Development Of Targeted Therapies Research Articles

Transcript-Targeted Therapy Based on RNA Interference and Antisense Oligonucleotides: Current Applications and Novel Molecular Targets

The development of novel target therapies based on the use of RNA interference (RNAi) and antisense oligonucleotides (ASOs) is growing in an exponential way, challenging the chance for the treatment of the genetic diseases and cancer by hitting selectively targeted RNA in a sequence-dependent manner. Multiple opportunities are taking shape, able to remove defective protein by silencing RNA (e.g., Inclisiran targets mRNA of protein PCSK9, permitting a longer half-life of LDL receptors in heterozygous familial hypercholesteremia), by arresting mRNA translation (i.e., Fomivirsen that binds to UL123-RNA and blocks the translation into IE2 protein in CMV-retinitis), or by reactivating modified functional protein (e.g., Eteplirsen able to restore a functional shorter dystrophin by skipping the exon 51 in Duchenne muscular dystrophy) or a not very functional protein. In this last case, the use of ASOs permits modifying the expression of specific proteins by modulating splicing of specific pre-RNAs (e.g., Nusinersen acts on the splicing of exon 7 in SMN2 mRNA normally not expressed; it is used for spinal muscular atrophy) or by downregulation of transcript levels (e.g., Inotersen acts on the transthryretin mRNA to reduce its expression; it is prescribed for the treatment of hereditary transthyretin amyloidosis) in order to restore the biochemical/physiological condition and ameliorate quality of life. In the era of precision medicine, recently, an experimental splice-modulating antisense oligonucleotide, Milasen, was designed and used to treat an 8-year-old girl affected by a rare, fatal, progressive form of neurodegenerative disease leading to death during adolescence. In this review, we summarize the main transcriptional therapeutic drugs approved to date for the treatment of genetic diseases by principal regulatory government agencies and recent clinical trials aimed at the treatment of cancer. Their mechanism of action, chemical structure, administration, and biomedical performance are predominantly discussed.

Metastatic Adenoid Cystic Carcinoma: Genomic Landscape and Emerging Treatments.

Adenoid cystic carcinoma (ACC) is a heterogeneous cancer that commonly develops in the salivary glands. Approximately 40 to 50% of patients with ACC develop recurrence and/or metastasis. Although most patients with ACC have slow-growing disease, a subset experiences aggressive disease with early visceral and/or bone metastasis. Thus far, there is no consensus on the best time to start palliative treatment in patients with indolent disease. The only systemic therapies available for recurrent or metastatic ACC are cytotoxic agents and multikinase inhibitors targeting vascular endothelial growth factor receptor, and both types of therapy have modest activity. Studies integrating proteomics, genomics, and clinical data have revealed distinct molecular ACC subtypes, ACC-I and ACC-II, with ACC-I generally associated with more aggressive disease biology. ACC-I tumors were enriched for NOTCH1-activating mutation and upregulation of MYC and MYC targets, while ACC-II tumors exhibited upregulation of TP63 and receptor tyrosine kinases. These findings highlight the importance of patient selection for surveillance and targeted therapy development in ACC. In recent clinical trials of targeted therapy in ACC, patients are being selected according to tumor molecular profile (e.g., presence of NOTCH-activating mutations), which represents a major advance in the field. Ongoing collaborative research focusing on the development of novel therapeutic strategies for ACC patients based on disease biology will increase the drug armamentarium and improve survival outcomes for these patients in dire need.

Somatic mutations in VEXAS Syndrome and Erdheim-Chester disease: Inflammatory myeloid diseases

Somatic mutations have been increasingly identified as etiologic for many hematologic and autoinflammatory disorders. VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic) syndrome and Erdheim-Chester disease (ECD), a type of histiocytosis, can be classified as inflammatory myeloid diseases, characterized by systemic inflammation and multi-organ disease with predisposition to myeloid malignancies. VEXAS is a novel disease caused by UBA1 mutations that was first discovered using a genotype-driven approach (genotype was used to identify patients with undiagnosed inflammatory diseases). Since the initial description, many VEXAS cases have been reported and disease phenotype is expanding rapidly. In contrast, ECD was first characterized in the 1930s based on patients’ phenotype, and only recently found to be caused by recurrent somatic mutations in the MAPK pathway (traditional phenotype-driven approach). The discovery of these mutations and development of target therapies have revolutionized the treatment of patients with histiocytosis, particularly ECD. Here we discuss the impact of causal and associated somatic mutations in VEXAS and ECD at both clinical and molecular levels.

