bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2784: A robust copy number based clonality classification for multiple pulmonary tumors in routine pathology practice by shallow next generation sequencing of formalin fixed clinical specimens
Authors: Barbosa Bárbara Andrade , Radonic Teodora , Kim Yongsoo , Ylstra Bauke
Published: June 2022

Multiple synchronous or metachronous independent tumors in one patient, particularly those with thoracic involvement, can be as high as 20%. Defining whether a patient has multiple primary cancers (MPC) or metastatic disease is an urgent clinical need due to differences in treatment strategies, with curative intend for MPCs. Discerning the clonal and non-clonal relationship of two lesions for metastases and MPCs, respectively, has been done since the 1970s by analyzing histological characteristics, which nowadays are complemented with genomic features. However, one or few mutation features are insufficient to unequivocally diagnose tumor clonality, due to intratumoral heterogeneity and common mutations. We and others have demonstrated convincing clonality classification by genome-wide copy number aberrations (CNAs) analysis, which is the recommendation from the International Association for the Study of Lung Cancer (IASLC), independent of histological differences or similarities. Notwithstanding, a gold standard clonality assessment is currently lacking such that quality evaluation and calibration of the diagnostic tests are not possible. Gold-standard data: Genetic data from non-clonal cancer samples has become virtually endless by means of Whole Exome and Whole Genome sequencing (WES and WGS) since the mutational discrepancies by WES between two MPCs from a single lung cancer patient is the same as those of two primary cancers from different patients. Therefore, publicly available WES or WGS data from tumors of different lung cancer patients can serve as a gold standard reference for non-clonality. A gold standard reference for clonal samples can be derived from WES data from multiple biopsies of one patient, i.e. from the TRACERx initiative. These datasets now allow for the first time to calibrate CNA clonality testing against a gold standard and calculate the sensitivity and specificity of multiple lung tumors in one patient. Diagnostic test: Shallow Whole Genome Sequencing (sWGS) based genome-wide CNA analysis on Formalin-Fixed Paraffin-Embedded (FFPE) specimens has been used as the diagnostic routine at the Cancer Center Amsterdam (CCA), Amsterdam UMC over the past 5 years. The pipeline was calibrated against gold standard clonal and non-clonal WES data. We present sensitivity and specificity for an adequate, fast, and more economic clonality test based on genome-wide CNAs.

Citation Format: Bárbara Andrade Barbosa, Teodora Radonic, Yongsoo Kim, Bauke Ylstra. A robust copy number based clonality classification for multiple pulmonary tumors in routine pathology practice by shallow next generation sequencing of formalin fixed clinical specimens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2784.
Abstract 2803: A renewable IHC control tool CytoSections࣪ for defining MAGEA3, MAGEA4, and MAGEA9 antibody specificity
Authors: Zewdu Eden B. , Gonzalez Rachel M. , Ma Yan , Ling Derek , Hu Xiaomin , Ren Qi , Zhang Ranran , Lei Li-Hui , Allampallam Krishnan , Yin Dezhong , Liu Xuan , Fu Wei
Published: June 2022

Melanoma-associated antigen gene A (MAGEA) family proteins are expressed in a variety of tumors with each MAGEA protein having unique roles in cancer pathogenesis. The MAGEA members lack of expression in normal tissue and their role in cancer make them well suited for targeted cancer immunotherapy. Thus, knowing which MAGEA protein expression exists in a tumor is important. However, the MAGEA family shares sequence similarity that makes it difficult to find antibodies that are specific to just one family member. It is also challenging to source tissue generally with HIPAA regulation and even more so to get tissue representation of all 12 MAGEA family members. Here we present CytoSections as an alternative to patient control tissues. CytoSections are formalin fixed, paraffin embedded (FFPE) section of over-expression cell pellets. These cells were transfected with expression plasmids coding for genes of individual MAGES family. In this study, CytoSections are used to screen antibodies to all 12-member MAGEA family of proteins. The MAGEA family members were initially shown positive by targeted protein expression in western and immunohistochemistry using DDK tag antibody. Then multiple MAGEA3, MAGEA4, MAGEA9 antibodies were assessed by IHC to determine their specificity to their intended target. Using MAGEA family as a pilot project, we show that CytoSections is a verified, reproducible, and renewable alternative to human control tissues and serves as an ideal tool for antibody specificity assessment.

Citation Format: Eden B. Zewdu, Rachel M. Gonzalez, Yan Ma, Derek Ling, Xiaomin Hu, Qi Ren, Ranran Zhang, Li-Hui Lei, Krishnan Allampallam, Dezhong Yin, Xuan Liu, Wei Fu. A renewable IHC control tool CytoSections࣪ for defining MAGEA3, MAGEA4, and MAGEA9 antibody specificity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2803.
Abstract 2816: A novel in vivo model to simultaneously assess efficacy and toxicity of chimeric antigen receptor T-cell immunotherapy
Authors: Yang Jiwon , Lee Won , Jiao Jing , Cai Danying , Gustafson Heather , Gardner Rebecca , Cheng Mingshan , Keck James G.
Published: June 2022

