Early Carcinogenesis Research Articles

Genomes and epigenomes of matched normal and tumor breast tissue reveal diverse evolutionary trajectories and tumor-host interactions.

Normal tissues adjacent to the tumor (NATs) may harbor early breast carcinogenesis events driven by field cancerization. Although previous studies have characterized copy-number (CN) and transcriptomic alterations, the evolutionary history of NATs in breast cancer (BC) remains poorly characterized. Utilizing whole-genome sequencing (WGS), methylation profiling, and RNA sequencing (RNA-seq), we analyzed paired germline, NATs, and tumor samples from 43 individuals with BC in Hong Kong (HK). We found that single-nucleotide variants (SNVs) were common in NATs, with one-third of NAT samples exhibiting SNVs in driver genes, many of which were present in paired tumor samples. The most frequently mutated genes in both tumor and NAT samples were PIK3CA, TP53, GATA3, and AKT1. In contrast, large-scale aberrations such as somatic CN alterations (SCNAs) and structural variants (SVs) were rarely detected in NAT samples. We generated phylogenetic trees to investigate the evolutionary history of paired NAT and tumor samples. They could be categorized into tumor only, shared, and multiple-tree groups, the last of which is concordant with non-genetic field cancerization. These groups exhibited distinct genomic and epigenomic characteristics in both NAT and tumor samples.Specifically, NAT samples in the shared-tree group showed higher number of mutations, while NAT samples belonging to the multiple-tree group showed a less inflammatory tumor microenvironment (TME), characterized by a higher proportion ofregulatory Tcells (Tregs) and lower presence of CD14 cell populations. In summary, our findings highlight the diverse evolutionary history in BC NAT/tumor pairs and the impact of field cancerization and TME in shaping the genomic evolutionary historyof tumors.

Global loss of promoter-enhancer connectivity and rebalancing of gene expression during early colorectal cancer carcinogenesis.

Although three-dimensional (3D) genome architecture is crucial for gene regulation, its role in disease remains elusive. We traced the evolution and malignant transformation of colorectal cancer (CRC) by generating high-resolution chromatin conformation maps of 33 colon samples spanning different stages of early neoplastic growth in persons with familial adenomatous polyposis (FAP). Our analysis revealed a substantial progressive loss of genome-wide cis-regulatory connectivity at early malignancy stages, correlating with nonlinear gene regulation effects. Genes with high promoter-enhancer (P-E) connectivity in unaffected mucosa were not linked to elevated baseline expression but tended to be upregulated in advanced stages. Inhibiting highly connected promoters preferentially represses gene expression in CRC cells compared to normal colonic epithelial cells. Our results suggest a two-phase model whereby neoplastic transformation reduces P-E connectivity from a redundant state to a rate-limiting one for transcriptional levels, highlighting the intricate interplay between 3D genome architecture and gene regulation during early CRC progression.

ARP2/3 complex affects myofibroblast differentiation and migration in pancreatic ductal adenocarcinoma.

The ARP2/3 complex, which orchestrates actin cytoskeleton organization and lamellipodia formation, has been implicated in the initiation of pancreatic ductal adenocarcinoma (PDAC). This study aims to clarify its impact on the activity of cancer-associated fibroblasts (CAFs), key players in PDAC progression, and patient outcomes. Early pancreatic carcinogenesis was modeled in p48Cre; LSL-KrasG12D mice with caerulein-induced pancreatitis, complemented by in vitro studies on human immortalized pancreatic stellate cells (PSCs) and primary PDAC-derived CAFs. Data were gained from microarray analysis, RNA sequencing (RNA-seq), and single-cell RNA sequencing (sc-RNA-seq), with subsequent bioinformatics analysis. We uncovered a specific transcriptional signature associated with fibroblast migration in early pancreatic carcinogenesis and linked it to poor survival in patients with PDAC. A pivotal role of the ARP2/3 complex in CAF migration was identified. Inhibition of the ARP2/3 complex markedly decreased CAF motility and induced significant morphological changes in vitro. Furthermore, its inhibition also hindered TGFβ1-mediated myofibroblastic CAF differentiation but had no effect on IL-1-mediated inflammatory CAF differentiation. Our findings position the ARP2/3 complex as central to the migration and differentiation of myofibroblastic CAF. Targeting this complex presents a promising new therapeutic avenue for PDAC treatment.

Obesity Promotes Marrow-Derived Myeloid Cell Accumulation while Exercise Reduces Proliferative Signaling in Colon Cancer.