Value of a Signature of Immune-Related Genes in Predicting the Prognosis of Melanoma and Its Responses to Immune Checkpoint Blocker Therapies.

Melanoma is becoming increasingly common worldwide, with high rates of transformation into malignancy compared to other skin lesions. The prognosis of patients with melanoma at an advanced stage is highly unsatisfying despite the development of immunotherapy, target therapy, or combinative therapy. The major barrier to exploiting immune checkpoint therapies and achieving the best benefits clinically is resistance that can easily develop if regimens are not selected appropriately. In this study, we investigated the possibility of using immune-related genes to predict patient survival and their responses to immune checkpoint blocker therapies with the expression profiles available at The Cancer Genome Atlas (TCGA) Program plus expression data from the Gene Expression Omnibus (GEO) for validation. A five gene signature that is highly correlated with the local infiltration of cytotoxic lymphocytes in the tumor microenvironment was identified, and a scoring model was developed with stepwise regression after multivariate Cox analyses. The score calculated strongly correlates with Breslow depth, and this model effectively predicts the prognosis of patients with melanoma, whether primary or metastasized. It also depicts the heterogenous immune-related nature of melanoma by revealing different predicted responses to immune checkpoint blocker therapies through its correlation to tumor immune dysfunction and exclusion (TIDE) score.

Frequent productive cough: Symptom burden and future exacerbation risk among patients with asthma and/or COPD in the NOVELTY study

IntroductionPersistent cough with sputum production is an important clinical trait in chronic obstructive pulmonary disease (COPD). We defined “frequent productive cough” based on 2 questions from the St George's Respiratory Questionnaire (SGRQ) and sought to determine its occurrence and associated outcomes in patients with physician-assigned asthma and/or COPD from the NOVELTY study. MethodsFrequent productive cough was defined as cough and sputum production most or several days/week for the past 3 months (scoring ≥3 for both SGRQ questions). Relationships with baseline disease characteristics and exacerbations over 12 months’ follow-up were examined using logistic regression. ResultsBaseline SGRQ data were available for 7125 patients, of whom 31.3% had frequent productive cough. It was more common in asthma+COPD (38.8%) and COPD (38.1%) than asthma (25.0%), increasing with physician-assessed severity, and in current versus former and never smokers. Patient-reported symptomatic worsening was more common in patients with versus without frequent productive cough. Reduced post-bronchodilator FEV1 (odds ratio [OR] per 10% decrement 1.14 [95% confidence interval 1.11–1.16]) and history of pollutant exposure at home/work (OR 1.50 [1.33–1.69]) were associated with frequent productive cough in all diagnoses. Patients with baseline frequent productive cough were more likely to have ≥1 exacerbation over the subsequent 12 months (OR 1.71 [1.52–1.93]), including exacerbations requiring hospital admission and those treated with oral corticosteroids. ConclusionsFrequent productive cough represents an important indicator of adverse clinical outcomes across asthma and/or COPD. Research into the underlying pathologic mechanisms is required to support targeted therapy development. Clinicaltrials.govNCT02760329.

