Lung Colorectal Cancer Research Articles

Targeting KAT7 inhibits the progression of colorectal cancer.

Rationale: Colorectal cancer (CRC) is a leading cause of cancer-related mortality. Epigenetic modifications play a significant role in the progression of CRC. KAT7, a histone acetyltransferase, has an unclear role in CRC. Methods: In this research, we analyzed the expression of KAT7 in CRC patients and its correlation with prognosis using the GEO database, western blot, and immunohistochemistry. We assessed the impact of KAT7 on CRC cell functions through cell viability, colony formation, flow cytometry, scratch, and transwell assays. Mechanistic insights were obtained via RNA sequencing and ChIP-qPCR. Additionally, we evaluated the effects of KAT7 on CRC growth and metastasis in vivo using mouse subcutaneous tumor and lung metastasis models. Results: In this study, we discovered an upregulated KAT7 signaling pathway in CRC and its association with poor patient survival. Knockdown of KAT7 promotes apoptosis and inhibits proliferation, migration, and invasion of CRC cells. Conversely, KAT7 overexpression enhanced these cellular processes. In vivo assays confirmed that knockdown of KAT7 can inhibit CRC proliferation and lung metastasis. Mechanistically, KAT7 acetylated histone H3 at lysine 14 (H3K14) to enhance MRAS transcription, which activated the MAPK/ERK pathway and promoted tumorigenesis. The enzymatic function of KAT7 as an acetyltransferase is crucial for the advancement of colorectal cancer. In KAT7 knockdown CRC cells, re-expression of KAT7, but not an acetyltransferase-deficient mutant, rescued MRAS expression, ERK phosphorylation, and CRC tumorigenesis. Conclusion: We found that KAT7 is highly expressed in CRC patients, and those with high KAT7 expression have a worse prognosis. KAT7 enhances MRAS gene transcription by promoting H3K14 acetylation, thereby activating the MAPK/ERK pathway and promoting malignant phenotypes of CRC. In summary, KAT7 represents a promising target for CRC therapy.

Yanghe decoction inhibits inflammation-induced lung metastasis of colorectal cancer.

Positive deficiency and cancer toxicity are the main pathogenesis of colorectal cancer (CRC) lung metastasis. Yanghe decoction (YHD), a traditional Chinese medicine, has the effects of warming yang, tonifying blood, dispersing cold and clearing stagnation, adopting a treatment method that combines supporting the right and dispelling the wrong, which has remarkable efficacy in anti-tumor.Although, its precise mechanism of inhibiting the metastasis of colorectal cancer to the lung is still poorly understood. This study aimed to elucidate the antitumor properties of YHD within the context of colorectal cancer lung metastasis. Ultrahigh-performance liquid chromatography coupled with mass spectrometry (UHPLC-MS) was utilized to analyze the chemical composition of YHD. The anticancer activity of YHD was evaluated in a CRC lung metastasis mouse model by quantifying pulmonary metastatic nodules. The effects of YHD on CRC cell proliferation, apoptosis, cell cycle progression, and invasion were assessed using CCK-8 assays, flow cytometry, and Transwell assays. YHD-mediated immune modulation in tumor-bearing mice was evaluated by analyzing antitumor immunity, immunosuppressive cells, and cytokines in peripheral blood and tumor tissue. Gut microbiota analysis was conducted to determine the impact of YHD on the gut microbiota in mice. Our analysis identified 1801 chemical markers in YHD. CFA exacerbated lung metastasis in CRC, whereas oral administration of YHD significantly mitigated this effect, as evidenced by the reduced number of metastatic lung nodules in CRC tumor-bearing mice. In vitro experiments demonstrated that YHD inhibits CRC cell proliferation, induces apoptosis, and suppresses invasion. In the lung tissues of mice with CRC metastasis treated with CFA, there was a significant reduction in NK cells and IL-21, along with an increase in M2 macrophages and IL-6. Following YHD treatment, there was a notable increase in NK cells and IL-21, accompanied by a decrease in M2 macrophages and IL-6 in lung tissues. YHD administration was also associated with an increase in beneficial bacterial species such as Bacillus and a decrease in deleterious bacterial species such as Oscillibacter. Our findings demonstrate that YHD inhibits lung metastasis in CRC by suppressing CRC cell proliferation and invasion, in addition to modulating the tumor microenvironment to favor antitumor immunity. These results provide a scientific basis for the clinical application of YHD in the treatment of CRC patients.

Colonization of stenotrophomonas and its associated microbiome between paired primary colorectal cancers and their lung metastatic tumors (experimental studies).

The role of gut microbiome in colorectal cancer (CRC) lung metastases is still unclear. Herein, we collected primary colorectal tumors, matched lung metastasis, and adjacent normal colon from seven patients for 16S rRNA sequencing to evaluate microbiome signature in distant site metastasis and analyze prognosis. Fecal samples from 19 non-metastases CRC patients and 11 CRC lung metastases patients were analyzed for validation. Our efforts led to a discovery of comprehensive microbiome in CRC lung metastatic tumors, characterized by a decreased microbial diversity and an increase bacterial interaction between gram-negative bacteria Stenotrophomonas and Brevundimonas . The primary-metastatic tumor pairs harbored highly similar microbial composition in both genus and species level, especially the high level of Stenotrophomonas maltophilia . Correspondingly, the S. maltophilia levels were positively associated with poor prognosis and cancer progression. Our work highlights the great potential of S. maltophilia and its associated microbiome as biomarkers in pathogenesis and prognosis of CRC lung metastases.

