Tag Archives: Molecule

Simultaneous mutation of two nonessential genes can lead to the death of cancer cells | Instant News


Ludwig cancer research, has revealed a new instance in which simultaneous mutation of two nonessential genes;none of which are vital for the survival of cells can cause cancer cell death.

Headed by member of the Ludwig San Diego Richard Kolodner and published in the current The proceedings of the National Academy of Sciences, the study also shows that it is a deadly combination, or “synthetic lethality” can be reproduced in a drug-like molecule that can be used to treat cancer.

The development and FDA approval of a new generation of drugs called PARP inhibitors, for the treatment of malignant tumors with defects in tumor suppressor genes BRCA1 and BRCA2 that cause breast cancer, ovarian and many other cancers, have generated significant interest in using synthetic lethal interactions to develop cancer treatments.

Scientists, including group Kolodner, are on the hunt for other synthetic lethal interactions in cancers. “PARP inhibitors are a major step forward, but they are not perfect. Patients can become resistant to them, so there’s always a need for new and better treatments.”

Building from research done on yeast cells, Kolodner and his colleagues found that disabling or removing FEN1 gene of mammals, which is essential for DNA replication and repair, is fatal to cancer cells, mutated forms of the genes BRCA1 and 2.

We have provided information that should make people think FEN1 as a potential interesting therapeutic target and showed how yeast can be used to predict a number of synthetic lethal interactions, which can then be tested in a bona FIDE cancer cell lines with genetic instruments”.

Richard Kolodner, Professor, Professor, Department of cellular and molecular medicine, University of California, San Diego

In previous work with yeast Saccharomyces as a model to identify and study genes that maintain the integrity of the genome, Kolodner and his colleagues found that the RAD27 gene, and of synthetic lethal interactions with the 59 other nonessential genes of yeast.

Two such genes, it should be noted RAD51 and RAD52 play a role in recombination of DNA.

FEN1 is a close analogue or homologue, RAD27 in mammals. Based on their studies of yeast, Kolodner and his colleagues predicted that FEN1 synthetic lethal interactions with BRCA1 and BRCA2, which function in the same biochemical reactions in mammals, as RAD51 and RAD52 to do in yeast.

To test this hypothesis, they synthesized four FEN1-blocking molecules and used the best of them, S8, to suppress the activity of FEN1 in tumor cell lines with or without BRCA mutations. C8 proved to be an effective killer of BRCA-mutant cells.

Then they demonstrated that genetic disorders FEN1 expression had the same effect that S8 did for the breast cancer gene-mutant cells, confirming that the S8 worked, causing synthetic lethality.

Finally, the researchers instilled in C8-C8 sensitive and-resistant tumors in mice and showed that C8 significantly inhibited the growth of C8-sensitive tumors, but not in C8-resistant tumors.

Interestingly, not all cancer cell lines and tumors that responded to treatment C8 was deficient BRCA, K, indicating that FEN1 and synthetic lethal interactions with other genes as well.

These results reveal FEN1 as a novel target for drugs for the treatment of various malignant tumors by induction of synthetic lethality.

They also demonstrate that yeast-based screens provide a powerful tool to accelerate the discovery of synthetic lethal interactions for potential therapeutic value;it is an ongoing project in the laboratory Kolodner.

.



image source

Seaweed extract outperforms remdesivir in inhibiting SARS-cov-2 in studies of cells | Instant News


In the test of antiviral efficacy against the virus that causes COVID-19, an extract from edible seaweed, has exceeded remdesivir, the current standard antiviral used to combat the disease. Heparin, a common blood thinner, heparin and variant deprived of its anticoagulant properties, is performed at the same level with remdesivir in inhibiting SARS-cov-2 infection in mammalian cells.

Published online today in Opening Cellsthe study is the latest example of the lure of the strategy, researchers from the center for biotechnology and interdisciplinary studies (CBIS) at Rensselear Polytechnic Institute are developing against viruses such as the coronavirus that triggered the current global health crisis.

The spike protein on the surface of SARS-cov-2 clips on the ACE-2 receptor, a molecule on the surface of human cells. After installation, the virus inserts its genetic material into the cell by hijacking the cell machinery for the production of replica viruses. But the virus can just as easily be convince to lock the bait molecule that offers such devices. Neutralized the virus will be trapped and eventually degraded naturally.

Previous studies have shown that this technique works the decoy in the breeding of wild other viruses, including Dengue, zika and influenza A. to listen to the researchers discuss their findings by watching this video.

We learn to block viral infection, and this knowledge we’ll need if we want to confront the pandemic. The reality is that we have no good antivirals. To protect yourself against future pandemics, we need an Arsenal of approaches that we can quickly adapt to emerging viruses.”