Abstract 6021: Establishment and characterization of cisplatin-resistant nasopharyngeal carcinoma (NPC) patient-derived xenograft

Abstract Background: NPC is particularly common in Southern China and Southeast Asia. The standard treatment for NPC is radiotherapy with or without adjuvant chemotherapy. Despite the high 5-year overall survival rate (85.6%) in early-stage patients, 15% to 20% of the patients still experience recurrence or metastasis. The development of new targeted therapy on NPC is still lagging behind. Considered that phase I clinical trials often recruit late-stage patients who previously received multiple treatments, lack of drug-resistant models may be one of the possible hindrances in targeted therapy development. In-vivo preclinical NPC models are very limited for research. Given the close association with Epstein-Barr virus (EBV) infection in NPC, only four EBV positive (EBV +ve) PDXs and one EBV +ve cell line are available for research. Our lab has been engaging in establishing new NPC models. Previously, we had successfully established four new EBV +ve PDXs: Xeno23, Xeno32, Xeno47 and Xeno76. To establish a more clinically relevant models for preclinical studies and drug resistance research, we continuously treat the PDX transplanted mice with cisplatin. And we successfully established the first cisplatin-resistant Xeno76 (Xeno76-CR) NPC PDX. Method: A 1 mm3 PDX tissues were implanted subcutaneously in Nude mice. Once the tissue started growing into ~100 mm3 in size, 3-5 mg/kg cisplatin was injected intraperitoneally weekly. Once the end point has met, the tumor tissue was repeatedly implanted into another mice, while the rest of the tissues were used for further evaluation. Result: The growth rate of Xeno76-CR has no significant difference with Xeno76 in Nude mice. We confirmed that Xeno76-CR is resistant to cisplatin in-vivo. When treating both Xeno76 and Xeno76-CR with 3 mg/kg cisplatin weekly, there is statistically significant 85.4% tumor growth inhibition in Xeno76, but no significant inhibition effect in cisplatin-treated Xeno76-CR. EBV infection of Xeno76-CR was confirmed with EBER staining. We have also studied whether Xeno76-CR is drug- or specifically cisplatin-resistant. We tested the Xeno76-CR with ixazomib, an oral proteasome inhibitor that can inhibit nuclear factor-kB (NF-kB) signalling pathway, and found significant 57% tumor growth inhibition in-vivo. Conclusion: We have established the first and only EBV +ve cisplatin-resistant NPC PDX, Xeno76-CR, that is a valuable and clinically relevant model for targeted therapies preclinical studies and resistant research. Single-cell RNA sequencing will be performed to examine if stem cell plays any role in cisplatin-resistance. Further genome and transcriptome expression profile and characterization studies are still ongoing. Citation Format: Ka Yee Li, Katie Sze Wai Fung, Chanping You, Yim Ling Yip, Victor Ho Fun Lee, George Sai Wah Tsao. Establishment and characterization of cisplatin-resistant nasopharyngeal carcinoma (NPC) patient-derived xenograft [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 6021.

Identifying the potential role of curcumin analogues as anti-breast cancer agents; an in silico approach

BackgroundBreast cancer ranks top among newly reported cancer cases and most of the women suffers from breast cancer. Development of target therapy using phytochemicals with minimal side effects is trending in health care research. Phytochemicals targets complex multiple signalling events in cancer and are pleiotropic in nature. Thus, the present study was conducted to check the effectivity of curcumin analogues (Capsaicin, Chlorogenic acid, Ferulic acid, Zingerone, Gingerol) against the receptors that are expressed in breast cancer cells and prove its ethno-medicinal value by using bioinformatic tools and softwares like PDB, Patch Dock, PubChem, Chimera and My Presto.ResultOut of the various curcumin analogues studied, Ferulic acid showed best binding affinity with all the breast cancer cell specific receptors (FGF, MMP9, RNRM1, TGF-beta, DHFR, VEGF and aromatase) which was confirmed through the docking studies.ConclusionThe current work was a preliminary step towards screening suitable drug candidate against breast cancer using in silico methods. This information can be used further to carry out in vivo studies using selected natural analogues of curcumin as a suitable drug candidate against breast cancer saving time and cost.

