bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2161: Quinolinic acid phosphoribosyl transferase (QPRT) is an essential liability of non-small cell lung cancer
Authors: Kashfi Hossein , Ward Nathan , Ilter Didem , Drapela Stanislav , Falzone Aimee , Gardell Stephen , DeNicola Gina M. , Gomes Ana P.
Published: April 2023

Lung cancer is the second most diagnosed type of cancer and the leading cause of cancer-related mortality. 85% of all lung cancer cases are non-small cell lung cancer (NSCLC), a large proportion of which are only diagnosed at advanced stages (II-IV) due to lack of early diagnostic techniques. Standard first-line treatment for late-stage inoperable NSCLC still is chemotherapy regiments. However, most NSCLC patients fail to respond to these regimens or relapse upon treatment. Thus, there is a pressing need to identify novel therapeutic targets that can more effectively treat NSCLC. Quinolinic acid phosphoribosyl transferase (QPRT) is a rate-limiting enzyme in the tryptophan catabolic pathway which fuels de novo NAD+ production, whose expression as previously been shown to have a prognostic value in certain cancers. Here, we demonstrate that high QPRT levels is a feature of a large proportion of NSCLC cell lines and that QPRT induction occurs in genetically engineered models of NSCLC (KP and KL) when compared to the normal lung. Considering the critical role of NAD+ levels to enable high rates of proliferation, we hypothesized that QPRT induction enables lung cancer cells to thrive. Strikingly, QPRT knockdown in a panoply of NSCLC cell lines results in pronounced suppression of tumor cells both in 2D and 3D conditions. Interestingly, we observe that QPRT suppression does not affect proliferation or cell cycle progression but rather due to pronounced induction of cell death. Surprisingly, while we observed DNA damage upon QPRT suppression, NAD+ levels were not affected by QPRT suppression indicating that QPRT’s effects in NSCLC are independent of its contribution to the NAD+ pools. Together, our results point for QPRT as a novel and effective therapeutic target for NSCLC.

Citation Format: Hossein Kashfi, Nathan Ward, Didem Ilter, Stanislav Drapela, Aimee Falzone, Stephen Gardell, Gina M. DeNicola, Ana P. Gomes. Quinolinic acid phosphoribosyl transferase (QPRT) is an essential liability of 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 2161.
Abstract 2361: Prevalence of mature tertiary lymphoid structures and association with tumor mutational burden in patients with solid tumors
Authors: Brunet Maxime , Vanhersecke Lucile , Loarer Francois Le , Soubeyran Isabelle , Italiano Antoine
Published: April 2023

Background: Mature Tertiary Lymphoid Structures (mTLS) are organized lymphoid structures containing B-lymphocytes admixed to CD23+ follicular dendritic cells, which play a key role in antitumor immune response. We and others have demonstrated that presence of mature TLS is higly predictive of benefot of anti-PD1/PDL1 antibodies in patients with solid tumors (Vanhersecke et al Nature Cancer 2021, Italiano et al Nature Medicine 2022). However, the relative prevalence of mature TLS in the different tumor types has not been investigated with an uniform methodological approach.

Methods: We screened the presence of mature TLS (mTLS) on tumor samples from patients included in an institutional molecular profiling program (BIP, NCT02534649). mTLS were screened using HES, CD20 (clone L26, Ventana) and CD23 (clone 1B12, Novocastra) stainings, performed on serial sections. TLS were defined as lymphoid aggregates of CD20+ B-lymphocytes of more than 50 immune cells within 1mm of the tumor front. mTLS were defined as TLS containing either a visible germinal center on HES or a meshwork of CD23+ follicular dendritic cells or isolated CD23+ dendritic cell if the cell harbored a morphology fitting with dendritic differentiation. If plasma sample of patient was available, bTMB was assessed by Foundation Medicine, with bTMB-high defined as at least 10 mutations/megabase (mut/Mb).

Results: We characterized 1,393 samples. mTLS were present in 467 cases (33.5%) across all tumor histotypes: 36.2% of carcinoma (N = 392/1080 cases), 24.8% of sarcoma (N =69/209 cases) and 25% of neuroendocrine tumor (N= 4/12 cases).

Among 569 cases with evaluable bTMB, the proportion of patients with bTMB-high was significantly higher in the mTLS group (16.7%, N=31/185) compared to absence of mTLS group (8.5%, N=30/353) (p=0.036).

Conclusion: mTLS are present in variable proportions in almost all tumor types even in those considered as resistant to current immune checkpoint inhibitors.