Over the past few years, chimeric antigen receptor T (CAR T)-cell therapy has emerged as a novel treatment option for certain hematologic malignancies. Several CAR T-cell therapies produced highly efficacious responses from patients with hematologic malignancies. However, wider adoption of CAR T-cell therapy is challenged due to the potential development of life-threatening toxicities including cytokine release syndrome (CRS). CRS symptoms can range from mild fever to life-threatening events, including death. Preclinical animal models to assess the efficacy and toxicity of CAR T-cell therapies have been lacking. Here, we developed a novel mouse model to assess the efficacy and toxicity of CAR T-cell therapy simultaneously. Two different PBMC humanized NSG™ variants were used. NSG-MHC Class I/II double knock-out strain is known to have a delayed onset of GvHD, and NSG-SGM3xIL15 strain shows a higher engraftment level of human NK and myeloid cells, in addition to human T cells. Mice were humanized using PBMCs and treated with autologous or allogeneic CAR T cells, and efficacy and toxicity were assessed. Compared to the control treatment, CD19 CAR T-cells caused a decrease in the human CD19+ cell population in the blood and spleen, and induced cytokine release. Both T-cell and myeloid cytokine releases, including IFNgamma, IL-10, RANTES, and MIP-1alpha, were induced in our animal models. The in vivo platform can also be used to determine individual PBMC/CAR T donor differences and show the extent of efficacy and toxicity of each PBMC donor treated with autologous and allogeneic CAR T-cell therapy. We further confirmed anticancer efficacy using luciferase-tagged human B-cell lymphoma Raji tumor cells, which express a high level of CD19. In summary, we have developed a novel in vivo model to test the efficacy and toxicity of both autologous and allogeneic CAR T-cell therapy simultaneously.

Citation Format: Jiwon Yang, Won Lee, Jing Jiao, Danying Cai, Heather Gustafson, Rebecca Gardner, Mingshan Cheng, James G. Keck. A novel in vivo model to simultaneously assess efficacy and toxicity of chimeric antigen receptor T-cell immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2816.
Abstract 2988: Proteasomal deubiquitinases represent an attractive therapeutic target in head and neck squamous cell carcinomas (HNSCC)
Authors: Morgan Ethan L. , Toni Tiffany , Yang Xinping , Cheng Hui , Viswanathan Ramya , Chen Zhong , Van Waes Carter
Published: June 2022

Despite recent advances in treatment options for Human Papillomavirus (HPV)- head and neck squamous cell carcinoma (HNSCC), the overall survival (OS) rate for HNSCC is low, demonstrating the need for novel therapies for these cancers. Targeting the ubiquitin-proteasome system (UPS) has emerged as a potential target for the development of novel anti-cancer therapies. Bortezomib, a first generation proteasome inhibitor clinically approved for the treatment of Multiple Myeloma, has previously been demonstrated to induce tumor regression in a subset of HNSCC patients through the inhibition of canonical NFκB activity; however, the failure of Bortezomib to inhibit other pro-survival pathways such as MAPK and STAT3 signalling resulted in heterologous responses. A recently developed alternative to proteasome inhibitors is the small molecule b-AP15, which inhibits the proteasomal deubiquitinases USP14 and UCHL5, thus allowing more specificity and less toxicity than proteasome inhibitors.

Here, we show b-AP15 inhibits proliferation and colony formation in multiple HPV- and HPV+ HNSCC cell lines, with minimal effects in normal oral keratinocytes. Furthermore, b-AP15 induces significant caspase-dependent apoptosis. Using siRNA targetting USP14 and UCHL5, only USP14 depletion significantly reduced cell proliferation and colony formation. Mechanistically, b-AP15 treatment reduces TNFα-induced NFκB activity and the expression of pro-survival proteins such as cIAP2 and TRAF2, promoting the formation of the TNFR complex II and sensitizing cells to TNFα-induced cell death. Finally, TCGA data demonstrates that USP14 is highly expressed in HNSCC when compared with normal tissue and is significantly correlated with worse overall survival. Together, we have identified that inhibition of proteasomal deubiquitinases inhibits the proliferation and survival of HNSCC cells and enhanced TNFα-induced cell death via the inhibition of NFκB activity. Our data suggest that combination therapies with b-AP15 could potentially offer a clinical benefit in HNSCC patients by promoting TNFα-induced cytotoxicity. Further studies will focus on the mechanism by which USP14 regulates NFκB signalling in HNSCC cells and the effect of b-AP15 activity in combination with radiation treatment in in vivo mouse xenograft models.

Citation Format: Ethan L. Morgan, Tiffany Toni, Xinping Yang, Hui Cheng, Ramya Viswanathan, Zhong Chen, Carter Van Waes. Proteasomal deubiquitinases represent an attractive therapeutic target in head and neck squamous cell carcinomas (HNSCC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2988.
Abstract 3157: KLF5 defines the secretory lineage and governs sphingolipid metabolism in the skin
Authors: Ge Yejing
Published: June 2022

Adult stem cells contribute to tissue regeneration and wound repair through cellular programs that can be hijacked by cancer cells. We previously reported such a phenomenon in skin, where squamous cell carcinomas adopt a plastic phenotype so called “stem cell lineage infidelity”, entailing a wide-spread co-expression of otherwise lineage restricted genes, exemplified by hair follicle factor SOX9 and interfollicular epidermal factor KLF5. Occurring transiently in wounds and persists in cancer, stem cell lineage infidelity is essential to drive wound repair and tumorigenesis. However, it remains unclear what were the functional mediators of lineage infidelity factors. We thus set out to investigate one of its defining factors, KLF5. Employing genome-wide transcriptome and epigenome profiling, we found KLF5 specifies the epidermal lineage in the skin and is essential for skin maturation in vivo. Strikingly, KLF5 binds to the genomic vicinity of, and transcriptionally controls, multiple key enzymes catalyzing rate-limiting steps of epidermal sphingolipid secretion, whose disruption led to loss of lipid ultrastructural integrity, compromised secretory machinery and selective decline of high complexity sphingolipids in the skin. Remarkably, lipid-rich dietary intervention was able to reverse skin defects caused by KLF5 ablation and KLF5 is prevalently disrupted in skin barrier and inflammatory diseases. Our findings suggest KLF5 serves as a master secretory lineage factor and a transcriptional governor of sphingolipid metabolism in the developing and diseased skin, warranting future investigation of its functional underpinning in wound repair and cancer.

