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BRCA2’s Relation with Aldehydes Leads to a Higher Risk of Developing Prostate Cancer Than BRCA1

This retrospective analysis research paper discusses why the difference between the responses of aldehyde exposure from patients with a heterozygous germline mutation in BRCA2 and BRCA1 leads to a higher risk of prostate cancer in BRCA2 mutant carriers than BRCA1. 

Abstract

Although two of the most significant breast cancer susceptibility proteins, BRCA1 and BRCA2, interact with each other in the same DNA damage repair (DDR) pathway (BRCA1 works in checkpoint activation and DNA repair, and BRCA2 plays a major role in homologous recombination), they react differently when exposed to aldehydes. Aldehydes, RCHO, have a more significant impact on heterozygous BRCA2  as mutation carriers than heterozygous BRCA1 mutation carriers because they directly interact with the BRCA2 proteins. Workers in the chemical industry, alcohol drinkers, and users of products with aldehydes in them are all at a higher risk of being exposed to aldehydes, specifically formaldehyde. Aldehydes directly interact with BRCA2 because the formaldehyde destabilizes and stalls DNA replication forks, increasing the genomic instability which increases mutational risk. On the other hand, aldehydes do not directly interact with BRCA1, so there is no increase in the risk of prostate cancer for heterozygous BRCA1 carriers. In addition, studies have shown that heterozygous BRCA1 carriers’ primary cells are defective in a stalled replication fork repair function, so aldehydes might not have much of an impact on them. This retrospective analysis research paper discusses why the difference between the responses of aldehyde exposure from patients with a heterozygous germline mutation in BRCA2 and BRCA1 leads to a higher risk of prostate cancer in BRCA2 mutant carriers than BRCA1

Introduction

Faulty BRCA1/2 genes are two of the most common causes of developing breast, prostate, pancreatic, and other types of cancer. They are genes that guide the production of tumor suppressor proteins. Everyone is born with two copies of this gene, one from each parent, and variants in these genes can cause cancer at an early age. Even if you inherit a faulty copy of the gene from one of your parents, you can still possess a good copy because embryos can not develop if both copies happen to be mutated. Overall, BRCA1/2’s primary function is DDR (1). Wildtype BRCA1 and BRCA2 genes both participate in mitophagy, cell cycle checkpoints regulation, transcriptional regulation, and homologous recombination (2). Although BRCA1 and BRCA2 have many similarities, there are also some differences in their functions. Unlike BRCA2, BRCA1 plays a vital role in embryonic development (3) and is a co-regulator of androgen receptors (4). BRCA1 works with RAD50, NBS, MRE1, and RAD51 for DNA repair, BARD1 and BAP5 for the ubiquitination cycle, HDAC and RB for chromatin structure regulation, CHK1, CHK2, TPX2, and NuMA for cell cycle regulation, and c-Myc, ZBRK1, E2F1, and CtlP for transcriptional regulation (5). It controls spindle formation and centromere numbers (6). It is also a pleiotropic DNA damage response protein meaning it has a role in DDR and makes sure the cell does not go through the cell cycle with mistakes in the DNA (7). Early on in homologous recombination, BRCA1 works with nucleases and coordinates DNA end resection to form the single-stranded DNA (8). BRCA1 carriers are also most likely to develop triple-negative breast cancer (9). BRCA2’s role is to regulate RAD51, a key enzyme in homologous recombination, filament formation and activity, and cytokinesis (10) (11). BRCA2 also takes part in telomere