Colorectal Tumors Research Articles

Photothermal Responsive Microcarriers Encapsulated With Cangrelor and 5-Fu for Colorectal Cancer Treatment.

Localized chemotherapy is emerging as a potential strategy for cancer treatment due to its low systemic toxicity. However, the immune evasion of tumor cells and the lack of an intelligent design of the delivery system limit its clinical application. Herein, photothermal responsive microcarriers are designed by microfluidic electrospray for colorectal tumor treatment. The microcarriers loaded with Cangrelor, 5-FU and MXene (G-M@F/C+NIR) show sustained delivery of antiplatelet drug Cangrelor, thus inhibiting the activity of platelets, interactions of platelet-tumor cell, as well as the tumor cells invasion and epithelial-mesenchymal transition (EMT). In addition, the sustained delivery of chemotherapeutics 5-FU and the photothermal effect provided by MXene enable the microcarriers to inhibit tumor cells proliferation and migration. In vivo studies validate that the G-M@F/C+NIR microcarriers significantly inhibites tumor growth, decreased the expression of Ki-67 in tumor cells and vascular endothelial growth factor (VEGF) in the tumor microenvironment, while increased the expression of E-cadherin. It is believe that by means of the proposed photothermal responsive microcarriers, the synergistic strategy of platelet inhibition, chemotherapy, and photothermal therapy can find practical applications in cancer treatment.

Efficacy of endoscopic submucosal dissection assisted by metal-clip pocket creation traction for colorectal tumors

To evaluate the clinical efficacy of endoscopic submucosal dissection (ESD) assisted by metal-clip pocketcreation traction for treatment of colorectal tumors. We retrospectively analyzed the clinical data of 244 patients with colorectal tumors undergoing colorectal ESD treatment between January, 2019 and December, 2022, including 169 patients receiving ESD without metal-clip pocket-creation traction (N-ESD group) and 75 with traction-assisted ESD (M-ESD group). Propensity score matching was used to screen the patients using general clinical data as the covariates for matching. Operative time, surgical resection outcome indicators, incidence of adverse events, and histopathological diagnosis indicators were compared between the two groups of patients after matching. The median operative time was significantly shorter in M-ESD group than in N-ESD group (20.0 [15.0, 30.0] vs 30 [20.0, 45.0] min, P=0.008). No significant difference was found in the en bloc resection rate (100% vs 98.6%), complete resection rate (97.3% vs 96%) and radical resection rate (97.3% vs 96%) between the two groups (P>0.05). The incidence of adverse events was low in both groups and showed no significant difference between them (P>0.05). In patients with colorectal tumors, the use of metal-clip pocket-creation traction can shorten the operative time of ESD although it does not significantly reduce the surgical resection rate or incidence of adverse events.

Improved visibility of colorectal tumor by texture and color enhancement imaging with indigo carmine.

Accurate diagnosis and early resection of colorectal polyps are important to prevent the occurrence of colorectal cancer. However, technical factors and morphological factors of polyps itself can lead to missed diagnoses. Image-enhanced endoscopy and chromoendoscopy (CE) have been developed to facilitate an accurate diagnosis. There have been no reports on visibility using a combination of texture and color enhancement imaging (TXI) and CE for colorectal tumors. To investigate the visibility of margins and surfaces with the combination of TXI and CE for colorectal lesions. This retrospective study included patients who underwent lower gastrointestinal endoscopy at the Toyoshima Endoscopy Clinic. We extracted polyps that were resected and diagnosed as adenomas or serrated polyps (hyperplastic polyps and sessile serrated lesions) from our endoscopic database. An expert endoscopist performed the lower gastrointestinal endoscopies and observed the lesion using white light imaging (WLI), TXI, CE, and TXI + CE modalities. Indigo carmine dye was used for CE. Three expert endoscopists rated the visibility of the margin and surface patterns in four ranks, from 1 to 4. The primary outcomes were the average visibility scores for the margin and surface patterns based on the WLI, TXI, CE, and TXI + CE observations. Visibility scores between the four modalities were compared by the Kruskal-Wallis and Dunn tests. A total of 48 patients with 81 polyps were assessed. The histological subtypes included 50 tubular adenomas, 16 hyperplastic polyps, and 15 sessile serrated lesions. The visibility scores for the margins based on WLI, TXI, CE, and TXI + CE were 2.44 ± 0.93, 2.90 ± 0.93, 3.37 ± 0.74, and 3.75 ± 0.49, respectively. The visibility scores for the surface based on WLI, TXI, CE, and TXI + CE were 2.25 ± 0.80, 2.84 ± 0.84, 3.12 ± 0.72, and 3.51 ± 0.60, respectively. The visibility scores for the detection and surface on TXI were significantly lower than that on CE but higher than that on WLI (P < 0.001). The visibility scores for the margin and surface on TXI + CE were significantly higher than those on CE (P < 0.001). In the sub-analysis of adenomas, the visibility for the margin and surface on TXI + CE was significantly better than that on WLI, TXI, and CE (P < 0.001). In the sub-analysis of serrated polyps, the visibility for the margin and surface on TXI + CE was also significantly better than that on WLI, TXI, and CE (P < 0.001). TXI + CE enhanced the visibility of the margin and surface compared to WLI, TXI, and CE for colorectal lesions.

