Mois : juillet 2024

Scientists have developed a conjugate of a drug and a nucleus-targeting antibody that can attack multiple types of cancer cells without targeting a particular antigen [1].

The anti-nuclear missile

Antinuclear antibodies (ANA) are usually associated with autoimmune diseases, such as lupus, where those antibodies attack cellular nuclei, binding to nucleic acids and other local molecules. ANAs’ ability to penetrate cellular membranes has drawn oncologists’ attention because ANAs can be conjugated with drugs and deliver them into tumor cells.

Some ANAs have an interesting Trojan horse-like strategy for sneaking into cells. They bind to extracellular nucleic acids and short chunks of DNA (nucleosomes) and piggyback on salvage pathways that transport those useful molecules back into cells [2]. In this new study, scientists from Yale university used this mechanism to attack cancer.

In fast-growing tumors, some cells do not get enough oxygen and nutrients and die off. This process, called tumor necrosis, is indicative of the tumor’s aggressiveness.

Those dead cells emit a lot of nucleosides, which are precursor molecules to nucleotides, DNA’s building blocks. Living cancer cells employ transporter proteins to bring those nucleosides back in where they can be reused.

Exploiting this effect, the researchers developed an antinuclear antibody-drug conjugate (ANADC) that targets those tumor-specific “nucleoside junkyards” and hitches a ride into tumor cells with those same transporter proteins. Upon uptake by the cell, ANADCs are cleaved by the protein cathepsin B, and the drug (the division-preventing agent monomethyl auristatin E, or MMAE) is released into the cytoplasm. The researchers describe their invention as “an anti-nuclear missile.”

Tumor growth suppressed

The researchers tested their invention on the U87 cell line, which is an established model of brain cancer. Treatment with ANADC caused a drastic decrease in the cells’ viability. Conversely, other approaches, such as an antibody-drug conjugate based on a non-antinuclear antibody, failed to achieve noticeable cytotoxicity.

In a mouse model of triple-negative breast cancer, the treatment completely suppressed tumor growth, while antibodies and non-antinuclear antibody-drug conjugates did not. The mice did not experience weight loss, which suggests a lack of off-target toxicity. ANADC was still effective, although not as drastically, in a model of colon cancer.

Crossing the blood-brain barrier

The toughest test, however, was brain tumors. Those hide behind the blood-brain barrier (BBB), which is impervious to most antibodies and antibody-drug conjugates. Nucleoside transporters, however, know how to traverse it.

The researchers confirmed that in a mouse U87 model of glioma, ANADC were indeed able to cross the BBB using the nucleoside transporter ENT2 and to target tumors. The treatment significantly increased the length of survival, although it did not cure the cancer completely. Importantly, advanced brain cancers are among the deadliest and fastest growing. No significant off-target deposition of ANADC was detected, suggesting high anti-cancer specificity.

According to the researchers, their invention has important advantages over current treatments, the most obvious one being that it does not target tumor cells via a specific antigen. Identifying the antigen that can be targeted in oncology is a complex and multifaceted process due to tumors’ genetic heterogeneity and other factors [3]. Pending further improvements, ANADC can potentially become an effective and fast off-the-shelf option for treating multiple cancer types.

The ANADC developed here targets the nucleic acid exhaust released by necrotic tumor cells and exploits mechanisms of nucleoside salvage by live cancer cells in the area as a DNA-seeking “antinuclear missile”. Antibody localization to extracellular nucleic acid waste helps mitigate concerns over target antigen depletion during therapy as tumor cell turnover and death yield a continuously renewing source of nucleic acids to draw ANADC to tumor microenvironments.

Literature

[1] Cao, F., Tang, C., Chen, X., Tu, Z., Jin, Y., Turk, O. M., … & Hansen, J. E. (2024). Cathepsin B Nuclear Flux in a DNA-Guided “Antinuclear Missile” Cancer Therapy. ACS Central Science.

[2] Weisbart, R. H., Chan, G., Jordaan, G., Noble, P. W., Liu, Y., Glazer, P. M., … & Hansen, J. E. (2015). DNA-dependent targeting of cell nuclei by a lupus autoantibody. Scientific Reports, 5(1), 1-6.

[3] Balibegloo, M., Keshavarz-Fathi, M., & Rezaei, N. (2021). Tumor Antigen Identification for Cancer Immunotherapy. Cancer Immunology: Bench to Bedside Immunotherapy of Cancers, 53-59.

In iScience, researchers have explained how physical mechanics can alter mitochondrial function in a way that leads to osteoarthritis.