Citation Format: Yejing Ge. KLF5 defines the secretory lineage and governs sphingolipid metabolism in the skin [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3157.
Abstract 3208: Angiogenesis inhibitors strongly synergize with therapeutics targeting tumor metabolism
Authors: Khadka Sunada , Lin Yu-Hsi , Ackroyd Jeffery , Arthur Kenisha , Barekatain Yasaman , Poral Anton , Tran Theresa , Behr Eliot , Chen Yining , Georgiou Dimitra , Depinho Ronald , Muller Florian
Published: June 2022

Angiogenesis inhibition has become a mainstay of oncology despite having fallen short of its early promise. As originally envisioned, angiogenesis inhibition would cut off the blood supply, deprive tumor cells of key nutrients, leading to their demise. In practice, while there is evidence that tumors under angiogenesis treatment do in fact exhibit some degree of metabolic stress, this is stress is not sufficient to induce significant cancer cell death. We posit that the full potential of angiogenesis inhibition can be realized by the combination of angiogenesis inhibition with emerging tumor metabolism targeting therapies. Because tumors under angiogenesis inhibition are already in a state of nutrient stress, the effects of metabolically targeted therapies such as amino acid depletion (e.g. asparginase, methionine restriction), inhibitors of stress adaption (AMPK and GCN2 inhibitors) or energy metabolism (e.g. IACS-010759, Metformin, POMHEX) stand to dramatically increase in potency whilst remaining selective for (angiogenic) tumor versus (non-angiogenic) normal tissue. Here, we provide proof-of-principal for this thesis. First, we performed metabolomic profiling of angiogenesis-inhibited tumors, which corroborates a state of nutrient stress in angiogenesis-inhibited tumors. Second, we demonstrate dramatic anti-neoplastic synergy (effectively curing of xenografted tumor-bearing mice, irrespective of initial tumor size), without enhanced adverse toxicities, between the anti-VEGFA antibody Avastin and enolase inhibitor HEX. The same results were recapitulated with the VEGFR inhibitor Tivozanib and HEX and the enolase inhibitor could be substituted with the Oxphos inhibitor IACS-010759, with similar effects. The synergy was observed in a broad range of tumor types, even those without clear genetic susceptibilities. Together, these results suggest that angiogenesis inhibitors synergize broadly with cancer therapies targeting metabolism, allowing the realization of the full potential of these previously disappointing drugs. Our results warrant systematic combination clinical trials between angiogenesis inhibitors and established, as well as emerging anti-metabolic cancer therapies.

Citation Format: Sunada Khadka, Yu-Hsi Lin, Jeffery Ackroyd, Kenisha Arthur, Yasaman Barekatain, Anton Poral, Theresa Tran, Eliot Behr, Yining Chen, Dimitra Georgiou, Ronald Depinho, Florian Muller. Angiogenesis inhibitors strongly synergize with therapeutics targeting tumor metabolism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3208.
Abstract 3227: Prognostic role of systemic inflammation in colon and rectal cancer patients: Results from the ColoCare Study
Authors: Ose Jennifer , Lin Tengda , Himbert Caroline , Warby Christy A. , Hardikar Sheetal , Boehm Juergen , Gigic Biljana , Schrotz-King Petra , Schneider Martin , Ulrich Alexis B. , Shibata David , Figueiredo Jane C. , Siegel Erin M. , Li Christopher I. , Toriola Adetunji T. , Ulrich Cornelia M.
Published: June 2022

Purpose: Inflammation and angiogenesis are hallmarks of cancer development and progression. No studies have prospectively examined association of angiogenesis-related biomarkers with clinical outcomes in colon and rectal cancer.

Patients and Methods: In pre-surgery serum samples from n=426 non-metastatic colorectal cancer (CRC) patients (stage I-III), we investigated associations of inflammatory (e.g., CRP, SAA, sICAM-1, sVCAM-1) and angiogenesis biomarkers (VEGF-A, VEGF-D) with overall survival (OS), disease-free survival (DFS), and risk of recurrence. We computed hazard ratios (HR) and 95% confidence intervals (CI). Analyses were adjusted for age, sex, body mass index, stage, tumor site, and study site, and also exploratory stratified by tumor site (colon vs. rectum).

Results: N=65 patients (15%) were deceased and n=59 patients (15%) had a recurrence after a median follow-up of 31 months. Overall, doubling of CRP was associated with a ~24% increase in risk of death (OS: HRlog2: 1.24; 95% CI: 1.11-1.40), a 19% increase in risk of recurrence: HRlog2: 1.19; 95% CI: 1.08-1.32, and a non-statistically-significant 12% increase for DFS HRlog2: 1.12; 95% CI: 0.98-1.28. Similar associations were observed for SAA. Doubling of sICAM-1 was associated with a 4-fold increase in risk of death (OS: HRlog2: 4.05; 95% CI: 2.35-6.98). For the angiogenesis marker VEGF-D, significant heterogeneity was observed in analyses stratified by tumor site: doubling was associated with a 3-fold increase in risk of death for rectal cancer (OS: HR: 3.26; 95% CI: 1.58-6.70) and a 22% reduction in mortality for colon cancer (OS: HR: 0.78; 95% CI: 0.35-1.73; pheterogenity <0.001). Similar heterogeneity was observed in associations of VEGF-D with DFS and risk of recurrence, although not statistically significant (pheterogeneity<0.10).

