Long-term survivals of immune checkpoint inhibitors as neoadjuvant and adjuvant therapy in dMMR/MSI-H colorectal and gastric cancers.

BACKGROUND: The long-term survival benefit of immune checkpoint inhibitors (ICIs) in neoadjuvant and adjuvant settings is unclear for colorectal cancers (CRC) and gastric cancers (GC) with deficiency of mismatch repair (dMMR) or microsatellite instability-high (MSI-H). METHODS: This retrospective study enrolled patients with dMMR/MSI-H CRC and GC who received at least one dose of neoadjuvant ICIs (neoadjuvant cohort, NAC) or adjuvant ICIs (adjuvant cohort, AC) at 17 centers in China. Patients with stage IV disease were also eligible if all tumor lesions were radically resectable. RESULTS: In NAC (n = 124), objective response rates were 75.7% and 55.4%, respectively, in CRC and GC, and pathological complete response rates were 73.4% and 47.7%, respectively. The 3-year disease-free survival (DFS) and overall survival (OS) rates were 96% (95%CI 90-100%) and 100% for CRC (median follow-up [mFU] 29.4 months), respectively, and were 84% (72-96%) and 93% (85-100%) for GC (mFU 33.0 months), respectively. In AC (n = 48), the 3-year DFS and OS rates were 94% (84-100%) and 100% for CRC (mFU 35.5 months), respectively, and were 92% (82-100%) and 96% (88-100%) for GC (mFU 40.4 months), respectively. Among the seven patients with distant relapse, four received dual blockade of PD1 and CTLA4 combined with or without chemo- and targeted drugs, with three partial response and one progressive disease. CONCLUSION: With a relatively long follow-up, this study demonstrated that neoadjuvant and adjuvant ICIs might be both associated with promising DFS and OS in dMMR/MSI-H CRC and GC, which should be confirmed in further randomized clinical trials.

Genome-wide CRISPR screening identifies the pivotal role of ANKRD42 in colorectal cancer metastasis through EMT regulation.

Colorectal cancer (CRC), a pervasive and lethal malignancy of gastrointestinal cancer, imposes significant challenges due to the occurrence of distant metastasis in advanced stages. Understanding the intricate regulatory mechanisms driving CRC distant metastasis is of paramount importance. CRISPR-Cas9 screening has emerged as a powerful tool for investigating tumor initiation and progression. However, its application in studying CRC distant metastasis remains largely unexplored. To establish a model that faithfully recapitulates CRC liver metastasis in patients, we developed an in vivo genome-wide CRISPR-Cas9 screening approach using a spleen-injected liver metastasis mouse model. Through comprehensive screening of a whole-genome sgRNA library, we identified ANKRD42 as a pivotal regulatory gene facilitating CRC liver metastasis. Analysis of the TCGA database and our clinical cohorts unveiled heightened ANKRD42 expression in metastases. At the cellular level, the attenuation of ANKRD42 impaired the migration and invasion processes of tumor cells. In vivo experiments further validated these observations, highlighting the diminished liver metastatic capacity of tumor cells upon ANKRD42 knockdown. To unravel the specific mechanisms by which ANKRD42 regulates CRC distant metastasis, we leveraged patient-derived organoid (PDO) models. Depleting ANKRD42 in PDOs sourced from liver metastases precipitated the downregulation of pivotal genes linked to epithelial-mesenchymal transition (EMT), including CDH2 and SNAI2, thereby effectively suppressing tumor metastasis. This study not only establishes a conceptual framework but also identifies potential therapeutic avenues for advanced-stage distant metastasis in CRC patients.

Influential upregulation of KCNE4: Propelling cancer associated fibroblasts-driven colorectal cancer progression.