Arenobufagin inhibits lung metastasis of colorectal cancer by targeting c-MYC/Nrf2 axis

Colorectal cancer (CRC) is one of the commonest cancers worldwide. Metastasis is the most common cause of death in patients with CRC. Arenobufagin is an active component of bufadienolides, extracted from toad skin and parotid venom. Arenobufagin reportedly inhibits epithelial-to-mesenchymal transition (EMT) and metastasis in various cancers. However, the mechanism through which arenobufagin inhibits CRC metastasis remains unclear. This study aimed to elucidate the molecular mechanisms by which arenobufagin inhibits CRC metastasis. Wound-healing and transwell assays were used to assess the migration and invasion of CRC cells. The expression of nuclear factor erythroid-2-related factor 2 (Nrf2) in the CRC tissues was assessed using immunohistochemistry. The protein expression levels of c-MYC and Nrf2 were detected by immunoblotting. A mouse model of lung metastasis was used to study the effects of arenobufagin on CRC lung metastasis in vivo. Arenobufagin observably inhibited the migration and invasion of CRC cells by downregulating c-MYC and inactivating the Nrf2 signaling pathway. Pretreatment with the Nrf2 inhibitor brusatol markedly enhanced arenobufagin-mediated inhibition of migration and invasion, whereas pretreatment with the Nrf2 agonist tert‑butylhydroquinone significantly attenuated arenobufagin-mediated inhibition of migration and invasion of CRC cells. Furthermore, Nrf2 knockdown with short hairpin RNA enhanced the arenobufagin-induced inhibition of the migration and invasion of CRC cells. Importantly, c-MYC acts as an upstream modulator of Nrf2 in CRC cells. c-MYC knockdown markedly enhanced arenobufagin-mediated inhibition of the Nrf2 signaling pathway, cell migration, and invasion. Arenobufagin inhibited CRC lung metastasis in vivo. Together, these findings provide evidence that interruption of the c-MYC/Nrf2 signaling pathway is crucial for arenobufagin-inhibited cell metastasis in CRC. Collectively, our findings show that arenobufagin could be used as a potential anticancer agent against CRC metastasis. The arenobufagin-targeted c-MYC/Nrf2 signaling pathway may be a novel chemotherapeutic strategy for treating CRC.

Abstract 1331: MiNK-215, an IL-15 armored FAP-targeting CAR iNKT cell therapy, effectively treats human organoid models of treatment-refractory MSS colorectal cancer (CRC) liver metastases

Abstract The presence of liver metastases in patients with microsatellite-stable (MSS) or mismatch repair proficient (pMMR) colorectal cancer (CRC) is associated with poor response to current pharmacological treatments, including immune checkpoint blockade (ICB). The tumor microenvironment (TME) of CRC liver metastases is characterized by a highly immunosuppressive phenotype that includes poor immunogenicity, a paucity of dendritic cells, elimination of tumor-specific effector T cells, and an abundance of immunosuppressive intrahepatic Kupffer cells and myeloid-derived suppressor cells. This underscores the urgent need for innovative therapeutic approaches targeted specifically to patients with CRC liver metastases. Here we describe MiNK-215, an IL-15 armored FAP-targeting CAR iNKT cell therapy, derived from the AgenT-797 allogeneic iNKT platform, as a potential novel therapeutic approach for patients with CRC liver metastases. To better model ICB refractory human MSS CRC liver metastases, we developed an ex vivo human organoid model that recapitulates the histological and immunological features of human disease. Organoids were comprised of MSS CRC cells engineered to express nuclear GFP and HLA-A2-NY-ESO-1, hepatocytes, Kupffer cell-enriched non-parenchymal cells, and HLA-matched peripheral blood mononuclear cells (PBMCs) supplemented with NY-ESO-1-reactive T cells. Organoid models of CRC lung metastases, which are traditionally more responsive to ICB therapy were also developed to compare the models to clinical experience and further validate their usefulness. Models were treated with Fc-enhanced anti-CTLA-4 (botensilimab) and anti-PD-1 (balstilimab) antibodies, or MiNK-215 to assess tumor killing and immune activation. Botensilimab and balstilimab enhanced T cell activation and cytotoxicity of CRC tumor cells in the absence of “normal” liver cells and in lung organoids. However, they were less effective in organoid models of CRC with liver metastases consistent with clinical observations. In contrast, MiNK-215 potently enhanced tumor killing by T cells in models of CRC with liver metastases. Tumor killing in CRC liver organoid models by MiNK-215 was associated with depletion of immune suppressive FAP-expressing stellate cells and increased CD8+ T cell infiltration. Our findings demonstrate that in treatment-refractory CRC-liver metastatic organoid models, MiNK-215 overcomes the limitations of ICB therapy to home to sites of disease, reprogram the TME, recruit tumor-reactive T cells and enhance tumor killing. Ex-vivo human CRC liver and lung metastatic organoid models are a novel platform that can be leveraged to rapidly identify new therapeutic approaches for potential clinical evaluation. Citation Format: Shanmugarajan Krishnan, Bishnu Joshi, Justin Keith, Ryan Frazier, Barbara Kalinowska, Jin San Choi, Stephanie Sanders, Shannon Boi, Olivier Le Tonqueze, Nick Kushner, Kah Teong Soh, Robert Stein, Tyler J. Curiel, Marc A. van Dijk, Enoch Kim, Eleni Chantzoura, Nils-Petter Rudqvist, Dhan Chand. MiNK-215, an IL-15 armored FAP-targeting CAR iNKT cell therapy, effectively treats human organoid models of treatment-refractory MSS colorectal cancer (CRC) liver metastases [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 1331.

