<p>Clinicopathologic and molecular summary information for <i>BRAF</i>-mutant patients in the combined cohort, stratified by functional classes 1, 2, and 3. The presence of concomitant pathogenic mutations in <i>KRAS</i>, <i>NRAS</i>, or <i>NF1</i> is shown for each class. In addition to the data in the table, a single instance of co-occurring class 2 and 3 <i>BRAF</i> mutations was found. Unclassified mutations are assumed to be passenger changes for the purpose of our analysis. WT <i>BRAF</i> colorectal cancers within the combined cohort are included for reference. Percentages shown in parentheses reflect the proportion of total samples with available data for a given variable.</p>
\n \n\n \n \n<p>Supplementary Table 4. Summary data reclassified using a positional based system based on proximity to functional domains, which may hold value in exploring atypical BRAF variants not yet functionally assessed (Classes A, B, and C). A single instance of co-occurring class 2 and 3 BRAF is omitted from the total counts yet presented as MSS with no additional clinical data available.</p>
\n \n\n \n \n<p>Survival analysis of <i>BRAF</i>-mutant colorectal cancer classes: multivariate and univariate mixed-effects Cox proportional hazards models. Model I compares <i>BRAF</i> classes across all patients irrespective of additional Ras mutation; model II compares class 1 without Ras vs. class 3 with Ras mutations; model III examines additional Ras mutation status within class 3. All models include study as a random effect. Both univariate and multivariate analyses were performed on identical patient cohorts per model to ensure direct comparability.</p>
\n \n\n \n \n<p>mRNA expression of the EGFR ligands <i>AREG</i> and <i>EREG</i>. <b>A,</b> Expression of <i>AREG</i> and <i>EREG</i> is significantly increased in the distal colorectum vs. proximal colon across the INTERMPHEN (<i>n</i> = 119 proximal and <i>n</i> = 236 distal colorectum) and TCGA COADREAD (<i>n</i> = 18 proximal and <i>n</i> = 19 distal/rectal). Log<sub>2</sub>FC and significance values obtained via DESeq2 analysis (see Materials and Methods). <b>B,</b> Expression of <i>EREG</i> is significantly increased in class 3 <i>BRAF</i>-mutant colorectal cancers vs. class 1 in the absence of a concomitant Ras mutation (<i>P</i>.adj < 0.005), which may reflect the distal colorectum bias of class 3 mutations. (Created with <a href=\"http://BioRender.com\" target=\"_blank\">BioRender.com</a>.)</p>
\n \n\n \n \n<p>Supplementary Figure 5. Atypical KRAS mutations and additional Ras pathway mutations. A CONSORT-style diagram shows the breakdown of tumours with atypical KRAS mutations by accompanying mutations in KRAS, BRAF, NRAS or NF1.</p>
\n \n\n \n \n<p>Classes of <i>BRAF</i> mutation exhibit distinct clinicopathologic and molecular characteristics. <b>A,</b> Subdivision of the large bowel into the proximal and distal colorectum. Proportions of colorectal cancers in each location are shown by <i>BRAF</i> mutation class. <b>B,</b> CMS classifications of <i>BRAF</i>-mutant colorectal cancers by mutation class. The differences between class 2 or 3 versus class 1 (V600E) were highly significant (<i>P</i> < 0.001). No significant difference was observed between classes 2 and 3 (<i>P</i> = 0.270). <b>C</b> and <b>D,</b> Sanger sequencing chromatograms highlighting the spatial co-occurrence of <i>BRAF</i> and additional Ras pathway mutations in two colorectal cancers. Two regions of each cancer were microdissected, DNA was extracted, and relevant exons were PCR-amplified prior to Sanger sequencing. Illustrative results are shown. The co-occurrence of <i>BRAF</i><sup>G466V</sup> and <i>NRAS</i><sup>G12D</sup> can be observed in tumor 1 (<b>C</b>), and of <i>BRAF</i><sup>D594G</sup> and <i>KRAS</i><sup>L19F</sup> in tumor 2 (<b>D</b>). This analysis does not exclude fine-scale spatial mixing of distinct subclones but is consistent with the two mutations tested being present in the same tumor cells. (Created with <a href=\"http://BioRender.com\" target=\"_blank\">BioRender.com</a>.)</p>
\n \n\n \n \n<p><i>BRAF</i> functional domains and comparison between the 1-2-3 and A-B-C <i>BRAF</i> classification systems. The 1-2-3 <i>BRAF</i> mutation classification is based on <i>in vitro</i> assays of Ras pathway activity. However, as it has not been validated by the analysis of native mutations in isogenic cells, we explored, in parallel, a simpler classification based on the mutation location in functional domains: A, codon 600; B, other active site mutations flanking codon 600 (581\u2013601); and C, ATP-binding site mutations (464\u2013469). RBD, Ras-binding domain. (Created with <a href=\"http://BioRender.com\" target=\"_blank\">BioRender.com</a>.)</p>