Demographic characteristics and survival in young onset colorectal neuroendocrine tumors

Demographic characteristics and survival in young onset colorectal neuroendocrine tumors

Colorectal Cancer: Is it Still a Disease of the Elderly?

&lt;b&gt;&lt;br&gt;Introduction:&lt;/b&gt; Colorectal cancer is becoming an increasingly significant health issue, being one of the more commonly diagnosed malignancies. Colorectal tumors account for 10% of all malignant cancers in women and 12% in men. Incidence is higher in the male population, especially among younger individuals. It is commonly believed that colorectal cancer is predominantly associated with advanced age. However, colorectal surgeons, who specialize in the treatment of this type of cancer, are observing a growing number of cases among middle-aged and younger individuals.&lt;/br&gt; &lt;b&gt;&lt;br&gt;Aim:&lt;/b&gt; The aim of our study was to investigate whether colorectal cancer still predominantly affects elderly individuals, how frequently it is diagnosed in younger patients, and whether the location of tumors in the intestines of younger patients aligns with data from elderly individuals.&lt;/br&gt; &lt;b&gt;&lt;br&gt;Materials and methods:&lt;/b&gt; The study was conducted retrospectively and included a cohort of 1771 patients who underwent surgical procedures due to colorectal cancer between 2012 and 2015 at the Department of General and Colorectal Surgery at the Medical University of Łódź and between 2014 and 2017 at the Department of General Surgery with a Division of Surgical Oncology at the District Health Center in Brzeziny. Data were analyzed regarding the frequency of colorectal cancer occurrence by age, tumor location in different age groups, and disease stage according to age. Age groups included &lt;40 years, 41-50 years, 51-70 years, and &gt;70 years.&lt;/br&gt; &lt;b&gt;&lt;br&gt;Results:&lt;/b&gt; The study encompassed a total of 1771 patients, with 988 (55.79%) being males and 783 (44.21%) females. The mean age of the patients was 65.27 11.12 years. The highest number of cases was observed in the age range of 60-70 years and 70-80 years. It was found that colorectal tumors in males more frequently occurred on the left side of the colon and rectum, while in females, they were more commonly located on the right side of the colon, which was statistically significant (P = 0.007). Younger age groups of patients (&lt;40 years, 40-50 years) had a similar male-to-female ratio, whereas in age groups above 50 years, males significantly outnumbered females (P = 0.049). The study revealed that in the group of patients below 40 years of age, an advanced stage of colorectal cancer was significantly more common; stage D occurred over twice as often as in the 51-70 age group and over three times as often as in the &gt;70 age group.&lt;/br&gt; &lt;b&gt;&lt;br&gt;Conclusions:&lt;/b&gt; The incidence of colorectal cancer in Poland is steadily increasing, with a growing number of diagnoses in younger individuals. Research findings demonstrate that males, especially those in younger age groups, are at a higher risk of developing colorectal cancer. A higher disease stage is more frequently observed in younger patients, possibly due to delayed diagnosis and symptomatic treatment. Screening programs should be adjusted to the changing age groups at higher risk. Our study underlines the need to raise public awareness regarding colorectal cancer, particularly among the younger population.&lt;/br&gt.

Cationic micelle delivery of a multi-epitope vaccine candidate derived from tumor-associated antigens, causing regression in established CT26 colorectal tumors in mice.

Among all the cancers, colorectal cancer (CRC) has the third mortality rank in both genders. Cancer vaccines have shown promising results in boosting patients' immune systems to fight cancer. Using the IEDB database, we predicted mouse MHC-I (H2-Ld) binding epitopes from four tumor-associated antigens (APC, KRAS, TP53, and PIK3CA) and designed a multi-epitope vaccine. We expressed the candidate vaccine and encapsulated it into the cationic micelle with polyethyleneimine conjugated to oleic acid as its building blocks. We studied tumor inhibition effect, cytokine production, and lymphocyte proliferation in the mouse CRC model after vaccination. Our finding illustrated significant tumor growth inhibition in mouse models treated with the candidate nanovaccine. Besides the significant release of IFN-γ and IL-4 by immunized mouse spleen T-lymphocytes, T-cell proliferation assay results confirmed effective immune response after the vaccination. These results demonstrate the potential therapeutic effects of nanovaccines and could be a possible approach to CRC immunotherapy.