Obesity increases colon cancer risk that has been previously linked to marrow-derived myeloid cells. We previously demonstrated that exercise training (EX) prevents colon cancer initiation, potentially through reduced myelopoiesis. However, it remains unknown whether early myeloid cell accumulation and inflammation in the colon precedes carcinogenesis with high-fat diet (HFD)-induced obesity, and if EX can attenuate these effects. We hypothesized that obesity would promote colon carcinogenesis that was preceded by myeloid cell accumulation and inflammation that would be attenuated by EX. C57BL/6 mice were randomized to a HFD or control (CON) diet for 8 weeks. The HFD mice switched to CON diet and all mice were given intraperitoneal injections of azoxymethane (AOM) to induce colon cancer and randomized into EX or sedentary (SED) conditions. HFD mice developed more aberrant crypt foci (ACF), a marker for early carcinogenesis, compared to CON (p < 0.01), and EX developed fewer ACF compared to SED (p < 0.0001). Marrow-derived (p < 0.001) CD206+ macrophages were elevated in HFD compared to CON at study week 16 (p < 0.01). Marrow-derived CD206- macrophages (p < 0.05) and marrow-derived (p < 0.05) CD206+ macrophages were more abundant in HFD compared to CON at study week 42. EX did not alter colon immune cell populations. β-Catenin protein was higher in HFD compared to CON at study week 42 (p < 0.05), and STAT3 protein content was lower at study week 28 with EX compared to SED (p < 0.05). The results suggest that obesity promotes colon ACF formation, potentially through early inflammatory myeloid cell accumulation. Despite attenuating ACF, EX did not alter myeloid cell accumulation in the colon, suggesting that EX inhibits ACF formation through alternative mechanisms which may include reduced β-Catenin and STAT3 signaling.

Exploring the Potential of Terpenoids as a Possible Treatment for Cancer: Structure-Activity Relationship and Mechanistic Studies.

Cancer stands as a significant global health challenge due to its mortality rates and the complexities involved in its treatment. Addressing issues, such as metastasis, recurrence, chemoresistance, and treatment-related toxicity, remains pivotal in cancer therapy advancement. Therefore, exploration of novel therapeutic agents has emerged as a priority. As the risk of cancer continues to rise, effective measures must be taken to combat it. One promising approach is to explore natural remedies, such as terpenoids, which have demonstrated anticancer activity. Utilizing terpenoids could aid in the development of potent compounds to fight cancer. By studying the structural makeup of various terpenoid derivatives from previous research, we can identify which structural groups are essential for their anticancer activity. This understanding of the structure-activity relationship is crucial for developing new, effective anticancer agents based on terpenoids. Terpenoids, a diverse class of plant-derived secondary metabolites composed of multiple isoprene units, have garnered attention for their potential anticancer and pharmacological qualities. Some terpenoids exhibit notable anticancer effects by concentrating on several stages of cancer development. They show promise in blocking the initiation of early carcinogenesis by the induction of cell cycle arrest, the inhibition of cancer cell differentiation, and the induction of apoptosis. This study delves into the investigation of specific terpenoids showcasing promising anticancer activity against prevalent malignancies, including breast, colon, ovarian, and lung cancers. The study also explores the relationship between the structure and activity of these compounds, which sheds light on how effective they are against a variety of cancer cell types. The comprehensive discussion centres on elucidating terpenoids with substantial potential for combating diverse cancer types, offering insights into their structural features and promising anticancer mechanisms.

Characterization of Extractable and Non-Extractable Phenols and Betalains in Berrycactus (Myrtillocactus geometrizans) and Its Chemoprotective Effect in Early Stage of Colon Cancer In Vivo.

This research identified the bioactive compounds and antioxidant capacity of the extractable (EP) and non-extractable (NEP) polyphenol fractions of berrycactus (BC). Additionally, the effects of BC and its residue (BCR) on preventing AOM/DSS-induced early colon carcinogenesis were evaluated in vivo. Male Sprague Dawley rats were randomly assigned to six groups (n = 12/group): healthy control (C), AOM/DSS, BC, BCR, BC+AOM/DSS, and BCR+AOM/DSS. NEP was obtained through acid hydrolysis using H2SO4 and HCl (1 M or 4 M). The HCl-NEP fraction exhibited the highest total phenolic and flavonoid content, while condensed tannins were more abundant in the H2SO4-NEP fraction. A total of 33 polyphenols were identified by UPLC-QTOF-MSE in both EP and NEP, some of which were novel to BC. Both NEP hydrolysates demonstrated significant total antioxidant capacity (TEAC), with HCl-NEP exhibiting the highest ORAC values. The BC+AOM/DSS and BCR+AOM/DSS groups exhibited fewer aberrant crypt foci (p < 0.05), reduced colonic epithelial injury, and presented lower fecal β-glucuronidase activity, when compared to AOM/DSS group. No differences in butyric acid concentrations were observed between groups. This study presents novel bioactive compounds in EP and NEP from BC that contribute to chemopreventive effects in early colon carcinogenesis, while reducing fecal β-glucuronidase activity and preserving colonic mucosal integrity.