Jonathan Dordick, the lead researcher and Professor of chemical and biological engineering rensselaer Polytechnic Institute

In Opening Cells paper tests of antiviral activity in three variants with heparin (heparin, trisulfated heparin, and without anticoagulant low molecular weight heparin) and two fucoidan (EPI-27 and RPI-28) derived from marine algae. All five compounds are long chains of sugar molecules, known as sulfated polysaccharides, structural conformation that the results of the binding study published earlier this month in anti-virus research is proposed as an effective bait.

Scientists have conducted studies of dose-response is known as the EC50 is the abbreviation for effective concentration of compound that inhibits 50% of viral infectivity — with each of the five compounds on mammalian cells. According to the results the EC50 is given in molar concentration, the lower the value of the signal more powerful composition.

IRTS-27 gave an EC50 value of approximately 83 molar, while similar to the previously published and independent in a test tube remdesivir on the same mammalian cells gave EC50 770 molar. Heparin gave EC50 of 2.1 microns, or about one-third as active as remdesivir, and the anticoagulant heparin gave similar EC50 5.0 µm, about one-fifth as active as remdesivir.

A separate test detected a toxic effect on the cells in any of the compounds, even at the highest concentration.

“What we want is a new way to get an infection,” said Robert Linhardt, Professor Rensselaer chemistry and chemical biology, which cooperates with the Dordick to develop a bait strategy. “Current thinking is that COVID-19 infection starts in the nose, and any of these substances could be the basis for a nasal spray. If you could just treat the infection, or early treatment even before you have the infection, you would have a way to block it before it enters the body”.

Added Dordick that compounds from algae “can serve as the basis for the oral approach, and delivery to address potential gastrointestinal infections.”

In the study of SARS-cov-2 sequencing data, Dordick and Linhardt identified several motifs on the structure of the spike protein, which is promised to Fit compatible with heparin, as a result, it was confirmed during the mandatory study. The spike protein is heavily encrusted in glycans, it is a fixture that protects it from human enzymes that can degrade it, and prepares it for binding with the specific receptor on the cell surface.

“This is a very complex mechanism that we, frankly, don’t know all the details, but we get more information,” said Dordick. “The only thing that was clear from this study is that the larger the molecule, the better suited you. More successful connections of large sulfated polysaccharides that offer a larger number of sites on the molecules to trap the virus.”

Molecular simulation-based binding study showed the sites on the spike protein where heparin can interact, increasing the prospects for such sulfated polysaccharides.

“This is a fascinating study of Professor Dordick and Linhardt among several ongoing studies in CBIS, as well as elsewhere in Rensselaer, for solving problems COVID-19 pandemic using new therapeutic approaches and repurpose existing drugs,” said CBIS Director Deepak Vashishth.

“Sulfated polysaccharides can effectively inhibit the SARS-cov-2 in vitro” was published in Opening Cells supported by the National research Foundation of Korea. In Rensselaer, Dordick and Linhardt were United in the study of Paul S. Kwon, Seok-Joon Kwon, Jin goes, fuming Zhang, and Kate Fraser and researchers at the Korea research Institute of Bioscience and biotechnology in Cheongju, Republic of Korea, and Zhejiang University of technology in Hangzhou, China.

Source:

Journal reference:

Kwon, S. P., et al. (2020) sulfated polysaccharides can effectively inhibit the SARS-cov-2 in vitro. The Opening Of The Cell. doi.org/10.1038/s41421-020-00192-8.

.



image source

Single-cell RNA-seq answers key questions in islet cell biology and diabetes research | Instant News


The pancreas is a stomach organ that produces digestive enzymes and hormones that regulate blood sugar levels. This hormone-producing function is localized on the island of Langerhans, which forms a group of various types of endocrine cells.

Among these are beta cells, which produce the hormone insulin which is needed to lower glucose levels (a type of sugar) in our blood, as well as alpha cells, which produce glucagon hormones whose job is to increase glucose levels in the blood.

Type 1 diabetes is a chronic disease where the immune system mistakenly attacks and destroys pancreatic insulin-producing beta cells. Regenerative medicine aims to replenish beta cell mass, and thus support and ultimately replace current insulin replacement therapy.

Changes in island composition, including lack of beta cell function and beta cell dedifference, also contribute to type II diabetes.

Therefore, a deeper understanding of the identity and crosstalk of various islet cell types leads to better characterization of both forms of diabetes and can contribute to the development of new therapeutic concepts.