Prostate Bladder Kidney Head and neck Gynecologic (except ovary) Ovary mTLS+% of patients (N) 34% (28) 33% (19) 25% (6) 53% (17) 33% (19) 44% (26) mTLS-% of patients (N) 66% (55) 67% (38) 75% (18) 47% (15) 67% (39) 55% (32) Upper gastro intestinal tract Pancreatic Biliary tract Breast Thyroid Sarcoma mTLS+% of patients (N) 37% (19) 50% (13) 40% (21) 30% (38) 49% (24) 25% (69) mTLS-% of patients (N) 63% (33) 50% (13) 60% (32) 70% (90) 51% (25) 75% (209)

Citation Format: Maxime Brunet, Lucile Vanhersecke, Francois Le Loarer, Isabelle Soubeyran, Antoine Italiano. Prevalence of mature tertiary lymphoid structures and association with tumor mutational burden in patients with solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2361.
Abstract 2368: Pilot characterization of the salivary microbiome in pediatric cancer patients
Authors: Garduño Alejandra Beatriz Cervantes , García Milca de la O , Nagano Jesús Adolfo Yamamoto , Cardos Rocio Del Socorro Cardenas , Hidalgo Liliana Velasco , García Blanca Olivia Almazán , Celis Gabriela Elisa Mercado
Published: April 2023

Microbial populations in the oral cavity have been linked with some malignant diseases, however, the evidence in this regard is poor. Oral microorganisms increase cytokine production and other mediators may be related to carcinogenesis. However, the role of the oral microbiome and its taxonomic composition in pediatric cancer has not been studied. We report a pilot study to assess to compare the microbial diversity of saliva in pediatric patients with cancer and without cancer. Materials and methods: Total saliva samples were collected from 3 cases and 3 controls with cancer, with prior informed consent and assent. Metagenomic sequencing was performed and data was analyzed using CosmosID bioinformatics software. Results: the mean age for the cases is 8.67 years and for the 8-year-old controls, the percentage by sex was 33.3% male and 66.6% female. In the shared analysis of bacterial diversity we found a total of 93 genera, the three genera with the highest abundance in the cases were: Prevotella, Streptococcus and Morococcus. A total of 153 species with differential abundance were identified, in the cases the species with the highest abundance were: Neisseria mucosa, Vellodela dispar, Prevotella melaninogenica and Morococcus cerebrosus. We found differences in bacterial diversity and abundance between cases and controls. The main species that we found in greater abundance in the cases have been related to various processes in the oral cavity, Streptococcus sp increases child caries, Prevotella melaninogenica a broad profile of resistance to families of antibiotics, and Morococcus cerebrosus has been associated with the mineralization of the supragingival dental plaque. Finally Vellodela dispar has been associated with bacteremia in cancer patients. In this pilot study a small number of samples were analyzed, the differences in bacterial diversity should be confirmed by conducting studies with a larger number of samples.

Citation Format: Alejandra Beatriz Cervantes Garduño, Milca de la O García, Jesús Adolfo Yamamoto Nagano, Rocio Del Socorro Cardenas Cardos, Liliana Velasco Hidalgo, Blanca Olivia Almazán García, Gabriela Elisa Mercado Celis. Pilot characterization of the salivary microbiome in pediatric cancer 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 2368.
Abstract 237: Coincidence of InDels with missense mutations: 0, 1 or 2 artifacts
Authors: Craven Kelly E. , Fischer Catherine G. , Jiang LiQun , Pallavajjala Aparna , Lin Ming-Tseh , Eshleman James R.
Published: April 2023

The purpose of this study was to explain a serendipitously discovered next generation sequencing (NGS) artifact. While reviewing clinical cases, we found a case of a large deletion that also contained multiple missense mutations. This phenotype was unknown to several molecular pathologists in our group. We reviewed 46 clinical cases with short insertions or deletions (InDels) (1-30 bases) or FLT3 internal tandem duplications (ITDs) (6-183 bases) of solid or hematologic malignancy processed with a clinical NGS assay and identified misaligned reads, ranging from 3 to 100% of reads showing mismapped bases. Reads that straddled the InDel with sufficient numbers of bases on both sides were correctly anchored both upstream and downstream of the InDel, and were called correctly. However, reads that ended in the middle of the InDel were incorrectly mapped onto the deletion, thereby producing artifactual missense mutations. The frequency of mismapped bases increased with InDel size, and the VAF of the InDel can be markedly underestimated. The ratio of forward to reverse reads was commonly skewed for the mismapped base compared to the ratio for the wildtype base at that position. ABRA2 was able to correct 41 to 100% of the reads with mismapped bases and led to absolute increases in the VAF from 1 to 61% along with correction of all of the SBSs except for two cases. Knowledge of the various artifacts commonly seen in next generation sequencing is essential to safely sign out molecular pathology cases.

Citation Format: Kelly E. Craven, Catherine G. Fischer, LiQun Jiang, Aparna Pallavajjala, Ming-Tseh Lin, James R. Eshleman. Coincidence of InDels with missense mutations: 0, 1 or 2 artifacts [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 237.
Abstract 2414: Early biological impacts of radiotherapy-activated NBTXR3 nanoparticles
Authors: Silva Jordan Da , Bienassis Célia , Paris Sebastien
Published: April 2023