Identification and prognostic analysis of candidate biomarkers for lung metastasis in colorectal cancer.

Colorectal cancer (CRC) is one of the most prevalent types of malignant tumors. It's vital to explore new biomarkers and potential therapeutic targets in CRC lung metastasis through adopting integrated bioinformatics tools. Multiple cohort datasets and databases were integrated to clarify and verify potential key candidate biomarkers and signal transduction pathways in CRC lung metastasis. DAVID, STRING, UALCAN, GEPIA, TIMER, cBioPortal, THE HUMAN PROTEIN ATLAS, GSEA 4.3.2, FUNRICH 3.1.3, and R 4.2.3 were utilized in this study. The enriched biological processes and pathways modulated by the differentially expressed genes (DEGs) were determined with Gene Ontology, Kyoto Encyclopedia of Genes and Genomes. The search tool Retrieval of Interacting Genes and Cytoscape were used to construct a protein-protein interaction network among DEGs. Four hundred fifty-nine colorectal primary cancer and lung metastatic gene expression profiles were screened from 3 gene expression profiles (GSE41258, GSE68468, and GSE41568). Forty-one upregulated genes and 8 downregulated genes were identified from these 3 gene expression profiles and verified by the transcriptional levels of hub genes in other GEO datasets and The Cancer Genome Atlas database. Two pathways (immune responses and chemokine receptors bind chemokines), 13 key DEGs, 6 hub genes (MMP3, SFTPD, ABCA3, CLU, APOE, and SPP1), and 2 biomarkers (APOE, SPP1) with significantly prognostic values were screened. Forty-nine DEGs were identified as potential candidate diagnostic biomarkers for patients with CRC lung metastasis in present study. Enrichment analysis indicated that immune responses and chemokine receptors bind chemokines may play a leading role in lung metastasis of CRC, and further studies are needed to validate these findings.

Kaempferol inhibits colorectal cancer metastasis through circ_0000345 mediated JMJD2C/β-catenin signalling pathway

BackgroundRecurrence and metastasis are the main causes of disease deterioration in colorectal cancer (CRC) patients, yet efficient therapeutic strategies are lacking. Natural compounds for efficient antitumour therapeutics are becoming increasingly prominent. Kaempferol, one of the main components of flavonoids in plants, displays a variety of pharmacological activities. Our preliminary experiments suggested that kaempferol could inhibit CRC metastasis and is significantly associated with the β-catenin signalling pathway. Moreover, we also defined the regulatory roles of JMJD2C in β-catenin signalling in our previous work. PurposeThis study aims to reveal the mechanism by which kaempferol inhibits CRC progression and regulates the JMJD2C/β-catenin signalling pathway. MethodsThe migratory capabilities of CRC cells after kaempferol intervention were measured by scratch wound healing and transwell assays. Circ_0000345 knockdown CRC stable cell lines were generated by lentivirus infection. The possible mechanism of kaempferol on circ_0000345 was verified by molecular-protein docking and verification program cellular thermal shift assay (CETSA). A dual luciferase reporter gene assay was carried out for the targeting relationship among circ_0000345, miR-205-5p and JMJD2C. Fluorescence in situ hybridization (FISH) was performed to determine the expression of circ_0000345 in tumour tissues. A pulmonary metastatic model of CRC in vitro was built to assess the antimetastatic effect and mechanism of kaempferol in vivo. ResultsIn vitro, kaempferol inhibits the ability to migrate of CRC cells by reducing the activation of the JMJD2C/β-catenin signalling pathway. MiR-205-5p is a key bridge for kaempferol to inhibit the expression of JMJD2C. The function of miR-205-5p is impeded by circ_0000345, which shows higher expression levels in human metastatic CRC tissues than nonmetastatic CRC tissues, and its formation is regulated by the RNA-binding proteins HNRNPK and HNRNPL. Mechanistically, kaempferol physically interacts with HNRNPK and HNRNPL to suppress JMJD2C by downregulating the expression of circ_0000345. In vivo, kaempferol suppresses CRC lung metastasis. Kaempferol inhibits the activation of JMJD2C/β-catenin signalling through reducing the expression of circ_0000345 in the CRC lung metastasis model. ConclusionCirc_0000345 enhances activation of the JMJD2C/β-catenin signalling pathway through miR-205-5p to promote CRC metastasis. Kaempferol inhibits CRC metastasis through the circ_0000345-mediated JMJD2C/β-catenin signalling pathway, and this effect is influenced as a direct consequence of the binding of kaempferol with HNRNPK and HNRNPL. This provides promising therapeutic and/or adjuvant agents for advanced CRC and sheds light on the multifaceted role of phytomedicine in cancer.

Radiofrequency ablation of lung metastases of colorectal cancer: could early radiomics analysis of the ablation zone help detect local tumor progression?