Molecular and immunohistochemical study of APC exon 16 and its possible role in colorectal carcinoma development

BackgroundColorectal cancer ranks second as a cause of cancer deaths. Mutations in the adenomatous polyposis coli (APC) gene, especially in exon 16, could contribute to colorectal carcinoma development. This study explored the correlations between APC gene exon 16 variations/expression and colorectal carcinoma progression. MethodsIn a case-control study, blood samples from 150 colorectal carcinoma patients and 50 healthy volunteers were analyzed by PCR and sequencing for APC exon 16 variations. The APC protein expression on tissue samples was evaluated by immunohistochemistry and statistical analyses were used to examine clinicopathological correlations. ResultsThe sequencing analysis revealed a mutation in exon 16 of the APC gene (rs459552) in 36 % of colorectal cancer cases while absent in all non-cancer controls. Subgroup analysis by tumor grade showed higher prevalence of mutant allele in Grade II and Grade III cases, with frequencies reaching 60.0 % and 69.2 %, respectively, compared to a substantially lower prevalence of 29.4 % in Grade I patients. Immunohistochemistry showed no significant correlation between this mutation and APC expression. APC positivity proportions were 25.5 % in Grade I tumors (n = 26/102) versus 17.1 % in Grade II (n = 6/35) and 46.2 % in Grade III (n = 6/13), showing a non-significant trend of reduced positivity in higher grade tumors (p>0.05). ConclusionsThe frequency of APC exon 16 mutation (rs459552) rose significantly with increasing tumor grade. Similarly, although not statistically significant, the percentage of APC positive staining increased with poorer tumor differentiation, rather than declining. Therefore, the APC exon 16 mutation and expression analysis provides insights into colorectal cancer progression, with the rs459552 mutation correlating with grade and may promoting aggression.

CSRP1 expression is associated with a mesenchymal, stroma-rich tumor profile and poor prognosis in colon cancer.

Cysteine and glycine-rich protein 1 (CSRP1) is involved in the cysteine-rich protein family and is a marker of smooth muscle lineages. In colon cancer, the expression of this gene is associated with poor prognosis. In this study, the aim was to reevaluate its prognostic relationship in independent cohorts and explore potential underlying biological processes that are linked to aggressive behavior in tumors with high CSRP1 expression, such as epithelial-to-mesenchymal transition (EMT), stromal fractions in the tumor microenvironment, and consensus molecular subtypes (CMSs). RNA sequencing (RNAseq)-, microarray-, and single-cell RNAseq (scRNAseq)-based transcriptomic data were obtained from public databases. The EMT score was calculated based on the expression of E-cadherin and vimentin genes using a previously published method. The stromal score generated by the ESTIMATE method was utilized for the analysis of correlation with the CSRP1 expression. The scRNAseq data were analyzed via the Seurat R package. The immunohistochemistry-based protein level expression of CSRP1 was evaluated using the Human Protein Atlas database. Lower CSRP1 expression was noted in colon tumors compared to normal colon tissue. Patients with a high CSRP1 expression had shorter recurrence-free, overall, and disease-specific survivals in the GSE39582 and GSE17536 datasets (p < 0.05). The methylation level of the CSRP1 gene was negatively correlated (r = -0.57, p < 0.0001) with CSRP1 expression in The Cancer Genome Atlas colon adenocarcinoma dataset. CSRP1 expression was positively correlated with the expression of mesenchymal markers, EMT score, and stromal score obtained via the ESTIMATE method. CMS4 colon tumors had a significantly higher CSRP1 expression compared to other CMSs. Analysis of the scRNAseq data revealed that CSRP1 was expressed by epithelial cells and cancer-associated fibroblasts in the colorectal tumor microenvironment, which was also confirmed by the protein expression data from the Human Protein Atlas database. CSRP1 expression is associated with CMS4, a more mesenchymal stroma-rich molecular profile, and poor prognosis in colon cancer.

Familial uveal melanoma and other tumors in 25 families with monoallelic germline MBD4 variants.