The Interplay among Wnt/β-catenin Family Members in Colorectal Adenomas and Surrounding Tissues.

The colorectal adenoma undergoes neoplastic progression via the normal epithelium-adenoma-adenocarcinoma sequence as reported in the Vogelgram. The hazard of developing a tumor is deeply associated with the number and size of adenomas and their subtype. Adenomatous polyps are histologically categorized as follows: approximately 80-90% are tubular, 5-15% are villous, and 5-10% are tubular/villous. Given the higher risk of a malignant transformation observed in tubular/villous adenomas, patients diagnosed with adenomatous polyposis are at an improved risk of developing CRC. The Wnt/β-catenin pathway plays a key role in the onset of colorectal adenoma; in particular, intestinal cells first acquire loss-of-function mutations in the APC gene that induce the formation of adenomas. Wnt/β-catenin pathway APC, Wnt3a, Wnt5a, LEF1, and BCL9 genes and protein expression analyses were conducted by qRT-PCR and western blot in 68 colonic samples (polyps and adjacent mucosa) from 41 patients, of which 17 were affected by FAP. Ten normal colonic mucosal samples were collected from 10 healthy donors. In this study, both the APC gene and protein were less expressed in the colon tumor compared to the adjacent colonic mucosa. Conversely, the activated β-catenin was more expressed in polyps than in the adjacent mucosa. All results confirmed the literature data on carcinomas. A statistically significant correlation between Wnt3a and BCL9 both in polyps and in the adjacent mucosa underlines that the canonical Wnt pathway is activated in early colon carcinogenesis and that the adjacent mucosa is already altered. This is the first study analyzing the difference in expression of the Wnt/β-catenin pathway in human colorectal adenomas. Understanding the progression from adenomas to colorectal carcinomas is essential for the development of new therapeutic strategies and improving clinical outcomes with the use of APC and β-catenin as biomarkers.

Diagnostic value of 8-OHdG, 8-iso-PGF2α and TNF-α levels in blood for early carcinogenesis of erosive oral lichen planus.

Oral cancer has a high worldwide incidence and mortality rate showing an upward trend year by year, predominantly occurring in emerging countries. Oral squamous cell carcinoma (OSCC) is one of the main types of oral cancer, accounting for more than 90% of all cases in oral cancer. To evaluate the diagnostic value of 8-hydroxy-2'-deoxyguanosine (8-OHdG), 8-iso-Prostaglandin F2alpha (8-iso-PGF2α) and tumor necrosis factor (TNF)-α as biomarkers in the early carcinogenesis of erosive oral lichen planus (EOLP) by measuring their levels in the blood of patients with EOLP and oral squamous cell carcinoma (OSCC). A total of 69 patients were enrolled in this case-control study [including an OSCC group (n= 23), an EOLP group (n= 23), and an age- and gender-matched healthy control group (n= 23)]. Blood levels of 8-OHdG, 8-iso-PGF2α and TNF-α were measured using enzyme-linked immunosorbent assay (ELISA). Statistical differences in these indicators among the three groups were analyzed. Plasma levels of 8-OHdG and 8-iso-PGF2α in the OSCC group were significantly higher than those in both the EOLP group and the control group (all P< 0.05); no significant statistical difference was found between the EOLP group and the control group. Serum levels of TNF-α in both the OSCC and EOLP groups were elevated compared with the control group, showing significant differences among all three groups (all P< 0.05). Receiver operating characteristic (ROC) curves revealed that plasma 8-OHdG and 8-iso-PGF2α levels and serum TNF-α levels had diagnostic effects on early carcinogenesis in EOLP patients. When these indicators were combined for diagnosis, the diagnostic effect was enhanced, with an area under the ROC curve (AUC) values of 0.819. 8-OHdG, 8-iso-PGF2α and TNF-α may serve as biological indicators for monitoring the early carcinogenesis of EOLP, and the diagnostic effect was augmented when these indicators were combined.

High-grade serous ovarian cancer (HGSOC) with fallopian tube involvement.

