Toxic effects of chemotherapy are caused by cell-free chromatin fragments that emerge from the dying cells

A large body of work by researchers at the Tata Memorial Centre (TMC), Mumbai has found conclusive evidence in cell lines, animal studies and two phase-2 clinical trials that cell-free chromatin fragments that are released from dying cells have the ability to readily enter healthy cells in the body and integrate into their genomes and trigger DNA damage by breaking the DNA double-strand, cause apoptosis (programmed cell death) and inflammation. Based on these studies, a team led by Dr. Indraneel Mittra, Professor Emeritus, Department of Surgical Oncology at TMC had proposed that “repeated and lifelong assault on healthy cells by cell-free chromatin fragments may be the underlying cause of ageing and multiple age-related disorders including cancer”. They also found that using a combination of two nutraceuticals — resveratrol and copper — minimises the damaging effects of cell-free chromatin fragments.

The researchers have collected sufficient evidence to prove that the toxic effects of chemotherapy are primarily caused by the cell-free chromatin fragments that emerge from dying cells and continue the vicious cycle of killing healthy cells and causing inflammation. This finding challenges the dogma that toxicity caused by chemotherapy is due to the direct damaging effect of drugs on healthy cells.

Based on preclinical studies that showed promising results when the resveratrol-copper combination was used, Dr. Mittra and other researchers at TMC tested it on 25 patients undergoing autologous bone marrow transplantation for multiple myeloma and 30 patients with advanced gastric cancer. In both studies, the toxicity of chemotherapy was far less among patients who received the resveratrol-copper combination.

Evidence from pre-clinical studies also suggests that resveratrol-copper nutraceuticals may prevent the metastatic spread of cancer. The researchers had earlier reported that cell-free chromatin particles released from dying cancer cells are potentially oncogenic. Studies in mice undertaken by the team showed that oncogenes contained in the cell-free chromatin fragments were carried by blood and had accumulated in the brain cells in mice. Though the oncogenes in the brain can potentially lead to metastasis, the study did not investigate whether the dissemination of oncogenes to brain cells indeed led to the development of metastases.

TMC statement

A statement issued on March 1 by TMC categorically stated that the effectiveness of resveratrol-copper for reducing chemotherapy toxicity or increasing cures in cancer patients “remains to be established and is currently under investigation”. The statement was released after Dr. Rajendra Badwe, a senior researcher at TMC told NDTV that the nutraceutical to reduce the toxicity of chemotherapy will be available by June this year after approval by the FSSAI. Several media reports wrongly reported that the nutraceutical to be approved by FSSAI is for preventing cancer metastasis.

Based on extensive pre-clinical studies and two phase-2 clinical trials, there is sufficient evidence to believe that resveratrol-copper would reduce chemotoxicity in patients with cancer, Dr. Mittra tells The Hindu in an email. “Evidence from pre-clinical studies suggests that resveratrol-copper combination may prevent metastatic spread. Randomised trials are now needed to establish the quantum of benefit [in reducing chemotoxicity]. The results of at least one randomised trial should hopefully be available in about a year’s time. In the meantime, and once resveratrol-copper is certified as a nutraceutical, it would be up to any individual to take it for [any of] the above indications if he/she so desires,” Dr. Mittra says.

Despite the absence of clinical trials supporting the use of a resveratrol-copper combination to prevent metastasis, Dr. Mittra does not see any problem in patients using the nutraceutical to prevent cancer metastasis as soon as it becomes available. “Once resveratrol-copper is approved as a nutraceutical anyone should be free to take it. Although at this point, we only have pre-clinical evidence that it prevents metastasis if someone decides to take resveratrol-copper for this indication, being a non-toxic combination, it is unlikely to cause any harm,” he says.

“Resveratrol-copper is probably a nutraceutical product with the most scientific evidence to back it for various conditions. For example, our pre-clinical studies have shown that it retards ageing parameters, such as getting rid of amyloid plaques which are responsible for Alzheimer’s disease. Resveratrol-copper also prevents telomere shortening, inflammation, DNA damage, and chromosomal abnormalities and reduces blood levels of cholesterol and glucose. So resveratrol-copper may have multiple other health benefits to the person who decides to take it,” Dr. Mittra adds. “I have personally been taking resveratrol-copper tablets for the past three years without any side effects. The mother of an oncologist at TMC who is suffering from Alzheimer’s disease has been taking resveratrol-copper tablets for several months now with remarkable improvement in her cognitive score and general mental state.”

On why the researchers are trying to get the combination approved by FSSAI as a nutraceutical and not as a drug by the drug regulator CDSCO, Dr. Mittra says TMC did not take the nutraceutical route willingly. “We had tied up with an industry partner who was sourcing resveratrol and copper from two different vendors for making the resveratrol-copper tablets in a specific formulation. The industry partner was unable to get their resveratrol-copper formulation approved by DCGI since the latter insisted that the respective vendors from whom resveratrol and copper were being sourced register their products with DCGI from their own GMP facilities. This the vendors were unwilling to do. As a result, the industry partner handed over the project to their subsidiary who are involved in the nutraceutical industry.”

Treating cancer is expensive, costing up to several lakh rupees depending on the type of cancer, the treatment options available, and the treatment setting (public or private). It can also take time, removing an individual from their work and family for extended periods, and be painful.

Sometimes, while an individual may have successfully forced a cancer into remission, there may be a risk of relapse. One way this happens is when a few cancer cells are able to resist the drugs used to destroy them: they lay in wait and produce a show of strength later. Understanding this resistance could eliminate the different ways in which it happens, and reduce the odds of a relapse.