When simple physics affects biology

Previous work has pinpointed abnormal mechanical loading, which occurs when joints are placed under excessive stresses in ways that they were not meant to handle, as a key driver of osteoarthritis [1]. This phenomenon can lead to the death or senescence of chondrocytes, the cells responsible for creating cartilage [2].

The microenvironment of the extracellular matrix (ECM) plays a strong role in the metabolism of chondrocytes [3]. This microenvironment can cause these cells’ mitochondria to uptake calcium [4], and in osteoarthritis, mineral particles begin to form on the joints, after which fibers and packed material are gradually pushed into the joints [5].

Mitochondrial dysfunction brought on by mechanical stresses has been found to be a core component of osteoarthritis [6], and these researchers have noted a downstream pathway: the stressed mitochondria communicate with the nucleus in a way that leads to the demethylation of H3K27me3, a key component of epigenetics [7].

The problems with stiffness

For their first experiment, the researchers used a gel substrate at three different stiffnesses and then grew an established line of chondrocytes in it. The stiffer the gel was, the more calcium was taken up by the chondrocytes. Genes related to stress in protein processing machinery (the endoplasmic reticulum) were upregulated as well.

The cells grown in stiffer conditions also had a decrease in Col2a1, a gene related to the formation of more complex molecules (anabolism), while Mmp13, a gene related to the breaking down of those molecules (catabolism), was increased. This implies that the same thing may be happening in vivo: that increased stress encourages chondrocytes to break down, rather than form, cartilage. Chondrocytes under stiffer conditions are also more prone to death by apoptosis.

The researchers also observed how mitochondria fail with increasing stiffness. At the lightest stiffness, mitochondria formed a normal network, doubling that stiffness broke up that network, and tripling it caused mitochondrial fragments and rings instead of a network of any kind. Mitochondrial division-related proteins increased, and fusion-related proteins decreased, with stiffness; additionally, the mitochondria were less efficient at producing energy.

Intracellular calcium was found to be key to this process. When intracellular calcium was removed, mitochondria under high-stiffness conditions functioned very closely to those under low-stiffness conditions, and they regained some of their energy production capacity.

Pinpointing the dysfunction

The increase in calcium released from the mitochondria increased their membranes’ permeability, which led to an increase in the expression of Phf8. This, the researchers found, was directly related to the demethylation of H3K27me3. Silencing Phf8 prevented this increase in demethylation brought on by stiffness.

These findings were confirmed in mice. An RNA silence of Phf8.was injected into the joints.of mouse model of osteoarthritis. The injected mice had more stable cartilage, more normal mitochondrial fusion and fission, and less demethylation of H3K27me3.

While there was no human testing of Phf8 involved, the researchers did confirm, through cartilage samples taken from human volunteers, that stiffness of the ECM was directly related to the progression of osteoarthritis.

This is an exploratory study, and there was no drug discovery involved; it may prove impractical to use Phf8 as a clinical target, and there may be side effects to this approach. However, in addition to approaches that focus on ECM stiffness itself, such as the well-known problem of cross-linked collagen, targeting how chondrocytes respond to this stiffness may be valuable in treating this crippling ailment.

Literature

[1] Katz, J. N., Arant, K. R., & Loeser, R. F. (2021). Diagnosis and treatment of hip and knee osteoarthritis: a review. Jama, 325(6), 568-578.

[2] Chang, S. H., Mori, D., Kobayashi, H., Mori, Y., Nakamoto, H., Okada, K., … & Saito, T. (2019). Excessive mechanical loading promotes osteoarthritis through the gremlin-1–NF-κB pathway. Nature communications, 10(1), 1442.

[3] Peng, Z., Sun, H., Bunpetch, V., Koh, Y., Wen, Y., Wu, D., & Ouyang, H. (2021). The regulation of cartilage extracellular matrix homeostasis in joint cartilage degeneration and regeneration. Biomaterials, 268, 120555.

[4] Li, X., Kordsmeier, J., Nookaew, I., Kim, H. N., & Xiong, J. (2022). Piezo1 stimulates mitochondrial function via cAMP signaling. FASEB journal: official publication of the Federation of American Societies for Experimental Biology, 36(10), e22519.

[5] Jiang, W., Liu, H., Wan, R., Wu, Y., Shi, Z., & Huang, W. (2021). Mechanisms linking mitochondrial mechanotransduction and chondrocyte biology in the pathogenesis of osteoarthritis. Ageing research reviews, 67, 101315.