Conclusion: Our data suggest that some biomarkers of inflammation and angiogenesis are prognostic markers for stage I-III CRC patients, with potential differences by tumor site for angiogenesis markers.

Citation Format: Jennifer Ose, Tengda Lin, Caroline Himbert, Christy A. Warby, Sheetal Hardikar, Juergen Boehm, Biljana Gigic, Petra Schrotz-King, Martin Schneider, Alexis B. Ulrich, David Shibata, Jane C. Figueiredo, Erin M. Siegel, Christopher I. Li, Adetunji T. Toriola, Cornelia M. Ulrich. Prognostic role of systemic inflammation in colon and rectal cancer patients: Results from the ColoCare Study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3227.
Abstract 3281: DNMT targeting enhances vulnerability of glioblastoma cells to MNK inhibition
Authors: Mazewski Candice , Eckerdt Frank , Baran Aneta , Fischietti Mariafausta , Vagadia Purav P. , Perez Ricardo E. , James Charles D. , Schiltz Gary E. , Platanias Leonidas C.
Published: June 2022

Many factors complicate therapeutic strategies for glioblastoma (GBM), including the existence of the blood brain barrier and a heterogenous population of difficult to treat glioma stem cells. Innovative strategies targeting novel pathways alone or in combination are needed for sustainable therapeutic improvements. The MAPK pathway has been implicated in many cancers. MAPK interacting kinases (MNK1 and MNK2) are downstream of MAPKs and phosphorylate the eukaryotic translation initiation factor 4E (eIF4E), a protein involved in translation of oncogenic mRNAs. We have previously established pharmacological MNK inhibition as a promising strategy for GBM. However, most currently available MNK inhibitors lack specificity and exhibit off-target effects. We developed novel selective MNK inhibitors that show MNK inhibition specificity in GBM established cell lines as well as patient-derived cell lines propagated under stem cell permissive conditions as 3-D neurospheres. MNK inhibitors reduced cell viability and neurosphere growth. Our previous work with MNK inhibitors showed involvement in negative feedback loops activated with treatment of other pharmacological agents, so we conducted a high-throughput screening to identify potential targets for combination treatment. One of the top hits was a DNA methyltransferase (DNMT) inhibitor that enhanced MNK inhibitor antineoplastic effects in GBM cells. Dual MNK and DNMT inhibition synergistically reduced neurosphere growth in 3-D glioma stem-like cells. The combination promoted apoptosis in the mesenchymal glioma stem-like cells as shown through flow cytometry and increased expression of cleaved PARP, cleaved caspase 3, and Bax. Also, DNMT targeting enhanced the viability reduction effects of siRNA mediated MNK1 knockdown in GBM cells. This combination of our novel MNK inhibitor with DNMT inhibition elicited antineoplastic benefits in both 2-D cultures and 3-D glioma stem cell-like populations, demonstrating a potential novel therapeutic strategy in GBM.

Citation Format: Candice Mazewski, Frank Eckerdt, Aneta Baran, Mariafausta Fischietti, Purav P. Vagadia, Ricardo E. Perez, Charles D. James, Gary E. Schiltz, Leonidas C. Platanias. DNMT targeting enhances vulnerability of glioblastoma cells to MNK inhibition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3281.
Abstract 3744: Differential Orthopedia Homeobox (OTP) expression in pulmonary carcinoids is associated with changes in DNA methylation
Authors: Moonen Laura , Mangiante Lise , Leunissen Daphne J. , Lap Lisa M. , Gabriel Aurelie , Hillen Lisa M. , Roemen Guido M. , Koch Alexander , van Engeland Manon , Dingemans Anne-Marie C. , Foll Matthieu , Alcala Nicolas , Fernandez-Cuesta Lynnette , Derks Jules L. , Speel Ernst-Jan M.
Published: June 2022

Introduction: Limited number of tumor types have been examined for Orthopedia Homeobox (OTP) expression. In pulmonary carcinoids, loss of expression is a strong indicator of poor prognosis. Here, we investigated OTP expression in 37 different tumor types, and the association between OTP expression and DNA methylation levels in lung neuroendocrine neoplasms.

Methods: We analyzed publicly available multi-omics data (whole-exome-, whole-genome-, RNA sequencing, Epic-850K-methylation array) of 58 typical-, 27 atypical carcinoids, 69 large cell neuroendocrine carcinoma and 51 small cell lung cancer patients and TCGA (The Cancer Genome Atlas) data of 33 tumor types. 850K-methylation analysis was cross validated using targeted pyrosequencing on 35 carcinoids.

Results: Results showed bimodality of OTP expression in carcinoids (OTPhigh versus OTPlow group, likelihood-ratio test p=1.5x10-2), with the OTPhigh group specific to pulmonary carcinoids while absent from all other cohorts analyzed. Significantly different DNA methylation levels were observed between OTPhigh and OTPlow carcinoids in 12/34 OTP infinium probes (fdr<0.05 & β-value effect size>0.2). Overall, OTPlow carcinoids harbor a high DNA methylation level as compared to OTPhigh carcinoids. OTPlow carcinoids showed a significantly worse overall survival (logrank test p=0.0052). Gene set enrichment analysis for somatically mutated genes associated with hallmarks of cancer showed robust enrichment of three hallmarks in the OTPlow group, i.e., sustaining proliferative signaling, evading growth suppressor, and genome instability and mutation.

