11 research outputs found
Elucidating the impact of CD4+ T cells on tumour progression in patients with colorectal cancer
In recent years, substantial evidence has been generated demonstrating the importance
of the immune system in preventing and controlling the growth of many cancers,
including colorectal adenocarcinomas. In particular, populations of tumour-specific
effector T cells appear to play a crucial role in restricting the generation and
expansion of transformed neoplastic cells. However, the fact that tumours continue to
grow in the presence of a seemingly intact immune system suggests that these
responses are often inadequate.
CD4+Foxp3+ T regulatory cells (Tregs) have been shown to play a key role in
modulating the immune system by keeping immune responses to self-antigens in
check, thereby preventing autoimmunity. These cells also appear to be employed by
tumours to protect against recognition and eradication, and have been demonstrated to
impinge upon the anti-tumour immune response in humans. Furthermore, it appears
that the tumour microenvironment facilitates the development of highly
immunosuppressive T cells, which may also allow subsequent tumour progression.
In colorectal cancer, the relationship between Tregs and tumour progression is
less clear – despite their well-documented ability to impinge on anti-tumour immune
responses, increased tumour infiltrates have also been associated with prolonged
survival. In this thesis, the phenotype and function of CD4+ T cells derived from
PBMC, colon and tumour samples were analysed for suppressive markers by FACS,
and anti-tumour responses by IFN-γ ELISpot. CRC patients with more advanced
tumours responded to fewer epitopes and generated a significantly weaker epitopespecific
T cell response to the oncofoetal antigen, 5T4 than healthy donors. Human
depletion experiments both in vitro and in vivo indicated suppression by Foxp3+
regulatory CD4+ T cells. These cells were found in abundance amongst tumourinfiltrating
lymphocytes; however, another equally prominent population of IL-10 and
TGF-β-producing CD4+Foxp3- T cells were found to be >50-fold more suppressive.
Thus, a major caveat to cancer immunotherapy is the suppressive tumour
microenvironment, which contributes to the selective decline of measurable antitumour
CD4+ T cell responses as tumours progress. These responses were enhanced
in metastatic CRC patients by depleting regulatory T cells.
It is hoped such findings will augment our understanding of how anti-tumour
CD4+ T cells are activated and conversely regulated, with the intention of designing
better treatment for patients with colorectal cancer
Highly prevalent colorectal cancer-infiltrating LAP+ Foxp3- T cells exhibit more potent immunosuppressive activity than Foxp3+ regulatory T cells
Although elevated CD4+Foxp3+ regulatory T cell (Treg) frequencies within tumors are well documented, the functional and phenotypic characteristics of CD4+Foxp3+ and CD4+Foxp3− T cell subsets from matched blood, healthy colon, and colorectal cancer require in-depth investigation. Flow cytometry revealed that the majority of intratumoral CD4+Foxp3+ T cells (Tregs) were Helios+ and expressed higher levels of cytotoxic T-lymphocyte antigen 4 (CTLA-4) and CD39 than Tregs from colon and blood. Moreover, ~30% of intratumoral CD4+Foxp3− T cells expressed markers associated with regulatory functions, including latency-associated peptide (LAP), lymphocyte activation gene-3 (LAG-3), and CD25. This unique population of cells produced interleukin-10 (IL-10) and transforming growth factor-β (TGF-β), and was ~50-fold more suppressive than Foxp3+ Tregs. Thus, intratumoral Tregs are diverse, posing multiple obstacles to immunotherapeutic intervention in colorectal malignancies
Prevalence and architecture of de novo mutations in developmental disorders.
The genomes of individuals with severe, undiagnosed developmental disorders are enriched in damaging de novo mutations (DNMs) in developmentally important genes. Here we have sequenced the exomes of 4,293 families containing individuals with developmental disorders, and meta-analysed these data with data from another 3,287 individuals with similar disorders. We show that the most important factors influencing the diagnostic yield of DNMs are the sex of the affected individual, the relatedness of their parents, whether close relatives are affected and the parental ages. We identified 94 genes enriched in damaging DNMs, including 14 that previously lacked compelling evidence of involvement in developmental disorders. We have also characterized the phenotypic diversity among these disorders. We estimate that 42% of our cohort carry pathogenic DNMs in coding sequences; approximately half of these DNMs disrupt gene function and the remainder result in altered protein function. We estimate that developmental disorders caused by DNMs have an average prevalence of 1 in 213 to 1 in 448 births, depending on parental age. Given current global demographics, this equates to almost 400,000 children born per year
Bi-allelic Loss-of-Function CACNA1B Mutations in Progressive Epilepsy-Dyskinesia.