homeostasis, cell cycle regulation, chromosome segregation during mitosis, facilitating mitophagy, and many more functions in the cytoplasm (12). This gene plays a vital role in the stabilization of the replication fork as well (2). Mutations in BRCA2 are the most common cause of prostate cancer (13). Prostate cancer is the second most common neoplasm and is the second most common cancer in men. It usually occurs in older men as it is the most common risk factor and forms in the prostate tissue, a gland in the male reproductive system (14). Thirteen out of every one hundred American men develop prostate cancer, and about two to three of them will die from it. African-Americans have the highest chance of developing prostate cancer with two times the rate of mortality of other men. Family history is also associated with a higher risk of developing prostate cancer. Some examples include having more than one first-degree family member with prostate cancer, being diagnosed when you were fifty-five or younger, or being diagnosed while other family members had breast, ovarian, or pancreatic cancer. Doctors have used some methods to treat prostate cancer: a prostatectomy, removing the prostate, and external or internal radiation therapy. (15) Prostate cancer can spread outside of the prostate gland, which can cause it to be very deadly. (16) BRCA1 mutation carriers have a lower risk of developing prostate cancer at about 3.4%. (17). However, together, BRCA1/2 mutation carriers have anywhere from an 11% to 33% increased risk of developing prostate cancer. (18) Aldehydes have been suspected to play a role in a higher risk of prostate cancer when associated with mutated BRCA2 genes versus BRCA1. Aldehydes are reactive and organic compounds that can be used as building blocks to build other chemicals that make products like perfumes, resins, dyes, detergent, soap, and some other organic acids (19). Aldehydes can be cytotoxic, mutagenic, and carcinogenic. Too much aldehyde exposure can lead to lousy aldehyde metabolism, which could potentially cause cancer (20). Too many aldehydes in your body can lead to damaged mucous membranes, scarred tissue, headaches, a hangover-like state, faster heartbeat, and stomach problems. In addition, compared to other tissues, acetaldehydes have the most effect on your brain and can cause memory loss (21). They can also oxidize to form carboxylic acids, making them irritate your skin (22). Aldehydes can be found in the air because of pollution from automobiles, industrial waste, fossil fuels being burnt, paint, and surgical smoke. Aldehydes can also be present in food or drinks and be manufactured (19) (23). Out of everything, aldehydes are most likely found in environmental sources and natural objects like cinnamaldehyde, vanillin, roses, citronella, vanilla, orange rind, and acrolein (23) (19). They can be formed within your system without outside exposure by lipid peroxidation, carbohydrate or metabolism ascorbate autoxidation, amine oxidases, cytochrome P-450s, or myeloperoxidase-catalyzed metabolic activation (24). We need to understand aldehydes’ impact on BRCA1 and BRCA2 because, from this information, we can understand a potential cause for why BRCA2 mutant carriers are more likely to develop prostate cancer. It is important to know why patients with BRCA2 mutants are more likely to develop prostate cancer because understanding the difference could potentially inform treatment approaches and help carriers avoid certain lifestyles. Knowing that BRCA2 mutations play a significant role in prostate cancer will push people to be more cautious, and beyond that, if they know why it is more likely to cause prostate cancer than BRCA1 mutations, they will know what to avoid. 

