cancers worldwide (12.4% of all cancers globally), followed by cancers of the female breast (11.6%), colorectum (9.6%), prostate (7.3%), and stomach (4.9%). Lung cancer was also the leading cause of cancer death, with an estimated 1.8 million deaths (18.7%), followed by colorectal (9.3%), liver (7.8%), female breast (6.9%), and stomach (6.8%) cancers. Breast cancer and lung cancer were the most frequent cancers in women and men, respectively (both cases and deaths). Incidence rates (including NMSC) varied from four-fold to five-fold across world regions, from over 500 in Australia/New Zealand (507.9 per 100,000) to under 100 in Western Africa (97.1 per 100,000) among men, and from over 400 in Australia/New Zealand (410.5 per 100,000) to close to 100 in South-Central Asia (103.3 per 100,000) among women. The authors examine the geographic variability across 20 world regions for the 10 leading cancer types, discussing recent trends, the underlying determinants, and the prospects for global cancer prevention and control. With demographics-based predictions indicating that the number of new cases of cancer will reach 35 million by 2050, investments in prevention, including the targeting of key risk factors for cancer (including smoking, overweight and obesity, and infection), could avert millions of future cancer diagnoses and save many lives worldwide, bringing huge economic as well as societal dividends to countries over the forthcoming decades.

Ensemble and Transformer Encoder-based Models for the Cervical Cancer Classification Using Pap-smear Images

Cervical cancer poses a health concern for women globally ranking as the seventh most common disease and the fourth most frequent cancer among women. The classification of cytopathology images is utilized in diagnosing this condition with a focus on automating the process due to potential human errors in manual examinations. This study presents an approach that integrates transfer learning, ensemble learning and a transformer encoder to classify cervical cancer using pap-smear images from the SIPaKMeD dataset. By combining these methods human involvement in the classification task is minimized. Initially individual models based on transfer learning are. Their unique characteristics are combined to create an ensemble model. This ensemble model is then input into the proposed transformer encoder specifically utilizing the Vision Transformer (ViT) model. The results highlight the effectiveness of this methodology. The VGG16 model demonstrates accuracy of 97.04% and an F1 score of 97.06% when applied to classifying five categories using the SIPaKMeD dataset. However surpassing this performance is the learning model, with an accuracy of 97.37%. Notably outperforming all models is the transformer encoder model achieving an accuracy of 97.54%.Through the utilization of transfer learning, ensemble learning and the transformer encoder model this research introduces a method, for automating the classification of cervical cancer. The findings underscore the capability of the suggested approach to enhance the precision and effectiveness of diagnosing cancer.

FNDC5 inhibits malignant growth of human cervical cancer cells via restraining PI3K/AKT pathway.

Cervical cancer (CxCa) is the fourth most frequent cancer in women. This study aimed to determine the role and underlying mechanism of fibronectin type III domain-containing protein 5 (FNDC5) in inhibiting CxCa growth. Experiments were performed in human CxCa tissues, human CxCa cell lines (HeLa and SiHa), and xenograft mouse model established by subcutaneous injection of SiHa cells in nude mice. Bioinformatics analysis showed that CxCa patients with high FNDC5 levels have a longer overall survival period. FNDC5 expression was increased in human CxCa tissues, HeLa and SiHa cells. FNDC5 overexpression or FNDC5 protein not only inhibited proliferation, but also restrained invasion and migration of HeLa and SiHa cells. The effects of FNDC5 were prevented by inhibiting integrin with cilengitide, activating PI3K with recilisib or activating Akt with SC79. FNDC5 inhibited the phosphorylation of PI3K and Akt, which was attenuated by recilisib. PI3K inhibitor LY294002 showed similar effects to FNDC5 in HeLa and SiHa cells. Intravenous injection of FNDC5 (20 μg/day) for 14 days inhibited the tumor growth, and reduced the proliferation marker Ki67 expression and the Akt phosphorylation in the CxCa xenograft mouse model. These results indicate that FNDC5 inhibits the malignant phenotype of CxCa cells through restraining PI3K/Akt signaling. Upregulation of FNDC5 may play a beneficial role in retarding the tumor growth of CxCa.