Monoallelic germline MBD4 pathogenic variants were recently reported to cause a predisposition to uveal melanoma, associated with a specific tumor mutational signature and good response to immunotherapy. Monoallelic tumor pathogenic variants have also been described in brain tumors, breast cancers, and myxofibrosarcomas, whereas biallelic germline MBD4 pathogenic variants have been involved in a recessive hereditary adenomatous polyposis and a specific type of acute myeloid leukemia. We analyzed MBD4 for all patients with a diagnosis of uveal melanoma at Institut Curie since July 2021 and in the 3240 consecutive female probands explored at the Institut Curie for suspicion of predisposition to breast cancer between July 2021 and February 2023. We describe 25 families whose probands carry a monoallelic germline pathogenic variant in MBD4. Eighteen of these families presented with uveal melanoma (including a case patient with multiple uveal melanoma), and 7 families presented with breast cancer. Family histories showed the first familial case of uveal melanoma in monoallelic MBD4 pathogenic variant carriers and other various types of cancers in relatives, especially breast, renal, and colorectal tumors. Monoallelic MBD4 pathogenic variant may explain some cases of familial and multiple uveal melanoma as well as various cancer types, expanding the tumor spectrum of this predisposition. Further genetic testing in relatives combined with molecular tumor analyses will help define the tumor spectrum and estimate each tumor's risk.

Interferon gamma immunoPET imaging to evaluate response to immune checkpoint inhibitors.

We previously developed a 89Zr-labeled antibody-based immuno-positron emission tomography (immunoPET) tracer targeting interferon gamma (IFNγ), a cytokine produced predominantly by activated T and natural killer (NK) cells during pathogen clearance, anti-tumor immunity, and various inflammatory and autoimmune conditions. The current study investigated [89Zr]Zr-DFO-anti-IFNγ PET as a method to monitor response to immune checkpoint inhibitors (ICIs). BALB/c mice bearing CT26 colorectal tumors were treated with combined ICI (anti-cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and anti-programmed death 1 (PD-1)). The [89Zr]Zr-DFO-anti-IFNγ PET tracer, generated with antibody clone AN18, was administered on the day of the second ICI treatment, with PET imaging 72 hours later. Tumor mRNA was analyzed by quantitative reverse-transcribed PCR (qRT-PCR). We detected significantly higher intratumoral localization of [89Zr]Zr-DFO-anti-IFNγ in ICI-treated mice compared to untreated controls, while uptake of an isotype control tracer remained similar between treated and untreated mice. Interestingly, [89Zr]Zr-DFO-anti-IFNγ uptake was also elevated relative to the isotype control in untreated mice, suggesting that the IFNγ-specific tracer might be able to detect underlying immune activity in situ in this immunogenic model. In an efficacy experiment, a significant inverse correlation between tracer uptake and tumor burden was also observed. Because antibodies to cytokines often exhibit neutralizing effects which might alter cellular communication within the tumor microenvironment, we also evaluated the impact of AN18 on downstream IFNγ signaling and ICI outcomes. Tumor transcript analysis using interferon regulatory factor 1 (IRF1) expression as a readout of IFNγ signaling suggested there may be a marginal disruption of this pathway. However, compared to a 250 µg dose known to neutralize IFNγ, which diminished ICI efficacy, a tracer-equivalent 50 µg dose did not reduce ICI response rates. These results support the use of IFNγ PET as a method to monitor immune activity in situ after ICI, which may also extend to additional T cell-activating immunotherapies.

Integrating scRNA-seq and bulk RNA-seq to characterize infiltrating cells in the colorectal cancer tumor microenvironment and construct molecular risk models.

Colorectal cancer (CRC) is a malignancy that is both highly lethal and heterogeneous. Although the correlation between intra-tumoral genetic and functional heterogeneity and cancer clinical prognosis is well-established, the underlying mechanism in CRC remains inadequately understood. Utilizing scRNA-seq data from GEO database, we re-isolated distinct subsets of cells, constructed a CRC tumor-related cell differentiation trajectory, and conducted cell-cell communication analysis to investigate potential interactions across cell clusters. A prognostic model was built by integrating scRNA-seq results with TCGA bulk RNA-seq data through univariate, LASSO, and multivariate Cox regression analyses. Eleven distinct cell types were identified, with Epithelial cells, Fibroblasts, and Mast cells exhibiting significant differences between CRC and healthy controls. T cells were observed to engage in extensive interactions with other cell types. Utilizing the 741 signature genes, prognostic risk score model was constructed. Patients with high-risk scores exhibited a significant correlation with unfavorable survival outcomes, high-stage tumors, metastasis, and low responsiveness to chemotherapy. The model demonstrated a strong predictive performance across five validation cohorts. Our investigation involved an analysis of the cellular composition and interactions of infiltrates within the microenvironment, and we developed a prognostic model. This model provides valuable insights into the prognosis and therapeutic evaluation of CRC.

