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The distribution of carcinoembryonic antigen (CEA) in colorectal cancer (CRC) differs from that in normal colorectal tissue, being found on all borders of the cell membrane and hence enabling access to intravenous antibody, making CEA a good target for antibody-based therapy. The distinctive anti-CEA antibody, PR1A3, binds only membrane-bound CEA. Humanised PR1A3 (hPR1A3) was assessed both in vitro cytotoxicity and binding assays with colorectal cancer cell lines expressing varying levels of CEA. Human peripheral blood mononuclear cells (PBMCs) and purified natural killer (NK) cells were used as effectors. The in vitro assays demonstrated hPR1A3 CEA-specific binding and antibody-dependent and CEA-specific killing of human colorectal cancer cell lines by human PBMCs. The effect increased with increasing concentration of antibody and surface CEA, and was lost by using the parent murine IgG1 PR1A3. Killing was also blocked by antibody to the Fc-gammaIIIA receptor. Purified human NK cells were effective at much lower effector:target ratios than unfractionated PBMCs, indicating that NK cells were the main mediators of hPR1A3-based CEA-specific killing. The results support the development of hPR1A3 for therapy of colorectal cancer. [ABSTRACT FROM AUTHOR]

Elevated cancer metabolism releases lactic acid and CO 2 into the under-perfused tumor microenvironment, resulting in extracellular acidosis. The surviving cancer cells must adapt to this selection pressure; thus, targeting tumor acidosis is a rational therapeutic strategy to manage tumor growth. However, none of the major approved treatments are based explicitly on disrupting acid handling, signaling, or adaptations, possibly because the distinction between acid-sensitive and acid-resistant phenotypes is not clear. Here, we report pH-related phenotypes of sixty-eight colorectal cancer (CRC) cell lines by measuring i) extracellular acidification as a readout of acid production by fermentative metabolism and ii) growth of cell biomass over a range of extracellular pH (pHe) levels as a measure of the acid sensitivity of proliferation. Based on these measurements, CRC cell lines were grouped along two dimensions as "acid-sensitive"/"acid-resistant" versus "low metabolic acid production"/"high metabolic acid production." Strikingly, acid resistance was associated with the expression of CEACAM6 and CEACAM5 genes coding for two related cell-adhesion molecules, and among pH-regulating genes, of CA12 . CEACAM5/6 protein levels were strongly induced by acidity, with a further induction under hypoxia in a subset of CRC lines. Lack of CEACAM6 (but not of CEACAM5) reduced cell growth and their ability to differentiate. Finally, CEACAM6 levels were strongly increased in human colorectal cancers from stage II and III patients, compared to matched samples from adjacent normal tissues. Thus, CEACAM6 is a marker of acid-resistant clones in colorectal cancer and a potential motif for targeting therapies to acidic regions within the tumors., Competing Interests: Competing interests statement:The authors declare no competing interest.

The naked mole rat (NMR), Heterocephalus glaber, the longest-living rodent, provides a unique opportunity to explore how evolution has shaped adult stem cell (ASC) activity and tissue function with increasing lifespan. Using cumulative BrdU labelling and a quantitative imaging approach to track intestinal ASCs (Lgr5 + ) in their native in vivo state, we find an expanded pool of Lgr5 + cells in NMRs, and these cells specifically at the crypt base (Lgr5 +CBC ) exhibit slower division rates compared to those in short-lived mice but have a similar turnover as human LGR5 +CBC cells. Instead of entering quiescence (G0), NMR Lgr5 +CBC cells reduce their division rates by prolonging arrest in the G1 and/or G2 phases of the cell cycle. Moreover, we also observe a higher proportion of differentiated cells in NMRs that confer enhanced protection and function to the intestinal mucosa which is able to detect any chemical imbalance in the luminal environment efficiently, triggering a robust pro-apoptotic, anti-proliferative response within the stem/progenitor cell zone., (© 2023. The Author(s).)