BACKGROUND: Colorectal cancer (CRC) is a malignancy of remarkable heterogeneity and heightened morbidity. Cancer associated fibroblasts (CAFs) are abundant in CRC tissues and are essential for CRC growth. Here, we aimed to develop a CAF-related classifier for predicting the prognosis of CRC and identify critical pro-tumorigenic genes in CAFs. METHOD: The mRNA expression and clinical information of CRC samples were sourced from two comprehensive databases, The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO). Using a weighted gene co-expression network analysis (WGCNA) approach, CAF-related genes were identified and a CAF risk signature was developed through the application of univariate analysis and the least absolute shrinkage and selection operator (LASSO) Cox regression model. EdU cell proliferation assay, and transwell assay were performed to detect the oncogenic role of KCNE4 in CAFs. RESULTS: We constructed a prognostic CAF model consisting of two genes (SFRP2 and KCNE4). CRC patients were classified into low- and high-CAF-risk groups using the median CAF risk score, and patients in the high-CAF-risk group had worse prognosis. Meanwhile, a higher risk score for CAFs was associated with greater stromal and CAF infiltrations, as well as higher expression of CAF markers. Furthermore, TIDE analysis indicated that patients with a high CAF risk score are less responsive to immunotherapy. Our further experiments had confirmed the strong correlation between KCNE4 and the malignant phenotypes of CAFs. Moreover, we had shown that KCNE4 could actively promote tumor-promoting phenotypes in CAFs, indicating its critical role in cancer progression. CONCLUSION: The two-gene prognostic CAF signature was constructed and could be reliable for predicting prognosis for CRC patients. Moreover, KCNE4 may be a promising strategy for the development of novel anti-cancer therapeutics specifically directed against CAFs.

Realizing Ultrabroadband NIR-II Emission and Wide-Range Wavelength Tuning in Cr4+-activated ABO2 (A = Li, Na; B = Al, Ga) Phosphors.

Cr4+-activated phosphors are important candidate materials for NIR-II light sources, but providing a suitable lattice coordination environment for Cr4+ and achieving long wavelength broadband emission remains a challenge. In this work, a series of Cr4+-activated ABO2 (A = Li, Na; B = Al, Ga) phosphors were successfully prepared. Due to the presence of only tetrahedral coordination structures available for Cr4+ to occupy in the matrix crystal ABO2, the valence state and luminescence stability of Cr4+ are effectively guaranteed. Through the cation substitution design of A-site (Na → Li) and B-site (Ga → Al), the [BO4] tetrahedron is distorted and expanded, which degrades the symmetry of the Cr4+ coordination crystal field. Consequently, the central wavelength of the Cr4+ emission peak is tuned from 1280 to 1430 nm, and the fwhm is significantly extended from 257 to 355 nm. Thebroadband NIR-II light sources constructed with LiAlO2: 0.03Cr4+ and NaGaO2: 0.03Cr4+ phosphors verify their important potential applications in nondestructive testing and biological imaging.

PTCH1 mutation promotes antitumor immunity and the response to immune checkpoint inhibitors in colorectal cancer patients.

Immunotherapy has emerged as an effective therapeutic strategy for various cancers, including colorectal cancer (CRC), but only a subset of MSI-H patients can benefit from such therapy. Patched1 (PTCH1) is a frequently altered gene in CRCs and its mutations contribute to unregulated Hedgehog (Hh) signaling. In the study, we evaluated the association of PTCH1 mutations with CRC immunity based on our single-center cohort and multiple cancer genomic datasets. Among 21 enrolled patients, six (28.6%) harbored a PTCH1 mutation based on WES analyses. In CRC patients, the PTCH1 mutation subgroup experienced a higher durable clinical benefit rate than the PTCH1 wild-type subgroup (100% vs. 40%, P = 0.017). In addition, patients with the PTCH1 mutation experienced greater progression-free survival (PFS, P = 0.037; HR, 0.208) and overall survival (OS, P = 0.045; HR, 0.185). A validation cohort from the MSKCC also confirmed the correlation between PTCH1 mutation and better prognosis (P = 0.022; HR, 0.290). Mechanically, diverse antitumor immune signatures were more highly enriched in PTCH1-mutated tumors than in PTCH1 wild-type tumors. Furthermore, PTCH1-mutated tumors had higher proportions of CD8 + T cells, activated NK cells, and M1 type macrophage infiltration, as well as elevated gene signatures of several steps in the cancer-immunity cycle. Notably, the PTCH1 mutation was correlated with tumor mutational burden (TMB), loss of heterozygosity score, and copy number variation burden. Our results show that the mutation of PTCH1 is a potential biomarker for predicting the response of CRC patients to immunotherapy.

Mutations of PI3K-AKT-mTOR pathway as predictors for immune cell infiltration and immunotherapy efficacy in dMMR/MSI-H gastric adenocarcinoma.