Conclusion: High OTP expression is a unique feature of pulmonary carcinoids with a favorable prognosis. In poor prognostic patients, OTP expression is lost, most likely due to changes in DNA methylation levels.

Citation Format: Laura Moonen, Lise Mangiante, Daphne J. Leunissen, Lisa M. Lap, Aurelie Gabriel, Lisa M. Hillen, Guido M. Roemen, Alexander Koch, Manon van Engeland, Anne-Marie C. Dingemans, Matthieu Foll, Nicolas Alcala, Lynnette Fernandez-Cuesta, Jules L. Derks, Ernst-Jan M. Speel. Differential Orthopedia Homeobox (OTP) expression in pulmonary carcinoids is associated with changes in DNA methylation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3744.
Abstract 4027: Dual MEK and AKT inhibition suppresses pancreatic cancer growth and migration
Authors: Jack Jarrid , Pierce Alexandra , Bye Bailey , Walsh McKinnon , Chalise Prabhakar , VanSaun Michael N.
Published: June 2022

Pancreatic cancer is one of the deadliest cancers, due to late diagnosis and very few available therapeutic treatments. The Kras gene is frequently mutated in pancreatic cancer, but previous clinical trials using RAS inhibitors have proven ineffective. Additionally, many drug treatments targeting the MAPK pathway have shown little success due to the inherent drug resistance of pancreatic cancer cells and to acquired resistance through activation of alternative proliferative pathways such as JAK-STAT and PI3K-AKT. Our pathway analysis after MEK inhibition in pancreatic cancer cells displayed sustained and/or upregulation of AKT activity in multiple cell lines. Therefore, we hypothesize that a combination treatment of Trametinib and Omipalisib would block two prominent mitogenic pathways (MEK and AKT, respectively) to suppress cancer cell proliferation and migration. Multiple analyses including Western blot, clonogenic, 5-Ethynyl-2’-Deoxyuridine (EDU), and scratch migration assays were employed to determine the augmentation of cellular function in murine and anthropic pancreatic cancer cell lines after single or combination treatment with Trametinib and/or Omipalisib. Results demonstrated that application of Omipalisib alone was successful at blocking pAKT but failed to suppress pERK. Conversely, Trametinib alone selectively inhibited pERK but did not affect pAKT levels. The combination treatment successfully suppressed both pathways in relatively low quantities, indicating their efficacy as a dual therapeutic. Dual therapy was further effective at inhibiting cell growth as evidenced by clonogenic and EdU assays. Recovery and closure from an in vitro scratch wound was also significantly inhibited with combination of Omipalisib and Trametinib. In vivo studies demonstrated that both the pancreatic xenograft mean tumor growth and final tumor size were significantly reduced in response to the combination treatment compared to vehicle. In conclusion, dual therapy with Omipalisib and Trametinib showed a greater anti-tumor efficacy than treatment with either drug alone. Currently, we are determining the prolonged effectiveness of this combination treatment program in genetically-engineered mouse models. If successful, a pharmacological application of Omipalisib and Trametinib has the potential to provide a beneficial therapeutic option to pancreatic cancer patients.

Citation Format: Jarrid Jack, Alexandra Pierce, Bailey Bye, McKinnon Walsh, Prabhakar Chalise, Michael N. VanSaun. Dual MEK and AKT inhibition suppresses pancreatic cancer growth and migration [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4027.
Abstract 4129: Descriptors of primary cutaneous follicular helper T-cell lymphoma (PCFHTCL): Analysis of a pooled database of a rare entity
Authors: Haddad Philip A. , Hammoud Dalia , Gallagher Kevin
Published: June 2022

Introduction: Primary Cutaneous Follicular Helper T-cell Lymphoma (PCFHTCL) is a rare cutaneous T-cell lymphoma that was recently described. PCFHTCL is comprised of small to medium lymphocytes that express CD4, CD19, Bcl6, PD1, ICOS, CXCR5, and CXCL13. While it tends to have mostly an indolent course, occasionally PCFHTCL may display a more aggressive pattern. We conducted this analysis to explore the key clinicopathologic characteristics of this rare and little-known entity.

Methods: In order to study the demographic characteristics, molecular and immunohistochemical signatures, therapeutic interventions, survival, and prognostic factors, we compiled a pooled database of 57 cases. Kaplan-Meier survival curves were constructed. Cox proportional-hazards model and Log-rank tests were used to assess the influence of demographic and clinicopathologic factors on overall survival and disease-free survival.

Results: A total of 57 patients with confirmed PCFHTCL were identified. The median age was 57 years with a peak incidence between ages 53 and 64. There was a male predominance with an M:F ratio of 1.25. PCFHTCL commonly involved the head (54%), followed by the torso (24%) and limbs (12%). Sixteen percent of the lesions were solitary and mostly involved the head. The most common pattern was nodular/tumorous. Histologically, PCFHTCL involved mainly the dermis with occasional focal epidermotropism (27%), folliculotropism (43%), syringotropism (7%), and subcutaneous extension (14%). Extracutaneous involvement comprised lymph nodes (11%), bone marrow (4%), and the peripheral blood (2%). The median duration of symptoms prior to diagnosis was 5 months. The six-year disease-free survival is around 85%. Surgical resection alone resulted in durable remission in 40% of the cases. Recurrent disease was successfully salvaged with re-resection, radiation, and chemotherapy. While solitary lesions tended to have a better disease-free survival, it did not reach statistical significance.