Acidic environments reduce the intracellular pH (pHi) of most cells to levels that are sub-optimal for growth and cellular functions. Yet, cancers maintain an alkaline cytoplasm despite low extracellular pH (pHe). Raised pHi is thought to be beneficial for tumor progression and invasiveness. However, the transport mechanisms underpinning this adaptation have not been studied systematically. Here, we characterize the pHe-pHi relationship in 66 colorectal cancer cell lines and identify the acid-loading anion exchanger 2 (AE2, SLC4A2) as a regulator of resting pHi. Cells adapt to chronic extracellular acidosis by degrading AE2 protein, which raises pHi and reduces acid sensitivity of growth. Acidity inhibits mTOR signaling, which stimulates lysosomal function and AE2 degradation, a process reversed by bafilomycin A1. We identify AE2 degradation as a mechanism for maintaining a conducive pHi in tumors. As an adaptive mechanism, inhibiting lysosomal degradation of AE2 is a potential therapeutic target., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Cancer evolution consists of a stepwise acquisition of genetic and epigenetic changes, which alter the gene expression profiles of cells in a particular tissue and result in phenotypic alterations acted upon by natural selection. The recurrent appearance of specific genetic lesions across individual cancers and cancer types suggests the existence of certain "driver mutations," which likely make up the major contribution to tumors' selective advantages over surrounding normal tissue and as such are responsible for the most consequential aspects of the cancer cells' gene expression patterns and phenotypes. We hypothesize that such mutations are likely to cluster with specific dichotomous shifts in the expression of the genes they most closely control, and propose GMMchi, a Python package that leverages Gaussian Mixture Modeling to detect and characterize bimodal gene expression patterns across cancer samples, as a tool to analyze such correlations using 2 × 2 contingency table statistics., Results: Using well-defined simulated data, we were able to confirm the robust performance of GMMchi, reaching 85% accuracy with a sample size of n = 90. We were also able to demonstrate a few examples of the application of GMMchi with respect to its capacity to characterize background florescent signals in microarray data, filter out uninformative background probe sets, as well as uncover novel genetic interrelationships and tumor characteristics. Our approach to analysing gene expression analysis in cancers provides an additional lens to supplement traditional continuous-valued statistical analysis by maximizing the information that can be gathered from bulk gene expression data., Conclusions: We confirm that GMMchi robustly and reliably extracts bimodal patterns from both colorectal cancer (CRC) cell line-derived microarray and tumor-derived RNA-Seq data and verify previously reported gene expression correlates of some well-characterized CRC phenotypes., Availability: The Python package GMMchi and our cell line microarray data used in this paper is available for downloading on GitHub at https://github.com/jeffliu6068/GMMchi ., (© 2022. The Author(s).)

Growth of cancer cells in vitro can be attenuated by genetically inactivating selected metabolic pathways. However, loss-of-function mutations in metabolic pathways are not negatively selected in human cancers, indicating that these genes are not essential in vivo. We hypothesize that spontaneous mutations in 'metabolic genes' will not necessarily produce functional defects because mutation-bearing cells may be rescued by metabolite exchange with neighboring wild-type cells via gap junctions. Using fluorescent substances to probe intercellular diffusion, we show that colorectal cancer (CRC) cells are coupled by gap junctions assembled from connexins, particularly Cx26. Cells with genetically inactivated components of pH regulation ( SLC9A1 ), glycolysis ( ALDOA ), or mitochondrial respiration ( NDUFS1 ) could be rescued through access to functional proteins in co-cultured wild-type cells. The effect of diffusive coupling was also observed in co-culture xenografts. Rescue was largely dependent on solute exchange via Cx26 channels, a uniformly and constitutively expressed isoform in CRCs. Due to diffusive coupling, the emergent phenotype is less heterogenous than its genotype, and thus an individual cell should not be considered as the unit under selection, at least for metabolite-handling processes. Our findings can explain why certain loss-of-function mutations in genes ascribed as 'essential' do not influence the growth of human cancers., Competing Interests: SM, JM, AH, JK, EB, AH, GA, WB, PS No competing interests declared, (© 2022, Monterisi et al.)