BACKGROUND: A significant subset of mismatch repair-deficient (dMMR)/microsatellite instability-high (MSI-H) gastric adenocarcinomas (GAC) are resistant to immune checkpoint inhibitors (ICIs), yet the underlying mechanism remains largely unknown. We sought to investigate the genomic correlates of the density of tumor-infiltrating immune cells (DTICs) and primary resistance to ICI treatment. METHODS: Four independent cohorts of MSI-H GAC were included: (i) the surgery cohort (n = 175) with genomic and DTIC data, (ii) the 3DMed cohort (n = 32) with genomic and PD-L1 data, (iii) the Cancer Genome Atlas (TCGA) cohort (n = 73) with genomic, transcriptomic, and survival data, and (iv) the ICI treatment cohort (n = 36) with pre-treatment genomic profile and ICI efficacy data. RESULTS: In the dMMR/MSI-H GAC, the number of mutated genes in the PI3K-AKT-mTOR pathway (NMP) was positively correlated with tumor mutational burden (P < 0.001) and sensitivity to PI3K-AKT-mTOR inhibitors and negatively correlated with CD3+ (P < 0.001), CD4+ (P = 0.065), CD8+ (P = 0.004), and FOXP3+ cells (P = 0.033) in the central-tumor rather than invasive-margin area, and the transcription of immune-related genes. Compared to the NMP-low (NMP = 0/1) patients, the NMP-high (NMP ≥ 2) patients exhibited a poorer objective response rate (29.4% vs. 85.7%, P < 0.001), progression-free survival (HR = 3.40, P = 0.019), and overall survival (HR = 3.59, P = 0.048) upon ICI treatment. CONCLUSIONS: Higher NMP was identified as a potential predictor of lower DTICs and primary resistance to ICIs in the dMMR/MSI-H GAC. Our results highlight the possibility of using mutational data to estimate DTICs and administering the PI3K-AKT-mTOR inhibitor as an immunotherapeutic adjuvant in NMP-high subpopulation to overcome the resistance to ICIs.

Phase I study of the recombinant humanized anti-HER2 monoclonal antibody-MMAE conjugate RC48-ADC in patients with HER2-positive advanced solid tumors.

PURPOSE: RC48 contains the novel humanized anti-HER2 antibody hertuzumab conjugated to MMAE via a cleavable linker. A phase I study was initiated to evaluate the toxicity, MTD, PK, and antitumor activity of RC48 in patients with HER2-overexpressing locally advanced or metastatic solid carcinomas, particularly gastric cancer. PATIENTS AND METHODS: This was a 2-part phase I study. Successive cohorts of patients received escalating doses of RC48 (0.1 mg/kg, 0.5 mg/kg, 1.0 mg/kg, 2.0 mg/kg, 2.5 mg/kg, and 3.0 mg/kg). Dose expansion proceeded at the dose of 2.0 mg/kg Q2W. The efficacy and safety set included all patients who received at least one dose of RC48. RESULTS: Fifty-seven patients were enrolled, the MTD was unavailable due to termination of 3.0 mg/kg cohort; 2.5 mg/kg Q2W was declared the RP2D. RC48 was well tolerated, the most frequent grade 3 or worse TRAEs included neutropenia (19.3%), leukopenia (17.5%), hypoesthesia (14.0%), and increased conjugated blood bilirubin (8.8%). Four deaths occurred during the whole study, three of which were believed to be related to RC48. Overall, ORR and DCR were 21.0% (12/57) and 49.1% (28/57). Notably, patients who were HER2 IHC2+/FISH- responded similarly to those who were IHC2+/FISH+ and IHC3+, with ORRs of 35.7% (5/14), 20% (2/10), and 13.6% (3/22), respectively. In patients who were pretreated with HER2-targeted drugs, RC48 also showed promising efficacy, with ORR of 15.0% (3/20) and DCR of 45.0% (9/20). CONCLUSION: RC48 was well tolerated and showed promising antitumor activity in HER2-positive solid tumors, including gastric cancer with HER2 IHC 2+/FISH- status. CLINICAL TRIAL INFORMATION: NCT02881190.

Baseline derived neutrophil-to-lymphocyte ratio as a prognostic biomarker for non-colorectal gastrointestinal cancer patients treated with immune checkpoint blockade.