Single cell transcriptomes are powerful techniques for characterizing cellular identity. Previously, CeMM researchers from the groups Christoph Bock and Stefan Kubicek at CeMM published the first single cell transcriptome from primary human pancreatic islet cells.

Advances in technology have enabled applications for the generation of global single cell atlase human and mouse transcript. Despite this progress, the single cell approach remains technologically challenging given the very small amount of RNA used in the experiment. Therefore, it is important to ensure the quality and purity of the resulting single cell transcriptome.

CeMM researchers in two laboratories who contributed unexpectedly identified high hormone expression in non-endocrine cell types, both in their own dataset and published single cell studies.

They set out to explain whether this would be the result of contamination by RNA molecules, for example from dying cells, and how they could be removed to obtain a more reliable dataset.

Such contamination appears to be present in RNA-seq single cell data from most tissues but is most visible on pancreatic islets. Islet endocrine cells are exclusively devoted to the production of a single hormone, and insulin in beta cells and glucagon in alpha cells is expressed to a higher level than a typical “household” gene.

Thus, the redistribution of these transcripts to other cell types is very clear. Based on these observations, their goal is to develop, validate and apply methods to experimentally determine and computationally eliminate the contamination.

In their investigation, the CeMM researchers used prickly cells of different cell types, both rat and human samples, which they added to their pancreatic islet samples. Importantly, the transcriptomes of these spike cells are fully characterized.

This enables them to internally and accurately control the level of RNA contamination in a single RNA-seq cell, providing that the human transcripts detected in mouse spike-in cells are contaminated RNA.

In this way, they found that the sample had a contamination rate of up to 20%, and was able to determine the contamination in each sample. They then developed a new bioinformatics approach to computationally eliminate contaminated readings from single cell transcriptomes.

Having now obtained the “decontamination” transcriptome, from which false signals have been removed, they proceed to characterize how cellular identities in different cell types respond to treatment with three different drugs.

They found that small molecular blockers of the FOXO1 transcription factor induced dedifferentiation of both alpha and beta cells.

Next, they studied artemeter, which has been found to reduce alpha cell function and can induce insulin production in both in vivo and in vitro studies. Effects of species-specific drug species and cell types.

In alpha cells, a small proportion of cells increase insulin expression and gain aspects of beta cell identity, both in rat and human samples. Importantly, the researchers found that in human beta cells, there was no significant change in insulin expression, whereas in mouse islands, beta cells reduced insulin expression and overall beta cell identity.

This study is the result of interdisciplinary collaboration from the laboratories of Stefan Kubicek and Christoph Bock at CeMM with Patrick Collombat at the Institute of Biology Valrose (France).

This is the first study to apply single cell sequencing to analyze dynamic drug responses in intact isolated tissue, which benefits from the high quantitative accuracy of the decontamination method.

Thus not only provides a new method for single cell decontamination and a very quantitative single cell analysis of drug responses in intact tissue, but also answers current questions that are important in islet cell biology and diabetes research. These findings could open up potential therapeutic avenues for treating type 1 diabetes in the future.

.



image source

Symptomatic COVID-19 patients tend to be smokers | Instant News


When the COVID-19 pandemic rages in many parts of the world, a new study shows that smoking may not play an important role in increasing the severity of the disease in these patients. This study was published on a preprinted server medRxiv* in May 2020.

It is now known that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus that causes COVID-19 disease, is more likely to cause severe pneumonia which often causes severe acute respiratory distress, multi-organ dysfunction, or both, causing Dead. Predisposing factors for serious or critical illness include older age, underlying medical conditions such as asthma, high blood pressure, diabetes, and cardiovascular disease.

However, does active smoking increase the risk? Little is known about the role of this factor, which is driving current research.

Smoking Can Increase Viral Entry – But What Is It?

Smoking increases the expression of angiotensin-converting enzyme 2 (ACE2) molecules in smokers’ tissues. ACE2 is known as the main receptor for viral attachment and entry into host cells in humans. Logically, it seems that smoking increases the risk of infection.

The SARS-CoV-2 virus binds to ACE-2 receptors in human cells, the initial stage of COVID-19 infection. Image Credit: Kateryna Kon / Shutterstock

However, some studies contradict this assumption, on the contrary showing the opposite. In Chinese studies, for example, only about 1.4% to 12.6% of COVID-19 patients are smokers. Likewise, only about 5% of New York patients, who are part of a very severe outbreak, are smokers.

Compared with national smoking statistics in the two countries, respectively around 25% and 17%, as taken from https://worldpopulationreview.com/countries/smokingrates-by-country, this is so low that the absence of direct links becomes clear. Current research is driven by the need to examine the existence of an inverse relationship between smoking and the possibility of infection with COVID-19.