Since many decades, radiotherapy (RT) is worldwide used and crucial therapeutic approach for the treatment of cancers. Nowadays, about 50% of cancer patients will receive radiotherapy during their treatment. Nevertheless, the use of RT is limited by the damage induced by ionizing radiation on healthy tissues. To overcome this constraint, we have developed high electron density of functionalized hafnium oxide nanoparticles (NBTXR3), allowing a high probability of interaction with incoming ionizing radiation, increasing energy dose deposit within cells. Many preclinical data have shown that RT-activated NBTXR3 (NBTXR3+RT) increase cancer cell destruction as well as better control treated tumor growth, compared to RT alone. In clinic, the benefit of NBTXR3+RT compared to RT alone has been demonstrated for patients with locally advanced Soft Tissue Sarcoma, in the randomized controlled phase II/III Act.in.Sarc study (NCT02379845). We also demonstrated in preclinical settings that, beyond the destruction of tumor cells, NBTXR3+RT was also able to induce an antitumor immune response capable to produce a significant abscopal effect, linked to the increase of CD8 infiltrates in tumors. It has been recently reported that NBTXR3+RT can also restore anti-PD1 efficacy in an anti-PD1 resistant mouse model. At the cancer cell level, we reported that NBTXR3+RT could modulate the immunopeptidome of cancer cells, induce the activation of the cGAS-STING pathway and increase the expression of ICD biomarkers. All these data indicate that NBTXR3+RT has the ability to transform the tumor into an in situ vaccine more efficiently than RT alone and could have important implications for the use of NBTXR3+RT in combination with immunotherapy. Nevertheless, we currently have relatively few elements on the early cellular events that could trigger or explain the above reported performances of NBTXR3+RT, compared to RT alone. To address this question, we have carried out a series of in vitro studies (cytometry, microscopy) on different cancer models (human and murine). We have thus identified that the vast majority of NBTXR3 nanoparticles are located in lysosomes after endocytosis by tumor cells. Once activated by RT, we detected lysosomal membrane permeabilization (LMP) only in cells treated with NBTXR3+RT, but not in cells only treated by RT. Following LMP, we measured a significant increase of in lipid peroxidation. These first elements allow a better understanding of the early biological consequences induced by NBTXR3+RT, compared to RT alone.

Citation Format: Jordan Da Silva, Célia Bienassis, Sebastien Paris. Early biological impacts of radiotherapy-activated NBTXR3 nanoparticles [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 2414.
Abstract 2417: Liver regeneration after radiation therapy: clinical observations and a new rat model
Authors: Su Tingshi , Zhao Weidong , Li qing , Wu Qiaoyuan
Published: April 2023

Background and Purpose: Radiation-induced liver damage, a dose-limiting complication of radiotherapy for liver tumors, has been focused mainly on prevention and treatment but has largely ignored the molecular mechanisms underlying the liver’s regenerative capacity after irradiation. Whether liver regeneration compensates for radiation-induced liver damage is of great clinical significance. This study aims to observe clinical liver regeneration and the potential molecular mechanism in rat models after irradiation.

Methods: To stimulate the liver regeneration response, we delineated a liver protection area in the patient’s normal liver to protect it from excessive irradiation, and designed a rat model in which large single fractions of 25 Gy or 15 Gy of radiation were delivered to 1/2 or 2/3 of the total liver volume to investigate liver regeneration after irradiation. We used proteomics analysis and western blotting assays to investigate liver regeneration signaling and molecular mechanisms.

Results: Four patients’ pairs of liver specimens from areas of high irradiation and regeneration (protected liver) were collected. Proteomics analysis showed enrichment in several pathways, with some of the greatest changes observed in PI3K-Akt signaling, further confirmed in liver-irradiated rat models. After irradiation, the liver injury index was significantly elevated on day-1 and gradually recovered to normal levels by day-8 in rat models. The model of a single 25-Gy fraction delivered to half of the liver volume best represented the clinically similar phenomenon of liver regeneration in the protected liver. We tested the effects of an Akt/mTOR activator (carbamazepine), which enhanced regeneration, and an Akt/mTOR inhibitor (temsirolimus), which inhibited regeneration. Finally, liver regeneration was enhanced in rats treated with temsirolimus followed by carbamazepine.

Conclusions: Our results suggest that protecting the normal liver outside the irradiated area enhances the probability of successful regeneration to compensate for radiation-induced liver injury. PI3K/AKT/mTOR is essential in liver regeneration after irradiation, and carbamazepine may enhance liver regeneration after irradiation in rats.

Citation Format: Tingshi Su, Weidong Zhao, qing Li, Qiaoyuan Wu. Liver regeneration after radiation therapy: clinical observations and a new rat 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 2417.
Abstract 2439: RPL27 augments growth and stemness properties in colorectal cancer through PLK1 signaling
Authors: Park Keon Uk , Hwang Ilseon , Ryoo Hun Mo , Lee Yun-Han , Park So-Young
Published: April 2023

Background: In 2020, colorectal cancer (CRC) had the second highest mortality rate worldwide and was the third most common cancer type among various carcinomas, accounting for 10.0% and 9.4% of all carcinomas, respectively. Although the expression of ribosomal protein L27 (RPL27) is upregulated during CRC development, oncogenic roles of RPL27 have not yet been elucidated. We aimed in this study to investigate the effect of RPL27 targeting on CRC growth and whether RPL27 gained an extra-ribosomal function during CRC development.

Methods: Human CRC cell lines, HCT116 and HT29, were transfected with RPL27-specific siRNA and tested for growth inhibition and apoptotic induction using MTS, FACS and Western blotting. To obtain insights into the molecular changes in response to RPL27 knockdown, global changes in gene expression were examined using RNA sequencing.