\n \n\n \n \n<p>Supplementary Table 6. Logistic regression model of atypical BRAF mutation versus V600E status with covariates MSI status, age, sex, and tumour location.</p>
\n \n\n \n \n<div>Abstract<p><i>BRAF</i> mutations in colorectal cancer comprise three functional classes: class 1 (V600E) with strong constitutive activation, class 2 with pathogenic kinase activity lower than that of class 1, and class 3 which paradoxically lacks kinase activity. Non\u2013class 1 mutations associate with better prognosis, microsatellite stability, distal tumor location, and better anti-EGFR response. An analysis of 13 colorectal cancer cohorts (<i>n</i> = 6,605 tumors) compared class 1 (<i>n</i> = 709, 10.7% of colorectal cancers), class 2 (<i>n</i> = 31, 0.47%), and class 3 (<i>n</i> = 81, 1.22%) mutations. Class 2\u2013mutant and class 3\u2013mutant colorectal cancers frequently co-occurred with additional Ras pathway mutations (29.0% and 45.7%, respectively, vs. 2.40% in class 1; <i>P</i> < 0.001), often at atypical sites (<i>KRAS</i> noncodon 12/13/61, <i>NRAS</i>, or <i>NF1</i>). Ras pathway activation was highest in class 1 and lowest in class 3, with a greater distal expression of EGFR ligands (amphiregulin/epiregulin) supporting weaker <i>BRAF</i> driver mutations. Unlike class 1 mutants, class 3 tumors resembled chromosomally unstable colorectal cancers in mutation burdens, signatures, driver mutations, and transcriptional subtypes, whereas class 2 mutants displayed intermediate characteristics. Atypical <i>BRAF</i> mutations were associated with longer overall survival than class 1 mutations (HR = 0.25; <i>P</i> = 0.011) but lost this advantage in cancers with additional Ras mutations (HR = 0.94; <i>P</i> = 0.86). This study supports the suggestion that class 3 <i>BRAF</i> mutations amplify existing Ras signaling in a two-mutation model and that the enhancement of weak/atypical Ras mutations may suffice for tumorigenesis, with potentially clinically important heterogeneity in the class 2/3 subgroup.</p><p><b>Implications:</b> The heterogeneous nature of <i>BRAF</i>-mutant colorectal cancers, particularly among class 2/3 mutations which frequently harbor additional Ras mutations, highlights the necessity of comprehensive molecular profiling.</p></div>
\n \n\n \n \n<p>Supplementary Figure 3. Molecular subtyping of BRAF-mutant CRCs into the consensus molecular subtypes of CRC (CMS). Left: CMS groupings within BRAF classes 1 to 3. Right: CMS groups when stratified with the A-B-C positional classifier.</p>
\n \n\n \n \n<p>Suplementary Table 3a-b. Summarised BRAF mutation characteristics per cohort following the 1-2-3 classification system (Table 3a) and the A-B-C classification system (Table 3b). Table cells with values less than 5 have been censored to facilitate Genomics England reporting restrictions. An uncensored version of Supplementary Table 3 is available within the Genomics England 100,000 Genomes Project Research Environment upon request.</p>
\n \n\n \n \nMany patients with cancer do not benefit from currently approved immune checkpoint inhibitors (ICIs), suggesting that additional immunomodulation of the immunosuppressive tumor microenvironment (TME) is required. MTL-CCAAT enhancer-binding protein alpha (CEBPA) specifically upregulates the expression of the master myeloid transcription factor, CEBPA, relieving myeloid-driven immunosuppression. Here, we report the safety, tolerability, pharmacokinetics, and efficacy of MTL-CEBPA in combination with pembrolizumab in patients with advanced solid tumors that typically show ICI resistance. Multimodal exploratory analyses of paired patient biopsies demonstrate biological changes associated with the combination treatment of MTL-CEBPA and pembrolizumab, including increased infiltration of T\u00a0cell and antigen-presenting cells supporting conversion from an immune-desert toward a more immune-inflamed TME. Patients with disease stabilization demonstrate reductions in immunosuppressive myeloid cells post treatment. Collectively, these data support a role for MTL-CEBPA in reducing immunosuppression in the TME. This study was registered at ClinicalTrials.gov (NCT04105335).
