Epigenomics And Cancer Heterogeneity

There can be considerable heterogeneity in cancers, and this can contribute to progression and resistance to therapy.  This heterogeneity can arise not only from DNA sequence changes (i.e., mutation) but also from epigenetic changes - changes in the modifications of chromosomes - on a single cell level, making one cell different than another with respect to gene expression, even if the gene sequences remain the same.  A paper of interest, abstract:

Cell-to-cell heterogeneity is a major driver of cancer evolution, progression, and emergence of drug resistance. Epigenomic variation at the single-cell level can rapidly create cancer heterogeneity but is difficult to detect and assess functionally.

RESULTS: We develop a strategy to bridge the gap between measurement and function in single-cell epigenomics. Using single-cell chromatin accessibility and RNA-seq data in K562 leukemic cells, we identify the cell surface marker CD24 as co-varying with chromatin accessibility changes linked to GATA transcription factors in single cells. Fluorescence-activated cell sorting of CD24 high versus low cells prospectively isolated GATA1 and GATA2 high versus low cells. GATA high versus low cells express differential gene regulatory networks, differential sensitivity to the drug imatinib mesylate, and differential self-renewal capacity. Lineage tracing experiments show that GATA/CD24hi cells have the capability to rapidly reconstitute the heterogeneity within the entire starting population, suggesting that GATA expression levels drive a phenotypically relevant source of epigenomic plasticity.
CONCLUSION: Single-cell chromatin accessibility can guide prospective characterization of cancer heterogeneity. Epigenomic subpopulations in cancer impact drug sensitivity and the clonal dynamics of cancer evolution.

Identifying these epigenetic changes can be the first step in developing novel approaches to reverse the changes and/or target those cells containing such changes, for therapeutic benefit.

Colon Cancer Mutations Associated With The Gut Microbiota

An extremely important study on colon cancer concludes: “We found statistically significant associations between loss-of-function mutations in tumor genes and shifts in the abundances of specific sets of bacterial taxa, suggestive of potential functional interaction.”  Further, the correlation between bacterial types and tumor mutations allowed for predictions about interactions between the mutations and the downstream signaling pathways affected by these mutations.  One can expect future work to more clearly delineate the molecular mechanisms of interaction between the microbiota and cancer mutations.  In the meantime, once again – for the thousandth time it seems – the importance of the gut microbiota is emphasized.  Abstract:

Variation in the gut microbiome has been linked to colorectal cancer (CRC), as well as to host genetic variation. However, we do not know whether, in addition to baseline host genetics, somatic mutational profiles in CRC tumors interact with the surrounding tumor microbiome, and if so, whether these changes can be used to understand microbe-host interactions with potential functional biological relevance. Here, we characterized the association between CRC microbial communities and tumor mutations using microbiome profiling and whole-exome sequencing in 44 pairs of tumors and matched normal tissues. We found statistically significant associations between loss-of-function mutations in tumor genes and shifts in the abundances of specific sets of bacterial taxa, suggestive of potential functional interaction. This correlation allows us to statistically predict interactions between loss-of-function tumor mutations in cancer-related genes and pathways, including MAPK and Wnt signaling, solely based on the composition of the microbiome. In conclusion, our study shows that CRC microbiomes are correlated with tumor mutational profiles, pointing towards possible mechanisms of molecular interaction.

Three Is Enough

Colorectal cancer is a prime example of the multiple hit hypothesis, a cancer that is driven by the accumulation of sequential mutations in key genes.  Here is a study in mice asserting that only three mutations in key pathways were sufficient to drive a metastatic form of this cancer in the animal model.  Abstract:

Colorectal cancer (CRC) is driven by the accumulation of driver mutations, but the contributions of specific mutations to different steps in malignant progression is not fully understood. In this study, we generated mouse models harboring different combinations of key CRC driver mutations (Apc, Kras, Tgfbr2, Trp53, Fbxw7) in intestinal epithelial cells to comprehensively investigate their roles in the development of primary tumors and metastases. Apc∆716 mutation caused intestinal adenomas and combination with Trp53R270H mutation or Tgfbr2 deletion induced submucosal invasion. The addition of KrasG12D mutation yielded EMT-like morphology and lymph vessel intravasation of the invasive tumors. In contrast, combinations of Apc∆716 with KrasG12D and Fbxw7 mutation was insufficient for submucosal invasion but still induced EMT-like histology. Studies using tumor-derived organoids showed that KrasG12D was critical for liver metastasis following splenic transplantation, when this mutation was combined with either Apc∆716 plus Trp53R270H or Tgfbr2 deletion, with the highest incidence of metastasis displayed by tumors with a Apc∆716 KrasG12D Tgfbr2-/- genotype. RNAseq analysis of tumor organoids defined distinct gene expression profiles characteristic for the respective combinations of driver mutations, with upregulated genes in Apc∆716 KrasG12D Tgfbr2-/- tumors found to be similarly upregulated in specimens of human metastatic CRC. Our results show how activation of Wnt and Kras with suppression of TGF-β signaling in intestinal epithelial cells is sufficient for CRC metastasis, with possible implications for the development of metastasis prevention strategies.