To determine whether radiomics data can predict local tumor progression (LTP) following radiofrequency ablation (RFA) of colorectal cancer (CRC) lung metastases on the first revaluation chest CT. This case-control single-center retrospective study included 95 distinct lung metastases treated by RFA (in 39 patients, median age: 63.1 years) with a contrast-enhanced CT-scan performed 3 months after RFA. Forty-eight radiomics features (RFs) were extracted from the 3D-segmentation of the ablation zone. Several supervised machine-learning algorithms were trained in 10-fold cross-validation on reproducible RFs to predict LTP, with/without denoising CT-scans. An unsupervised classification based on reproducible RFs was built with k-means algorithm. There were 20/95 (26.7%) relapses within a median delay of 10 months. The best model was a stepwise logistic regression on raw CT-scans. Its cross-validated performances were: AUROC = 0.72 (0.58-0.86), area under the Precision-Recall curve (AUPRC) = 0.44. Cross-validated balanced-accuracy, sensitivity and specificity were 0.59, 0.25 and 0.93, respectively, using p = 0.5 to dichotomize the model predicted probabilities (vs 0.71, 0.70 and 0.72, respectively using p = 0.188 according to Youden index). The unsupervised approach identified two clusters, which were not associated with LTP (p = 0.8211) but with the occurrence of per-RFA intra-alveolar hemorrhage, post-RFA cavitations and fistulizations (p = 0.0150). Predictive models using RFs from the post-RFA ablation zone on the first revaluation CT-scan of CRC lung metastases seemed moderately informative regarding the occurrence of LTP. Radiomics approach on interventional radiology data is feasible. However, patterns of heterogeneity detected with RFs on early re-evaluation CT-scans seem biased by different healing processes following benign RFA complications.

MicroOrganoSpheres as a clinically applicable precision oncology platform for the discovery of novel therapies in colorectal cancer.

3592 Background: Patient-derived models of cancer, such as cell lines, patient-derived organoids, and patient-derived xenografts, are useful models of patient response in the clinic. However, these models are often not clinically applicable within the time periods necessary to inform clinical decision making, as they can take weeks to months to develop. An ideal platform using patient-derived models would be generated from a core biopsy with a subsequent drug screen within 10-14 days to minimize delay in therapy. We recently reported the development of MicroOrganoSpheres (MOS) that can be used in drug screens within 14 days of obtaining a biopsy. In the current study, we use this MOS system as precision oncology platform in colorectal cancer (CRC) to identify new therapies and predict response to therapy. Methods: CRC patient tissue samples were collected under a Duke Institutional Review Board approved protocol (Pro00089222). Resections or biopsies were mechanically and enzymatically digested to obtain a single cell suspension. Cells were then plated in Matrigel at a ratio of 20,000 cells:5 µL Matrigel to establish “mini-bulk” organoid cultures. After establishment for 5-7 days, cultures were harvested with subsequent generation of MOS at a ratio of 50 cells per MOS. After growing for 3-4 days, MOS were used for dose-response curves using oxaliplatin, SN38, and 5-Fluorouracil (5-FU) as well as high-throughput drug screens with the NCI Approved Oncology Drugs Set VI library. Results: We developed and optimized a MOS pipeline on over 50 CRC specimens, including 9 primary rectal, 35 primary CRC, 12 CRC liver metastasis, and 1 CRC lung metastasis lines with a success rate of 80% and an average of 10-21days from biopsy to MOS generation. The high success of generating CRC MOS in a clinically applicable time frame led to the next phase of the project where a total of 10 CRC MOS were tested against standard of care chemotherapy agents used in CRC (oxaliplatin, irinotecan and 5-FU) as well as the NCI Approved Oncology Drugs Set VI within 14-21days of establishment. We noted a range of sensitivity of approximately 100-fold for standard of care agents. The most sensitive drugs found in the high-throughput screen were Bortezomib, Carfilzomib, and Panobinostat and the most resistant were Gefitinib, Chlorambucil, and Procarbazine hydrochloride. Conclusions: These results demonstrate that our MOS pipeline can be used as a precision oncology platform within a clinically applicable time frame to potentially guide therapy. We are now in the process of correlating drug response in MOS to patient outcome data and these findings will be presented at the annual meeting.

Abstract 5735: Novel KRAS G12D degrader ASP3082 demonstrates in vivo, dose-dependent KRAS degradation, KRAS pathway inhibition, and antitumor efficacy in multiple KRAS G12D-mutated cancer models