\n \n\n \n \nOBJECTIVE: To identify and analyse the interventions delivered opportunistically in secondary or tertiary medical settings, focused on improving routine vaccination uptake in children and young people. DESIGN: Scoping review. SEARCH STRATEGY: We searched CINAHL, Web of Science, Medline, Embase and Cochrane Database of Systematic Reviews for studies in English published between 1989 and 2021 detailing interventions delivered in secondary or tertiary care that aimed to improve childhood vaccination coverage. Title, abstract and full-text screening were performed by two independent reviewers. RESULTS: After deduplication, the search returned 3456 titles. Following screening and discussion between reviewers, 53 studies were included in the review. Most papers were single-centre studies from high-income countries and varied considerably in terms of their study design, population, target vaccination, clinical setting and intervention delivered. To present and analyse the study findings, and to depict the complexity of vaccination interventions in hospital settings, findings were presented and described as a sequential pathway to opportunistic vaccination in secondary and tertiary care comprising the following stages: (1) identify patients eligible for vaccination; (2) take consent and offer immunisations; (3) order/prescribe vaccine; (4) dispense vaccine; (5) administer vaccine; (6) communicate with primary care; and (7) ongoing benefits of vaccination. CONCLUSIONS: Most published studies report improved vaccination coverage associated with opportunistic vaccination interventions in secondary and tertiary care. Children attending hospital appear to have lower baseline vaccination coverage and are likely to benefit from vaccination interventions in these settings. Checking immunisation status is challenging, however, and electronic immunisation registers are required to enable this to be done quickly and accurately in hospital settings. Further research is required in this area, particularly multicentre studies and cost-effectiveness analysis of interventions.
\n \n\n \n \nPurpose: The purpose of this study is to analyze the relationship between nuclear charge (Z), atomic mass (A), LET (linear energy transfer for maximal relative biological effectiveness (RBE)) for accelerated ions based on the hypothesis that for each ion, LETU is related to their nuclear radius. Methods: Published LETU data for proton, helium, carbon, neon, silicon, argon, and iron ions and their Z and A numbers are fitted by a power law function (PLF) and compared with PLF based on atomic cross-sections and nuclear dimensions for spherical or spheroidal atomic nuclei. The PLF allows for isoeffective RBE estimations for different ions at any value of LET based on the LETU estimations. For any two ions, A and B, and a specified bioeffect obtained at LETA, the equivalent isoeffective LETB, is estimated using (Formula presented.). Results: The data-fitting program provided the following results: (Formula presented.), and (Formula presented.), where 78.1 and 86.6 keV.\u03bcm\u22121 are the proton LETU values (i.e., without proton cellular range limit considerations). Goodness-of-fit tests are similar for each model, but the proton estimations differ. These exponents are lower than 0.66 and 0.33 (those for nuclear cross-sections and spherical nuclear radii, respectively), but suggest prolate nuclear shapes in most of the ions studied. Worked examples of estimating isoeffective LET values for two different ions are provided. Conclusions: The fitted power law relationships between LETU and Z or A are broadly equivalent and compatible with prolate nuclear shapes. These models may offer a more rational basis for future ion-beam radiobiology research.
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