Short tandem repeat (STR) polymorphisms are traditionally assessed by measuring allele lengths via capillary electrophoresis (CE). Massively parallel sequencing (MPS) reveals differences among alleles of the same length, thus improving discrimination, but also identifying groups of alleles likely related by descent. These may have relatively restricted geographical distributions and thus MPS could detect population structure more effectively than CE-based analysis. We addressed this question by applying an MPS multiplex, the Promega PowerSeq™ Auto/Mito/Y System prototype, to 362 individuals chosen to represent a wide geographical spread from the People of the British Isles (PoBI) cohort, which represents at least three generations of local rural ancestry. As well as 22 autosomal STRs (aSTRs; equivalent to PowerPlex Fusion loci) the system sequences 23 Y-STRs (the PowerPlexY 23 loci) and the control region (CR) of mitochondrial DNA (mtDNA), allowing population structure to be compared across biparentally and uniparentally inherited segments of the genome. For all loci, F ST -based tests of population structure were done based on historical, linguistic, and geographical partitions, and for aSTRs the clustering algorithm STRUCTURE was also applied. STRs were considered using both length and sequence. Sequencing increased aSTR allele diversity by 87.5% compared to CE-based designations, reducing random match probability to 1.25E-30, compared to a CE-based 6.72E-27. Significant population structure was detectable in just one pairwise comparison (Central/South East England compared to the rest), and for sequence-based alleles only. The 362 samples carried 308 distinct mtDNA CR haplotypes corresponding to 13 broad haplogroups, representing a haplotype diversity of 0.9985 ( ± 0.0005), and a haplotype match probability of 0.0043. No significant population structure was observed. Y-STR haplotypes belonged to ten broad predicted Y-haplogroups. Allele diversity increased by 33% when considered at the sequence rather than length level, although haplotype diversity was unchanged at 0.999969 ( ± 0.000001); haplotype match probability was 2.79E-03. In contrast to the biparentally and maternally inherited loci, Y-STR haplotypes showed significant population structure at several levels, but most markedly in a comparison of regions subject to Anglo-Saxon influence in the east with the rest of the sample. This was evident for both length- and sequence-based allele designations, with no systematic difference between the two. We conclude that MPS analysis of aSTRs or Y-STRs does not generally reveal stronger population structure than length-based analysis, that UK maternal lineages are not significantly structured, and that Y-STR haplotypes reveal significant population structure that may reflect the Anglo-Saxon migrations to Britain in the 6th century., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Unlike most cell types, many cancer cells survive at low extracellular pH (pHe), a chemical signature of tumors. Genes that facilitate survival under acid stress are therefore potential targets for cancer therapies. We performed a genome-wide CRISPR-Cas9 cell viability screen at physiological and acidic conditions to systematically identify gene knockouts associated with pH-related fitness defects in colorectal cancer cells. Knockouts of genes involved in oxidative phosphorylation (NDUFS1) and iron-sulfur cluster biogenesis (IBA57, NFU1) grew well at physiological pHe, but underwent profound cell death under acidic conditions. We identified several small-molecule inhibitors of mitochondrial metabolism that can kill cancer cells at low pHe only. Xenografts established from NDUFS1 -/- cells grew considerably slower than their wild-type controls, but growth could be stimulated with systemic bicarbonate therapy that lessens the tumoral acid stress. These findings raise the possibility of therapeutically targeting mitochondrial metabolism in combination with acid stress as a cancer treatment option., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Purpose: The molecular pathogenesis of solid tumour was first assessed in colorectal cancer (CRC). To date, ≤ 100 genes with somatic alterations have been found to inter-connectively promote neoplastic transformation through specific pathways. The process of colorectal carcinogenesis via genome landscape is reviewed on the basis of an adenoma-to-carcinoma sequence, as shown by serial histological and epidemiological observations., Methods: The relevant literatures from PubMed (1980-2021) have been reviewed for this article., Results: The major routes of CRC development, chromosomal instability (CIN), microsatellite instability (MSI), and the serrated route either via CIN or MSI, proceed through the respective molecular pathway of colorectal carcinogenesis. Particular aspects of CRC carcinogenesis can also be determined by evaluating familial CRCs (FCRC) and genotype-phenotype correlations. Specific causative gene alterations still leave to be identified in several FCRCs. Otherwise, recently verified FCRC can be particularly notable, for example, EPCAM-associated Lynch syndrome, polymerase proofreading-associated polyposis, RNF43-associated polyposis syndrome or NTHL1 tumour syndrome, and hereditary mixed polyposis syndrome. The oncogenic landscape is described, including representative pathway genes, the three routes of carcinogenesis, familial CRCs, genotype-phenotype correlations, the identification of causative genes, and consensus molecular subtypes (CMS)., Conclusion: Whole genome research using multi-gene panels (MGPs) has facilitated high through-put detection of previously unidentified genes involved in colorectal carcinogenesis. New approaches designed to identify rare variants are recommended to consider their alterations implicated in the molecular pathogenesis., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)