The occurrence of non-epileptic hyperkinetic movements in the context of developmental epileptic encephalopathies is an increasingly recognized phenomenon. Identification of causative mutations provides an important insight into common pathogenic mechanisms that cause both seizures and abnormal motor control. We report bi-allelic loss-of-function CACNA1B variants in six children from three unrelated families whose affected members present with a complex and progressive neurological syndrome. All affected individuals presented with epileptic encephalopathy, severe neurodevelopmental delay (often with regression), and a hyperkinetic movement disorder. Additional neurological features included postnatal microcephaly and hypotonia. Five children died in childhood or adolescence (mean age of death: 9 years), mainly as a result of secondary respiratory complications. CACNA1B encodes the pore-forming subunit of the pre-synaptic neuronal voltage-gated calcium channel Cav2.2/N-type, crucial for SNARE-mediated neurotransmission, particularly in the early postnatal period. Bi-allelic loss-of-function variants in CACNA1B are predicted to cause disruption of Ca2+ influx, leading to impaired synaptic neurotransmission. The resultant effect on neuronal function is likely to be important in the development of involuntary movements and epilepsy. Overall, our findings provide further evidence for the key role of Cav2.2 in normal human neurodevelopment.MAK is funded by an NIHR Research Professorship and receives funding from the Wellcome Trust, Great Ormond Street Children's Hospital Charity, and Rosetrees Trust. E.M. received funding from the Rosetrees Trust (CD-A53) and Great Ormond Street Hospital Children's Charity. K.G. received funding from Temple Street Foundation. A.M. is funded by Great Ormond Street Hospital, the National Institute for Health Research (NIHR), and Biomedical Research Centre. F.L.R. and D.G. are funded by Cambridge Biomedical Research Centre. K.C. and A.S.J. are funded by NIHR Bioresource for Rare Diseases. The DDD Study presents independent research commissioned by the Health Innovation Challenge Fund (grant number HICF-1009-003), a parallel funding partnership between the Wellcome Trust and the Department of Health, and the Wellcome Trust Sanger Institute (grant number WT098051). We acknowledge support from the UK Department of Health via the NIHR comprehensive Biomedical Research Centre award to Guy's and St. Thomas' National Health Service (NHS) Foundation Trust in partnership with King's College London. This research was also supported by the NIHR Great Ormond Street Hospital Biomedical Research Centre. J.H.C. is in receipt of an NIHR Senior Investigator Award. The research team acknowledges the support of the NIHR through the Comprehensive Clinical Research Network. The views expressed are those of the author(s) and not necessarily those of the NHS, the NIHR, Department of Health, or Wellcome Trust. E.R.M. acknowledges support from NIHR Cambridge Biomedical Research Centre, an NIHR Senior Investigator Award, and the University of Cambridge has received salary support in respect of E.R.M. from the NHS in the East of England through the Clinical Academic Reserve. I.E.S. is supported by the National Health and Medical Research Council of Australia (Program Grant and Practitioner Fellowship)
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The contribution of X-linked coding variation to severe developmental disorders
Abstract: Over 130 X-linked genes have been robustly associated with developmental disorders, and X-linked causes have been hypothesised to underlie the higher developmental disorder rates in males. Here, we evaluate the burden of X-linked coding variation in 11,044 developmental disorder patients, and find a similar rate of X-linked causes in males and females (6.0% and 6.9%, respectively), indicating that such variants do not account for the 1.4-fold male bias. We develop an improved strategy to detect X-linked developmental disorders and identify 23 significant genes, all of which were previously known, consistent with our inference that the vast majority of the X-linked burden is in known developmental disorder-associated genes. Importantly, we estimate that, in male probands, only 13% of inherited rare missense variants in known developmental disorder-associated genes are likely to be pathogenic. Our results demonstrate that statistical analysis of large datasets can refine our understanding of modes of inheritance for individual X-linked disorders
Heterozygous Variants in KMT2E Cause a Spectrum of Neurodevelopmental Disorders and Epilepsy.