Literature data present new studies about precancerous lesions of pelvic serous carcinoma that originate from the tubal secretory cells. It has long been thought that ovarian cancer cannot be prevented by prophylactic screening or surgery. In recent years, gynecologists have adapted to new principles and so, during routine hysterectomies in perimenopausal women for benign uterine pathologies, salpingo-oophorectomy is performed as a prophylactic approach. The purpose of our article was to draw attention to the association between abnormal fallopian tube pathology and the presence of serous ovarian neoplasia in perimenopausal women at risk. We report the case of a 45-year-old woman who had unspecific symptoms of abdominal pain and loss of appetite and weight. A pelvic magnetic resonance imaging was performed, and an ovarian mass was detected. Our case shows that the fallopian tube can be the primary point of origin for a pelvic disease, therefore prevention is possible with early computed tomography scan and annual ultrasound. The patient presented with a T1c staging post-surgery and her chances of survival could have decreased if she had postponed medical examination longer. We found a significant increase in the absolute number of tubal secretory cells in patients with ovarian neoplasia, which supports the assumption that serous tubal intraepithelial carcinoma lesions are found especially in the serous ovarian type. Our article is a strong suggestion that serous ovarian cancer originates from the fallopian tube and can potentially serve as a sensitive biomarker for early serous carcinogenesis within the fallopian tube.

Transcript-Level Biomarkers of Early Lung Carcinogenesis in Bronchial Lesions

Premalignant lesions within the bronchial epithelium signify the initial phases of squamous cell lung carcinoma, posing challenges for detection via conventional methods. Instead of focusing solely on gene expression, in this study, we explore transcriptomic alterations linked to lesion progression, with an emphasis on protein-coding transcripts. We reanalyzed a publicly available RNA-Seq dataset on airway epithelial cells from 82 smokers with and without premalignant lesions. Transcript and gene abundance were quantified using kallisto, while differential expression and transcript usage analysis was performed utilizing sleuth and RATs packages. Functional characterization involved overrepresentation analysis via clusterProfiler, weighted coexpression network analysis (WGCNA), and network analysis via Enrichr-KG. We detected 5906 differentially expressed transcripts and 4626 genes, exhibiting significant enrichment within pathways associated with oxidative phosphorylation and mitochondrial function. Remarkably, transcript-level WGCNA revealed a single module correlated with dysplasia status, notably enriched in cilium-related biological processes. Notable hub transcripts included RABL2B (ENST00000395590), DNAH1 (ENST00000420323), EFHC1 (ENST00000635996), and VWA3A (ENST00000563389) along with transcription factors such as FOXJ1 and ZNF474 as potential regulators. Our findings underscore the value of transcript-level analysis in uncovering novel insights into premalignant bronchial lesion biology, including identification of potential biomarkers associated with early lung carcinogenesis.

Current and Future Trends of Colorectal Cancer Treatment: Exploring Advances in Immunotherapy.

Cancer of the colon and rectum (CRC) has been identified among the three most prevalent types of cancer and cancer-related deaths for both sexes. Even though significant progress in surgical and chemotherapeutic techniques has markedly improved disease-free and overall survival rates in contrast to those three decades ago, recent years have seen a stagnation in these improvements. This underscores the need for new therapies aiming to augment patient outcomes. A number of emerging strategies, such as immune checkpoint inhibitors (ICIs) and adoptive cell therapy (ACT), have exhibited promising outcomes not only in preclinical but also in clinical settings. Additionally, a thorough appreciation of the underlying biology has expanded the scope of research into potential therapeutic interventions. For instance, the pivotal role of altered telomere length in early CRC carcinogenesis, leading to chromosomal instability and telomere dysfunction, presents a promising avenue for future treatments. Thus, this review explores the advancements in CRC immunotherapy and telomere-targeted therapies, examining potential synergies and how these novel treatment modalities intersect to potentially enhance each other's efficacy, paving the way for promising future therapeutic advancements.

The evolution of lung adenocarcinoma precursors is associated with chromosomal instability and transition from innate to adaptive immune response/evasion.

Studying lung adenocarcinoma (LUAD) early carcinogenesis is challenging, primarily due to the lack of LUAD precursors specimens. We amassed multi-omics data from 213 LUAD and LUAD precursors to identify molecular features underlying LUAD precancer evolution. We observed progressively increasing mutations, chromosomal aberrations, whole genome doubling and genomic instability from precancer to invasive LUAD, indicating aggravating chromosomal instability (CIN). Telomere shortening, a crucial genomic alteration linked to CIN, emerged at precancer stage. Moreover, later-stage lesions demonstrated increasing cancer stemness and decreasing alveolar identity, suggesting epithelial de-differentiation during early LUAD carcinogenesis. The innate immune cells progressively diminished from precancer to invasive LUAD, concomitant with a gradual recruitment of adaptive immune cells (except CD8+ and gamma-delta T cells that decreased in later stages) and upregulation of numerous immune checkpoints, suggesting LUAD precancer evolution is associated with a shift from innate to adaptive immune response and immune evasion mediated by various mechanisms.