Background

Aldehydes have a more significant impact on BRCA2 than BRCA1. Formaldehyde exposure causes heterozygous BRCA2 truncations to be sensitive to BRCA2 haploinsufficiency. People who inherit a faulty BRCA2 copy have reduced BRCA2 protein levels. The levels go down below the amount that is required for sufficient DNA repair (25). One way to help fight against the problems caused by formaldehyde is the use of ribonuclease H11. Ribonuclease H11 helps improve the instability in the replication fork and chromosomal aberrations. BRCA2 not functioning correctly causes mutagenesis during DNA replication through R-loops (26). Aldehyde dehydrogenase (ALDH2) is a mitochondrial enzyme that is one of the most strongly associated genes with alcoholism. ALDH2 decreases the risk of alcohol toxicity by increasing acetaldehyde levels for one of two reasons: either the acetaldehyde is oxidized slower, or the ethanol is oxidized faster. When a variant of ALDH is produced, it can cause an imbalance between acetaldehyde and ethanol oxidation, leading to changes in the acetaldehyde concentration (27). ALDH2 metabolizes aldehydes to make them less toxic, and if you are frequently exposed to aldehydes and have a poor aldehyde metabolism, which could happen by inheriting genetic variants of ALDH2, then you have the potential risk of developing cancer. Exposure to aldehydes causes BRCA2 proteins to lose their function and break down our body’s defense mechanism, meaning that they cannot repair DNA. Aldehydes cause BRCA2 proteins to lose their function because of the replicative stress caused by formaldehyde. Aldehydes induce DNA damage that BRCA2 haploinsufficiency cannot repair, which leads to more mutagenesis (and eventual cancer development). Since BRCA2 genes are extra sensitive to aldehydes compared to other proteins, it is common for there to be a disruption in their function. A test has been done in which they used two cell line models, genetically engineered cells and cells from patients with a faulty copy of the BRCA2 gene, to understand the effect of aldehydes on BRCA2 (25). Oxidative metabolism produces replication stress which triggers genetic instability from BRCA1/2 (7). ALDH metabolizes internal and external aldehydes and mitigates oxidative stress. It also helps abrogate oxidative stress and helps it resist against chemotherapeutic agents (28). This shows that although aldehydes can be extremely dangerous, ALDH is actually helping BRCA2 proteins away from oxidative stress. Heterozygous BRCA2 mutations that are also exposed to formaldehyde have more genomic instability because of the replicative stress that the formaldehyde causes. Formaldehyde exposure also induces selective proteasomal degradation of BRCA2, leading to an unstable DNA replication fork that causes stalled DNA (29). One more thing that formaldehyde exposure causes are induced haploinsufficiency. Heterozygous BRCA2 carriers have BRCA2 protein levels that are already down by 50%, and exposure to aldehydes makes it go down by another 20%, causing the induced haploinsufficiency. (8) BRCA1 mutations are also sensitive to acetaldehyde because of defective homologous recombination. However, when BRCA1 has secondary mutations, it allows it to have proficient homologous recombination. (30) Similar to BRCA2 mutation carriers, when BRCA1 is exposed to formaldehyde, DNA damage and stalled DNA replication results. However, the cells of heterozygous BRCA1 carriers are defective in a stalled replication fork repair function. Formaldehyde also causes a defect in the DNA replication fork. (8) This may explain the reason behind aldehydes having a greater impact on BRCA2 mutant carriers. Since BRCA1 carriers already have a defect in the stalled replication fork repair function, formaldehyde does not change anything for them as they are already adapted to compensate for this loss, and therefore will not make it any worse. This is extremely important in understanding the difference between BRCA1/2 when associated with aldehydes. If BRCA1 is unaffected, and BRCA2 is affected, then BRCA2 will be more likely to contribute to prostate cancer (6). The information gathered in turn means that cells with BRCA1 mutations are already able to adapt to the type of replication fork stalling that is induced by aldehydes, so they are better adapted to withstand that kind of effect versus BRCA2 mutant cells that do not have adaptive mechanisms to deal with the addition of stalled replication forks, leading to more mutagenesis and increased risk for cancer.

Method

This research paper is a retrospective analysis meaning the conclusion was discovered based on pre-existing research. BRCA2 mutation carriers have a higher chance of developing prostate cancer than BRCA1 mutation carriers. Why though? Especially since they interact with each other in the same pathway, why does a mutation in one increase the risk of developing prostate cancer over the other? To figure out the answers to these questions, I started by reading about our key players, which are BRCA1, BRCA2, and prostate cancer. After going through many primary literature articles and reviews, I started to understand the similarities and differences between the two types of breast cancer genes. I used PubMed as my search engine. As I continued researching, I came across a review about an experiment conducted by Professor Ashok Venkatraman about aldehydes concerning BRCA2 and prostate cancer. After I looked deeper into this, I realized that there was definitely a connection here, but what about BRCA1 and aldehydes? There is no direct relation between BRCA1 and aldehydes, but they perform a similar function when BRCA1 is mutated, which could explain why aldehydes do not affect them. By the time I was done performing my research, I had been through multiple resources about aldehydes, prostate cancer, BRCA1/2, and all of their connections.