Comprehensive analysis of FGFR mutations in Chinese cancer patients.

e15117 Background: Fibroblast growth factor receptors (FGFRs) are highly conserved transmembrane tyrosine kinase receptors (RTKS) that regulating several cellular processes such as proliferation, differentiation, migration and survival. In recent years, some studies have documented that signaling through FGFRs impact progression of cancer and the survival of patients with cancer. As a potential therapeutic target, exploring the mutation frequency of FGFRs in more different cancers are necessary. Here we performed a comprehensive analysis of the frequency and types of FGFRs mutations in Chinese cancer patients and provided reference for targeted therapy development. Methods: From January 2020 to December 2021, a total of 4,102 patients with nine common cancers were included in the study. The next-generation sequencing was utilized for detecting genetic mutations. Results: A total 328 patients (7.9%) with FGFR mutation were identified. The FGFR 3 was the most frequently gene in our cohort. Among all the cancer patients, the highest frequency of FGFR mutations was observed in bladder cancer (32.5%) and the percentages in FGFR1, FGFR2 and, FGFR3 were 2.7%, 1.35%, and 6.76% respectively. Furthermore, the proportion of FGFR mutation in lung carcinoma, cholangiocarcinoma, colorectal carcinoma, melanoma, breast carcinoma and ovarian carcinoma was 8.64%, 11.58%, 13%, 16.7%, 9.62% and 10.81% respectively. Additionally, FGFR mutation have been detected in other tumor types, including renal cell carcinoma (5.63%), prostate cancer (5.45%), and gastric carcinoma (13.72%). Conclusions: The present study provided the comprehensive FGFR mutation data of Chinese cancer patients. A high frequency mutation in FGFR have been found in Chinese patients with different cancers, which mean that patients with multiple cancer types might benefit from development of efficient EGFR inhibitors.

4-Aryl-N-phenylpyrimidin-2-amines targeting EGFR-tyrosine kinase attenuated EGFR-expressing cell lines

Target therapies have been widely developed to combat various diseases and cancer. Epidermal Growth Factor Receptor is still a currently therapeutic target for solid tumor. Aberration of EGFR activity or expression reflect disease progression and poor prognosis. Recently, several newly synthesized 4-aryl-N-phenylpyrimidin-2-amines with some modifications at R2 selectively elicited cytotoxicity against A549. Therefore, harboring EGFR expression would be reasonable for the assessment. Herein, the 4-aryl-N-phenylpyrimidin-2-amines derivatives; N-(3-{[4-(4-methoxyphenyl) pyrimidin-2-yl]amino}phenyl), 3-{[4-(3-methoxyphenyl) pyrimidin-2-yl] amino}benzene-1-sulfonamide.(13g), methanesulfonamide (13c), 3-[(4-phenylpyrimidin-2-yl)amino] benzene-1-sulfonamide (13f) and 3-(benzene-1-sulfonamide) (5) were selected as promising derivatives for targeted-EGFR analysis. Kinase enzymatic-based assay exhibited IC50 values of 5.61, 31.92, 73.80 and 0.79 ​nM of each derivative, respectively whilst 41.50 ​nM of Gefitinib. Molecular docking deciphered the mode of binding of each derivative in ATP-binding site greater than gefitinib. Although, (13g) demonstrated the highest binding free energy among the other but not in ATP-binding site as the others does that related to its IC50 values. (13c), (13f), and (5) therefore were subjected for cell-based analysis. A549 and A431 were used as wtEGFR-expressing cells for cell-based assay. (13c), (13f), and (5) had high toxicity towards in both cells while (5) had much more toxicity on A431. (13c), (13f) and (5) not only induced apoptosis in a dose-dependent manner but reduced clonogenic formation greater than gefitinib. Migration of EGF-stimulated A431 was significantly delayed by the compounds over time but they could not delay EGF-stimulated A549 migration. Taken together, (13c), (13f) and (5) would be a newly synthesized derivative by targeting wtEGFR-expressing cells that attenuated EGFR-driven cancer hallmark leading to targeted therapy development.