PSMC2 promotes glioma progression by regulating immune microenvironment and PI3K/AKT/mTOR pathway

BackgroundGlioma, the most frequent and malignant central nervous system (CNS) cancer, has a bad outcome. Proteasome 26S subunit ATPase 2 (PSMC2) is an essential part of the 26S proteasome and promotes the development of several tumors. However, the pathway and function of PSMC2 in glioma have not been unelucidated. MethodsThis study analyzed PSMC2 expression in glioma tissues and its predictive significance for patients. We examined the link between PSMC2 and DNA methylation, immune cell infiltration, tumor immune cycle, immune cell homeostasis, and immune checkpoints. Subsequently, immunohistochemistry and in vitro trials were employed to validate the expression, prognostic potential, and function of PSMC2 in glioma. The mechanisms of PSMC2 in glioma were further explored. ResultsOur study revealed that PSMC2 expression increased in glioma tissues contrasted with healthy tissues, and patients with high PSMC2 glioma exhibited poor overall survival (OS) compared to the low-PSMC2 group. Immune profile analysis revealed that PSMC2 was positively related to immunosuppressive cell infiltration and immune checkpoints and adversely related to the cancer immune cycle and immune cell homeostasis. In cell-based investigations, the inhibition of PSMC2 was found to effectively suppress the aggressiveness and proliferation of glioma cell lines while also enhancing cell cycle arrest and promoting cell death. Gene Set Enrichment Analysis (GSEA), Gene Set Variation Analysis (GSVA), and in vitro experiments showed that PSMC2 promoted glioma development through the PI3K/AKT/mTOR pathway. ConclusionsPSMC2 was upregulated in glioma and promoted cancer progression by modulating the tumor immune microenvironment, cancer cell biological behavior, immune cell homeostasis, and the PI3K/AKT/mTOR pathway, providing a new option to treat glioma.

The haemoglobin levels of patients with prostate cancer treated with radical radiotherapy

Introduction: The worldwide estimate points to prostate cancer as the second most frequent cancer in men worldwide, where radiotherapy has been an important modality of curative treatment in managing this neoplasia. The prevalence and impact of cancer-related anemia are not widely known, and it may be overlooked or considered clinically significant. It is believed that the outcomes and toxicity of radiotherapy in prostate cancer treatment may be influenced by blood hemoglobin levels. Objective: To evaluate hemoglobin levels in patients with prostate cancer before and after radiotherapy treatment. Methods: Retrospective, longitudinal study of patients diagnosed with malignant prostate neoplasms undergoing external radiotherapy. Hemoglobin levels were measured before radiotherapy and after treatment completion (20-40 days). Anemia was defined by the World Health Organization as men with values less than 13 g/dl. Prostate cancer recurrence risk classification was based on the National Comprehensive Cancer Network risk criteria. Results: Forty patients were evaluated with measurement of Hemoglobin levels before and after radiotherapy treatment. The mean age was 67 years. According to the risk group classification, 4 patients were low-risk (10%), 27 patients were intermediate-risk (67.5%), and 9 patients were high-risk (22.5%). The mean pre-radiotherapy Hemoglobin levels were 13.3 g/dL, and the mean post-radiotherapy Hemoglobin levels were 10.4 g/dL, p < 0.0001. Conclusion: In this study, a reduction in serum Hemoglobin levels was observed after radiotherapy, highlighting the need for better monitoring of these patients.