Results

Due to an experiment led by Professor Ashok Venkatraman, it can be concluded that aldehydes cause BRCA2 proteins to lose their function and not be able to repair DNA causing cancer. He and his research group used two cell line models: genetically engineered cells and cells from patients with faulty copies of the BRCA2 gene to understand their connection. He overexpressed BRCA2. (25). Their first attempt was to recapitulate the sensitivity by knocking out BRCA using CRISPR/Cas9. They took a cell line that did not have this mutation and examined its response to aldehydes. From this test, they learned that it was not very sensitive. However, if they were to mutate or knock down the gene, then it sensitized the cells to aldehydes causing them to die. When you have a cell line that does not have a BRCA2 deficiency, the cells are fine even if they are exposed to aldehydes. However, if they do have a BRCA2 deficiency, then they are sensitive to aldehydes. When you want to test the function of a protein, you should consider what happens when you knock it down, or if it is knocked down, and what it becomes sensitive to that it might not have been sensitive to before. They looked at models in which it was not deficient and was deficient (has the mutation), and because they were genetically engineered, they were able to control the expression of both BRCA1 and BRCA2 using a DOX inducible system. Within the DNA, they genetically engineered a section and inserted a gene called a siRNA (short interrupting RNA). They genetically engineered a siRNA that was for BRCA1 and BRCA2 (two different siRNAs). siRNA is under doxycycline inducer, so after you have inserted this gene, treating the cells with doxycycline (an antibiotic) turns on transcription of whatever the gene is under the DOX inducible promoter. This will only be expressed and made into a protein if you use doxycycline. After this, the siRNA is going to bind to the mRNA and form a double-stranded RNA. The siRNA is particular and perfectly matches BRCA1/2. Cells do not like double-stranded RNA, so when it happens, it gets destroyed by a proteasome. This system is how they knocked down BRCA1/2. It is not permanent, and the expression is not entirely stopped. The amount of expression is described by how much DOX you use. First, they knocked down BRCA1 in a dish of cells, and then they treated it with aldehydes. They had a parental cell line (control group) and their treatment groups. They tested just BRCA1 and then just BRCA2 in terms of aldehydes versus no aldehydes. They then compared these results and were able to say that aldehydes impacted the mutants much more significantly than they did the parental line. The vehicle, whatever your treatment is dissolved in (DMSO in this case), and the other is the treatment group (aldehydes). Through the tests, we see that BRCA2‘s viability percentage gets lower and lower as it has higher exposure to aldehydes. BRCA1‘s percentage also goes down, but not as much as BRCA2‘s. (31) They looked at the viability under these two treatment groups and compared the results. The first main conclusion from the studies includes that cells that are homologous recombination deficient are hypersensitive to acetaldehyde. The next one is that an ALDH inhibitor known as disulfiram aims for BRCA1/2 deficient cells. Acetaldehyde and disulfiram induce severe replication stress in BRCA2. The acetaldehyde hypersensitivity of the cells that lack BRCA2 leads to non-effective DNA replication, checkpoint activation, G2/M arrest, and apoptosis. Acetaldehyde treatment invoked homologous repair, potentially explaining the reduced survival rate of cells without BRCA2. The mouse models used for the experiment show us that acetaldehyde treatment prevents growth in tumors deficient in the BRCA1/2 proteins. Upon treatment by acetaldehydes or disulfiram, the replication was significantly reduced with the absence of BRCA2. The use of disulfiram is good because if you treat it with an aldehyde inhibitor, it will not be able to create the breaks in DNA that need to be repaired. They used an alcohol aversive agent because it almost serves as a treatment. It only kills the cancer cells that are deficient (leads to selective killing of BRCA2 deficient cells). They could use this as a potential therapy. Even though people at a higher risk of exposure may develop cancer because it leads to a higher risk of gaining a mutation, you could treat it and cause so much DNA damage that the cells do not just cause mutations, but they have so many breaks that they die. 

(32)

Discussion

The results from this experiment allowed me to hypothesize that although BRCA1 and BRCA2 interact in the same pathway, they do not react the same when exposed to aldehydes. Exposure to aldehydes causes our bodies’ defense mechanisms to stop working, so the damaged DNA cannot be repaired. Although this can happen in any cell, BRCA2 genes are more sensitive to aldehydes, so they are more affected. Aldehydes are more likely to affect BRCA2 because although BRCA1/2 are both affected by aldehydes, acetaldehyde and disulfiram induce severe replication stress in BRCA2. One problem with this experiment is that overexpression is imperfect since you end up having more copies of that gene in the cell than you would in a clinical setting, so you may get some off-target effects. After this experience, some more things that should be looked into are how we can prevent aldehydes from affecting BRCA2 mutation carriers and what we can do to treat people who have been affected. It is also essential to understand how we can prevent people from being exposed to aldehydes in general. Aldehyde exposure can lead to way more problems than prostate cancer, so we must consider all possibilities. In conclusion, aldehyde exposure is hazardous in that it affects BRCA2 mutation carriers even more and leads them to be more likely to develop prostate cancer than BRCA1 mutation carriers. 

Nihitha Reddy, Youth Medical Journal 2022

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By Nihitha Reddy

Hi, my name is Nihitha Reddy! I am a sophomore at Tahanto Regional High School in central Massachusetts. I am interested in the field of public health.

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