Sequential Delivery of Aptamer and STING Agonist Shuttled by Graphene Oxide

Stimulator of interferon genes (STING) is a prospective target spot of immunological therapy. The development of STING is still in the preliminary stage and full of challenges. such as lack of tumor specificity, and high-dose-induced T-cell apoptosis. Herein, the STING agonists were selectively delivered and released in the tumor through an aptamer (Apt) modified graphene oxide (GO). The STING agonist was released controllably from GO on account of the acidic tumor microenvironment, and then binds specifically to the STING protein, aiming to active STING pathway and stimulate immunosuppressive factors secretion via phosphorylating TBK1 and IFN IRF3, and secreta high-level IL-6. The resulting immunotherapy exhibited significant inhibition of colorectal tumors, and the producing stimulus was sufficient to activate plentiful enough T-cells with adaptability to inhibit the growth and metastasis of tumors. This work helps to promote sustaining study on the targeted delivery of STING agonists and enhanced activation of the STING pathway, and the translation of novel understanding to clinical applications will enhance the efficiency of immunotherapy for cancer with STING agonists.

Engineering Clinically Relevant Probiotics with Switchable "Nano-Promoter" and "Nano-Effector" for Precision Tumor Therapy.

Probiotics have the potential as biotherapeutic agents for cancer management in preclinical models and human trials by secreting antineoplastic or immunoregulatory agents in the tumor microenvironment (TME). However, current probiotics lack the ability to dynamically respond to unique TME characteristics, leading to limited therapeutic accuracy and efficacy. Although progress has been made in customizing controllable probiotics through synthetic biology, the engineering process is complex and the predictability of production is relatively low. To address this, here, for the first time, this work adopts pH-dependent peroxidase-like (POD-like) artificial enzymes as both an inducible "nano-promoter" and "nano-effector" to engineer clinically relevant probiotics to achieve switchable control of probiotic therapy. The nanozyme initially serves as an inducible "nano-promoter," generating trace amounts of nonlethal reactive oxygen species (ROS) stress to upregulate acidic metabolites in probiotics. Once metabolites acidify the TME to a threshold, the nanozyme switches to a "nano-effector," producing a great deal of lethal ROS to fight cancer. This approach shows promise in subcutaneous, orthotopic, and colitis-associated colorectal cancer tumors, offering a new methodology for modulating probiotic metabolism in a pathological environment.

Tumor Segmentation in Colorectal Ultrasound Images Using an Ensemble Transfer Learning Model: Towards Intra-Operative Margin Assessment

Tumor boundary identification during colorectal cancer surgery can be challenging, and incomplete tumor removal occurs in approximately 10% of the patients operated for advanced rectal cancer. In this paper, a deep learning framework for automatic tumor segmentation in colorectal ultrasound images was developed, to provide real-time guidance on resection margins using intra-operative ultrasound. A colorectal ultrasound dataset was acquired consisting of 179 images from 74 patients, with ground truth tumor annotations based on histopathology results. To address data scarcity, transfer learning techniques were used to optimize models pre-trained on breast ultrasound data for colorectal ultrasound data. A new custom gradient-based loss function (GWDice) was developed, which emphasizes the clinically relevant top margin of the tumor while training the networks. Lastly, ensemble learning methods were applied to combine tumor segmentation predictions of multiple individual models and further improve the overall tumor segmentation performance. Transfer learning outperformed training from scratch, with an average Dice coefficient over all individual networks of 0.78 compared to 0.68. The new GWDice loss function clearly decreased the average tumor margin prediction error from 1.08 mm to 0.92 mm, without compromising the segmentation of the overall tumor contour. Ensemble learning further improved the Dice coefficient to 0.84 and the tumor margin prediction error to 0.67 mm. Using transfer and ensemble learning strategies, good tumor segmentation performance was achieved despite the relatively small dataset. The developed US segmentation model may contribute to more accurate colorectal tumor resections by providing real-time intra-operative feedback on tumor margins.

Correlations between MSH2 and MSH6 Concentrations in Different Biological Fluids and Clinicopathological Features in Colorectal Adenocarcinoma Patients and Their Contribution to Fast and Early Diagnosis of Colorectal Adenocarcinoma.