Evaluation of miR-130b-3p and miR-375 levels and telomere length with telomerase activity in prostate cancer

Prostate cancer (PC) is the most frequent cancer in males, as well as the second highest cause of cancer-related deaths in men. Differences in expression levels of miRNAs were linked with prostat cancer pathogenesis. qPCR was used to evaluate the expression of miR-130b-3p and miR-375 in Benign Prostate Hyperplasia (BPH (n = 20) and PC (n = 22, pre- and post-operative) patients plasma. Relative telomere lengths (RLTs) in genomic DNA isolated from plasma were measured with qPCR, and telomerase activity analyzed by the ELISA method. PSA levels of PC patients were greater than of BPH patients (p = 0.0473). miR-130b-3p and miR-375 levels were significantly lower in pre-operative specimens of PC patients according to BPH (p = 0,0362, p = 0.0168, respectively). Similarly, post-operative miR-375 levels were lower in PC patients than in BPH patients (p = 0.1866). BPH patients had shorter RTLs than PC patients in both pre- (p=0.0438) and post-operative (p=0.0297) specimens. Telomerase activity was higher in PC patients than BPH(p = 0.0129). Interestingly, telomerase activity was further increased after surgery (p = 0.0003). We aim to identify the levels of miR-130b-3p and miR-375 expression and their relationship with telomerase activity in PC patients. Our data suggest that miRNAs and telomere length (TL) with telomerase activity may play a role in regulating prostate tumorgenesis and may be used as biomarkers for PC diagnosis.

Abstract 3709: Single cell analysis from CSF of patients with suspected leptomeningeal metastasis

Abstract Introduction: Leptomeningeal metastases (LM) are diagnosed in up to 10% of metastatic cancer patients, the most frequent cancers being carcinomas of breast and lung or melanoma. Due to a dismal prognosis and an overall median survival of only four months new approaches for molecular analysis could help to identify individual treatment options. Material and Methods: In this study, we collected 28 cerebrospinal fluid (CSF) samples from patients with different primary tumors but suspected LM. CSF was divided in two parts for standard neuropathologic cytomorphology and detection of circulating tumor cells (CTCs) by using the CellSearch® CTC Kit, respectively. Subsequently, single Cytokeratin-positive CTCs were isolated for whole genome amplification by Ampli1TM and molecular analysis. Additionally, we were able to isolate cell-free DNA (cfDNA) from CSF in a subset of patients which we amplified by an adapted Ampli1TM library protocol. In a small subset of patients, we collected slices of formalin-fixed paraffin embedded (FFPE) primary tumors and systemic metastases for dissociation and isolation of pure subpopulations by DEPArrayTM technology. Isolated CTCs, cfDNA and subpopulations of tumor and stromal cells were analyzed for copy number variations (CNV) applying the Ampli1TM LowPass protocol on the Illumina MiSeqTM platform. Results: By comparing results from classical cytomorphology and CellSearch® workflow, we could find high concordance (24/28 samples) with a slightly higher tumor cell detection rate by CellSearch® (11 positive cases vs. 9 by cytomorphology). Importantly, we could confirm the malignant origin of isolated cells by CNV analysis in all patients positive in CellSearch® assay. Moreover, by comparing single cell data with cfDNA analysis of a subset of 20 samples, we could also detect a higher sensitivity for tumor detection in CTC-based analysis (8/20 cases positive) in comparison to cfDNA-based analysis (5/20 cases positive). In a comprehensive analysis of multiple samples of an individual patient, including CTCs and cfDNA from CSF at different time points, CTCs from blood and biopsies from multiple visceral metastasis, we could identify divergent genomic evolution between tumor cells from the central nervous syste Conclusion: In this proof-of-concept study we show that a liquid biopsy-based approach for single cell analysis from CSF holds potential for innovative diagnostic assays for patients with suspected LM. Combining CTC quantification with molecular single cell analysis could enable us to define predictive tests to select novel treatment options for patients with LM in the future. Citation Format: Caecilia Koestler, Giancarlo Feliciello, Florian Lueke, Kathrin Weidele, Saida Zoubaa, Peter Hau, Tobias Pukrop, Markus J. Riemenschneider, Christoph A. Klein, Bernhard Polzer. Single cell analysis from CSF of patients with suspected leptomeningeal metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3709.