BACKGROUND: Biomarkers in non-colorectal gastrointestinal (GI) cancer patients receiving immune checkpoint blockades (ICBs) are still limited. METHODS: Data were prospectively collected from a discovery cohort (n = 53) and a validation cohort (n = 107) in patients with non-colorectal GI cancer receiving ICB, as well as a chemotherapy-only cohort (n = 171). System inflammatory markers and derived neutrophil-to-lymphocyte ratio (dNLR) were determined as biomarkers by univariate and multivariate analyses. RESULTS: A higher level of dNLR (cutoff = 3) was associated with shorter overall survival (OS) in discovery and validation cohorts. In pooled cohort, disease control rate (DCR) (28% vs. 48.1%) was associated with dNLR (p = .017). In univariate analysis, original tumor site, tumor histopathology, number of metastases, and dNLR were correlated with OS. In multivariate analysis, higher dNLR level was correlated with reduced OS (10.43 months vs. 4.20 months, p < .001). In chemotherapy-only cohort, dNLR was also correlated with DCR and OS. CONCLUSION: Higher dNLR level was correlated with worse outcomes, suggesting that dNLR may help risk-group stratification and assist disease management strategies as a prognostic biomarker for non-colorectal GI patients receiving ICB.

Synthesis and biological activity of benzoylcyclohexanedione herbicide SYP-9121.

Benzoylcyclohexanedione herbicides work by inhibiting 4-hydroxyphenylpyruvate dioxygenase which was the last new target site introduced for herbicides. In an attempt to find new 4-hydroxyphenylpyruvate dioxygenase inhibitors with high efficacy and selectivity, a novel benzoylcyclohexanedione compound SYP-9121 was synthesized and studied in greenhouse and field. In the greenhouse, SYP-9121 showed broad spectrum herbicidal activity and good safety to maize. Its control of barnyard grass, crabgrass, redroot pigweed, purslane, dayflower and night shade was equivalent to that of the commercial herbicide mesotrione. Three field trials in summer maize showed that SYP-9121 could efficiently control both grass and broadleaf weeds with good selectivity. Herbicidal activity of SYP-9121 was comparable to that of mesotrione.

Secondary structure of expansion segment D1 in LSU rDNA from Arachnida and its phylogenetic application in Eriophyoid mites and in Acari.

An increasing number of researchers have applied secondary-structure based multiple alignments of rDNA genes in phylogeny. These studies mostly depended on a few valuable divergent domains in LSU and SSU rDNA. Yet other divergent domains, e.g. D1, were poorly investigated and rarely used. However, these domains might contain additional evolutionary data and play a vital role in DNA-based phylogenetic study. Here, we investigated all available D1 sequences of Arachnida taxa and predicted corresponding secondary structures to help identify homologous positions in the D1 region. Long insertions were found exclusive to Eriophyoidea and folded into three newly proposed helices. Non-Acari taxa were all GC rich. In Acari, most Trombidiformes and all Mesostigmata (Parasitiformes) taxa were AT rich and Ixodida (Parasitiformes) GC rich; however there was no consistent base bias in Sarcoptiformes sequences. For Eriophyoid mites, genera Cecidophyopsis and Aceria were both well supported in MP, NJ, ME and ML tress based on D1 sequences, and clusters of Cecidophyopsis species were identical with former study. This demonstrated that the D1 region could act as a valuable molecular marker in phylogenetic reconstruction of Eriophyoidea. Additionally, D1 has been proven suitable in phylogenetic analysis at the family and genus level in Acari, but not in Opiliones.

WALL-ASSOCIATED KINASE Like 14 regulates vascular tissue development in Arabidopsis and tomato.

Initiation of plant vascular tissue is regulated by transcriptional networks during development and in response to environmental stimuli. The WALL-ASSOCIATED KINASES (WAKs) and WAK-likes (WAKLs) are cell surface receptors involved in cell expansion and defence in cells with primary walls, yet their roles in regulation of vascular tissue development that contain secondary walls remains unclear. In this study, we showed tomato (Solanum lycopersicum) SlWAKL2 and the orthologous gene in Arabidopsis thaliana, AtWAKL14, were specifically expressed in vascular tissues. SlWAKL2-RNAi tomato plants displayed smaller fruit size with fewer seeds and vascular bundles compared to wild-type (WT) and over-expression (OE) lines. RNA-seq data showed that SlWAKL2-RNAi fruits down-regulated transcript levels of genes related to vascular tissue development compared to WT. Histological analysis showed T-DNA insertion mutant wakl14-1 had reduced plant stem length with fewer number of xylem vessels and interfascicular fibres compared to WT, with no significant differences in cellulose and lignin content. Mutant wakl14-1 also showed reduced number of vascular bundles in fruit. A proWAKL14::mCherry-WAKL14 fusion protein was able to complement wakl14-1 phenotypes and showed mCherry-WAKL14 associated with the plasma membrane. In vitro binding assays showed both SlWAKL2 and AtWAKL14 can interact with pectin and oligogalacturonides. Our results reveal novel roles of WAKLs in regulating vascular tissue development.