Th1 and Th17 cells are resistant towards T cell activation-induced downregulation of CD6

BackgroundThe cell surface molecule CD6 is a modulator of T cell receptor (TCR) signaling. Recently, it has been reported that CD6 is downregulated on CD4+ T cells following T cell activation. This mechanism could limit the efficacy of anti-CD6 therapeutical antibodies. MethodsWe analyzed CD6 expression on activated and non-activated Th1 cells and Th17 cells by flow cytometry. ResultsOur experiments confirmed a significant downregulation of CD6 on IFNγ- and IL17-negative CD4+ T cells from healthy individuals and from patients with rheumatoid arthritis following T cell activation with anti-CD3 and anti-CD28 antibodies. In contrast, CD6 expression remained stable on activated Th17 cells and Th1 cells. ConclusionsTh1 and Th17 cells are resistant towards T cell activation-induced downregulation of CD6. These findings are relevant for the future development of CD6 targeting therapies and show that CD6 expression is differentially regulated in CD4+ T cell subsets.

Genetic evidence supports the development of SLC26A9 targeting therapies for the treatment of lung disease

Over 400 variants in the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) are CF-causing. CFTR modulators target variants to improve lung function, but marked variability in response exists and current therapies do not address all CF-causing variants highlighting unmet needs. Alternative epithelial ion channel/transporters such as SLC26A9 could compensate for CFTR dysfunction, providing therapeutic targets that may benefit all individuals with CF. We investigate the relationship between rs7512462, a marker of SLC26A9 activity, and lung function pre- and post-treatment with CFTR modulators in Canadian and US CF cohorts, in the general population, and in those with chronic obstructive pulmonary disease (COPD). Rs7512462 CC genotype is associated with greater lung function in CF individuals with minimal function variants (for which there are currently no approved therapies; p = 0.008); and for gating (p = 0.033) and p.Phe508del/ p.Phe508del (p = 0.006) genotypes upon treatment with CFTR modulators. In parallel, human nasal epithelia with CC and p.Phe508del/p.Phe508del after Ussing chamber analysis of a combination of approved and experimental modulator treatments show greater CFTR function (p = 0.0022). Beyond CF, rs7512462 is associated with peak expiratory flow in a meta-analysis of the UK Biobank and Spirometa Consortium (p = 2.74 × 10−44) and provides p = 0.0891 in an analysis of COPD case-control status in the UK Biobank defined by spirometry. These findings support SLC26A9 as a therapeutic target to improve lung function for all people with CF and in individuals with other obstructive lung diseases.

Endothelial Dysfunction and Cardiovascular Risk in Obstructive Sleep Apnea: A Review Article.

Obstructive sleep apnea (OSA) is a respiratory condition during sleep caused by repeated pauses in breathing due to upper airway obstruction. It is estimated that OSA affects 30% of the population, but only 10% are well diagnosed due to the absence of a well-defined symptomatology and poor screening tools for early diagnosis. OSA is associated to an endothelial dysfunction inducing several biological responses such as hypoxia, hypercapnia and oxidative stress, among others. OSA also triggers respiratory, nervous, metabolic, humoral and immunity system activations that increase the possibility of suffering a cardiovascular (CV) disease. In this review, we expose different studies that show the relationship between OSA and endothelial dysfunction and its association with CV pathologies like hypertension, and we define the most well-known treatments and their limitations. Additionally, we describe the potential future directions in OSA research, and we report clinical features such as endothelial progenitor cell alterations that could act as biomarkers for the development of new diagnostic tools and target therapies.

LncRNA HOXC-AS3 promotes non-small-cell lung cancer growth and metastasis through upregulation of YBX1