Abstract KRAS is one of the most frequently mutated oncogenes in various cancers. Among KRAS mutations, KRAS G12D is the most frequent driver mutation and is found in approximately 34% of patients with pancreatic ductal adenocarcinoma (PDAC), 12% of patients with colorectal cancer (CRC), 4% of patients with lung adenocarcinoma, and in a subset of patients with other solid tumors. We have identified ASP3082 as a novel KRAS G12D degrader with high potency and selectivity. Here, we have evaluated in vivo antitumor activities and pharmacodynamic properties of ASP3082 in various KRAS G12D-mutated xenograft models. ASP3082 was intravenously administered to KRAS G12D-mutated cancer-xenograft-bearing mice, and plasma and tumors were collected at defined time points. The drug concentration and KRAS-related signal transduction were measured in the xenograft model. The in vivo efficacy of ASP3082 monotherapy was confirmed in multiple xenograft mouse models following intravenous administration. Once-weekly intravenous administration of ASP3082 induced dose-dependent and significant growth inhibition of KRAS G12D PDAC tumors, resulting in profound tumor regression without body weight loss. ASP3082 showed sustained concentrations in the xenograft tumors after a single intravenous administration and decreased KRAS G12D-mutated-protein levels according to the duration of the sustained ASP3082 concentrations. ASP3082 also demonstrated marked inhibition of extracellular signal-regulated kinase phosphorylation and its downstream genes, and potently induced cleavage of caspase 3. In addition, ASP3082 exhibited potent antitumor activities in not only PDAC but also CRC and non-small cell lung cancer KRAS G12D-mutated mouse models. These studies demonstrated that ASP3082 induced degradation of KRAS G12D protein, inhibition of KRAS downstream molecules, and an apoptotic response to show dose-dependent antitumor activity in multiple KRAS G12D-mutated cancer models. ASP3082 is a potential therapeutic agent for patients with tumors harboring the KRAS G12D mutation. Currently, a phase 1 clinical trial is underway in patients with previously treated, locally advanced or metastatic solid tumors with KRAS G12D mutation (NCT05382559). Citation Format: Takeyuki Nagashima, Tomohiro Yoshinari, Yoshihiro Nishizono, Mamoru Tasaki, Kohei Inamura, Hiroki Ishioka, Atsushi Suzuki, Fumio Osaki, Yosuke Yamanaka, Masahiko Hayakawa. Novel KRAS G12D degrader ASP3082 demonstrates in vivo, dose-dependent KRAS degradation, KRAS pathway inhibition, and antitumor efficacy in multiple KRAS G12D-mutated cancer models. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5735.

Circular RNA ACACA negatively regulated p53-modulated mevalonate pathway to promote colorectal tumorigenesis via regulating miR-193a/b-3p/HDAC3 axis.

This study aimed to explore the biological functions and underlying mechanism of circRNA acetyl-CoA carboxylase alpha (circACACA) in colorectal cancer (CRC). The RNA and protein levels were detected by qRT-PCR and western blot assays. The malignant capacities of CRC cells were analyzed by cell counting kit-8 (CCK-8), colony formation, flow cytometry, and transwell assays. The target relationship between miR-193a/b-3p and circACACA/histone deacetylase 3 (HDAC3) was determined by luciferase reporter assay and RNA immunoprecipitation. The binding of HDAC3 to the p53 promoter was validated by chromatin immunoprecipitation (ChIP). CRC cell growth and lung metastasis were evaluated in nude mice in vivo. High expression of circACACA was found in CRC tissues and cells, which was closely associated with the advanced tumor, lymph node, metastasis (TNM) stage, metastasis, and low overall survival rate. circACACA downregulation effectively delayed CRC cell proliferation and metastasis, but triggered apoptosis via inactivating the mevalonic acid (MVA) pathway. However, circACACA overexpression resulted in the opposite effects. Mechanistically, circACACA enhanced HDAC3 expression through sponging miR-193a/b-3p, which activated theMVA pathway via inhibiting the acetylation and transcription of p53. Moreover, rescue experiments confirmed that miR-193a/b-3p inhibition reversed the inhibitory effect of circACACA deficiency on CRC growth and metastasis. Moreover, circACACA overexpression-mediated malignant phenotypes of CRC cells were abrogated by HDAC3 knockdown. circACACA promoted CRC progression via regulating themiR-193a/b-3p/HDAC3/p53 axis to activate theMVA pathway, providing evidence for circACACA as a promising therapeutic target for CRC.

A CT-based radiomics approach to predict immediate response of radiofrequency ablation in colorectal cancer lung metastases.

To objectively and accurately assess the immediate efficacy of radiofrequency ablation (RFA) on colorectal cancer (CRC) lung metastases, the novel multimodal data fusion model based on radiomics features and clinical variables was developed. This case-control single-center retrospective study included 479 lung metastases treated with RFA in 198 CRC patients. Clinical and radiological data before and intraoperative computed tomography (CT) scans were retrieved. The relative radiomics features were extracted from pre- and immediate post-RFA CT scans by maximum relevance and minimum redundancy algorithm (MRMRA). The Gaussian mixture model (GMM) was used to divide the data of the training dataset and testing dataset. In the process of modeling in the training set, radiomics model, clinical model and fusion model were built based on a random forest classifier. Finally, verification was carried out on an independent test dataset. The receiver operating characteristic curves (ROC) were drawn based on the obtained predicted scores, and the corresponding area under ROC curve (AUC), accuracy, sensitivity, and specificity were calculated and compared. Among the 479 pulmonary metastases, 379 had complete response (CR) ablation and 100 had incomplete response ablation. Three hundred eighty-six lesions were selected to construct a training dataset and 93 lesions to construct a testing dataset. The multivariate logistic regression analysis revealed cancer antigen 19-9 (CA19-9, p<0.001) and the location of the metastases (p< 0.05) as independent risk factors. Significant correlations were observed between complete ablation and 9 radiomics features. The best prediction performance was achieved with the proposed multimodal data fusion model integrating radiomic features and clinical variables with the highest accuracy (82.6%), AUC value (0.921), sensitivity (80.3%), and specificity (81.4%). This novel multimodal data fusion model was demonstrated efficient for immediate efficacy evaluation after RFA for CRC lung metastases, which could benefit necessary complementary treatment.