High Twist expression can be an early event in lip carcinogenesis

AbstractObjectiveActinic cheilitis (AC) is a chronic lesion that usually precedes the onset of squamous cell carcinoma of the lip. Microscopically, AC may exhibit epithelial dysplasia (ED). Twist is a transcription factor that regulates the mesenchymal phenotype through negative E‐cadherin regulation. This study evaluated Twist and E‐cadherin immunoexpression in AC with different ED degrees.Materials and MethodsTwist and E‐cadherin immunoexpression was evaluated semiquantitatively in the epithelium of 86 cases of AC with different ED degrees. Data obtained were submitted to the Mann–Whitney test and Spearman's correlation test.ResultsImmunohistochemical analysis revealed nuclear and cytoplasmic Twist immunoreactivity in all epithelial layers except the cornified layer. In contrast, the membrane and cytoplasmic E‐cadherin immunoreactivity was observed in all epithelial layers except the cornified layer. No significance was observed between the scores of these proteins and the ED degree. It was observed a correlation between total Twist and total (p = 0.030) and membrane (p = 0.014) E‐cadherin and between nuclear Twist and cytoplasmic E‐cadherin (p = 0.030).ConclusionsThe results suggest that the high immunoexpression of Twist is an early lip carcinogenesis event. However, it appears not to influence the progression of ED in AC.

Modeling Radiation-Induced Epithelial Cell Injury in Murine Three-Dimensional Esophageal Organoids.

Esophageal squamous cell carcinoma (ESCC) is a deadly consequence of radiation exposure to the esophagus. ESCC arises from esophageal epithelial cells that undergo malignant transformation and features a perturbed squamous cell differentiation program. Understanding the dose- and radiation quality-dependence of the esophageal epithelium response to radiation may provide insights into the ability of radiation to promote ESCC. We have explored factors that may play a role in esophageal epithelial radiosensitivity and their potential relationship to ESCC risk. We have utilized a murine three-dimensional (3D) organoid model that recapitulates the morphology and functions of the stratified squamous epithelium of the esophagus to study persistent dose- and radiation quality-dependent changes. Interestingly, although high-linear energy transfer (LET) Fe ion exposure induced a more intense and persistent alteration of squamous differentiation and 53BP1 DNA damage foci levels as compared to Cs, the MAPK/SAPK stress pathway signaling showed similar altered levels for most phospho-proteins with both radiation qualities. In addition, the lower dose of high-LET exposure also revealed nearly the same degree of morphological changes, even though only ~36% of the cells were predicted to be hit at the lower 0.1 Gy dose, suggesting that a bystander effect may be induced. Although p38 and ERK/MAPK revealed the highest levels following high-LET exposure, the findings reveal that even a low dose (0.1 Gy) of both radiation qualities can elicit a persistent stress signaling response that may critically impact the differentiation gradient of the esophageal epithelium, providing novel insights into the pathogenesis of radiation-induced esophageal injury and early stage esophageal carcinogenesis.

The value of LCI-based modified Kyoto classification risk scoring system in predicting the risk of early gastric cancer

Objective To study and compare the value of the Kyoto classification risk scoring system and the modified Kyoto classification risk scoring system based on linked color imaging (LCI) in predicting the risk of early gastric cancer. Methods One hundred and fifty patients with pathologically confirmed non-cardia early gastric cancer by endoscopic LCI and 150 non-gastric cancer patients matched for age and gender were included. Basic patient data and whole gastric endoscopic images under LCI were collected, and the images were scored according to the LCI-based Kyoto classification risk scoring system and the LCI-based modified Kyoto classification risk scoring system. Results Compared with the LCI-based Kyoto classification risk scoring system, the LCI-based modified Kyoto classification risk scoring system had a higher AUC for predicting the risk of early gastric cancer (0.723 vs. 0.784, p = 0.023), with a score of ≥3 being the best cutoff value for predicting the risk of early gastric cancer (sensitivity 61.33%, specificity 86.00%), and scores of 3 to 5 were significantly associated with early gastric carcinogenesis significantly (OR = 9.032, 95% CI: 4.995–16.330, p < 0.001). Conclusions Compared with the LCI-based Kyoto classification risk scoring system, the LCI-based Kyoto modified classification risk scoring system has a better value for predicting the risk of early gastric cancer, and the score of 3 to 5 is a high-risk factor for the risk of early gastric cancer development, which is more strongly correlated with the risk of early gastric cancer.