We delineate a KMT2E-related neurodevelopmental disorder on the basis of 38 individuals in 36 families. This study includes 31 distinct heterozygous variants in KMT2E (28 ascertained from Matchmaker Exchange and three previously reported), and four individuals with chromosome 7q22.2-22.23 microdeletions encompassing KMT2E (one previously reported). Almost all variants occurred de novo, and most were truncating. Most affected individuals with protein-truncating variants presented with mild intellectual disability. One-quarter of individuals met criteria for autism. Additional common features include macrocephaly, hypotonia, functional gastrointestinal abnormalities, and a subtle facial gestalt. Epilepsy was present in about one-fifth of individuals with truncating variants and was responsive to treatment with anti-epileptic medications in almost all. More than 70% of the individuals were male, and expressivity was variable by sex; epilepsy was more common in females and autism more common in males. The four individuals with microdeletions encompassing KMT2E generally presented similarly to those with truncating variants, but the degree of developmental delay was greater. The group of four individuals with missense variants in KMT2E presented with the most severe developmental delays. Epilepsy was present in all individuals with missense variants, often manifesting as treatment-resistant infantile epileptic encephalopathy. Microcephaly was also common in this group. Haploinsufficiency versus gain-of-function or dominant-negative effects specific to these missense variants in KMT2E might explain this divergence in phenotype, but requires independent validation. Disruptive variants in KMT2E are an under-recognized cause of neurodevelopmental abnormalities
PMO-088 A large proportion of colorectal tumour-infiltrating CD4+ T cells are suppressive irrespective of FOXP3 expression
Introduction The presence of increased numbers of CD3+ T cells in colorectal cancer (CRC) correlates with improved prognosis. However, it is difficult to measure anti-tumour responses in tumour-infiltrating lymphocytes (TILs) suggesting these cells are suppressed. Although we have demonstrated CD4+Foxp3+ regulatory T cells (Tregs) within the tumour and its stroma, the numbers are often low. We sought to identify phenotypic and functional characteristics of CD4+Foxp3− T cells to determine whether other regulatory populations exist within this environment.
Methods Tumour samples were obtained from CRC patients with different stages of malignancy. Fixed tumour samples were examined by immunofluoresence for CD3, CD8 and FoxP3. TILs from fresh tumour tissue were stained with a panel of 20 antibodies (including Helios, LAG-3, LAP) and examined by FACS.
Results Histology revealed tumours to be infiltrated by CD4+, CD8+ and Foxp3+ positive cells. Despite an increase in CD4+ and CD8+ T cells in advanced tumours, there was not always a concomitant increase in Foxp3+ cells. Flow cytometry revealed the majority of the Treg fraction was Helios+ (indicating thymically-derived) and expressed higher levels of CTLA-4 and CD39 than Tregs from colon and blood. However, 30% of “conventional” CD4+Foxp3− T cells express markers associated with Tregs including LAP (latency-associated peptide), LAG-3 and CD25 and were highly suppressive in vitro.
Conclusion Tumour-infiltrating CD4+ T cells are heterogeneous. A high percentage of these cells appear to have a regulatory function and include both Foxp3+ as well as FoxP3− T cells. Overcoming the suppressive environment of CRC is a major challenge for boosting anti-tumour immunity.
Competing interests None declared
T cell subsets and colorectal cancer: discerning the good from the bad
Tumor-specific T cells must overcome a multitude of suppressive mechanisms to destroy cancerous cells effectively. Furthermore, it appears that the tumor microenvironment facilitates the development of highly immunosuppressive T cells, which may also allow subsequent tumor progression. In colorectal cancer, the relationship between regulatory T cells (e.g. FoxP3(+) Tregs) and tumor prognosis and progression is less clear, despite their well-documented ability to impinge on anti-tumor immune responses. Here we explore our current knowledge of colorectal TIL heterogeneity, deciphering subsets which may be of benefit or detriment