Dosimetric Parameters are Associated with Local Control after SBRT for Colorectal Cancer Lung Metastases

<h3>Purpose/Objective(s)</h3> Recent data suggest that higher stereotactic body radiotherapy (SBRT) prescription doses, and hence biologically effective doses (BED), are associated with improved local control (LC) for patients who receive SBRT for colorectal cancer (CRC) oligometastasis. However, the immense heterogeneity of SBRT dose distributions, even at isoeffective prescription doses, has made it challenging to extrapolate reported prescription dose thresholds into contemporary plan optimization. Our objective was to characterize dosimetric predictors of LC after SBRT for CRC lung metastases. We hypothesized that the internal target volume (ITV) and planning target volume (PTV) doses covering 90% of the volume (D90) and minimum doses (Dmin) would be associated with local control. <h3>Materials/Methods</h3> This was a retrospective cohort study including patients who received SBRT to CRC lung metastases at a single cancer center between 2013-2019. Dosimetric parameters including the D90 and Dmin of both the ITV and PTV were collected and converted to BED using the linear quadratic model assuming an alpha/beta of 10 Gy. The Kaplan-Meier method was used to estimate LC, and univariate analyses were performed with the Cox proportional hazards regression model. <h3>Results</h3> 53 CRC lung metastases amongst 28 patients were included for analysis. Median dose and number of fractions were 50 Gy (IQR 50-54 Gy) and 5 (IQR 3-5) fractions, respectively, with a median BED of 100 Gy (IQR 100-151.2 Gy). Median follow-up after SBRT was 27 months (IQR 17 – 38 months). By last follow-up, 18 of 53 (34%) metastases had progressed, of which only 4 (22%) were isolated sites of first-progression. The 2-year LC was 66.0% (95% CI 54.3% – 81.6%). On univariate analysis, lung metastasis size (HR 1.601 per cm, 95% CI 1.052 – 2.436) and BEDs including the ITV D90 (HR 0.83 per 10 Gy, 95%CI 0.75–0.93, p = 0.002) and Dmin (HR 0.821 per 10 Gy, 95% CI 0.73–0.93, p = 0.002), and PTV D90 (HR 0.79, 95%CI 0.68–0.93, p = 0.004) and Dmin (HR 0.80, 95% CI 0.66–0.97, p = 0.025) were associated with LC. When BED dose thresholds were chosen based on bottom quartile, improved 2-year LC was associated with ITV D90 BED ≥118 Gy (84.6% vs. 21.4% p < 0.0001) and Dmin BED ≥ 114 Gy (81.5% vs. 30.5% p < 0.0001). This remained true among the 29 lesions treated with the most common prescription dose of 50 Gy in 5 fractions (90.0% vs. 22.2%, p < 0.0001 and 83.5% vs. 37.5%, p = 0.011, respectively). <h3>Conclusion</h3> BEDs of over 118 Gy to at least 90% of the ITV volume (D90) and a minimum dose of 114 Gy were associated with improved LC after SBRT for CRC lung metastases. These data may be informative in-patient selection and plan optimization, particularly in challenging circumstances where the target is adjacent to critical organs at risk. With the favorable LC achieved with dose-escalated SBRT and the infrequency of isolated in-field first-progression events, SBRT remains an excellent local treatment option for CRC lung metastasis.

Spread through Air Spaces (STAS) in Solitary Pulmonary Metastases from Colorectal Cancer (CRC).

Spread through air spaces (STAS) is a recently described route of tumor invasion associated with poor prognosis in primary lung cancer. Aim of this study was to investigate the presence of STAS and to assess its prognostic significance in patients undergoing pulmonary metastasectomy (PM) for solitary metastases from colorectal cancer (CRC). All 49 CRC patients (30 male and 19 female, median age 66 years) who underwent PM between January 2008 and December 2015 were retrospectively analyzed. STAS was identified in 26.5% (n = 13) of resected specimens. Location of pulmonary lesions (central vs. peripheral) was assessed based on the available computed tomography imaging (n = 47, 96%). STAS was detected in all five patients with central metastases (100%) versus 7 of 42 (17%) with peripheral metastases (p = 0.0001). Locoregional recurrence occurred in STAS-positive patients (n = 4 of 13 vs. n = 0 of 36), all STAS-negative patients remained recurrence-free (p = 0.003). Median number of alveoli with STAS involvement was four (range from 2 to 9). There was statistically positive relationship between the number of alveoli invaded with STAS and locoregional recurrence of metastases (p = 0.0001). The presence of STAS is not a factor affecting the 5-year overall survival rate (p = 0.6651). We identified STAS as a frequent finding in resected CRC lung metastases and found insignificant association with outcome.

Impact of the Ablative Margin on Local Tumor Progression after Radiofrequency Ablation for Lung Metastases from Colorectal Carcinoma: Supplementary Analysis of a Phase II Trial (MLCSG-0802)

PurposeTo explore what extent of ablative margin depicted by computed tomography (CT) immediately after radiofrequency (RF) ablation is required to reduce local tumor progression (LTP) for colorectal cancer (CRC) lung metastases. Materials and MethodsThis retrospective study was undertaken as a supplementary analysis of a previous prospective trial. Seventy patients (49 men and 21 women; mean age ± standard deviation, 64.9 years ± 10.6 years) underwent RF ablation for CRC lung metastases, and 95 tumors that were treated in the trial and followed up with CT at least 12 months after RF ablation were evaluated. The mean tumor size was 1.0 cm ± 0.5 cm. The ablative margin was estimated as the shortest distance between the outer edge of the tumor and the surrounding ground-glass opacity on CT obtained immediately after RF ablation. The impact of the ablative margin on LTP was evaluated using logistic regression analysis. Multivariate logistic regression analysis was also performed to identify the risk factors for LTP. The result was validated with multivariate logistic regression applying a bootstrap method (1,000 times resampling). ResultsThe mean ablative margin was 2.7 mm ± 1.3 (range, 0.4–7.3 mm). LTP developed in 6 tumors (6%, 6/95) 6–19 months after RF ablation. The LTP rate was significantly higher when the margin was less than 2 mm (P = .023). A margin of <2 mm was also found to be a significant factor for LTP (P = .048) on multivariate analysis and validated using the bootstrap method (P = .025). ConclusionsAn ablative margin of at least 2 mm is important to reduce LTP after RF ablation for CRC lung metastases.