Results: We observed that inhibition of RPL27 expression suppresses CRC cell growth and long-term colony formation through an induction of apoptotic cell death. Targeting RPL27 also significantly inhibited the growth of human CRC xenografts in nude mice. Noticeably, polo-like kinase 1 (PLK1), which is known to play an important role in mitotic cell cycle progression and stemness, was commonly down-regulated in both HCT116 and HT29 cells with RPL27 depletion. Western blot analysis confirmed that RPL27 silencing reduces the levels of PLK1 protein and G2/M-associated regulators, such as p-CDC25C, CDK1, and cyclin B1. Reflecting these molecular responses, knockdown of RPL27 reduced the migration and invasion abilities of CRC cells. In terms of CSC phenotypic changes, RPL27 knockdown suppressed sphere forming capacity of isolated CD133+ CSC population accompanying with the reduction in CD133 and PLK1 levels.

Conclusion: Taken together, these findings indicate that RPL27 contributes to promotion of CRC growth and stemness through PLK1 signaling and that targeting RPL27 could be a next-line therapeutic strategy for both primary CRC treatment and prevention of metastasis and/or recurrence.

Citation Format: Keon Uk Park, Ilseon Hwang, Hun Mo Ryoo, Yun-Han Lee, So-Young Park. RPL27 augments growth and stemness properties in colorectal cancer through PLK1 signaling [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 2439.
Abstract 2452: A study of AXIN2-regulatory mechanisms and their role in the stem cell origin of human CRC
Authors: Zhang Chi , Facey Caroline , Opdenaker Lynn , Boman Bruce
Published: April 2023

Colorectal cancer (CRC) originates from stem cell (SC) overpopulation, which is driven by constitutively activated WNT-signaling due to mutations in genes that encode components of the WNT-signaling pathway, particularly APC. WNT-signaling is regulated by the ꞵ-catenin destruction complex, which contains APC, AXIN1, AXIN2 and other proteins. Availability of AXIN1/2 proteins is the rate-limiting factor of destruction complex function.

The Gap in Our Knowledge is how to target AXIN in order to restore destruction complex function in APC-mutant colon tissues. A growing body of scientific evidence shows both AXIN1 & AXIN2 promote ꞵ-catenin degradation and AXINs are hub proteins involved in regulation of multiple cellular pathways that play a role in tumor phenotype. While AXIN2 expression is up-regulated in CRC, the increased level of AXIN2 is incapable of deterring CRC progression. This evidence indicates that AXIN2 is in a key pivotal position to therapeutically target WNT signaling whereby the availability of AXIN2 can be used to fine-tune ꞵ-catenin levels.

We HYPOTHESIZE that the intracellular level of AXIN2 and AXIN1 determines the degree of WNT-signaling activity in APC-mutant CRC cells and affects tumor cell phenotypic properties relevant to CRC progression. Accordingly, we have created siRNA-mediated AXIN2 knockdown and lentiviral-based AXIN2 overexpressing HT29 and SW480 cells to study the effect of modulating AXIN2 levels.

The RESULTS show that decreasing AXIN2 leads to increased WNT-signaling and increasing AXIN2 leads to increased expression of WNT-target genes. Moreover, increasing AXIN2 leads to diminished tumor cell phenotype as shown by: 1) lower cell proliferation rate; 2) up-regulated expression of SC markers (ALDH & LGR5) and increased SC population size (ALDEFLUOR+ or LGR5+); 3) down-regulated cell cycle rate; 4) down-regulated apoptosis. Moreover, our bioinformatics analyses of the effect of tankyrase inhibitors (TIs), which prevent AXIN degradation, indicate that efficacy of TIs against CRC cells depends on the APC genotype. Thus, our findings indicate that AXIN2, by down-regulating WNT-signaling, reduces cancer stem cell phenotype including decreased SC population size.

SIGNIFICANCE: discovering ways to target AXIN2-based mechanisms has vast clinical importance for developing new treatment against CRC.

Citation Format: Chi Zhang, Caroline Facey, Lynn Opdenaker, Bruce Boman. A study of AXIN2-regulatory mechanisms and their role in the stem cell origin of human CRC [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 2452.
Abstract 2461: The cell migration inhibitor dihydromotuporamine C regulates actin-myosin contractility and actin polymerization
Authors: von Kalm Laurence , Seavey Corey
Published: April 2023

The motuporamines are a promising class of anti-metastatic compounds. Dihydromotuporamine C (Motu33) has been shown to activate the small GTPase RhoA, however, little is known about subsequent downstream events leading to cell migration inhibition. In the present study, we investigated the mechanism of action of Motu33 and a synthetic derivative, Motu-(CH2)-33, in Drosophila by manipulating the gene dose of positive and negative regulators of actin dynamics. Consistent with previous findings, reduced gene dose of Rho1 (the Drosophila RhoA ortholog) attenuates motuporamine activity, confirming that RhoA/Rho1 is targeted by these compounds. Actin-myosin contraction is controlled by the RhoA-ROCK-myosin regulatory light chain (MRLC) pathway. Reduced gene dose of the myosin binding subunit of myosin phosphatase, a negative regulator of the RhoA-ROCK-MRLC pathway, enhances motuporamine activity indicating that the motuporamines stimulate actin-myosin contraction through activation of the myosin regulatory light chain. RhoA also activates diaphanous (dia) to control actin polymerization. Surprisingly, reduced gene dose of dia enhances motuporamine activity, suggesting that the motuporamines act on dia in a RhoA-independent manner. Reduction in gene dose of the Drosophila Rac orthologs also enhances motuporamine activity. In contrast, motuporamine activity is unaltered by reduced gene dose of slingshot (ssh) which acts to trigger actin severing and depolymerization. Since ssh is directly regulated by Rac, the enhanced activity of motuporamines observed when Rac gene dose is reduced may reflect an indirect mode of action on the Rac GTPases leading to increased RhoA activity. In summary, these findings demonstrate that motuporamines act through RhoA and diaphanous to regulate actin-myosin contractility and actin polymerization.