(1) Background: The human MutS homolog, hMSH2, is known to be involved in DNA mismatch repair and is responsible for maintaining the stability of the genome. When DNA damage occurs, MSH2 promotes cell apoptosis via the regulation of ATR/Chk2/p53 signal transduction, and MSH2 deficiency is also related to accelerated telomere shortening in humans. MSH2 missense mutations are involved in a defective DNA reparation process, and it can be implied in carcinogenesis, as it is already involved in well-known cancer-related syndromes such as Lynch syndrome. Human MSH6, which stands for mutS homolog 6, is a member of the MMR family that is responsible for the repair of post-replicative mismatched DNA bases. It is also one of the proteins with gene mutations that are associated with a high risk of developing Lynch syndrome, leading to a large series of tumors. (2) Methods: Patients and their clinical and pathological features were selected from the database of the project GRAPHSENSGASTROINTES and used accordingly, with ethics committee approval no. 32647/2018 awarded by the County Emergency Hospital from Targu-Mures. Analyses were conducted on whole blood, saliva, urine, and tumoral tissue samples using a stochastic method with stochastic microsensors. (3) Results: The results obtained using stochastic sensors were correlated with a series of macroscopic and microscopic pathological features for each sample type. Criteria or relationships were established for tumor location, vascular and perineural invasions, lymph node metastases, the presence of tumor deposits, and the presence of a mucus compound in the tumor mass. (4) Conclusions: The correlation between the concentrations of MSH2 in the four types of samples and the pathological features allowed for the fast characterization of a tumor, which can help surgeons and oncologists choose personalized treatments. Also, the colorectal tumor location was correlated with the concentration of MSH2 in whole blood, urine, and saliva. MSH6, which stands for mutS homolog 6, is not only useful in immunohistochemistry but in pathology practice as well. In this paper, the relationships between MSH6 levels in four biological fluids-whole blood, saliva, urine, and tissues-and tumor locations among the colorectal area, gross features, presence of a mucinous compound, molecular subtype, stroma features, and vascular invasions are presented.

Prevalence of PIK3CA Mutation Based on Tumour Locations Among Colorectal Cancer Patients in Bali

Colorectal tumour location is important in determining therapeutic strategies because the location of the tumour will affect the histological and molecular characteristics of the cancer. Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) is a gene known to be associated with colorectal cancer location. The prevalence of PIK3CA mutations in colorectal cancer (CRC) ranges from 10-20%. Although the prevalence is small, this gene has a potential as a predictive and prognostic biomarker in the management of CRC. 80% of PIK3CA mutations occur in exon 9 (codon 542 and 545) and exon 20 (codon 1047). This study aimed to determine the prevalence of PIK3CA gene mutations based on tumour location among colorectal cancer patients in Bali, especially in Prof. dr. I G.N.G. Ngoerah Hospital, Denpasar, Bali. DNA was isolated from formalin-fixed-paraffin embedded (FFPE) blocks of colorectal cancer samples which were stored in the Department of Pathology of Prof. dr. I G.N.G. Ngoerah Hospital. Mutation was detected using polymerase chain reaction (PCR) and direct sequencing. Data on age, gender, and tumour location were recorded from patient medical records. Our findings on 31 colorectal cancer samples showed 26 samples (96.3%) were found with heterozygous mutation in exon 9 with AC genotype, but no mutation was found on exon 20. Based on tumor location, among 26 samples with mutation, 16 samples (61.5%) were left colorectal cancer and 10 samples (38.5%) were right colorectal cancer. Further studies are needed to identify the association of this mutation with colorectal cancer location and the other clinicopathological aspects of CRC. Keywords: Colorectal cancer, PIK3CA gene mutation, PIK3CA exon 9, PIK3CA exon 20, colorectal cancer location, left-sided colorectal cancer, right-sided colorectal cancer

Abstract A146: FPI-2068: A novel anti-EGFR/cMET, alpha-particle emitting, radioimmunoconjugate for cancer therapy

Abstract Introduction: FPI-2068, a targeted alpha therapeutic (TAT), consists of a humanized EGFR and cMET targeting bispecific antibody (FPI-2053) radiolabeled with actinium-225 [225Ac]. FPI-2068 leverages the EGFR/cMET co-expression in cancer (high vs. low in normal tissues) and internalizing nature of the receptors to deliver radionuclides into tumor cells. The primary tumoricidal mechanism of FPI-2068 is double-strand DNA breaks (DSB) induced by alpha radiation, which lead to cell death. Methods: FPI-2068 consists of FPI-2053 conjugated with a DOTA-chelate and radiolabeled with [225Ac]. FPI-2071 is the lutetium-177 analogue. The binding and internalization of FPI-2071 was studied in vitro using cancer cell lines (HT29 colorectal and H292, H441, H1975, HCC827 lung cancers). In vivo studies were performed in corresponding xenograft models. For biodistribution studies FPI-2071 was injected intravenously into mice and tumor/organ radioactivity was quantified ex vivo at selected timepoints. For therapeutic studies, a single dose of FPI-2068 was administered to groups of mice over a range from 92.5 to 740 kBq/kg. Explanted HT29 and H441 tumors from FPI-2068 treated mice were analyzed by immunoblotting to detect activation of DDR and cell death pathways. Results: FPI-2071 demonstrated in vitro binding to all cell lines tested, followed by internalization and retention of the radioisotope into target cells. The biodistribution profile of FPI-2071 showed low normal organ uptake in all xenograft models tested, while tumor uptake varied. The peak tumor uptake in each model was: H292 (~73 %ID/g), H441 (~38 %ID/g), HT29 (~30 %ID/g), HCC827 (~25 %ID/g) and H1975 (20 %ID/g). Delivery of FPI-2068 to target xenografts resulted in anti-tumor efficacy; treatment was well-tolerated by animals. Sustained tumor regression (&amp;gt;28 days) was generally observed at doses of 370 kBq/kg and 740 kBq/kg across models. Lower doses were tumor growth suppressive. FPI-2068 treatment resulted in a dose-dependent increase in pATM and pRad50 expression indicating activation of DDR pathways in HT29 and H441 xenografts. Formation of DSB (pH2AX) and the induction of apoptosis (cleaved caspase 3) was also observed, consistent with the proposed primary mechanism of action. Conclusions: FPI-2068 demonstrated its anti-tumor efficacy in colorectal and lung tumor xenograft mouse models. Single-dose administration of FPI-2068 led to prolonged tumor regression. FPI-2068 caused activation of the DDR pathway as well as apoptosis, suggesting an inability of the cellular machinery to repair the DNA damage induced by the alpha radiation. These data support initiation of clinical trials in patients with solid tumors. Citation Format: Sadaf Aghevlian, Sean Collens, Julie Metcalf, Natalie Grinshtein, John Forbes, Frank Comer, Darshan Dalal, Puja Sapra, Eric Burak. FPI-2068: A novel anti-EGFR/cMET, alpha-particle emitting, radioimmunoconjugate for cancer therapy [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A146.