Abstract 1457: Decoding the molecular mechanisms related to mutations in the GNAQ/11 and BAP1 genes in ocular melanoma

Abstract Uveal melanoma (UM) is the most frequent intraocular cancer in adults and 50% of patients develop liver metastases for which no treatment is effective. According to The Cancer Genome Atlas, 92% of cases are characterized by early mutations in two genes coding for alpha subunits of G protein coupled receptors (GPCR): G protein subunit alpha q (GNAQ) or 11 (GNA11). These mutations are mostly found in the codon Q209 causing a constitutive activation of different signaling pathways linked to cellular proliferation such as MAPK and PKC. Furthermore, a bi-allelic inactivation of the BRCA1 associated protein-1 (BAP1) is found in 83% of metastatic UM and causes a loss of function of this protein. Our hypothesis is that the inactivation of compensatory pathways in the absence of these proteins and the understanding of pathways essential only to mutated cells will pave the way for the identification of better therapeutic candidates specific to UM cancer cells (UMCCs) to prevent metastatic progression and improve treatment options. The research objectives include the identification of compensatory signaling pathways in the absence of GNAQ/11 and BAP1 using classical CRISPR/Cas9, the discovery of essential molecular mechanisms for the survival of mutated UMCCs using Prime Editing, and the identification and characterization of novel therapeutic targets using 3D in vitro and in vivo models. First, the electroporation optimization using an eGFP plasmid demonstrated an efficiency of 40-50% for normal choroidal melanocytes (NCM) and 60-80% for UMCCs Mel285. Subsequent transfection with CRISPR complexes resulted in an average editing efficiency of approximately 20% for GNAQ/11 and 60% for BAP1. Currently, we have successfully established a GNAQ knockout (GNAQ-KO) Mel285 cell line. The characterization and validation of this cell line were performed through sequencing and Western blotting, confirming the creation of stop codons and the absence of GNAQ expression. We observed an overactivation of the ERK1/2 pathway and an upregulation of beta-catenin in the Mel285 GNAQ-KO line. Additionally, we noted a decrease in proliferation, adhesion, and migration on collagen I. For Prime editing, we constructed three plasmids with desired mutations. Preliminary tests in HEK293T cells showed an insertion efficiency of 4%, 20%, and 10% for GNAQ, GNA11, and BAP1, respectively, following a single treatment. Finally, in vitro testing has been started for inhibitors targeting compensatory pathways involving beta-catenin (WIKI4) and ERK1/2 (ASN007), in combination with the GNAQ/11 inhibitor (FR900359). These tests are being conducted on various mutant UMCCs for GNAQ/11.A better understanding of the altered pathways in our GNAQ/11 or BAP1 mutant isogenic cell lines will help to identify new drugs targeting specifically UM cells. Citation Format: Aurelie Fuentes-Rodriguez, Andrew Mitchell, Joël Rousseau, Jacques P. Tremblay, Solange Landreville. Decoding the molecular mechanisms related to mutations in the GNAQ/11 and BAP1 genes in ocular melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1457.

Abstract 235: Application of high-grade serous ovarian cancer organoids in functional precision medicine