Tumor Mutations and anti-PD-1 Immunotherapy Response

Inhibitors of factors that themselves inhibit an anti-tumor response are promising therapeutic agents.  Here we see once again that response to such agents is related to the number of mutations a tumor has.  The more mutations, the more neoantigens produced – the more aberrant proteins produced that can be recognized by the immune system once the “brakes” are off.  Excerpts (figure shown is from the paper, which is freely accessible online):

Inhibitors of programmed death 1 (PD-1) protein or its ligand (PD-L1) have shown remarkable clinical benefit in many cancers. One emerging biomarker of response to anti–PD-1 therapy is the tumor mutational burden (i.e., the total number of mutations per coding area of a tumor genome). This finding is supported by the clinical activity of anti–PD-1 therapy in colorectal cancer with mismatch repair deficiency, a tumor subtype with a high tumor mutational burden, as compared with the colorectal cancer subtype with mismatch repair proficiency, which has a significantly lower tumor mutational burden and a poor response to these agents. 

To evaluate the relationship between the tumor mutational burden and the objective response rate, we plotted the objective response rate for anti–PD-1 or anti–PD-L1 therapy against the corresponding median tumor mutational burden across multiple cancer…We observed a significant correlation between the tumor mutational burden and the objective response rate (P<0.001). The correlation coefficient of 0.74 suggests that 55% of the differences in the objective response rate across cancer types may be explained by the tumor mutational burden. 

Neoantigen Immunotherapy

T cell is the blue one on the right.

By Electron Microscopy Facility at The National Cancer Institute at Frederick (NCI-Frederick) - [1], Public Domain, https://commons.wikimedia.org/w/index.php?curid=407197

The mutations that accumulate in cancer can cause the production of abnormal protein products that could in theory be recognized by the immune system to fight cancer.  This is a field of study in cancer immunotherapy, and today we would like to link to two papers that show some potential advances in this field.  The first one is here. Abstract:

Effective anti-tumour immunity in humans has been associated with the presence of T cells directed at cancer neoantigens, a class of HLA-bound peptides that arise from tumour-specific mutations. They are highly immunogenic because they are not present in normal tissues and hence bypass central thymic tolerance. Although neoantigens were long-envisioned as optimal targets for an anti-tumour immune response, their systematic discovery and evaluation only became feasible with the recent availability of massively parallel sequencing for detection of all coding mutations within tumours, and of machine learning approaches to reliably predict those mutated peptides with high-affinity binding of autologous human leukocyte antigen (HLA) molecules. We hypothesized that vaccination with neoantigens can both expand pre-existing neoantigen-specific T-cell populations and induce a broader repertoire of new T-cell specificities in cancer patients, tipping the intra-tumoural balance in favour of enhanced tumour control. Here we demonstrate the feasibility, safety, and immunogenicity of a vaccine that targets up to 20 predicted personal tumour neoantigens. Vaccine-induced polyfunctional CD4+ and CD8+ T cells targeted 58 (60%) and 15 (16%) of the 97 unique neoantigens used across patients, respectively. These T cells discriminated mutated from wild-type antigens, and in some cases directly recognized autologous tumour. Of six vaccinated patients, four had no recurrence at 25 months after vaccination, while two with recurrent disease were subsequently treated with anti-PD-1 (anti-programmed cell death-1) therapy and experienced complete tumour regression, with expansion of the repertoire of neoantigen-specific T cells. These data provide a strong rationale for further development of this approach, alone and in combination with checkpoint blockade or other immunotherapies.

And here is another paper on the same subject. Abstract:

T cells directed against mutant neo-epitopes drive cancer immunity. However, spontaneous immune recognition of mutations is inefficient. We recently introduced the concept of individualized mutanome vaccines and implemented an RNA-based poly-neo-epitope approach to mobilize immunity against a spectrum of cancer mutations. Here we report the first-in-human application of this concept in melanoma. We set up a process comprising comprehensive identification of individual mutations, computational prediction of neo-epitopes, and design and manufacturing of a vaccine unique for each patient. All patients developed T cell responses against multiple vaccine neo-epitopes at up to high single-digit percentages. Vaccine-induced T cell infiltration and neo-epitope-specific killing of autologous tumour cells were shown in post-vaccination resected metastases from two patients. The cumulative rate of metastatic events was highly significantly reduced after the start of vaccination, resulting in a sustained progression-free survival. Two of the five patients with metastatic disease experienced vaccine-related objective responses. One of these patients had a late relapse owing to outgrowth of β2-microglobulin-deficient melanoma cells as an acquired resistance mechanism. A third patient developed a complete response to vaccination in combination with PD-1 blockade therapy. Our study demonstrates that individual mutations can be exploited, thereby opening a path to personalized immunotherapy for patients with cancer.