Citation Format: Laurence von Kalm, Corey Seavey. The cell migration inhibitor dihydromotuporamine C regulates actin-myosin contractility and actin polymerization [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 2461.
Abstract 2464: Novel models of breast cancer metastasis with different metastatic competence
Authors: Bhatt Akshita B. , Wang Ruishan , Jia Hong , Labelle Myriam
Published: April 2023

Metastasis is responsible for poor overall survival of cancer patients and a better understanding of the molecular mechanisms that drive metastasis is crucial for improving disease outcome. In mouse models, gene expression profiling of cancer cells isolated from metastases and from their matched primary tumors is an approach that has been successfully employed to identify biomarkers and drivers of metastasis. However, only few such tumor pairs have been derived from immunocompetent mouse models, limiting our ability to compare the metastatic potentials of these pairs in the context of an intact immune system. Moreover, most of the metastatic cancer cell lines currently available on the C57BL/6 background have a relatively low metastatic potential, complicating the evaluation of therapeutic interventions that may inhibit metastasis.

To address these critical limitations, we developed novel cell lines from distinct spontaneous lung metastases and corresponding primary breast tumors from C57BL/6 mice. Specifically, C57BL/6 mice were orthotopically injected with copGFP-expressing E0771 breast cancer cells and the tumors were allowed to grow for 28 days. Tumor cells at different stages of the metastatic cascade - primary tumor evolution, transit through the blood circulation (circulating tumor cells), survival at secondary site, dormancy, and metastatic outgrowth, were captured by collecting and culturing copGFP-positive cancer cell lines from the primary tumor, blood circulation, and metastatic lungs at the experimental endpoint. These cells were reintroduced orthotopically (mammary fat pad injection) and intravenously (tail-vein injection) to evaluate tumor growth and metastasis in vivo. Interestingly, cell lines derived from overt metastases displayed a wide array of metastatic competences, indicating that some metastases retain that ability to further metastasize while others do not. RNA sequencing of these cell lines identified genes and pathways that are significantly upregulated in highly metastatic vs. poorly metastatic cancer cell lines. Future studies will investigate whether modulation of these pathways in the primary tumor and CTCs can be used to predict metastatic recurrence and serve as a basis for developing novel biomarker-based therapeutic approaches for the treatment of metastatic disease. We also envision that the cell lines we generated will be useful for studies of the influence of host-tumor cells interactions on metastasis on the immunocompetent C57BL/6 background.

Citation Format: Akshita B. Bhatt, Ruishan Wang, Hong Jia, Myriam Labelle. Novel models of breast cancer metastasis with different metastatic competence [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 2464.
Abstract 2469: Investigating the effects of in vivo silencing of RBBP6 gene on tumor growth in the FVBN-Tg(MMTV PyVT)634MulJ breast cancer mouse model
Authors: Moela Pontsho , Govender Mayur , Ambele Melvin
Published: April 2023

Breast cancer is one of the most common causes of cancer mortality, with close to two million new cases and just over 600 000 cancer deaths projected in 2022 in the United States of America alone. Identifying prognostic markers for breast cancer is critical to reducing these numbers. Retinoblastoma binding protein 6 (RBBP6) is considered a potential cancer biomarker due to its association with cell proliferation and the fact that it is overexpressed at breast cancer sites where there is marked apoptosis and elevated p53. More information is emerging regarding the role of RBBP6 in cancer treatment, specifically its potential to sensitize cancer cells to radiation and certain chemotherapeutic agents. In vivo studies on RBBP6 in breast cancer mouse models are limited. Our study aims to investigate the molecular function and the prognostic potential of RBBP6 in breast cancer using MCF-7 cell line and the FVB/N-Tg(MMTV PyVT)634Mul/J breast cancer mouse model. The expressional effects of RBBP6 on cancer cell proliferation and metastatic tumour growth in cancer mice were examined in vitro and in vivo using RT-PCR, western blotting, and tumour size monitoring. The results demonstrate that RBBP6 mRNA expression was significantly higher in both local and distal tumours with a corresponding overexpression of angiogenic and metastatic key markers, VEGF164 and CD31, respectively. In addition, tumour growth reduced gradually over a period of two weeks in response to RBB6 gene knockdown. Our study indicated that RBBP6 may be a potential metastatic marker in breast tumours.