Conclusions: This study represents the largest cohort of this rare cutaneous T-cell entity. It describes the demographic and clinical patterns of disease presentation and provides a detailed update on the clinicopathologic features PCFHTCL.

Citation Format: Philip A. Haddad, Dalia Hammoud, Kevin Gallagher. Descriptors of primary cutaneous follicular helper T-cell lymphoma (PCFHTCL): Analysis of a pooled database of a rare entity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4129.
Abstract 413: WT RAS signaling is an essential therapeutic target in RAS mutated cancers
Authors: Sealover Nancy E. , Kortum Robert L.
Published: June 2022

Single-agent targeting of RAS-mutated cancers using RAF/MEK/ERK or PI3K/AKT pathway inhibitors is ineffective; blocking one pathway relieves negative feedback control of RTK signaling and hyperactivates parallel effector pathways. While combined MEK and PI3K inhibition is effective in preclinical models, toxicity of this combination prevents its clinical use. Alternative strategies for treating RAS-mutated cancers are essential. Therapeutics directly targeting mutated RAS proteins have the potential to spare normal cellular function thereby lessening overall toxicity. Unfortunately, direct RAS inhibition as a monotherapy does not fully block RAS effector signaling. RAS proteins show differential activation of RAF and PI3K pathways: KRAS potently activates RAF but poorly activates PI3K, whereas HRAS potently activates PI3K but poorly activates RAF. Further, mutant RAS inhibition relieves negative feedback controls leading to rapid hyperactivation of RTK−WT RAS signaling. A more comprehensive understanding of the interplay between mutant RAS and RTK−WT RAS signaling is essential to developing rational therapeutic approaches to treat RAS-mutated cancers. We found that WT RAS enhanced mutant RAS-driven transformation by activating RAS effectors poorly engaged by mutated RAS. Further, inhibition of RAS effectors activated poorly by mutant RAS synergized with and limited resistance to mutant HRAS and KRAS inhibitors. The mutant HRAS inhibitor tipifarnib blocked PI3K signaling and synergized with MEK inhibitors in HRAS-mutated cancer cell lines; covalent KRASG12C inhibitors blocked MEK signaling and synergized with PI3K inhibitors in KRASG12C-mutated cell lines. Synergy between inhibitors of mutant RAS and RAS effectors was dependent on intact RTK/WT RAS signaling and was lost in both RASless or SOSless MEFs. Upstream of RAS, the RASGEFs SOS1 and SOS2 showed unique and overlapping functions to promote mutant RAS-driven transformation. SOS1 was critical for mutant RAS (G12V) activation and SOS1 inhibition augmented the efficacy of mutant RAS inhibitors. RTK−SOS2−PI3K signaling protected cells from anoikis and mediated mutant KRAS-driven transformation. These results should inform the design of clinical trials for patients with RAS-mutated cancers.

Citation Format: Nancy E. Sealover, Robert L. Kortum. WT RAS signaling is an essential therapeutic target in RAS mutated cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 413.
Abstract 4134: Stromal FAP expression is associated with MRI visibility and patient survival in prostate cancer
Authors: Pellinen Teijo , Smith Kevin , Blom Sami , Turkki Riku , Hemmes Annabrita , Välimäki Katja , Eineluoto Juho , Kenttämies Anu , Nordling Stig , Kallioniemi Olli , Rannikko Antti , Mirtti Tuomas
Published: June 2022

Some clinically significant prostate cancers (PCa) are missed by magnetic resonance imaging (MRI). We asked if the tumor stroma in surgically treated primary PCa lesions positive or negative with MRI are different in their cellular and molecular properties, and whether the differences are reflected to the clinical course of the disease.

We profiled the stromal and immune cell composition of MRI-classified tumor lesions by applying multiplexed fluorescence immunohistochemistry (mfIHC) and automated image analysis in a clinical cohort of 343 patients (Cohort I). We compared stromal variables between MRI-visible, invisible lesions, and benign tissue and assessed the predictive significance for biochemical recurrence (BCR) and disease-specific survival (DSS) using Cox regression and Log rank analysis. Subsequently, we carried out a prognostic validation of the identified biomarkers in a population-based cohort of 319 patients (Cohort II).

MRI true-positive lesions are different from benign tissue and MRI false-negative lesions in their stromal composition. CD163-positive cells and fibroblast activation protein (FAP)-positive cells were more abundant in MRI true-positive than in MRI false-negative lesions or benign areas. In MRI true-visible lesions, a high proportion of stromal FAP-positive cells was associated with PTEN status and increased immune infiltration (CD8+, CD163+), and predicted elevated risk for BCR. High FAP phenotype was confirmed to be a strong indicator of poor prognosis in two independent patient cohorts using also conventional IHC.

The molecular composition of the tumor stroma may determine whether early prostate lesions are detectable by MRI and associates with survival after surgical treatment.