Helicobacter pylori and immunotherapy for gastrointestinal cancer.

Helicobacter pylori infection is associated with the risk of gastrointestinal (GI) cancers; however, its impact on immunotherapy for GI cancers remains uncertain. In this study, we included 10,122 patients who underwent 13C-urea breath tests. Among 636 patients with Epstein-Barr virus-negative microsatellite-stable gastric cancer (GC) who were treated with anti-PD-1/PD-L1 therapy, H. pylori-positive patients exhibited significantly longer immune-related progression-free survival (irPFS) compared with H. pylori-negative patients (6.97 months versus 5.03 months, p < 0.001, hazard ratio [HR] 0.76, 95% confidence interval [CI] 0.62-0.95, p = 0.015). Moreover, the H. pylori-positive group demonstrated a trend of 4 months longer median immune-related overall survival (irOS) than the H. pylori-negative group. H. pylori-positive GC displayed higher densities of PD-L1+ cells and nonexhausted CD8+ T cells, indicative of a "hot" tumor microenvironment. Transcriptomic analysis revealed that H. pylori-positive GC shared molecular characteristics similar to those of immunotherapy-sensitive GC. However, H. pylori-positive patients with DNA mismatch repair-deficient (dMMR)/microsatellite instability-high (MSI-H) colorectal adenocarcinoma and esophageal squamous cell carcinoma (ESCC) had shorter irPFS compared with H. pylori-negative patients (16.13 months versus not reached, p = 0.042, HR 2.26, 95% CI 1.13-4.50, p = 0.021 and 5.57 months versus 6.97 months, p = 0.029, HR 1.59, 95% CI 1.14-2.23, p = 0.006, respectively). The difference in irOS between H. pylori-positive and -negative patients had the same trend as that between dMMR/MSI-H colorectal adenocarcinoma and ESCC patients. We also identified a trend of shorter irPFS and irOS in H. pylori-positive liver cancer and pancreatic cancer patients. In summary, our findings supported that H. pylori infection is a beneficial factor for GC immunotherapy by shaping hot tumor microenvironments. However, in dMMR/MSI-H colorectal adenocarcinoma and ESCC patients, H. pylori adversely affects the efficacy of immunotherapy.

Claudin18.2-specific CAR T cells in gastrointestinal cancers: phase 1 trial final results.

Claudin18.2 (CLDN18.2) is highly expressed with the development of various malignant tumors, especially gastrointestinal cancers, and is emerging as a new target for cancer treatment. Satricabtagene autoleucel (satri-cel)/CT041 is an autologous chimeric antigen receptor (CAR) T cell targeting CLDN18.2, and the interim results of the CT041-CG4006 trial were reported in June 2022. Here we present the final results of this single-arm, open-label, phase 1 trial, which evaluated the safety and efficacy of satri-cel in patients with CLDN18.2-positive advanced gastrointestinal cancers. This trial included a dose-escalation stage (n = 15) and a dose-expansion stage in four different cohorts (total n = 83): cohort 1, satri-cel monotherapy in 61 patients with standard chemotherapy-refractory gastrointestinal cancers; cohort 2, satri-cel plus anti-PD-1 therapy in 15 patients with standard chemotherapy-refractory gastrointestinal cancers; cohort 3, satri-cel as sequential treatment after first-line therapy in five patients with gastrointestinal cancers; and cohort 4, satri-cel monotherapy in two patients with anti-CLDN18.2 monoclonal antibody-refractory gastric cancer. The primary endpoint was safety; secondary endpoints included efficacy, pharmacokinetics and immunogenicity. A total of 98 patients received satri-cel infusion, among whom 89 were dosed with 2.5 × 108, six with 3.75 × 108 and three with 5.0 × 108 CAR T cells. Median follow-up was 32.4 months (95% confidence interval (CI): 27.3, 36.5) since apheresis. No dose-limiting toxicities, treatment-related deaths or immune effector cell-associated neurotoxicity syndrome were reported. Cytokine release syndrome occurred in 96.9% of patients, all classified as grade 1-2. Gastric mucosal injuries were identified in eight (8.2%) patients. The overall response rate and disease control rate in all 98 patients were 38.8% and 91.8%, respectively, and the median progression-free survival and overall survival were 4.4 months (95% CI: 3.7, 6.6) and 8.8 months (95% CI: 7.1, 10.2), respectively. Satri-cel demonstrates therapeutic potential with a manageable safety profile in patients with CLDN18.2-positive advanced gastrointestinal cancer. ClinicalTrials.gov identifier: NCT03874897 .