Abstract High-grade serous ovarian cancer (HGSC) is the most frequent and lethal ovarian cancer type. Despite responding favorably to first-line therapy, most HGSC patients eventually relapse. HGSC is characterized by high interpatient genomic heterogeneity and few recurrent mutations that could be used as therapeutic biomarkers. Thus, except the introduction of PARP inhibitors for BRCA1/2-mutated patients, there has been little progress with personalizing HGSC treatment. Functional precision medicine - tailoring personalized therapies based on functional assays on patient-derived cells - has been applied in cancer clinical trials to address this issue. Organoids - three-dimensional cell cultures, recapitulating aspects of original tumors’ structure and function - have often been used as models in these efforts. However, inefficient and time-consuming organoid derivation protocols have been hindering the application of HGSC organoids in functional precision medicine clinical trials, due to the short time during which the therapy must be selected. To address this, we here aimed to establish whether we could predict second-line therapy drug sensitivities in relapsed HGSC using organoids from tumor material from first-line therapy. We have previously derived a collection of 23 organoid models from 16 patients, sampled sequentially at different times of their treatment (1). Here, for several organoid primary-relapse pairs, we performed drug screens using a library of 370 compounds (approved, in clinical development or tool compounds). We found that 90% of the hit compounds in the primary organoids also were hits in the matched relapse-derived organoid models. In several patients, we observed a pronounced cytotoxicity of several cyclin-dependent kinase (CDK) inhibitors, all of which potently inhibit CDK9, including two selective inhibitors (AZD4573 and VIP152) in clinical development. Screening of CDK9 inhibitors in an extended panel of organoid models revealed that most organoids are sensitive to CDK9 inhibition in nanomolar range. CDK9 inhibitors also prevented organoid outgrowth in long-term culture (>1.5 month) in drug-free medium. Taken together, we demonstrate that HGSC organoids from material sampled during first-line treatment predict most drug sensitivities of relapse-derived organoids. This suggests a feasibility of functional precision approach for selection of second-line therapy using only primary tumor material, circumventing the problem of limited time available for the therapy choice after tumor sampling. This work also highlights that CDK9 inhibitors, currently in clinical development for hematological malignancies, are worthwhile to be explored in HGSC. 1. Senkowski et al. A platform for efficient establishment and drug-response profiling of high-grade serous ovarian cancer organoids. Developmental Cell (2023) Citation Format: Wojciech Senkowski, Laura Gall-Mas, Matias M. Falco, Yilin Li, Kari Lavikka, Aura Crispino, Jaana Oikkonen, Daria Bulanova, Yan-Jun Chen, Elena M. Doncel, Karolin Voßgröne, Erdogan P. Erkan, Jun Dai, Anastasia Lundgren, Jutta Huvila, Kaisa Huhtinen, Johanna Hynninen, Sampsa Hautaniemi, Anna Vähärautio, Krister Wennerberg. Application of high-grade serous ovarian cancer organoids in functional precision medicine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 235.

Abstract 6346: IgG engineered to target PIGR-mediated transcytosis specifically targets intracellular oncoproteins and abrogated tumor growth

Abstract Despite their long half-life, therapeutic antibodies are considered ineffective against intracellular antigens due to their perceived inability to penetrate epithelial cells. Using recombinant antibodies targeting common mutations in oncogenes, we have demonstrated that dimeric IgA, but not the same antibody on an IgG backbone, penetrates human epithelial cancer cells through PIGR-dependent directional transcytosis, specifically neutralizing mutated oncodrivers and expelling antigens outside the cell, bound to secretory IgA. Accordingly, targeting of KRASG12D abrogated tumor cell proliferation in cell culture assays. In vivo, KRASG12D-specific dIgA1 limited the growth of RASG12D-mutated ovarian and lung carcinomas, including in syngeneic tumor-bearing immunocompetent mice. Moreover, dIgA targeting of KRASG12D restricted tumor growth more effectively than small-molecule KRASG12D inhibitors, supporting the potential of this approach for the treatment of human cancers. Since producing dimeric IgA is more challenging than producing monomeric IgG, and because the half-life of IgA is shorter (4-7 days) than IgG (21 days), we have developed a new approach combining KRASG12D specific IgG with a peptide that could join PIGR and penetrates tumor cells through transcytosis, that showed the same effect than IgA controlling tumor growth. Our results provide a rationale for developing modified IgG cell-penetrating antibodies to effectively target common mutations in intracellular oncogenes that drive many aggressive and frequent human cancers. In addition, further experiments suggest that these antibodies could be effectively used to target tumors that have become resistant to existing KRAS inhibitors. Citation Format: Carmen Maria Anadon Galindo, Luis U. Lopez-Bailon, Ricardo A. Chaurio, Jessica A. Mine, Noah E. Plappert, Jose R. Conejo-Garcia. IgG engineered to target PIGR-mediated transcytosis specifically targets intracellular oncoproteins and abrogated tumor growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6346.