Citation Format: Teijo Pellinen, Kevin Smith, Sami Blom, Riku Turkki, Annabrita Hemmes, Katja Välimäki, Juho Eineluoto, Anu Kenttämies, Stig Nordling, Olli Kallioniemi, Antti Rannikko, Tuomas Mirtti. Stromal FAP expression is associated with MRI visibility and patient survival in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4134.
Abstract 4156: BI-1808 - a first in class ligand-blocking αTNFR2 antibody for cancer immunotherapy
Authors: Mårtensson Linda , Cleary Kirsty , Holmkvist Petra , Kovacek Mathilda , Svensson Carolin , Semmrich Monika , Blidberg Therese , Demiri Mimoza , Dadas Osman , Borggren Marie , Pitic Vici , Lim Sean H. , Beers Stephen A. , Gertsson Susanne , Rohrberg Kristoffer S. , Karlsson Ingrid , McAllister Andres , Cragg Mark S. , Frendeus Björn , Teige Ingrid
Published: June 2022

The pleiotropic TNF-α:TNFR axis plays a central role in immune regulation. While the cellular expression of TNFR1 is broad, TNFR2 expression is mainly restricted to immune cells. The therapeutic potential of targeting TNFR2 for cancer treatment has been previously indicated and to gain further insight, we generated a wide panel of TNFR2-specific antibodies using the n-CoDeR F.I.R.S.T™ target and antibody discovery platforms. We identified antibodies against mouse or human TNFR2 that could block TNF-α binding and showed potent anti-tumor activity in several immune-competent models, both as single agent and in combination with anti-PD1. The mechanism-of-action was shown to be FcγR dependent and likely mediated through a combination of intra-tumor T reg depletion, CD8+ T cell expansion and modulation of innate immune cells e.g. neutrophils and myeloid cells. Toxicological studies of the human lead-candidate BI-1808 in non-human primates (NHP) demonstrated very good tolerability at doses up to 200 mg/kg, and in vitro studies with human cells showed no signs of harmful cytokine release. Furthermore, in vivo studies using immune competent mouse cancer experimental models showed a clear relationship between dose, receptor occupancy (RO) and efficacy. In both murine models and in the NHPs, soluble TNFR2 was clearly modulated by the treatment and closely correlated with RO. In addition, several changes in the immune cells (e.g. T cells and neutrophils) in the blood compartment were observed. Since January 2021, BI-1808 is evaluated in an ongoing Phase I/IIa trial. Similar to the preclinical studies, correlations between dose, RO and soluble TNFR2 has been clearly observed in the patients. In addition, and consistent with TNF-α:TNFR biology being conserved across the species, modulations of T cell and neutrophil numbers are also paralleled between the patients, NHPs and mice, increasing the likelihood of successful clinical translation of our findings.

Citation Format: Linda Mårtensson, Kirsty Cleary, Petra Holmkvist, Mathilda Kovacek, Carolin Svensson, Monika Semmrich, Therese Blidberg, Mimoza Demiri, Osman Dadas, Marie Borggren, Vici Pitic, Sean H. Lim, Stephen A. Beers, Susanne Gertsson, Kristoffer S. Rohrberg, Ingrid Karlsson, Andres McAllister, Mark S. Cragg, Björn Frendeus, Ingrid Teige. BI-1808 - a first in class ligand-blocking αTNFR2 antibody for cancer immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4156.
Abstract 5134: Biomarker concordance between matched primary and metastatic colorectal tumors
Authors: Yang Yun , Xin Ying , Song Jianning , Jing Chao , Zhao Baoyi , Huang Depei , Deng Wei
Published: June 2022

Background: Distant metastasis accounts for ~90% of deaths from colorectal cancer (CRC). The use of biomarkers for targeted therapy in metastatic CRC is well-established, requiring genetic testing of primary or metastatic biopsies. Here we aimed to assess the concordance of actionable alterations in matched primary and metastatic colorectal tumors.

Methods: Hybrid capture-based next-generation sequencing (NGS) were performed in a CAP/CLIA-approved laboratory (3DMed). NGS testing for SNV, InDel, CNV, MSI and TMB were performed on 19 primary tumors and 23 matched metastatic biopsies.

Results: 42 matched primary and metastatic tumor samples from 19 CRC patients were analyzed. 68.4% of the patients had only liver metastases, 5.3% ovarian metastases, 5.3% peritoneal metastases and 10.5% of the patients had more than one metastatic site. Mutational landscape showed that the primary samples and the metastases shared 80% of top 20 mutations. Biomarker concordance for KRAS was 89.5% (17/19), NRAS was 89.5% (17/19), BRAF was 94.7% (18/19), EGFR was 89.5% (17/19), BRCA2 was 94.7% (18/19) and MS status was 100% (19/19). Inconsistent RAS/BRAF status was found in two liver metastases and one ovarian metastasis. The frequencies of SRC amplification (P = 0.08) and ACVR2A mutation (P = 0.31) seem to be higher in the primary tumors, although these were not statistically significant. Conversely, mutations of NRAS, EGFR and MAP2K4 were detected only in the metastases. One patient harbored EGFR amplification in the ovarian metastasis, while this variant was not present in liver and primary sites. Copy number loss of MAP2K4 was found in the metastatic biopsies of two patients. Tumor mutation burden was comparable between the primary tumors and metastases (3.12 vs 2.17muts/Mb, P=0.29).

Conclusions: Our results demonstrates high concordance across multiple biomarkers between the primary and paired metastatic colorectal tumors. Molecular features of ovarian metastasis is more prone to inconsistent with primary sites than liver metastasis, which remains to be further verified in a large population.