Secondary structure prediction for complete rDNA sequences (18S, 5.8S, and 28S rDNA) of Demodex folliculorum, and comparison of divergent domains structures across Acari.

According to base pairing, the rRNA folds into corresponding secondary structures, which contain additional phylogenetic information. On the basis of sequencing for complete rDNA sequences (18S, ITS1, 5.8S, ITS2 and 28S rDNA) of Demodex, we predicted the secondary structure of the complete rDNA sequence (18S, 5.8S, and 28S rDNA) of Demodex folliculorum, which was in concordance with that of the main arthropod lineages in past studies. And together with the sequence data from GenBank, we also predicted the secondary structures of divergent domains in SSU rRNA of 51 species and in LSU rRNA of 43 species from four superfamilies in Acari (Cheyletoidea, Tetranychoidea, Analgoidea and Ixodoidea). The multiple alignment among the four superfamilies in Acari showed that, insertions from Tetranychoidea SSU rRNA formed two newly proposed helixes, and helix c3-2b of LSU rRNA was absent in Demodex (Cheyletoidea) taxa. Generally speaking, LSU rRNA presented more remarkable differences than SSU rRNA did, mainly in D2, D3, D5, D7a, D7b, D8 and D10.

Real-world treatment and outcomes of patients with metastatic BRAF mutant colorectal cancer.

BACKGROUND: BRAF mutation occurs in 5%-10% of metastatic colorectal cancers (mCRCs). Patients with BRAF mutant mCRC exhibit a specific metastatic pattern and poor prognosis. Survival outcomes are heterogeneous in cases of mCRC with a BRAF mutation. The optimal first-line therapy is still controversial. METHODS: We retrospectively reviewed the medical records of patients with mCRC between June 2010 and December 2021. Clinicopathologic characteristics, treatment and BRAF mutation testing results were collected. Patients with a BRAF mutation were included. Kaplan-Meier methods and log-rank tests were used to analyze and compare survival. Cox proportional hazards regression was used to establish the predictive nomogram model. RESULTS: Of the 4475 mCRC, 261 have a BRAF mutation, including 240 V600E and 21 non-V600E mutants. The median overall survival (OS) was 18.2 months in the BRAF V600E mutant group versus 38.0 months in the non-V600E mutant group (p = 0.022). ECOG score, tumor differentiation, liver metastasis, bone metastasis and primary tumor resection were independent prognostic factors for the OS of BRAF V600E mutant mCRC. A nomogram model was established using these factors. The median OS was 39.3 m, 18.2 m and 10.7 m for the low-risk, intermediate-risk and high-risk groups defined by this model, respectively (p < 0.0001). Patients who received first-line triplet chemotherapy ± bevacizumab had comparable progression free survival (PFS) and OS compared with those treated with doublets ± bevacizumab. CONCLUSION: BRAF V600E mutant mCRCs exhibit unfavorable and heterogeneous prognosis. The first-line intensive chemotherapy did not confer a marked impact on the PFS and OS.

Trastuzumab Combined with Irinotecan in Patients with HER2-Positive Metastatic Colorectal Cancer: A Phase II Single-Arm Study and Exploratory Biomarker Analysis.