NSCLC is common and is the primary cause of cancer-related deaths due to a lack of early diagnosis and its propensity for metastasis. The pathogenesis of NSCLC is still unclear. Here, we explored the molecular mechanisms underlying NSCLC development, focusing on the HOXC-AS3/YBX1/HOXC8 axis. Human NSCLC specimens and cell lines were used. qRT-PCR and western blotting were utilised to examine the levels of HOXC-AS3/YBX1/HOXC8. CCK-8, colony formation, scratch wound healing and Transwell assays were performed to evaluate cancer cell proliferation, migration and invasion. A nude mouse xenograft model was used to examine tumour growth and metastasis in vivo. RNA pull-down, chromatin immunoprecipitation, coimmunoprecipitation and dual-luciferase assays were applied to validate the interactions of HOXC-AS3/YBX1, MDM2/YBX1 and the YBX1/HOXC8 promoter. The levels of HOXC-AS3 and HOXC8 were increased in human NSCLC specimens and cells. Knockdown of HOXC-AS3 suppressed NSCLC cell proliferation, migration and invasion, as well as tumour growth and metastasis in vivo. HOXC-AS3 directly bound to YBX1 to suppress its ubiquitination mediated by MDM2. YBX1 bound to the HOXC8 promoter and enhanced its transcription. Knockdown of HOXC8 inhibited the effects of HOXC-AS3 overexpression on NSCLC. HOXC-AS3 promotes NSCLC growth and metastasis by stabilising YBX1 and thus increasing HOXC8 transcription. Our study indicates that the HOXC-AS3/YBX1/HOXC8 axis could serve as a biomarker for NSCLC diagnosis or as a target for therapy development.

Long Non-Coding RNA Cancer Susceptibility Candidate 9 Regulates the Malignant Biological Behavior of Nasopharyngeal Carcinoma Cells by Targeting miR-497-5p/Wnt3a/β-catenin Signaling Pathway.

Nasopharyngeal carcinoma (NPC) is a major kind of head and neck epithelial carcinoma. Increasing evidences reveal that long noncoding RNAs are considered as vital regulators in tumorigenesis and progression. Although previous studies have found that cancer susceptibility candidate 9 (CASC9) highly expresses in NPC, the underlying mechanisms need to be further studied. In this study, we found that CASC9 was overexpressed and associated with worse prognosis in NPC. CASC9 knockdown significantly inhibited the cell proliferation, migration and invasion in vitro and enhanced the sensitivity of tumor cells to cisplatin and paclitaxel. Mechanism research confirmed CASC9 regulated the malignant biological behavior of nasopharyngeal carcinoma cells by targeting miR-497-5p/Wnt3a/β-catenin signaling pathway. The present study might provide a novel mechanism for tumorigenesis and progression of NPC and contribute to the development of an effective molecular target therapy.

Artificial Intelligence-Assisted Serial Analysis of Clinical Cancer Genomics Data Identifies Changing Treatment Recommendations and Therapeutic Targets.

Given the pace of predictive biomarker and targeted therapy development, it is unknown whether repeat annotation of the same next-generation sequencing data can identify additional clinically actionable targets that could be therapeutically leveraged. In this study, we sought to determine the predictive yield of serial reanalysis of clinical tumor sequencing data. Using artificial intelligence (AI)-assisted variant annotation, we retrospectively reanalyzed sequencing data from 2,219 patients with cancer from a single academic medical center at 3-month intervals totaling 9 months in 2020. The yield of serial reanalysis was assessed by the proportion of patients with improved strength of therapeutic recommendations. A total of 1,775 patients (80%) had ≥1 potentially clinically actionable mutation at baseline, including 243 (11%) patients who had an alteration targeted by an FDA-approved drug for their cancer type. By month 9, the latter increased to 458 (21%) patients mainly due to a single pan-cancer agent directed against tumors with high tumor mutation burden. Within this timeframe, 67 new therapies became available and 45 were no longer available. Variant pathogenicity classifications also changed leading to changes in treatment recommendations for 124 patients (6%). Serial reannotation of tumor sequencing data improved the strength of treatment recommendations (based on level of evidence) in a mixed cancer cohort and showed substantial changes in available therapies and variant classifications. These results suggest a role for repeat analysis of tumor sequencing data in clinical practice, which can be streamlined with AI support.