Abstract 3387: Spatial analysis revealed distinct molecular and immune landscape in genetic versus carcinogen-induced lung adenocarcinoma precancer mouse models

Abstract Introduction: Our understanding of early carcinogenesis of lung adenocarcinoma (LUAD) is still rudimentary. Single-cell technologies have unveiled the intricate interaction between tumor cells and tumor microenvironment (TME) with unprecedented precision. However, the spatial context of single-cell phenotypes within TME during LUAD early carcinogenesis remain poorly understood. Here, we sought to characterize human-relevant murine LUAD precancer models to elucidate the molecular and immune changes in the spatial context during early LUAD carcinogenesis to inform lung cancer interception strategies. Methods: We have developed genetically engineered mouse models (GEMMs) (129S4 KrasG12D) and carcinogen-induced LUAD precancer model (CITM) (129S4 Urethane). Lung tissues were collected at various time points after induction and analyzed using whole exome sequencing (WES), imaging mass cytometry (IMC), and spatial transcriptomics (10x Visium). Results: Pathological image analysis demonstrated a significantly higher tumor burden (p&amp;lt;0.01) in the 129S4 KrasG12D model compared to the 129S4 Urethane model across different time points. WES revealed a progressive increase in tumor mutation burden (TMB) from precancers to invasive LUADs in both models, with the CITM model having significantly higher TMB (p&amp;lt;0.05) at matched timepoints than GEMM model. IMC assessed 39 biomarkers in 284 regions of interest (ROIs) and identified 1.4 million individual cells with 17 major cell types. Macrophages were the most prevalent immune cell type in TME, representing 9.00% of total cells in GEMM and 8.29% in CITM. The percent immune cells of the total cells progressively decreased from early-stage hyperplasia to late-stage adenocarcinoma in both models. The CITM exhibited a significantly (p&amp;lt;0.05) higher immune proportion than the GEMM model at hyperplasia and adenoma stages, with no significant difference at the invasive adenocarcinoma tumor cores between these 2 models. On the other hand, CITM had a significantly higher overall immune proportion (p&amp;lt;0.05) and the CD8 T cell and B cell proportions than GEMM tumor adjacent lung tissues at the adenocarcinoma stage leading to a significantly higher immune cell gradient between tumor adjacent lung tissues and tumor cores in CITM models. Spatial transcriptomics (ST) data demonstrated an increased presence of immune response signaling at the tumor adjacent lung tissues compared to the tumor core in the latest stage. Conclusion: Spatial analysis revealed different molecular and immune landscapes in the genetic and carcinogen-induced LUAD precancer models, leading to distinct cellular spatial arrangements within the tumor microenvironment in each model. Overall, innate immunity, particularly macrophage, probably plays critical roles during early carcinogenesis of LUAD regardless of carcinogen exposure. Citation Format: Bo Zhu, Pingjun Chen, Jian-Rong Li, Muhammad Aminu, Chenyang Li, Hong Chen, Yanhua Tian, Frank R. Rojas, Parra Cuentas Edwin Roger, Ou Shi, Monique B. Nilsson, Alissa Poteete, Shawna M. Hubert, Khaja B. Khan, Wei Lu, Xingzhi Song, Jianhua Zhang, Don L. Gibbons, Luisa M. Solis Soto, Ignacio I. Wistuba, John V. Heymach, Chao Cheng, Jia Wu, Jianjun Zhang. Spatial analysis revealed distinct molecular and immune landscape in genetic versus carcinogen-induced lung adenocarcinoma precancer mouse models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3387.

Abstract 6098: Spatial single-cell immune microenvironment analysis reveals the transition from innate to adaptive immune response during early lung adenocarcinoma carcinogenesis