PURPOSE: The human epidermal growth factor receptor 2 (HER2) is an established therapeutic target for various kinds of solid tumors. HER2 amplification occurs in approximately 1% to 6% of colorectal cancer. In this study, we aimed to assess the efficacy and safety of trastuzumab in combination with chemotherapy in HER2-positive metastatic colorectal cancer (mCRC). Materials and Methods: An open-label, phase II trial (Clinicaltrials.gov: NCT03185988) was designed to evaluate the antitumor activity of trastuzumab and chemotherapy in HER2-positive digestive cancers excluding gastric cancer in 2017. Patients from this trial with HER2-positive, KRAS/BRAF wild-type, unresectable mCRC were analyzed in this manuscript. Eligible patients were treated with trastuzumab (8 mg/kg loading dose and then 6 mg/kg every 3 weeks) and irinotecan (120 mg/m2 days 1 and 8 every 3 weeks). The primary endpoint was the objective response rate. RESULTS: Twenty-one HER2-positive mCRC patients were enrolled in this study. Seven patients (33.3%) achieved an objective res-ponse, and 11 patients (52.4%) had stable disease as their best response. The median progression-free survival (PFS) was 4.3 months (95% confidence interval, 2.7 to 5.9). Four of the 21 patients (19.0%) had grade 3 adverse events, including leukopenia, neutropenia, urinary tract infection, and diarrhea. No treatment-related death was reported. Exploratory analyses revealed that high tumor tissue HER2 copy number was associated with better therapeutic response and PFS. Alterations in the mitogen-activated protein kinase pathway, HER2 gene, phosphoinositide 3-kinase/AKT pathway, and cell cycle control genes were potential drivers of trastuzumab resistance in mCRC. CONCLUSION: Trastuzumab combined with chemotherapy is a potentially effective and well-tolerated therapeutic regimen in mCRC with a high HER2 copy number.

PD-1/PD-L1 Inhibitor Plus Chemotherapy Versus PD-1/PD-L1 Inhibitor in Microsatellite Instability Gastrointestinal Cancers: A Multicenter Retrospective Study.

PURPOSE: To investigate the efficacy of PD-1/PD-L1 inhibitors plus chemotherapy versus anti-PD-1/PD-L1 monotherapy in advanced microsatellite instability (MSI)/mismatch repair-deficient (dMMR) gastrointestinal cancers. METHODS: We retrospectively recruited patients with MSI/dMMR gastrointestinal cancer who received anti-PD-1/PD-L1 with or without chemotherapy and compared objective response rate (ORR), disease control rate (DCR), progression-free survival (PFS), and overall survival (OS) of PD-1/PD-L1 inhibitor plus chemotherapy (chemo-anti-PD-1/PD-L1 group) and PD-1/PD-L1 inhibitor alone (anti-PD-1/PD-L1 group). Propensity score-based overlap weighting analysis was conducted to adjust the baseline covariable imbalance. Sensitivity analysis was performed to confirm the stability of the results by propensity score matching and multivariable Cox and logistic regression models. RESULTS: A total of 256 patients were eligible, with 68 and 188 receiving chemo-anti-PD-1/PD-L1 and anti-PD-1/PD-L1, respectively. The chemo-anti-PD-1/PD-L1 group showed significant improvements versus the anti-PD-1/PD-L1 group in ORR (61.8% v 38.8%; P = .001), DCR (92.6% v 74.5%; P = .002), PFS (median PFS [mPFS], not reached [NR] v 27.9 months; P = .004), and OS (median OS [mOS], NR v NR; P = .014). After overlap weighting, the improvements tended to be more significant with chemo-anti-PD-1/PD-L1 versus anti-PD-1/PD-L1 in ORR (62.5% v. 38.3%; P < .001), DCR (93.8% v 74.2%; P < .001), PFS (mPFS, NR v 26.0 months; P = .004), and OS (mOS, NR v NR; P = .010). These results were solidified through sensitivity analysis. CONCLUSION: Chemo-anti-PD-1/PD-L1 is superior to anti-PD-1/PD-L1 in MSI/dMMR gastrointestinal cancers with improved efficacy.

Complete sequence analysis of 18S rDNA based on genomic DNA extraction from individual Demodex mites (Acari: Demodicidae).