Identification of the effects of COVID-19 on patients with pulmonary fibrosis and lung cancer: a bioinformatics analysis and literature review.

Coronavirus disease 2019 (COVID-19) poses a serious threat to human health and life. The effective prevention and treatment of COVID-19 complications have become crucial to saving patients’ lives. During the phase of mass spread of the epidemic, a large number of patients with pulmonary fibrosis and lung cancers were inevitably infected with the SARS-CoV-2 virus. Lung cancers have the highest tumor morbidity and mortality rates worldwide, and pulmonary fibrosis itself is one of the complications of COVID-19. Idiopathic lung fibrosis (IPF) and various lung cancers (primary and metastatic) become risk factors for complications of COVID-19 and significantly increase mortality in patients. Therefore, we applied bioinformatics and systems biology approaches to identify molecular biomarkers and common pathways in COVID-19, IPF, colorectal cancer (CRC) lung metastasis, SCLC and NSCLC. We identified 79 DEGs between COVID-19, IPF, CRC lung metastasis, SCLC and NSCLC. Meanwhile, based on the transcriptome features of DSigDB and common DEGs, we identified 10 drug candidates. In this study, 79 DEGs are the common core genes of the 5 diseases. The 10 drugs were found to have positive effects in treating COVID-19 and lung cancer, potentially reducing the risk of pulmonary fibrosis.

Study on the Action Mechanism of the Yifei Jianpi Tongfu Formula in Treatment of Colorectal Cancer Lung Metastasis Based on Network Analysis, Molecular Docking, and Experimental Validation.

Objective The lung is the second most common site of colorectal cancer (CRC) metastasis. This study aims to investigate the therapeutic effects and potential action mechanisms of Yifei Jianpi Tongfu formula (YJTF) in CRC lung metastasis in a comprehensive and systematic way by network analysis, molecular docking, and experimental verification. Methods The main ingredients in YJTF were screened from the Traditional Chinese Medicine System Pharmacology Database and Analysis Platform (TCMSP) and Traditional Chinese Medicine Integrated Database (TCMID), and the disease-related targets from the Online Mendelian Inheritance in Man (OMIM) and GeneCards and the compound-related targets from SwissTargetPrediction were collected. Then, Metascape was used for pathway annotation and enrichment analysis, and meanwhile, a protein-protein interaction (PPI) network was constructed. Molecular docking was carried out to investigate interactions between the active compounds and the potential targets. The in vivo effect of YJTF on CRC lung metastasis was observed in a tail vein injection mouse model. Results A total of 243 active compounds and 81 disease-related targets of YJTF were selected for analysis. The results of multiple network analysis showed that the core targets of YJTF were enriched onto various cancer-related pathways, especially focal adhesion and adherens junction. The results of molecular docking demonstrated that all core compounds (quercetin, kaempferol, luteolin, apigenin, and isorhamnetin) were capable of binding with AKT1, EGFR, SRC, ESR1, and PTGS2. Experimental validation in vivo demonstrated that YJTF combined with oxaliplatin could significantly reduce the number of lung metastases and improve the quality of life in mice. Further research suggested that YJTF inhibited CRC lung metastasis probably by modulating epithelial-to-mesenchymal transition (EMT). Conclusions According to the analysis, YJTF can be considered as an effective adjuvant therapy for CRC lung metastasis.

Long-term outcome following microwave ablation of lung metastases from colorectal cancer.

PurposeTo retrospectively evaluate the safety and efficacy of percutaneous computed tomography (CT)-guided microwave ablation (MWA) in colorectal cancer (CRC) lung metastases, and to analyze prognostic factors.Materials and methodsData were collected from 31 patients with CRC lung metastases from May 2013 to September 2017. They had removed the CRC, no extrapulmonary metastases, no more than three metastases in the lung, the maximum diameter of the lesions was ≤3 cm, and all the lung metastases could be completely ablated. The ablation procedures were performed using a KY-2000 microwave multifunctional therapeutic apparatus. Efficacy is assessed two to four weeks after ablation, and follow-up are performed every three months for two years. The primary outcome was overall survival (OS). The secondary outcomes were progression-free survival (PFS), and complications. Cox regression analysis was used for the evaluation of the statistical significance of factors affecting the end result of MWA therapy. The Kaplan–Meier method was used for estimation of survival rates.ResultsA total of 45 metastatic lung lesions from CRC in 31 patients were treated with CT-guided MWA procedures. The median OS was 76 months. The one, two, three, and five-year survival rates were 93.5%, 80.6%, 61.3%, and 51.6%, respectively. Multivariate analysis showed that the primary tumor from the rectum (P = 0.009) and liver metastases at the diagnosis of lung metastases (P = 0.043) were risk factors affecting OS, while PFS was a protective factor. The median PFS was 13 months. The maximum diameter of lung metastases lesions (P = 0.004) was a risk factor. The interval between pulmonary metastases and MWA (P=0.031) was the protective factor. Pneumothorax was observed in 13 out of 36 procedures. Four patients developed pneumothorax requiring drainage tube insertion. No patient deaths occurred within 30 days of ablation. Three out of 31 patients (9.67%) were found to have local recurrence of the original lung metastatic ablation foci.ConclusionMWA therapy may be safely and effectively used as a therapeutic tool for the treatment of selected CRC pulmonary metastases, and the prognosis is better in patients without liver metastases at the diagnosis of lung metastases.