Abstract 2673: Copper chelation overcomes the immunosuppressive neuroblastoma microenvironment to potentiate anti-GD2 antibody therapy

Abstract Neuroblastoma is the most frequent extra-cranial pediatric cancer with high-risk disease possessing a dismal five-year survival rate <50%. Antibody-based therapies targeting the surface disialoganglioside GD2 have shown promising anti-tumor activity in patients however efficacy is strongly hampered by immunosuppressive mechanisms present in the tumor microenvironment. Neuroblastomas typically feature low mutational burdens, limited major histocompatibility complex class I expression, increased immune checkpoint markers, sparse immune infiltration, and soluble mediators which drastically limit anti-tumor immunity. Given the emerging link between elevated copper levels and tumoral immune evasion, we explored copper chelation as a therapeutic strategy using the agents tetraethylenepentamine (TEPA) and the clinically approved analogue triethylenetetramine (TETA). Using the immunocompetent TH-MYCN model, we performed single-cell RNA sequencing supported by OPAL multiplex immunohistochemistry and cytokine profiling of resected tumors to assess cellular and molecular changes occurring after one week of TEPA treatment. Therapy was observed to successfully deplete intratumoral copper and reinvigorate anti-tumor immunity with increased infiltration and activity of pro-inflammatory immune cells, and was specifically found to induce egress and N1-polarisation of neutrophils. Further mechanistic studies in vitro revealed the sequestration of copper by neuroblastoma cells attenuated neutrophil function, which could be successfully reversed upon TEPA treatment. Findings propose a novel mechanism of immune evasion, highlighting copper chelation as a therapeutic strategy to counteract immunosuppression. Using the TH-MYCN model, TEPA was employed as a priming agent to effectively potentiate anti-GD2 antibody therapy and achieve durable tumor control. Importantly, this was associated with increased Fc-receptor-bearing natural killer and myeloid cells, which elicit tumor clearance via antibody-dependent cellular cytotoxicity. Currently indicated for the treatment of copper overload disorder Wilson’s disease, TETA was evaluated for potential repurposing as a novel immunomodulatory agent. Using the syngeneic NXS2 model, TETA exhibited an exceptional safety profile and combination therapy achieved durable tumor control with no relapses occurring after treatment cessation. This result was similarly associated with the infiltration of pro-inflammatory immune cells, including N1 neutrophils. Collectively, this study credentials copper chelation as a non-toxic strategy to disrupt the immunosuppressive tumor microenvironment and reinvigorate anti-tumour immunity. Findings provide crucial translational evidence for repurposing TETA to potentiate anti-GD2 antibody therapy and improve neuroblastoma patient responses. Citation Format: Jourdin R. Rouaen, Antonella Salerno, Fabio Luciani, Jayne Murray, Tyler Shai-Hee, Nicodemus Tedla, Michelle Haber, Toby N. Trahair, Orazio Vittorio. Copper chelation overcomes the immunosuppressive neuroblastoma microenvironment to potentiate anti-GD2 antibody therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2673.

Abstract 320: Development and automation of a streamlined targeted enrichment method for cancer mutation detection

Abstract Next Generation Sequencing (NGS) of DNA is a powerful tool for the detection of genetic variations, including single nucleotide polymorphisms, copy number variation, and small insertions/deletions. Target enrichment technologies enhance DNA sequencing by enabling users to analyze specific regions of interest only, which helps to effectively increase sequencing depth and sample throughput while minimizing cost. However, many targeted DNA library construction workflows are not optimal for routine genetic testing needs due to inconsistent, lengthy workflow. This poses significant challenge to application like cancer mutation profiling that requires detection sensitivity down to 1% or below due to heterogeneous nature of tumor tissue. In addition, high throughput testing with minimal hands on time is desirable with the rapid advance of precision medicine. Magnetic bead separations are commonly used between the individual reactions of targeted DNA library preparation workflows for cleanup and size selection. These bead clean up reactions are not only labor-intensive and tedious procedures but can also greatly contribute to sample-to sample variation especially when employing automated library preparation procedures. With the sophisticated QIAseq Targeted DNA Pro, we developed a novel streamlined targeted DNA workflow that greatly reduces handling variation by replacing the majority of bead clean up steps with enzymatic reactions, which now can be finished in as little as six hours. In combination with the newly developed QIAseq Normalizer kit, the protocol has been successfully implemented on the Hamilton NGS Star liquid handling platform resulting in up to 96 highly uniform, complex and ready-to-sequence libraries that can be effortless generated in one run. This complete automated solution saves researcher up to four hours of hands-on time. When using commercial reference samples, cancer mutations down to 1% were consistently detected with this automated workflow. The presented data demonstrate that this integrated and streamlined workflow does not only provide an automation friendly workflow but also allows to reliably identify low frequent cancer mutations. The applications presented here are for research use only. Not for use in diagnostic procedures. Citation Format: Zhong Wu, Michelle Baird, Cornelia Mechlen, Peter Hahn, Jonathan M. Shaffer. Development and automation of a streamlined targeted enrichment method for cancer mutation detection [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 320.