Citation Format: Pontsho Moela, Mayur Govender, Melvin Ambele. Investigating the effects of in vivo silencing of RBBP6 gene on tumor growth in the FVBN-Tg(MMTV PyVT)634MulJ breast cancer mouse 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 2469.
Abstract 2473: Tissue factor-induced fibrinogenesis and cancer cell clustering contribute to polyclonal peritoneal metastasis in diffuse-type gastric carcinoma
Authors: Miyazaki Makoto , Nagamura Yuko , Yamaguchi Hideki
Published: April 2023

Diffuse-type gastric carcinoma is characterized by highly invasive growth, massive fibrosis, and peritoneal metastasis. Peritoneal metastasis is the leading cause of death in diffuse-type gastric carcinoma; however, the underlying molecular mechanisms remain largely unknown. Recent studies have established a concept of polyclonal metastasis, in which metastases arise from circulating tumor cell clusters with high metastatic potential. Polyclonal metastasis is a cause of tumor heterogeneity that contributes to tumor progression and therapeutic resistance. However, the clonality of peritoneally metastasized tumors and the evidence for polyclonal metastasis in diffuse-type gastric carcinoma have been poorly elucidated. In this study, we utilized multicolor fluorescent lineage tracking to examine the clonality of peritoneal metastasis in mouse xenograft models. When peritoneal metastasis was induced by intraperitoneal or orthotopic injection of multicolored cancer cells, each peritoneally metastasized tumor exhibited multicolor fluorescence, showing that it contains multiclonal cancer cell populations. Peritoneal cytology and in situ fluorescent imaging of micrometastases revealed that multicolored cancer cell clusters formed within the peritoneal cavity and collectively attached to the peritoneum. In vitro, peritoneal lavage fluid or cleared ascitic fluid derived from cancer patients elicited cancer cell clustering, which was inhibited by anticoagulants. Immunofluorescence and immunoblotting revealed that cancer cell clusters formed in vitro and in vivo were associated with fibrin formation. Moreover, tissue factor knockout in cancer cells significantly reduced fibrin formation, cell clustering, peritoneal attachment, and peritoneal metastasis. In an in vitro adhesion assay, fibrin formation facilitated the cancer cell-mesothelial cell adhesion. Thus, these results demonstrate that cancer cells activate the coagulation cascade through tissue factor to form fibrin-mediated cell clusters and accelerate peritoneal attachment, leading to the development of polyclonal peritoneal metastasis. Our findings identify a novel mechanism of peritoneal metastasis that may be a potential therapeutic target.

Citation Format: Makoto Miyazaki, Yuko Nagamura, Hideki Yamaguchi. Tissue factor-induced fibrinogenesis and cancer cell clustering contribute to polyclonal peritoneal metastasis in diffuse-type gastric carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2473.
Abstract 2560: Post-translational modification of focal adhesion kinase (FAK) protein increases in breast cancer cells upon exposure to conditioned medium derived from in vitro and in vivo adipocytes
Authors: Mubtasim Noshin , Boren Caleb , Barr Benjamin , Gollahon Lauren
Published: April 2023

Under obese conditions, dysregulation of the normal secretion of adipokines from fat cells has been shown to drive cancer progression. However, the role adipokine dysregulation plays in driving metastasis is still unclear. Recent studies have reported the activation of metastasis-associated cell signaling pathways. One such pathway involves Focal Adhesion Kinase (FAK), a non-receptor cytoplasmic protein that acts as a signal transducer between receptor activation by adipokines and downstream effector proteins that control the biological phenomenon of metastasis. Nevertheless, the association between obesity-induced changes in adipocyte secretory factors and the subsequent increase in the transactivation of FAK through phosphorylation is unknown. To elucidate the potential relationship of adipocyte-secreted factors towards activating FAK, breast cancer cells with low (MCF7) and high (MDA-MB-231) metastatic potential were exposed to in vitro and in vivo, murine adipocyte-derived, conditioned medium. For the in vitro adipocyte model, 3T3-L1 MBX cells differentiated to obesity, with fatty acids supplementation, were used. For the in vivo model, adipocyte-secreted factors from organ-cultured visceral fat harvested from C3H/HeJ female mice fed an either high fat (46%) or control (11%) diets, were collected in the medium. Adipocyte secretions from the in vitro and in vivo adipocyte samples were determined using a mouse adipokine proteomic profiler array and ELISA kit. Under obese conditions, both the 3T3-L1 MBX cells and C3H/HeJ visceral fat were found to have significant changes in their adipokine secretions in comparison to controls. Results showed that the breast cancer cells got increased FAK autophosphorylation when exposed to either in vitro or in vivo-derived obese adipocyte-conditioned medium in comparison to the untreated breast cancer cells.

Citation Format: Noshin Mubtasim, Caleb Boren, Benjamin Barr, Lauren Gollahon. Post-translational modification of focal adhesion kinase (FAK) protein increases in breast cancer cells upon exposure to conditioned medium derived from in vitro and in vivo adipocytes [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 2560.
Abstract 2563: Identifying BCLAF1 as a downstream effector of p85β in ovarian cancer
Authors: Wang Panpan , Rao Ling , Mak Victor CY , Cheung Lydia WT
Published: April 2023