Abstract A093: Deep Cyclic Inhibition of the MAPK pathway with IMM-6-415, alone and in combination with encorafenib, demonstrates anti-tumor activity and tolerability in RAF mutant tumors in vivo

Abstract Introduction: A significant proportion of cancer patients have tumors addicted to uncontrolled MAPK signaling. Activating mutations in RAS or RAF are often directly responsible and have been observed in over 20% of human tumors1. Because MEK is downstream of RAS and RAF, it is an appealing drug target. However, MEK inhibitors have historically suffered from poor clinical durability, high toxicity and a susceptibility to pathway reactivation that has limited monotherapy activity in the RAS mutant setting. Unlike other MEK inhibitors, IMM-1-104 [NCT05585320] and IMM-6-415 are designed with distinctive features including both (1.) a unique target engagement mechanism that helps resist MAPK pathway reactivation and (2.) a pharmacokinetic (PK) profile that enables fast cadence deep cyclic inhibition (DCI). DCI drives pulsatile targeted inhibition that deprives tumor cells of a critical oncogenic pathway while limiting drug-related toxicities by affording normal cells an adequate PK recovery window between drug doses. To our knowledge, IMM-6-415’s preclinical activity is driven by the shortest drug plasma half-life (0.3-hours in mice) of any MEK inhibitor developed to date. Experimental Procedures: IMM-6-415 has already demonstrated promising activity in RAS-mutant xenograft tumor models (SITC 2022). Here, the antitumor activity of IMM-6-415 was evaluated in over 60 humanized 3D tumor growth assays (3D-TGA), which included 30 BRAF class I-mutant tumor models. Additionally, multiple drug-drug combinations have been explored, including vertical drug combinations with BRAF inhibitors. IMM-6-415, binimetinib and encorafenib were tested head-to-head as single agents and in combination with encorafenib in BRAFV600E melanoma and colorectal subcutaneous tumor xenograft models in female BALB/c nude mice. Summary of New Data: As monotherapy, IMM-6-415 demonstrated antitumor activity in over 50% (34 of 66) of the 3D-TGA models tested, including 30 BRAF-mutant preclinical models in which 19 (63%) showed activity. Therefore, we further explored both monotherapy and combination activity of IMM-6-415 in the A-375 (melanoma) and HT-29 (colorectal) BRAF V600E tumor xenograft models. Monotherapy treatment with encorafenib or IMM-6-415 displayed superior tumor growth inhibition (TGI) when compared to binimetinib. Furthermore, the combination of IMM-6-415 plus encorafenib prompted greater TGI with superior durability of response when tested head-to-head against the combination of binimetinib plus encorafenib in vivo at human equivalent doses for registered drugs. Conclusions: IMM-6-415 demonstrated promising activity and tolerability in preclinical models alone and in combination with encorafenib. In combination with encorafenib, IMM-6-415 achieved a greater TGI in vivo than the combination of encorafenib plus binimetinib in BRAFV600E colorectal cancer and melanoma tumor models, suggesting an opportunity for IMM-6-415 as monotherapy or in combination in BRAF mutant tumors. Citation Format: Anna Travesa, Mai Johnson, Peter King, Praveen Nair, Jason Funt, Sarah Kolitz, Kevin D Fowler, John Brothers II, Amy Axel, Scott Barrett, Benjamin J Zeskind, Brett Hall. Deep Cyclic Inhibition of the MAPK pathway with IMM-6-415, alone and in combination with encorafenib, demonstrates anti-tumor activity and tolerability in RAF mutant tumors in vivo [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A093.