Abstract 3468: Emerging lessons from patient-reported hospital discharge experiences among hospitalized cancer patients with frequent emergency department use

Abstract Background: While teamwork is essential to providing high-quality patient-centered care, challenges in interprofessional collaboration and decision-making in the hospital settings are common, especially for cancer patients. The purpose of this study was to identify emerging themes related to hospital discharge experiences among patients hospitalized for cancer who became frequent emergency department (ED) users following their hospital discharge. Design & Methods: A cohort of cancer patients discharged from an academic health center in Montreal (Canada) between October 2014 and November 2016 was assembled. Frequent ED (FED) users were identified as patients who had a > 4 ED visits in the year following hospital discharge using health administrative claims from the provincial universal health care program. Qualitative analysis of telephone interviews conducted with patients 30 days’ post-discharge were used for in-depth exploratory analyses to characterize hospital discharge experiences and transition process from the hospital to the community. Results: A cohort of 1253 cancer patients was formed. The mean age was 70.9 (SD=11.8) and the most frequent cancers included 488 (38.9%) respiratory and 309 (24.6%) upper digestive cancer. Overall, 14.5% (n=182) of patients became FED users. Content analyses revealed the most common emerging themes from the FED patients interviews on hospital discharge experiences. These included:1. Early hospital discharge putting patients at high risk of being re-admitted and going back to the ER shortly after that. Some patients mentioned post-discharge complications and emerging of new health issues that could have been avoided if patient was kept in the hospital for longer.2. Lack of communication between different specialists at the hospital. Some patients mentioned the help of a nurse as crucial during inpatient stays in maintaining communication between doctors. 3. Lack of communication of medications prescribed. Some patients mentioned the lack of communication of what doses was modified, what medications were stopped and which ones were newly prescribed.4. The need to schedule follow-up appointments at the time of hospital discharge. This becomes especially important for vulnerable patients who have been on pain medication during the hospital stay, affecting their cognitive abilities and making post-discharge planning more difficult. Conclusions: This study using integrated data from administrative claims and patient interviews provided insights into the challenges related to hospital discharge experiences and transition into community among hospitalized cancer patients with frequent emergency department use. Application of our findings could assist in hospital discharge preparation and improvement in healthcare delivery and health outcomes. Citation Format: Siyana Kurteva, Robyn Tamblyn. Emerging lessons from patient-reported hospital discharge experiences among hospitalized cancer patients with frequent emergency department use [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3468.

Abstract 4344: Comprehensive analysis of LINE-1 transposable elements in colorectal cancer