Citation Format: Yun Yang, Ying Xin, Jianning Song, Chao Jing, Baoyi Zhao, Depei Huang, Wei Deng. Biomarker concordance between matched primary and metastatic colorectal tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5134.
Abstract 5177: Lymph node ratio is superior to AJCC N stage for predicting recurrence in papillary thyroid carcinoma
Authors: Parvathareddy Sandeep K. , Siraj Abdul K. , Qadri Zeeshan , Ahmed Saeeda O. , Siraj Nabil , DeVera Felisa , Annaiyappanaidu Padmanaban , Sabido Maria A. , Haqawi Wael , Aldossari Hassan , Al-Kuraya Khawla S.
Published: June 2022

Objective: Recently, lymph node ratio (LNR) has emerged as an alternative to American Joint Committee on Cancer (AJCC) N stage, with superior prognostic value. The utility of LNR in Middle Eastern Papillary thyroid carcinoma (PTC) remains unknown. Therefore, we retrospectively analyzed a large cohort of 1407 PTC patients for clinico-pathological associations of LNR.

Methods: Receiver operating characteristics (ROC) curve was used to determine the cut-off for LNR. We also performed multivariate logistic regression analysis to determine whether LNR or AJCC N stage was superior in predicting recurrence in PTC.

Results: Based on ROC curve analysis, a cut-off of 0.15 was chosen for LNR. High LNR was significantly associated with adverse clinico-pathological characteristics such as male sex, extrathyroidal extension, lymphovascular invasion, multifocality, bilateral tumors, T4 tumors, lateral lymph node (N1b) involvement, distant metastasis, advanced tumor stage, ATA high risk category and tumor recurrence. On multivariate analysis, we found that LNR was a better predictor of tumor recurrence than AJCC N stage (Odds ratio: 1.96 vs. 1.30; p value: 0.0184 vs. 0.3831). We also found that LNR combined with TNM stage and ATA risk category improved the prediction of recurrence-free survival, compared to TNM stage or ATA risk category alone.

Conclusions: The present study suggests LNR is an independent predictor of recurrence in Middle Eastern PTC. Integration of LNR with 8th edition AJCC TNM staging system and ATA risk stratification will improve the accuracy to predict recurrence in Middle Eastern PTC and help in tailoring treatment and surveillance strategies in these patients.

Citation Format: Sandeep K. Parvathareddy, Abdul K. Siraj, Zeeshan Qadri, Saeeda O. Ahmed, Nabil Siraj, Felisa DeVera, Padmanaban Annaiyappanaidu, Maria A. Sabido, Wael Haqawi, Hassan Aldossari, Khawla S. Al-Kuraya. Lymph node ratio is superior to AJCC N stage for predicting recurrence in papillary thyroid carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5177.
Abstract 5230: Neoadjuvant programmed cell death protein 1 inhibitors combined with chemotherapy in resectable esophageal squamous carcinoma: an open-label, single-arm study
Authors: Jiang Bo , Yang Xin , Zhang Junling , Huang Mengli
Published: June 2022

Objective: Although neoadjuvant chemotherapy has been recommended for resectable ESCC patients, the 5-year overall survival rate was less than 50%. Immune checkpoint inhibitors have been shown to be efficient for advanced ESCC, while few studies focus on the neoadjuvant immunotherapy combined with chemotherapy. Herein, we designed a trial to evaluate the safety and efficacy of neoadjuvant PD-1 inhibitors in combination with chemotherapy for resectable ESCC.

Methods: All patients had treatment-naïve resectable ESCC (stage III) that were confirmed by histopathology. All patients were scheduled to receive surgery within 4-6 weeks after neoadjuvant treatment (2 cycles) consisting of PD-1 inhibitors (Camrelizumab 200mg or Sintilimab 200 mg or Tislelizumab 200 mg) combined with two conventional chemotherapy regimens (Cisplatin + paclitaxel). The primary endpoint was the major pathologic response (MPR). Secondary endpoints were disease-free survival (DFS), R0 resection rate and safety profile. Immune signature of immune cells were evaluated in surgical specimens using immunofluorescence.

Results: Between Aug. 1, 2020, and Oct. 1, 2021, we assessed 10 patients for screening, of whom 10 patients were enrolled. Neoadjuvant therapy was not associated with delays in surgery or increased surgical complications/mortality. The median follow-up was 5.0 months (range 2.0∼31.0 months). Ten (100.0%) successfully underwent R0 resection. Of the 10 evaluable patients, 6 (60.0%) were MPR, 4 (40.0%) were pathologic complete response (pCR). Imaging evaluation was feasible in all patients. Partial response (PR) was achieved in 1 (10.0%) and stable disease (SD) was observed in 8 (80.0%). The most common treatment-related grade 1-2 adverse events were anemia (5, 50.0%), nausea (3, 30.0%), vomiting (3, 30.0%), constipation (2, 20.0%) and myelosuppression (2, 20.0%). There was no grade 3-4 events and treatment-related deaths. The presence of CD3+ T cells, B cell, and CD56 bright were statistically comparable patient groups of MPR and pCR, with an increased tendency in pCR patients. In addition, macrophages, CD8+ T cells, PD-1+CD8+ T cell, Treg cells, and PD-L1+CD68+ cells in pCR patients showed a downward trend.

Conclusions: Neoadjuvant PD-1 inhibitors plus chemotherapies had manageable treatment-related toxic effects. This regimen induced pCR or MPR in 100.0% of resected tumor, demonstrating its antitumor efficacy in resectable ESCC.

Citation Format: Bo Jiang, Xin Yang, Junling Zhang, Mengli Huang. Neoadjuvant programmed cell death protein 1 inhibitors combined with chemotherapy in resectable esophageal squamous carcinoma: an open-label, single-arm study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5230.