How was the Smoking-COVID-19 Research conducted?

The researchers conducted a retrospective study of about 440 patients with COVID-19, all admitted sequentially to the tertiary level center in Parma, Italy. All of them have confirmed infection with nasopharyngeal swabs proved positive with the reverse transcriptase-polymerase chain reaction (rt-PCR).

The researchers took demographic, clinical, laboratory, and death data from hospital electronic health records. They also attempted to confirm smoking data with direct contact with patients or their relatives, in 423 of 441 cases.

What does the Cigarette Study show about COVID-19?

The results showed that about 62% of patients were male, and the median age was 71 years. 35% of patients died during their hospital stay, with 65% excluded after clinical recovery.

About 5% of them actively smoked at the time of the study, while 10% were smokers but had quit. The rest are not smokers.

With a closer analysis of clinical characteristics, it was observed that more men died from this disease. Although they constitute 62% of the total patients, they account for 72% of deaths and only 59% of survivors. Those who died were also older, at an average age of 76 years, compared to 67 years for those who survived. However, this is not statistically significant.

Again, the presence of certain medical diseases that coexist is significantly more common among deaths. For example, patients with a history of cardiovascular disease each contribute 21% and 10% of those who die and survive.

While hypertension was present in 61% and 56% of deaths and survivors, respectively, while diabetes was present in 26% and 18% respectively, this was not significant.

The median level of D-dimers, which is a marker of inflammation, is almost double the value in those who die compared to those who survive. There was a much smaller but significant increase in the median level of C-reactive protein, which also increased inflammation, among those who died.

However, with smoking, around 6% and 4% of those who die and live are current smokers, which means there is no significant difference. Similarly, the number of former smokers and never smokers was almost identical in the two groups.

What Does This Study Mean for People Affected by COVID-19?

Italy has experienced the second highest number of COVID-19 deaths in the European Union (EU). However, in this retrospective study, only 5% of smokers patients currently, even though the prevalence of the smoking population is 24% in Italy. This finding, therefore, agrees with previous research in China and the US.

This study did not include asymptomatic patients by design, and therefore the conclusions should be limited to cases of COVID-19 hospitalized or symptomatic.

The researchers said, “Current research shows that smokers can carry several types of protective mechanisms from symptomatic SARS-CoV-2 infections.” However, as scientists have warned, carefully designed controlled studies alone can validate this impression.

The mechanism underlying the low prevalence of smoking in the population of COVID-19 patients treated in this hospital is purely speculative at this time. For example, exposure to cigarette smoke can suppress the immune response, which contributes to low systemic inflammation compared to those who have never smoked.

By modulating the normal intensity and spectrum of the immune system against viruses, due to “continuous inflammatory insults,” cytokine storms may be less likely to occur in COVID-19 patients, making them less susceptible to severe or symptomatic disease.

The importance of ensuring the protective role for smoking in this pandemic is that, according to the researchers, “it can help uncover the molecular mechanisms underlying predisposition to SARS-CoV-2 infection, then also potentially being exploited by newly designed protective drugs. “

* Important Notification

medRxiv publish initial scientific reports that are not reviewed by colleagues and, therefore, should not be considered conclusive, guide clinical practice / health-related behaviors, or be treated as pre-existing information.

.



image source

CRISPR can help in kidney transplantation by detecting viruses | Instant News


An interesting new study was published in the journal Natural Biomedical Engineering in April 2020 reported excellent results with new tests for two common viruses that infect kidney transplant patients. The level of this virus increases during the acute phase of rejection.

Why is immunosuppressive needed?

Kidney transplantation is a life-saving but complicated procedure. There are many points where it can fail. Of all these, the most feared complication after successful surgery is organ rejection – when the patient’s body rejects a new kidney as a foreign object. The immune system starts producing antibodies to try and kill new kidneys. This not only removes the valuable donor kidney but can cause patient death.

To prevent this, doctors usually prescribe drugs that reduce the strength of the immune response, called immunosuppressive. This reduces the chance of rejection. But there is a downside.

Problems with immunosuppressive drugs

The disadvantage of such drugs is to disarm the system that allows us to function normally even when exposed to millions of bacteria, viruses, and other germs that, if left unchecked, will enter the body and endanger or even kill organisms. What makes them avoid is the immune system – the body’s natural defenses that are tuned. Sometimes it fails, such as when the body is exposed to new and unknown threats (such as COVID-19), or is taxed beyond its strength. With the immune system no longer operating at full power, the bridge pulls down – we are far more vulnerable to external threats. This is why doctors who carry out transplants have to follow a fine line between too little silencer (resulting in organ rejection), and too much (which allows infection to persist in the body). In addition, infections that occur in immunocompromised patients are just ordinary infections, which in healthy people can be easily removed.