The study for the first time attempted to accomplish 18S ribosomal DNA (rDNA) complete sequence amplification and analysis for three Demodex species (Demodex folliculorum, Demodex brevis and Demodex canis) based on gDNA extraction from individual mites. The mites were treated by DNA Release Additive and Hot Start II DNA Polymerase so as to promote mite disruption and increase PCR specificity. Determination of D. folliculorum gDNA showed that the gDNA yield reached the highest at 1 mite, tending to descend with the increase of mite number. The individual mite gDNA was successfully used for 18S rDNA fragment (about 900 bp) amplification examination. The alignments of 18S rDNA complete sequences of individual mite samples and those of pooled mite samples ( ≥ 1000mites/sample) showed over 97% identities for each species, indicating that the gDNA extracted from a single individual mite was as satisfactory as that from pooled mites for PCR amplification. Further pairwise sequence analyses showed that average divergence, genetic distance, transition/transversion or phylogenetic tree could not effectively identify the three Demodex species, largely due to the differentiation in the D. canis isolates. It can be concluded that the individual Demodex mite gDNA can satisfy the molecular study of Demodex. 18S rDNA complete sequence is suitable for interfamily identification in Cheyletoidea, but whether it is suitable for intrafamily identification cannot be confirmed until the ascertainment of the types of Demodex mites parasitizing in dogs.

Sequencing for complete rDNA sequences (18S, ITS1, 5.8S, ITS2, and 28S rDNA) of Demodex and phylogenetic analysis of Acari based on 18S and 28S rDNA.

Due to the difficulty of DNA extraction for Demodex, few studies dealt with the identification and the phyletic evolution of Demodex at molecular level. In this study, we amplified, sequenced, and analyzed a complete (Demodex folliculorum) and an almost complete (D12 missing) (Demodex brevis) ribosomal DNA (rDNA) sequence and also analyzed the primary sequences of divergent domains in small-subunit ribosomal RNA (rRNA) of 51 species and in large-subunit rRNA of 43 species from four superfamilies in Acari (Cheyletoidea, Tetranychoidea, Analgoidea, and Ixodoidea). The results revealed that 18S rDNA sequence was relatively conserved in rDNA-coding regions and was not evolving as rapidly as 28S rDNA sequence. The evolutionary rates of transcribed spacer regions were much higher than those of the coding regions. The maximum parsimony trees of 18S and 28S rDNA appeared to be almost identical, consistent with their morphological classification. Based on the fact that the resolution capability of sequence length and the divergence of the 13 segments (D1-D6, D7a, D7b, and D8-D12) of 28S rDNA were stronger than that of the nine variable regions (V1-V9) of 18S rDNA, we were able to identify Demodex (Cheyletoidea) by the indels occurring in D2, D6, and D8.

Molecular mechanisms underlying the resistance of BRAF V600E-mutant metastatic colorectal cancer to EGFR/BRAF inhibitors.

Background: Combinatorial inhibition of epidermal growth factor receptor (EGFR) and BRAF shows remarkable clinical benefits in patients with BRAF V600E-mutant metastatic colorectal cancer (mCRC). However, the tumor may inevitably develop resistance to the targeted therapy, thereby limiting the response rate and durability. This study aimed to determine the genetic alterations associated with intrinsic and acquired resistance to EGFR/BRAF inhibitors in BRAF V600E-mutant mCRC. Methods: Targeted sequencing of 520 cancer-related genes was performed in tumor tissues and in plasma samples collected from patients with BRAF V600E-mutant mCRC, who were treated with EGFR/BRAF ±â€„MEK inhibitors, before and after the targeted treatment. Clinical benefit was defined as an objective response or a stable disease lasting longer than the median progression-free survival (PFS). Results: In all, 25 patients with BRAF V600E-mutant mCRC were included in this study. Those with RNF43 mutations (n = 8) were more likely to achieve clinical benefit from EGFR/BRAF inhibitors than those with wild-type RNF43 (87.5% versus 37.5%, p = 0.034). Genetic alterations in receptor tyrosine kinase genes (n = 6) were associated with worse PFS (p = 0.005). Among the 23 patients whose disease progressed after the EGFR/BRAF-targeted therapy, at least one acquired resistance-related mutation was detected in 12 patients. Acquired mutations were most frequently observed in the mitogen-activated protein kinase pathway-related genes (n = 9), including KRAS (G12D and Q61H/R), NRAS (Q61L/R/K and amplification), BRAF (amplification), and MEK1 (K57T). MET amplification and PIK3R1 Q579fs mutation emerged in three patients and one patient, respectively, after disease progression. Conclusion: Multiple genetic alterations are associated with clinical benefits and resistance to EGFR/BRAF inhibitors in BRAF V600E-mutant mCRC. Our findings provide novel insights into strategies for overcoming resistance to EGFR/BRAF inhibitors in patients with BRAF V600E-mutant mCRC.