In a new study, published in Cell Reports on September 20, researchers from the Netherlands Cancer Institute investigated the resistance of some cancer cells to a drug called Taxol. They have reported that the culprit could be the location of a particular gene inside the cancer cells’ nucleus.

The oncologist’s challenge

A characteristic feature of cancer cells is that they divide rapidly, in uncontrolled fashion. Anti-cancer drugs – i.e. chemotherapeutic agents – work by stalling or blocking this proliferation. When the division of a cancer cell is arrested, it generally responds by triggering a pathway of programmed cell death, called apoptosis. So in this way, chemotherapy eliminates the cancer cells without affecting other non-cancerous cells nearby that are not dividing.

But this is also why chemotherapy deals a lot of collateral damage. Any tissue with a significant number of normal cells that are also dividing – such as cells in the digestive tract, the bone marrow, and hair follicles – are also affected by chemotherapeutic agents and suffer apoptosis. This cell death underlies the unpleasant side-effects of chemotherapy, such as painful inflammation of the oral cavity and the gut, and nausea, diarrhoea, anaemia, and hair loss.

An oncologist’s challenge is to find the dose of a drug that effectively kills cancer cells but whose side-effects are not unbearable for the patient. One way researchers have tried to achieve this is by developing antibody-drug conjugates (ADCs) against some cancers. An ADC is a drug attached to an antibody that recognises a protein found only on, or at least preferentially on, the cancer cells. This way, the antibody guides the chemotherapeutic drug to the cancer cells, where the drug begins its work. And, of course, non-cancer cells are bypassed.

The toxin-remover protein

As it happens, a small subset of cancer cells can still escape confrontation with the anti-cancer drug. This happens when these cells express elevated levels of a protein called P-gp – short for permeability glycoprotein. For a cell to produce P-gp, it uses information encoded in a gene called ABCB1.

Inside the cell, P-gp works like a pump, moving toxic compounds out. And in cells that make too much P-gp, the protein removes toxins well enough to flush the chemotherapeutic agents out as well. So the latter can’t accumulate to levels that arrest cell division and trigger apoptosis, allowing the cancer cell to live another day.

In fact, these surviving cells can allow the cancer to return after a period of remission.

Finding the mechanism of resistance

In the Cell Reports study, the researchers used cells from the human eye retinal pigment epithelium as a model to explore a small subset that expressed the P-gp protein and thus became resistant to the anti-cancer drug Taxol. They found that a cell’s sensitivity to Taxol, including its ability to resist Taxol’s anti-cancer effects, was related to the location of the ABCB1 gene inside the cell’s nucleus.

The nucleus is the part of the cell that houses the DNA and the associated proteins. A membrane called the nuclear envelope separates it from the rest of the cell. Genes are segments of a DNA molecule; when a gene is expressed, it means the cell can use it as a template to form molecules called RNA.

DNA and RNA share many chemical properties. The DNA contains the archival copy of a gene whereas the cell uses the RNA as the working copy. But only the RNA, and not the DNA, enters the cytoplasm – i.e. the rest of the cell – where it ‘instructs’ the cellular machinery on the way to link different amino acids to form the protein encoded by a gene.

In those retinal pigment epithelium cells that were sensitive to Taxol, the ABCB1 gene was found to be located close to the nuclear envelope. In cells that could resist the effects of Taxol, the gene had detached from the nuclear envelope and had moved further inside the nucleus. As a result, resistant cells exhibited a 100-fold increase in the amount of RNA corresponding to the ABCB1 gene compared to cells that remained sensitive to Taxol.

The P-gp efflux pump made from this RNA was responsible for Taxol-resistance.

Resisting the resistance

To identify what tethered the ABCB1 gene to the nuclear envelope in sensitive cells, the researchers turned different genes ‘off’ to see which one affected the proteins that the cell uses to make the envelope.

They zeroed in on a protein called lamin B receptor (LBR). According to the researchers, when the LBR protein was absent, a cell could activate the ABCB1 gene when it was exposed to Taxol. But when they deleted the gene used to make LBR, the cells didn’t increase ABCB1 expression right away; they had to be exposed to Taxol as well. So additional factors, instead of just LBR, help silence ABCB1 in the bulk population.

The researchers also studied the effect of depleting LBR from breast, head and neck, and lung cancer cells. Lung cancer cells expressed the RNA corresponding to ABCB1 to a high degree, and depleting LBR proteins didn’t further increase the fraction of Taxol-resistant cells. On the other hand, among breast cancer cells, depleting LBR increased the Taxol-resistant fraction – but not in the head and neck cancer cells.

Preferences among cells

Why do different cancers respond so differently to LBR depletion? An analogy from everyday life might help to understand. There are different ways to keep clothes dry in a bathroom: by hanging them on hooks, on towel rods or on a ledge. But not all bathrooms offer all options. In one with only a few hooks, there is a greater risk of clothes piled on a hook dropping to the floor.

We can rely less on hooks if there are rods and ledges as well. Similarly, the breast cancer cells may have depended more on LBR to tether genes to the nuclear envelope than the other cancer-cell types.

These findings highlight the need for more research to uncover the different ways in which cancer cells express or silence genes. By revealing how some cells develop Taxol-resistance, the study also opens the door for researchers to develop new ways to ensure anti-cancer drugs remain potent and patients recover faster.

The author is a retired scientist.