Doctors must monitor their patients very carefully to ensure that the drugs are at the right concentration to prevent any of these things from happening. Monitoring like this is usually done through kidney biopsy and blood tests – expensive, invasive, and time consuming.

New way

Urine tests are much simpler and cheaper than blood and kidney tests. And now, scientists have combined this modality with a powerful technology called CRISPR to sense the presence of molecules that indicate infection. CRISPR is more often associated with gene editing. However, current use can produce very sensitive diagnostic tools to detect early signs of rejection, just like the more common urine tests that filter urine sugar, pregnancy hormones, and the like.

3D illustration of the CRISPR-Cas9 genome editing system. Credit Illustration: Meletios Verras / Shutterstock

To adapt the urine test for this purpose, it must be able to detect nucleic acids – DNA or RNA. And this is why CRISPR is so useful because it is able to find small segments of nucleic acid in a specific order if there is a complementary piece of RNA guidance to help it. This is combined with an enzyme called Cas, which occurs in several forms, and makes pieces to isolate the sequence in question. It is also bound to a fluorescent molecule called a reporter, because it lights up when the target sequence is split.

Lateral flow strips showed three samples of patients who were negative for BK virus (13,14,15) and three samples of patients who were positive (16,17,18). The presence of the upper band shows positive test results. Image Credit: Michael Kaminski, MDC

Lateral flow strips showed three samples of patients who were negative for BK virus (13,14,15) and three samples of patients who were positive (16,17,18). The presence of the upper band shows positive test results. Image Credit: Michael Kaminski, MDC

Many researchers have proven that CRISPR can take useful diagnostic information on man-made samples, but clinical testing is rarely done. This involves achieving enough sensitivity to detect very low concentrations, which are usually found in biological samples.

Researcher Michael Kaminski commented: “The challenge is to get down to clinically meaningful concentrations. This really makes a big difference if you aim at a ton of synthetic targets in your test tube, compared to if you want to reach the level of a single molecule in a patient’s fluid. “

How is CRISPR assisted urine testing done?

Urine test kits or tests are carried out in two stages. The first involves amplification of viral DNA from the two most common opportunistic pathogens, namely, cytomegalovirus (CMV) and BK polyomavirus (BKV). This process means making many copies of a small amount of target DNA to get a concentration high enough that it can be detected by CRISPR. Amplification goes as far as is needed to allow detection of even one target molecule. This was achieved by a CRISPR-Cas13 program called SHERLOCK, which adapted the process for use with viral DNA.

The testing kit is a paper strip that acts somewhat like a pregnancy strip at home. Two lines indicate the presence of a virus when the strip is dipped in a sample that has been prepared for testing. If the target sequence exists but at a very small level, a pale second line may appear. To avoid this, they also designed smartphone applications to ensure an impartial strip analysis with results that reflect line intensity.

The steps are repeated this time using the CXCL9 biomarker, which signifies organ rejection. MRNA from the sample was isolated and then amplified before detecting it using CRISPR-Cas13.

The researchers perfected the technique, testing the analysis on more than 100 real-life kidney transplant samples. They found that it responded to very low concentrations of BKV or CMV, both of which were related to acute rejection responses mediated by cellular infiltration.

Embryonic renal culture ex-vivo. Image Credit: Michael Kaminski, MDC

Embryonic renal culture ex-vivo. Image Credit: Michael Kaminski, MDC

What happens next for CRISPR?

The researchers applied for a patent. Kaminski also plans to conduct clinical studies of sufficient size to make a meaningful contribution in the field. This will compare current tests with conventional methods to monitor these patients. Scientists are working on a smoother and less complicated test protocol, without having to heat up the sample before being analyzed.

Such dreams can bring tests to the level of testing at home, where one strip only needs to be tested to get quantitative results from various biomarkers. This will allow changes in each field compared to the baseline for each marker.

The researchers said, “Testing allows – through simple visualization – post-transplant monitoring for common opportunistic viral infections and graft rejection, and should facilitate monitoring at the post-transplant care site. Not only will this test be of benefit to others with a weak immune system, but CRISPR-mediated diagnostic capabilities can also lead to the development of new tests for other types of organ transplants as well. “

Journal reference:

Kaminski, M.M., Alcantar, M.A., Lape, I.T. et al. CRISPR-based test to detect opportunistic infections after transplant and for monitoring transplant rejection. Nat Biomed Eng (2020). https://doi.org/10.1038/s41551-020-0546-5

.



image source