Berberine inhibited the formation of metastasis by intervening the secondary homing of colorectal cancer cells in the blood circulation to the lung and liver through HEY2

BackgroundMetastasis is the leading cause of death in patients with colorectal cancer (CRC). The 5-year survival rate of CRC patients in whom the cancer has spread to distant sites is 13.5%. The most common sites of CRC metastasis are liver and lung. The principal therapies for CRC metastatic disease are surgery, but its benefits are limited. PurposeThis study aimed to reveal the regulatory mechanism of berberine on secondary homing of CRC cells to form metastatic focus. This was more valuable than the previous direct study of the migration and metastasis characteristics of CRC cells. MethodsIn this study, we used the functional enrichment analysis of differentially expressed genes after berberine treatment and investigated co-expression modules related with CRC metastasis by WGCNA. PPI and survival analyses of significant modules were also conducted. The biological functions of berberine in CRC lung and liver metastasis were investigated by a series of in vitro and in vivo experiments: MTT, colony formation and mouse tail vein injection. And we scanned through the entire extracellular domain of HEY2 protein for autodocking analysis with berberine. ResultsWe found the differentially expressed genes (DEGs) after berberine treatment were related with cancer progression and metastasis related pathways. Through WGCNA analysis, four cancer progression and metastasis related modules were detected. After PPI and survival analysis, we identified and validated HEY2 as a hub gene, high expression and poor survival at the metastatic stage. Functionally, berberine inhibited the survival, invasion and migration of CRC cells in vitro and in vivo. Mechanistically, berberine treatment down-regulated the expression of HEY2, metastasis related protein E-cadherin, β-catenin and Cyclin D1 during Mesenchymal epithelial transformation (MET). Berberine and HEY2 showed a significant interaction, and berberine binded to HEY2 protein at the residue HIS-99 interface with a hydrogen-bond distance of 1.9A. ConclusionsWe revealed that berberine could significantly inhibit the expression of hub gene HEY2 and metastasis related proteins E-cadherin and β-catenin and Cyclin D1 during MET in CRC lung and liver metastases. In total, HEY2 was a promising candidate biomarker for prognosis and molecular characteristics in CRC metastasis.

A clinical AI-driven multiplex immunofluorescence imaging pipeline to characterize tumor microenvironment heterogeneity.

3020 Background: Understanding the underlying heterogeneity of the tumor microenvironment (TME) on a single-cell level is becoming increasingly important to predict a patient’s response to immunotherapy. Conventional imaging methods can help reveal tissue heterogeneity, but are not optimal for identifying multiple cellular subpopulations or cellular interactions from a single slide image, limiting their use in clinical settings. Here, we present a clinical artificial intelligence (AI)-driven multiplex immunofluorescence (MxIF) imaging pipeline based on novel cell segmentation and cell typing methods to evaluate tumor cellular heterogeneity, immune cell composition, and cell-to-cell interactions. Methods: A machine learning (ML)-based cell segmentation algorithm was trained on a manually annotated dataset created from 219 different regions of interest (ROIs) that contained 85,991 cells from various tissues (colon, kidney, lung, lymph node, tonsil, and ureter). A dataset containing 58,676 cells from 146 ROIs was used for validation and accuracy was determined between automated and manually annotated images; accuracy was further evaluated by calculating the f1-score using available methods (DeepCell and Stardist). Marker stains with a low signal-to-noise ratio were automatically enhanced, allowing for adequate cell-to-cell interaction analysis. Results: An automated MxIF image processing workflow was developed. Validation of the trained cell segmentation model showed high accuracy (0.80 f1-score), demonstrating superior performance compared to other methods (DeepCell and Stardist - 0.55 and 0.78 f1-score, respectively). The pathologist-determined accuracy (0.84 mean f1-score) indicated a near-human performance of the developed method. Normalized expression values obtained from the cell typing model allowed automated cell recognition. We analyzed cellular heterogeneity across 3 regions of colorectal cancer (CRC), gastric cancer (GC), and non-small cell lung cancer (NSCLC) samples. While proportions of immune cells varied, proportions of malignant epithelial cells were stable across all regions of each sample, as concordant percentages of Ki67+ cells were identified (CRC-19%; GC-21%; NSCLC-5%). Analysis of cell-to-cell interactions and immune communities identified tumor-, immune-, and stromal-enriched communities in all tumor samples that were stable across regions. Conclusions: By analyzing complex tumor tissue at single-cell resolution with high accuracy, this AI-driven MxIF imaging technology is able to characterize tumor and microenvironment heterogeneity across cancer types. This novel AI-based tool is currently being integrated into several ongoing prospective clinical studies to aid in the development of predictive and prognostic biomarkers.