Abstract Introduction: Lung cancer remains the leading cause of cancer death worldwide. Although prevention holds the promise to reduce lung cancer mortality, the results from randomized trials have been disappointing, likely due to our rudimentary knowledge of early lung cancer carcinogenesis. Cancer initiation and progression involve complex processes dependent on the dynamic interplay of individual cells within the tumor microenvironment (TME). Our understanding of the multicellular composition, interaction, spatial distribution, and functional dynamics during the early evolution from precancer to invasive LUAD is limited. Methods: We performed spatial single cell immune microenvironment analysis of 10 normal, 40 AAH, 22 AIS, 18 MIA, and 34 IAC by Imaging Mass Cytometry (IMC) and analyzed high-dimensional images of 1,618 regions-of-interest (ROIs) using a 34-plex marker. Using machine learning approaches, we classified 4,828,879 cells into 14 major cell types, which were subjected to further analysis for multicellular composition, interaction, spatial distribution, and functional dynamics in LUAD precursors of different stages. Results: From AAH to AIS, MIA, and IAC, we observed a gradual escalation in the density (number of cells/mm2) of all immune subsets. Proportionally, adaptive immune cells exhibited a progressive increase, whereas innate immune cells demonstrated a concurrent decrease with the neoplastic progression from normal lung to AAH, AIS, MIA, and ADC. Notably, macrophages emerged as the predominant immune cells within the immune microenvironment of these LUAD precursors. Upon closer examination of subtypes, protumor M2 macrophages showed a gradual increase from AAH to AIS, MIA, and IAC, whereas anti-tumor M1 macrophages increased from normal to AIH, AIS and subsequently decreased in AIS, MIA, and IAC stages. These alterations resulted in a progressive increase in M2/M1 ratios from AAH to IAC stages. Furthermore, M2 macrophages exhibited higher heterotypic interactions with epithelial cells than M1 across LUAD precursors of different stages, especially in the later stages. Co-expression analysis revealed 5 modules that distinguishes each histological stage. Tim-3 module was significantly enriched in AAH lesions, while NK cell-related features were enriched at AIS stage, B cell-related features enriched at MIA stage and Ki-67-related adaptive immune features enriched in IAC stage. Conclusion: The spatial single cell immune microenvironment analysis revealed a transition from innate to adaptive immunity during initiation and early progression of LUAD progression and innate immunity may play critical roles during early LUAD development. Therefore, reprograming innate immunity such as repolarization of M2/M1 may provide avenues to intercept LUAD precursors to prevent invasive LUAD. Citation Format: Bo Zhu, Pingjun Chen, Muhammad Aminu, Junya Fujimoto, Lingzhi Hong, Andre L. Moreira, Jian-Rong Li, Yanhua Tian, Luisa M. Solis Soto, Parra Cuentas Edwin Roger, Ou Shi, Hong Chen, Frank R. Rojas, Eduardo Vilar, Anirban Maitra, Ken Chen, Nicolas Navin, Cara L. Haymacker, Vamsidhar Velcheti, Daniel H. Sterman, P Andrew Futreal, Don L. Gibbons, Ignacio I. Wistuba, John V. Heymach, Chao Cheng, Harvey I. Pass, Jia Wu, Jianjun Zhang. Spatial single-cell immune microenvironment analysis reveals the transition from innate to adaptive immune response during early lung adenocarcinoma carcinogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6098.

Abstract 7367: Dissecting cellular and molecular alterations in pre-invasive to invasive lung cancer progression at single-cell resolution

Abstract Non-small cell lung cancer (NSCLC) accounts for most cancer-related deaths worldwide, largely due to diagnosis at advanced metastatic stages. Increased implementation of low-dose computed tomography (CT)-guided screening has led to the identification of non-solid lesions or ground-glass opacities (GGOs) that are not visible on plain chest radiography and can harbor very early-stage tumors. Growth of the lesions and/or development of a solid component can indicate more aggressive behavior. Using the low-dose CT reduced lung cancer mortality by &amp;gt;20%, underscoring the need for early detection and intervention. However, the cellular and molecular determinants contributing to the malignant progression of those early preinvasive lesions remain poorly understood. To assess intratumoral heterogeneity and associated dynamic changes in the immune contexture during early lung carcinogenesis, we performed single-cell RNA-seq and Hyperion imaging on surgically resected freshly harvested tissues from 37 patients (matched solid and non-solid components of lesions and adjacent non-involved lungs), representing a spectrum from preneoplasia to invasive disease. Analysis of &amp;gt;300K cells identified cellular, transcriptomic, and metabolic alterations as well as tumor-stroma cross-talk pathways. Specifically, preinvasive to invasive disease progression was associated with significant changes in the immune landscape determined by an increase in exhausted CD4/CD8 T-cells, suppressive T-regs, polymorphonuclear myeloid-derived suppressor cells, and M2-like macrophages; and a decrease in cytotoxic CD8 T-cells, NK cells, and inflammatory cDC2 cells, suggesting that activation of immunosuppressive/anti-inflammatory microenvironment and suppression of immune activation microenvironment may enable tumor progression. Citation Format: Liron Yoffe. Dissecting cellular and molecular alterations in pre-invasive to invasive lung cancer progression at single-cell resolution [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7367.

Abstract 7324: Impact of BMI on immune pathways in preinvasive colorectal lesions