The Vices Of Men

By Vincent van Gogh - Scanned from Smoke: a global history of smoking (2004) ISBN 1-86189-200-4, Public Domain, https://commons.wikimedia.org/w/index.php?curid=3857076

The paper linked here is a fascinating review that looks at three major vices of men – drinking alcohol, being overweight/obese, and smoking tobacco – and how these activities can damage male fertility and also negatively affect the health of their offspring. Mechanisms by which these vices can affect fertility and the health of children include epigenetic changes to sperm DNA and non-coding RNA (changes to the modification to DNA and RNA, not including actual sequence mutation, that can affect gene expression), sperm DNA damage (which can cause actual sequence mutation affecting gene expression), changes in sperm chromatin structure (for example, changes in the chromosome structure that can affect which genes are expressed and which are silenced), and changes in seminal plasma (that can affect the function of the sperm cells themselves).  Abstract:

There is growing evidence from animal and human studies that demonstrate that acquired paternal traits can impair both a male's fertility and the health of his offspring, including advanced age, smoking, stress, trauma, under-nutrition, infection, toxin exposure, and obesity. Curiously, many of these factors manifest as impaired neurological, behavioural, and/or metabolic functioning in offspring. The underlying molecular mechanisms that respond to the paternal environment and act as vectors of intergenerational transmission are beginning to emerge. This review focuses on three vices of men (alcohol consumption, overweight/obesity, and tobacco smoking) that damage fertility and pose risks to offspring health. These vices are not only the three most prevalent but are also leading risk factors for death and disability adjusted life years (DALYs) worldwide. Clearly, any epigenetic/genetic alterations induced by the paternal exposures responsible for transmission need to escape/bypass the substantial post-fertilisation reprogramming that occurs during embryo development. For example paternal obesity alters the molecular composition of sperm, alters the developmental trajectory of resultant embryos, and increase the incidence of obesity and metabolic disorders in offspring. Mechanistic candidates of paternal programming include changes to the sperm epigenome (eg DNA methylation, histone/protamine modifications, and sperm borne small non-coding RNAs), increased sperm DNA damage, aberrant sperm DNA chromatin structure, and components of seminal plasma. Understanding the molecular mechanisms underpinning paternal programming may lead to the development of interventions designed to reduce the disease burden in future generations, who were born to fathers exposed to these initiating factors. Given that these vices are predominantly self-inflicted, interventions aimed at mitigating their consequences are readily identified.

Add this information to all the other reasons to avoid these vices.

"Divide and conquer" - does it work in cancer care

Fotogeniss -

Philip-ii-of-macedon

"Divide and conquer" may have worked for Philip II of Macedon; however, is it an effective strategy in understanding cancer development and providing cancer care?    

Have you seen the talk by Dr. Gary Fettke, "Nutrition and Cancer - Time to Rethink"? The talk leads to the conclusion that cancer is a disease of our metabolism. The chromosomal changes, including any gene mutations, seem to be bystanders in the unfolding madness of cancer development.

However, recent research indicates that all cancer cell changes (mutations and metabolic changes), the immediate environment of the cancer cells (influenced by external environmental and internal factors), and the metabolism at the organism level are interconnected, and contribute to the neoplastic growth.

It is not any single element, it is rather the congregation of factors that allows for the abnormality of cancer to emerge, persist, and eventually kill.

We should be mindful of the perils of reductionism in science and cancer prevention/treatment, and try to adopt a holistic view on cancer. Here are the precise definitions of the two approaches:

Reductionism,
the practice of analyzing and describing a complex phenomenon in terms of simpler, more fundamental phenomena...

versus...

Holism,
the theory that parts of a whole are in intimate interconnection, such that they cannot exist independently of the whole, or cannot be understood without reference to the whole...

For more examples on misleading reductionism see here.



ACTIONABLES:
Yes, we should be aware of not only what we eat but how we eat it.

Yes, we should stop drinking to death, and I am not even referring to drinking alcohol.

Yes, we should learn how to cook for ourselves, since the food industry is force-feeding us with loads of sugar.

Yes, you should educate yourself, and not rely on your doctors and mainstream media to do so.

Yes, all of us should be aware that it is easier to deal with a just-born, immature troublemaker than a fully established, mature enemy; in other words, cancer prevention is more effective than cancer treatment.

Yes, we should try to integrate mainstream and alternative approaches to cancer care.

And yes, for all of you cancer researchers and enthusiasts: analyzing a phenomenon by breaking it down to its elements/building blocks and addressing only a single element is not going to bring a success. Cancer prevention and treatment need to address all aspects of the cancer development.