The phosphatidylinositol 3-kinase (PI3K)/AKT pathway is dysregulated in a broad spectrum of human cancers. Genomic alterations of the pathway members can cause cell transformation and tumorigenesis. The class IA PI3K is a dimeric enzyme composed of a catalytic subunit and a regulatory subunit to initiate AKT activation, with the two major regulatory subunits being p85α and p85β. Apart from sharing a canonical function in binding to and stabilizing p110α, we and the others have demonstrated that p85α and p85β carry distinct activities. We have recently reported an oncogenic role of p85β in ovarian cancer cells. In this study, we analyzed multiple omics data to further explore the oncogenic mechanism of p85β. BCL2 Associated Transcription Factor 1 (BCLAF1) was identified as a downstream effector of p85β. We found that the protein level of BCLAF1 was consistently decreased upon PIK3R2 silencing in ovarian cancer cell lines, and that the decrease could be reversed after PIK3R2 overexpression. Importantly, knockdown of BCLAF1 reduced the oncogenicity of p85β, whereas BCLAF1 overexpression promoted cell viability, cell invasion and migration. Accompanied with the increase in total protein level was enhanced phosphorylation of multiple serine residues of BCLAF1. Functional analysis of the serine residues by expressing the phosphorylation-defective mutants suggested that Ser161, Ser512 and Ser531 are critical for BCLAF1 oncogenic functions in ovarian cancer. Together, the findings reveal a new downstream effector by which p85β exerts its oncogenic effects in ovarian cancer.

Citation Format: Panpan Wang, Ling Rao, Victor CY Mak, Lydia WT Cheung. Identifying BCLAF1 as a downstream effector of p85β in ovarian 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 2563.
Abstract 2573: A novel regulation of the Wnt/β-catenin pathway involving NLRP12 during colorectal tumorigenesis
Authors: Khan Shahanshah , Peng Lan , Lewis Cheryl M. , Gao Yunpeng , Mani Ram S. , Zaki Hasan
Published: April 2023

NLRP12, a member of the NOD-like receptor family, has been characterized as a negative regulator of NF-κB and MAPK pathways in immune cells. We previously demonstrated that Nlrp12-deficient mice are highly susceptible to azoxymethane plus dextran sodium sulfate induced colorectal tumorigenesis in mice. However, the precise mechanism of NLRP12-mediated protection against colorectal cancer (CRC) was poorly understood. RNA-seq analysis revealed that tumors of Nlrp12-deficient mice express higher levels of protooncogenes (cMyc, Ccnd1), Wnt target genes (Ctnnb1, Axin2, Yap1, Tcf3, Lgr5), matrix metalloproteinases, and epithelial to mesenchymal transition markers (Vim, Fn1, Foxc2, Zeb2, Ezh2) compared to wild-type tumor. Interestingly, higher expression of these genes in Nlrp12-deficient tumors was not linked to any inflammatory signaling pathways as there was no difference in the activation of NF-kB, ERK, JNK, AKT, and STAT3 signaling pathways between Nlrp12-/- and wild-type tumors. However, we observed higher activation of β-catenin in Nlrp12-deficient tumors. Using intestinal epithelial cell specific Nlrp12-conditional knockout mice we showed that Nlrp12 deficiency in intestinal epithelial cells leads to increased tumorigenesis, metastasis, and β-catenin activation. To further understand the role of NLRP12 in the regulation of the Wnt/β-catenin, we overexpressed NLRP12 in HEK293T cells and HCT116 cells followed by stimulation with Wnt ligand Wnt3A. NLRP12 overexpressed cells showed significantly reduced activation of β-catenin and expression of Wnt target genes. Consistently, knocking down of NLRP12 in HEK293T and HCT116 cells caused higher activation of the Wnt/β-catenin pathway. Further biochemical analysis revealed that NLRP12-mediated suppression of β-catenin activation is associated with reduced phosphorylation of GSK3β. Interestingly, human and mouse colorectal tumors showed reduced expression of NLRP12 along with increased activation of β-catenin and GSK3β phosphorylation. Altogether, these data suggest that NLRP12 is a potent negative regulator of the Wnt/β-catenin pathway in colorectal tumor cells, and the NLRP12-GSK3β axis could be a novel target for CRC treatment.

Citation Format: Shahanshah Khan, Lan Peng, Cheryl M. Lewis, Yunpeng Gao, Ram S. Mani, Hasan Zaki. A novel regulation of the Wnt/β-catenin pathway involving NLRP12 during colorectal tumorigenesis [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 2573.
Abstract 2575: A multi-omic screening approach identifies the E3-ubiquitin ligase DCAF13 as a novel vulnerability in lung adenocarcinoma
Authors: Wagner Vinona T. , Chang Yun-Chien , Giansanti Piero , Öllinger Rupert , Engleitner Thomas , Kuster Bernhard , Bassermann Florian
Published: April 2023