Abstract Background: Obesity is a risk factor for colorectal neoplasia, possibly mediated by metabolic dysregulation or chronic inflammation. Circulating inflammatory cytokines have been associated with an increased risk of CRC, but no studies have examined the impact of BMI on immune gene expression in preinvasive lesions. Our analysis combines data from 3 retrospective epidemiologic studies among patients with colorectal adenomas or serrated polyps. Using a cross-sectional design, we investigate whether BMI categories (obese (OB), overweight (OW), normal (NL)) are associated with T-helper gene pathways. Methods: For each patient, we include demographic, clinical, and specimen (archival FFPE for RNA isolation) data. Immune gene expression data was determined using the NanoString nCounter platform on a subset of cases (n=100) balanced by study site and lesion type from our cohort (n=630). Genes were normalized using a modified RUVseq methodology. We examined 4 pathways (Th1, Th2, Th17, Treg) defined by gene lists included in the NS immune v2. panel. Differences in pathway expression by BMI categories were analyzed using our distance-based methods (Wd* multivariate analysis of variance and TW2 pairwise method) adjusted for age &amp; sex. Using linear regression analysis, we generated coefficients (p-values) for OW and OB vs NL for genes within each pathway. Results: 86 samples passed QC. Th2 and Th1 pathways were significant for OW vs NL (p&amp;lt;0.05) whereas Th17 was marginally significant (&amp;lt;0.1). Several genes within the Th1, Th2, and Th17 pathways were enriched in OW vs. NL: e.g., IL2 (coeff. 0.77 (p=0.008), IL5 (coeff. 1.02, p=0.001), IL17F (coeff. 0.57, p=0.05), respectively. (Table) Conclusion: Enrichment of genes specific to the Th pathways in OW persons suggest that metabolic immune modulation may be important in early colorectal carcinogenesis. Differences in immune pathway and gene expression by BMI categories TW2 (p-value) Wd* (p-value) Pathways Normal (N = 22) Overweight (N = 32) Obese (N = 29) Th2 Ref. 0.017 0.15 0.038 Th1 Ref. 0.03 0.204 0.10 Th17 Ref. 0.073 0.125 0.103 MLR coefficient (p-value) Pathway Gene Overweight vs. Normal Obese vs. Normal ANOVA (p-value) Th2 IL2RA 0.34 (0.041) -0.11 (0.517) 0.010 IL4 0.58 (0.019) 0.22 (0.386) 0.052 IL5 1.02 (0.001) 0.41 (0.191) 0.004 NOTCH1 -0.05 (0.742) -0.34 (0.045) 0.078 Th1 IL12B 0.57 (0.031) 0.40 (0.136) 0.093 IL2 0.77 (0.008) 0.39 (0.179) 0.028 TBX21 0.46 (0.015) 0.19 (0.308) 0.044 Th17 IL17F 0.57 (0.050) 0.42 (0.157) 0.139 IL21 0.77 (0.004) 0.42 (0.131) 0.017 IL23R 0.29 (0.048) -0.008 (0.957) 0.047 IL6R 0.002 (0.990) 0.36 (0.039) 0.041 IRF4 -0.61 (0.007) -0.33 (0.149) 0.025 TGFB1 -0.26 (0.023) -0.19 (0.101) 0.069 Citation Format: Lauren R. Fanning, Bashir Hamidi, Ivan Korostenskij, Souvik Seal, Tami L. Crawford, Todd A. MacKenzie, Elizabeth L. Barry, Jessica E. Thaxton, Martha J. Shrubsole, Benjamin Vincent, John A. Baron, Alexander V. Alekseyenko, Kristin Wallace. Impact of BMI on immune pathways in preinvasive colorectal lesions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7324.

Significance of p53, cyclooxygenase-2, and epithelial cell adhesion molecule expression in hepatocellular carcinoma: an immunohistochemical study

Background Hepatocellular carcinoma (HCC) is a significant global health concern with a high mortality rate. To date, the most effective therapy for HCC is resection at an early tumor stage. However, tumor recurrence is common, and identifying key molecules facilitates the understanding of the pathogenesis of HCC and the prediction of prognosis to provide novel targets for anticancer therapy. Aim This study evaluated the expression of p53, cyclooxygenase-2 (COX-2), and epithelial cell adhesion molecule (EpCAM) in HCC and investigated their correlation with clinicopathological features and prognosis. Methods An Immunohistochemical analysis of p53, COX-2, and EpCAM was conducted on selected 51 HCC cases and adjacent noncancerous hepatic tissue. Results In the current study, p53, COX-2, and EpCAM expression were significantly higher in HCC cases than in the adjacent nontumor tissue (P=0.05, P=0.03, and P=0.041, respectively). P53, COX-2, and EpCAM were significantly overexpressed among patients with advanced stage (P=0.039, P=0.000, and P=0.016, respectively), large tumor size (P=0.004 and P=0.001) and poor disease-free survival (P=0.036, P=0.001, and P=0.000, respectively). P53 and EpCAM were significantly correlated with vascular invasion (P=0.045 and P=0.032) and higher grade (P=0.019 and P=0.033). While COX-2 was associated with well-differentiated HCC cases. There was no statistically significant correlation between p53 and COX-2 or, EpCAM, while COX-2 was directly correlated with EpCAM (r=0.001). Conclusion p53, COX-2, and EpCAM might have an important role in early carcinogenesis, progression of HCC, and poor prognosis, suggesting that the inhibition of these proteins may hold potential as a multitarget therapeutic approach in HCC patients.