Trends of Molecular Testing for Lung Cancer at the King Faisal Hospital, Kigali: Therapeutic and Survival Implications

IntroductionLung cancer is the leading cause of cancer mortality worldwide, both in high and low resource settings. Knowledge has been generated elsewhere regarding molecular subtyping and subsequent targeted therapy development, contributing substantially to patient survival. Little is known on the data around lung cancer and its treatment outcomes in Sub-Saharan Africa. This study describes the experience in lung cancer diagnosis, molecular and biomarker testing, and treatment for advanced cases in a single institution in East Africa, between the years 2019 and 2021. MethodsThis was a retrospective observational study evaluating patients with metastatic (stage IV) lung cancer. Data on patient demographics, histologic diagnosis, molecular and biomarker testing, and treatment details and outcomes were collected. Molecular test results were reported as positive if there were biomarkers identified (e.g., EGFR, ALK, programmed death-ligand 1), and patients who had negative test results were reported as negative for biomarkers. ResultsA total of 14 patients were diagnosed with having stage IV disease, and all were proposed to undergo molecular testing. For 12 (86%) patients who were able to have molecular testing done, EGFR and programmed death-ligand 1 were the most common with 66.7% (N = 8) of tissues with either finding. For all 14 patients, treatment changes were made for eight patients (57.1%) after being primarily placed on a combination of paclitaxel and carboplatin for an average of six cycles. Changing treatment significantly improved the 2-year overall survival (85% versus 25%, p = 0.0006). ConclusionsDespite being the number one cause of mortality, gains are being made in poor-resource settings to improve the survival of patients with advanced lung cancers. Limitations to this quest remain misdiagnosis and delayed diagnosis and resource constraints for both molecular testing and subsequent treatments.

Specific (sialyl-)Lewis core 2 O-glycans differentiate colorectal cancer from healthy colon epithelium.

Cells are covered with a dense layer of carbohydrates, some of which are solely present on neoplastic cells. The so-called tumor-associated carbohydrate antigens (TACAs) are increasingly recognized as promising targets for immunotherapy. These carbohydrates differ from those of the surrounding non-cancerous tissues and contribute to the malignant phenotype of the cancer cells by promoting proliferation, metastasis, and immunosuppression. However, due to tumor tissue heterogeneity and technological limitations, TACAs are insufficiently explored.Methods: A workflow was established to decode the colorectal cancer (CRC)-associated O-linked glycans from approximately 20,000 cell extracts. Extracts were obtained through laser capture microdissection of formalin fixed paraffin embedded tissues of both primary tumors and metastatic sites, and compared to healthy colon mucosa from the same patients. The released O-glycans were analyzed by porous graphitized carbon liquid chromatography-tandem mass spectrometry in negative ion mode.Results: Distinctive O-glycosylation features were found in cancerous, stromal and normal colon mucosal regions. Over 100 O-linked glycans were detected in cancerous regions with absence in normal mucosa. From those, six core 2 O-glycans were exclusively found in more than 33% of the cancers, carrying the terminal (sialyl-)LewisX/A antigen. Moreover, two O-glycans were present in 72% of the analyzed cancers and 94% of the investigated cancers expressed at least one of these two O-glycans. In contrast, normal colon mucosa predominantly expressed core 3 O-glycans, carrying α2-6-linked sialylation, (sulfo-)LewisX/A and Sda antigens.Conclusion: In this study, we present a novel panel of highly specific TACAs, based upon differences in the glycomic profiles between CRC and healthy colon mucosa. These TACAs are promising new targets for development of innovative cancer immune target therapies and lay the foundation for the targeted treatment of CRC.

A Comprehensive Analysis of the Thrombin Binding Aptamer Containing Functionalized Pyrrolo-2'-deoxycytidines.

Aptamers constitute an answer for the growing need for targeted therapy development. One of the most well-known representatives of this group of compounds is thrombin binding aptamers (TBA) targeted towards thrombin. The TBA inhibitory activity is determined by its spatial arrangement, which consists of two G-tetrads linked by two shorter TT loops and one longer TGT loop and folds into a unimolecular, antiparallel G-quadruplex structure. Interesting properties of the aptamer can be further improved via the introduction of a number of chemical modifications. Herein, a comprehensive analysis of the influence of pyrrolo-2’-deoxycytidine (Py-dC) and its derivatives on TBA physicochemical and biological properties has been presented. The studies have shown that the presence of modified residues at the T7 position of the TGT loop has only minor effects on TBA thermodynamic stability without affecting its folding topology. All analyzed oligomers exhibit anticoagulant properties, but only aptamer modified with a decyl derivative of Py-dC was able to inhibit thrombin activity more efficiently than unmodified, parental compounds. Importantly, the same compound also possessed the potential to effectively restrain HeLa cell line growth.