Lung adenocarcinoma (LuAD) is the most prevalent subtype of lung cancer but despite advances in targeted therapy approaches, LuAD patients still have a 5-year survival rate of less than 15%. This is largely owed to a high variability in cancer driver mutations, with only around 25% of patients being eligible for targeted therapy, highlighting the need for therapy approaches beyond the level of genetic modifications. In order to determine novel vulnerabilities of LuAD, we performed a CRISPR/Cas9-based genomic screen with a pooled sgRNA library targeting genes of the ubiquitin system. The screen identified the poorly characterized E3-ubiquitin ligase DCAF13 as an essential gene for LuAD cell survival. Data mining of LuAD patient data revealed that DCAF13 is significantly overexpressed in tumor versus normal tissue and that high DCAF13 expression correlates with adverse overall survival and progression free survival. Mechanistically, DCAF13 depletion leads to a decrease in cell proliferation and subsequent cell death, which can be rescued by apoptosis inhibition. Immunofluorescence revealed that DCAF13 localizes to the centrosome, cell-cell contacts and the nucleus of LuAD cells, indicating a potential role in transcription control. Functionally, DCAF13 is a predicted substrate recruiting unit of a CUL4-family ubiquitin ligase. Indeed, we show that DCAF13 binds to CUL4B and DDB1 to form an active ubiquitin ligase complex. In an effort to identify lung cancer specific relevant ubiquitylation substrates of DCAF13, we performed affinity-based interactome screens and non-affinity-based functional proteomics in LuAD cells. Cross-validation of these approaches revealed distinct tumor-suppressive transcription factors, which are known to play a role in tumorigenesis, embryogenesis and stem cell differentiation. Ongoing biochemical and transcriptomic analyses further determine the involvement of these transcription factors and DCAF13 in the establishment and maintenance of LuAD. Taken together, we nominate DCAF13 as a novel vulnerability in LuAD with potential implication in future targeted therapy strategies.

Citation Format: Vinona T. Wagner, Yun-Chien Chang, Piero Giansanti, Rupert Öllinger, Thomas Engleitner, Bernhard Kuster, Florian Bassermann. A multi-omic screening approach identifies the E3-ubiquitin ligase DCAF13 as a novel vulnerability in lung adenocarcinoma [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 2575.
Abstract 258: Whole genome analysis of hereditary leiomyomatosis and renal cell cancer identifies role of localized hypermutational events
Authors: Wu Selina , Boutros Paul C.
Published: April 2023

Hereditary leiomyomatosis and renal cell cancer (HLRCC) is a rare hereditary condition affecting over 1 in 200,000 people worldwide. Individuals with HLRCC are also at risk of developing an aggressive form of renal cell carcinoma (RCC), as 15-20% of patients develop type 2 papillary RCC associated with poor prognosis. Although RCC risk and cancer penetrance of this condition has been studied, whole genome sequencing of associated tumors has yet to be examined, and the landscape of structural variants of HLRCC is largely unknown. We have therefore performed whole genome sequencing of tumor/normal pairs of HLRCC patients to study the tumor molecular architecture in this form of RCC. Through germline variant and somatic mutation calling, we have identified specific somatic mutational drivers, including those at the structural variant level for the first time in HLRCC. We have also quantified both chromothripsis and kataegis in these tumors, identifying the specific role of localized hypermutational events. Similarly, using tumour subclonal reconstruction techniques, we have inferred the clonality of HLRCC-associated RCC tumors and elucidate the relative timing of key oncogenic events and mutational processes. We also compare each of these mutational features to their equivalents in other RCC tumour subtypes. These studies shed key light on the specific mutational processes and drivers of this rare hereditary malignancy and on its lethality.

Citation Format: Selina Wu, Paul C. Boutros. Whole genome analysis of hereditary leiomyomatosis and renal cell cancer identifies role of localized hypermutational events [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 258.
Abstract 2871: Spatial landscape of the tumor microenvironment in pancreatic ductal adenocarcinoma using multiplexed imaging and AI based analysis
Authors: Smith Michael J. , Huang Chi-Chou , Phan Tuan H. , Sasaki Hideki , White Katie O. , Heil-Chapdelaine Richard A. , Angus-Hill Melinda
Published: April 2023

Pancreatic tumors are highly heterogenous, often with more aggressive regions that are responsible for invasion and metastasis. These aggressive pancreatic ductal adenocarcinoma (PDAC) cells undergo epithelial to mesenchymal transition and create subregions in the tumor that evade treatments and provide a critical support niche for continued tumor growth and metastasis. Conventional chemotherapy plus radiation, or in advanced disease chemotherapy plus targeted drug therapy can lengthen patient survival. However, even in patients with local disease, the 5-year survival rate remains at around 40%, demonstrating that there is need for improved conventional and immune therapies for all pancreatic cancer patients. In this study, dozens of biomarkers have been used to probe tumor heterogeneity in pancreatic ductal adenocarcinoma (PDAC) tissue. We rely on the published finding that hypoxia gene expression is consistent in PDAC, regardless of tumor location, whether primary tumor or metastasis. With the spatial biology solution, Cell DIVETM and proprietary Spatial AI analytics software, dozens of biomarkers can be used to computationally reduce tumor heterogeneity, and to spatially define cells in the microenvironment within hypoxic and normoxic regions of PDAC, and in normal pancreas. We found that these hypoxic PDAC regions are devoid of immune cells in general, with the appearance of spatially co-localized populations of cell types consistent with tumor desmoplasia and inflammation. Taken together, multiplexed whole slide imaging and analysis enables spatially resolved whole tissue analysis of the tumor microenvironment, including new insights into spatial biology in the hypoxic PDAC environment.

Citation Format: Michael J. Smith, Chi-Chou Huang, Tuan H. Phan, Hideki Sasaki, Katie O. White, Richard A. Heil-Chapdelaine, Melinda Angus-Hill. Spatial landscape of the tumor microenvironment in pancreatic ductal adenocarcinoma using multiplexed imaging and AI based analysis [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 2871.