Abstract C071: HBI-2375, a selective inhibitor of MLL1-WDR5 interaction, possesses desirable preclinical characteristics in solid tumors in combination with checkpoint inhibitors and also in leukemias in the future clinical investigations

Abstract MLL1 (mixed lineage leukemia1) interaction with WDR5 (WD repeat domain 5) is critical for its histone methyl transferase (HMT) activity leading to transcriptional regulation of gene expression. MLL1 dysregulation such as its translocation at chromosome 11q23 results in generation of MLL1 fusion genes and leukemia development. Additionally, recent studies indicated a role for MLL1-WDR5 interaction in solid tumors progression via directly affecting cancer cells or by suppression of tumor immunity. Therefore, MLL1-WDR5 interaction is an important target in the discovery of anti-cancer drugs for both leukemia and solid tumors. HUYABIO recently identified HBI-2375 as a selective inhibitor of MLL1-WDR5 interaction. HBI-2375 showed potent inhibition of WDR5 (IC50=4.48 nM) and inhibited MV4-11 cellular proliferation (average IC50=3.15 µM) in CTG assays. In addition, HBI-2375 was stable in liver S9 and whole blood, with moderate to high plasma protein binding rates in multiple species, and acceptable hERG IC50 (17 µM). Furthermore, HBI-2375 displayed reasonable oral PK properties (Cmax, T1/2, AUC, and F%) in mouse and dog. In vivo, HBI-2375 has shown significant dose dependent anti-tumor activity in MV4-11 xenograft AML model along with reduction in methylation of H3K4 in the tumors harvested from the treated animals demonstrating the targeted disruption of HMT activity in cancer cells by HBI-2375. Furthermore HBI-2375 in combination with PD-1 mAb significantly inhibited tumor growth in MC38, colorectal tumor model, and in 3LL lung carcinoma syngeneic tumor models. Flow cytometry and IHC showed significant increase in the infiltration of CD8+ CTLs in the combination group further providing immune-driven mechanisms for greater efficacy of combination therapy. Gene expression analysis also identified genes that are involved in promoting immune system activity in the combination group. Therefore, HBI-2375 may represent a first-in-class MLL1-WDR5 inhibitor in the clinic. Citation Format: Farbod Shojaei, Che Fang, Jill M Ricono, Mireille Gillings. HBI-2375, a selective inhibitor of MLL1-WDR5 interaction, possesses desirable preclinical characteristics in solid tumors in combination with checkpoint inhibitors and also in leukemias in the future clinical investigations [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C071.

Abstract A016: Reprogramming of immune-enhancing neutrophils by subclinical low-dose endotoxin for the treatment of cancer

Abstract Despite the recent resurgence of the pioneering concept of “Coley’s toxin” in ushering in anti-cancer immune therapies highlight by check-point inhibitors and CAR-T approaches, fundamental mechanisms responsible for the immune-enhancing efficacy of innate low-dose “Coley’s toxin” remain poorly understood. This study aims to reveal the novel reprogramming of immune-enhancing neutrophils by super-low dose endotoxin conducive for innate immunity-based anti-cancer therapies. We perform scRNAseq analyses and reveal that neutrophils trained by super-low dose endotoxin (SL-LPS) adopt a unique immune-enhancing cluster characterized by CD177loCD11BloCD80hiCD40hi. Transfusion of SL-LPS trained neutrophils into recipient mice with AOM/DSS-inducing colorectal tumors exhibit potent efficacy in reducing tumor burden. SL-LPS trained neutrophils show relieved suppression of adaptive T cells as compared to un-trained neutrophils in vivo and in vitro. Mechanistically, SL-LPS enables the generation of immune-enhancing neutrophils through activating STAT5 and reducing innate suppressor IRAK-M. Herein, we present evidence revealing the key principle that underlying the immune-enhancing effects of super-low dose endotoxin, unrevealing the long-held mystery of low-dose “Coley’s toxin” in generically boosting host anti-tumor defense. Our mechanistic and translational observations not only reveal fundamental mechanisms for re-programming immune-enhancing neutrophils, but also provide a proof-of-principle in developing innate neutrophil-based anti-tumor therapeutics. Citation Format: Yao Zhang, Liwu Li. Reprogramming of immune-enhancing neutrophils by subclinical low-dose endotoxin for the treatment of cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr A016.