Abstract Background: Colorectal cancer (CRC) is the third most frequent cancer worldwide, and approximately a third of patients die from the disease. There is therefore an urgent need to better understand CRC biology. A particular feature in (colorectal) cancer is the activation of LINE-1 (L1) retrotransposons: DNA fragments which propagate to other places in the genome through an RNA intermediate. Per genome, about 100-150 genomic L1 copies are still retrotransposition-competent, but repressed in most somatic tissues. Their role in tumor development is currently unknown. Recently, the PCAWG consortium characterized the landscape of somatic L1 retrotranspositions in a series of primary cancers of different origin, and identified 124 recurrently active L1 source regions. However, only a limited number of these concerned CRC. Aim: This study aims to perform a comprehensive analysis of L1 activity in a large cohort of metastatic colorectal cancer. Methods: To this end, we retrieved data from the Hartwig Medical Foundation, comprising deep (106x) whole genome sequencing (WGS) data from metastatic CRC (n=745), to compare to breast (n=888) and lung cancer samples (n=640). We determined L1 activity as the number of somatic L1 insertions reported by the HMF DNA analysis pipeline and used the activity of each of the 124 L1 sources as previously reported by the PCAWG consortium. Results: 99% of mCRC samples (n=741) displayed at least 1 somatic L1 retrotransposition, with at least 10 retrotranspositions in 89% of samples. L1 retrotransposition is more prevalent in mCRC than in metastatic lung and breast cancer, in which only 40% and 20% had at least 10 retrotranspositions per sample, respectively. Each cancer type showed a unique profile of active L1 sources. We did not observe a clear preference of L1 insertions for a single genomic region. However, in CRC, the gene most frequently affected in an exon was APC (n=13), an important tumor-suppressor gene. An analysis of somatic SNVs and SVs confirmed an association of mutations in some known L1 regulators such as TP53 with increased L1 activity, whereas mutations in others showed no change in the number of somatic retrotranspositions. Conclusions: We conclude that somatic L1 retrotranspositions are highly prevalent in metastatic colorectal cancer. Presence of L1 insertions in APC suggests retrotranspositions start very early in CRC development and prompts further investigation into earlier CRC stages. Citation Format: Renske de Wit, Soufyan Lakbir, Caterina Buranelli, Gerrit A. Meijer, Sanne Abeln, Remond J. Fijneman. Comprehensive analysis of LINE-1 transposable elements in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4344.

Abstract 4778: Development of self-amplifying RNA vaccines targeting prostate tissue-restricted and tumor-associated antigens

Abstract Prostate cancer is the most frequent cancer type in men, with increasing incidence at advanced age. As this cancer is frequently slow growing, opportunities for cancer interception exist, particularly among men with low-grade tumors who frequently undergo active surveillance. A well-tolerated vaccine intervention would be ideal for this population. Self-amplifying RNA (saRNA) vaccines built on a replication-deficient alphavirus backbone such as VEEV, offer several advantages for vaccine development. Sequences for relatively large antigens can be encoded and efficiently expressed, at low vaccine doses. Practical and efficient delivery of the saRNA is possible with advanced nanoparticles such as cationic inorganic lipid nanoparticles (LION). Among candidate antigens for a prostate cancer interception vaccine, we selected prostein (SLC45A3), a protein whose natural expression is restricted to the prostate gland and prostate cancer and ERG, a member of the ETS family of transcription factors frequently involved in chromosomal translocations in prostate cancer. We generated saRNA constructs with several design strategies and demonstrated that upon in vitro transfection, both antigens are expressed as assessed by mass spectrometry and Western blot analysis. This was achieved using commercial transfection reagents and LION. Prostein expression was particularly notable in that this eleven-transmembrane protein could be detected in HEK-293 cells using flow-cytometry and cell-surface staining. Nearly 50% of the cell population stained positively. Immunogenicity of the vaccines was tested in mice. Following formulation with LION particles i.m. injection and boosting, saRNAs coding for prostein elicited strong cellular immunity as demonstrated by IFN-γ ELISPOT analysis. Detailed analysis of intracellular cytokine expression indicated a strong CD8 response. saRNAs coding for ERG also showed antigen-specific ELISPOTs. Using commercially available recombinant ERG protein, an ELISA was developed and a strong humoral response to the ERG saRNA vaccine was demonstrated. The saRNA vaccines will be further credentialled with additional preclinical studies including cancer preventive efficacy studies in genetically engineered mouse models of prostate cancer. Citation Format: Robert H. Shoemaker, Jesse H. Erasmus, Peter Berglund, Jason D. Marshall, Yuri Koboziev, Bradley M. Beckman, Brian D. Cholewa, Steven G. Reed. Development of self-amplifying RNA vaccines targeting prostate tissue-restricted and tumor-associated antigens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4778.