Alterations of Chromatin Regulators in the Pathogenesis of Urinary Bladder Urothelial Carcinoma.

Simple SummaryUrinary bladder cancer is one of the ten major cancers worldwide, with higher incidences in males, in smokers, and in highly industrialized countries. New therapies beyond cytotoxic chemotherapy are urgently needed to improve treatment of these tumors. A better understanding of the mechanisms underlying their development may help in this regard. Recently, it was discovered that a group of proteins regulating the state of chromatin and thus gene expression is exceptionally and frequently affected by gene mutations in bladder cancers. Altered function of these mutated chromatin regulators must therefore be fundamental in their development, but how and why is poorly understood. Here we review the current knowledge on changes in chromatin regulators and discuss their possible consequences for bladder cancer development and options for new therapies.Urothelial carcinoma (UC) is the most frequent histological type of cancer in the urinary bladder. Genomic changes in UC activate MAPK and PI3K/AKT signal transduction pathways, which increase cell proliferation and survival, interfere with cell cycle and checkpoint control, and prevent senescence. A more recently discovered additional category of genetic changes in UC affects chromatin regulators, including histone-modifying enzymes (KMT2C, KMT2D, KDM6A, EZH2), transcription cofactors (CREBBP, EP300), and components of the chromatin remodeling complex SWI/SNF (ARID1A, SMARCA4). It is not yet well understood how these changes contribute to the development and progression of UC. Therefore, we review here the emerging knowledge on genomic and gene expression alterations of chromatin regulators and their consequences for cell differentiation, cellular plasticity, and clonal expansion during UC pathogenesis. Our analysis identifies additional relevant chromatin regulators and suggests a model for urothelial carcinogenesis as a basis for further mechanistic studies and targeted therapy development.

Abstract PO-079: Proteomic profiling reveals subtype specific kinase expression in pancreatic cancer

Abstract Pancreatic adenocarcinoma (PDAC) remains an extremely lethal disease with few effective therapeutic options. Targeted therapy development has remained largely unsuccessful due to complex tumor heterogeneity and better molecular indicators of response are urgently needed. We have previously identified two distinct molecular tumor subtypes in PDAC, basal-like and classical, which are prognostic and dictate response to chemotherapy. We have also developed a patient derived xenograft (PDX) mouse model that maintains the integrity of patient tumor biology and recapitulates primary tumor subtypes in vivo. Here, we aim to utilize multiplexed kinase inhibitor beads and quantitative mass spectrometry (MIB-MS) to determine baseline kinase expression and adaptive kinome responses to therapy in PDAC PDX tumors. A total of 381 kinases were identified across all samples and tumor specific kinases were isolated by alignment to human peptides. Distinct kinase expression profiles were observed for basal-like and classical tumors. Basal-like tumors showed high expression of kinases involved in receptor tyrosine kinase activity, MAP kinase activity, and clathrin-dependent endocytosis. Classical subtype tumors showed increased expression of kinases involved in metabolism and cytoskeletal regulatory kinases. Differential protein expression was further validated by differential gene expression using RNAseq. MIB profiling of kinome response to EGFR inhibition by erlotinib indicated kinome reprogramming with upregulation of microtubule associated kinases and MAP kinase family members. Overall, these results suggest that basal-like and classical subtype tumors exhibit distinct kinome profiles that present possible subtype specific vulnerabilities to kinase inhibition. Stratifying kinase inhibitor therapies based on tumor subtype could improve response to targeted therapies in PDAC. Citation Format: Yi Xu, Michael East, Ashley Morrison, Gabriela Herrera, Laura Peng, Gary Johnson, Jen Jen Yeh. Proteomic profiling reveals subtype specific kinase expression in pancreatic cancer [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-079.