[6] Blanco, F. J., Rego, I., & Ruiz-Romero, C. (2011). The role of mitochondria in osteoarthritis. Nature Reviews Rheumatology, 7(3), 161-169.

[7] Peña-Oyarzun, D., Rodriguez-Peña, M., Burgos-Bravo, F., Vergara, A., Kretschmar, C., Sotomayor-Flores, C., … & Criollo, A. (2021). PKD2/polycystin-2 induces autophagy by forming a complex with BECN1. Autophagy, 17(7), 1714-1728.

A new mouse study has found that both germline knockout and late-life inhibition of the pro-inflammatory cytokine IL-11 lead to comparable and powerful healthspan and lifespan extension [1].

The inflammatory message

As part of inflammaging, the pro-inflammatory cytokines that act as messengers, transferring and amplifying inflammatory signals, generally increase with age.

Despite its links to cellular senescence, IL-11 is not the most well-known of these cytokines. However, this close relative of the better-researched IL-6 has been shown to stimulate important age-related metabolic pathways, which include the mechanistic target or rapamycin (mTOR) [2]. Using rapamycin and other means to Inhibit mTOR potently increases healthspan and lifespan in various animal models [3].

Its various effects have made this obscure cytokine the subject of a new study by an international team of scientists, which was published in Nature.

Multiple health benefits

First, the researchers confirmed IL-11’s age-related upregulation in mice. Old mice had more IL-11 across many cell types and tissues, most notably in liver cells (hepatocytes), fat cells (adipocytes), and muscle cells (myocytes) in skeletal muscle.

To study the effects of IL-11, the researchers created genetically modified mice with the IL-11-producing gene knocked out (IL-11-KO). Despite missing the protein, the modified animals showed improved metabolism and lower levels of the senescent markers p16 and p21.

Two-year-old IL-11-KO mice had lower body weights, decreased fat mass, and increased lean mass. They also had attenuated serum triglyceride and cholesterol levels compared to controls.

Two markers of liver damage, ALT and AST, increased with age in wild-type mice but not in the genetically modified ones. Digging deeper into the molecular mechanisms behind those health benefits, the researchers found that mTOR was significantly downregulated in the study group, while AMPK, a kinase that mitigates mTOR, was upregulated.

IL-11 knockout also slowed down telomere attrition and the loss of mitochondrial DNA, which is a marker of mitochondrial health. Interestingly, average body temperature was slightly higher in the study group, hinting at differences in metabolism.

While genetic manipulations to the germ line are a neat way to investigate a protein’s function, they are hardly translatable into actual therapies. The researchers created a more realistic setting by treating 75-week-old mice (about equivalent to 55-year-old humans) with IL-11-blocking antibodies.

Despite the late start, the treated mice of both sexes quickly lost fat. Their frailty scores, full-body strength, and glucose metabolism improved significantly. Like the IL-11-KO mice, they had better glucose control and liver function, and slightly increased body temperature. Levels of phosphorylated mTOR plummeted, and of AMPK, a kinase that downregulates mTOR, increased.

Late start, mighty life extension

Most importantly, IL-11-deficient mice also received a lifespan boost. Genetic deletion of IL-11 increased median lifespan of both sexes by almost 25%, while late-life IL-11 inhibition increased median lifespan in males by 22.5% and in females by 25%.

These results are impressive and on par with some of the best life-extending interventions in mice currently known to science. The fact that the late-life treatment was practically as impactful as lifelong genetic deletion is particularly stunning. Importantly, while many known geroprotectors increase lifespan mostly or solely in one sex, the difference was minuscule here.

However, mice are not like humans in terms of causes of death. Lab mice usually die from cancer. The researchers report a much lower incidence of cancer in treated mice, but this is unlikely to be the leading cause of the life extension since IL-11 inhibition also produced a wide array of lifelong health improvements.

Inhibition of IL-11 increased lifespan in both male and female mice. The magnitude of lifespan extension remains to be fully determined but current data suggest that anti-IL-11 therapy given in late life increases median lifespan by more than 20% in both sexes. In these experiments, anti-IL-11 was injected in mice from 75 weeks of age (human equivalent to approximately 55 years of age) and it remains to be seen whether administration to older mice has similar effects and/or if short term anti-IL-11 therapy is effective for lifespan extension, as seen for rapamycin.

Literature

[1] Widjaja, A.A., Lim, WW., Viswanathan, S. et al. (2024). Inhibition of IL-11 signalling extends mammalian healthspan and lifespan. Nature.

[2] Widjaja, A. A., Viswanathan, S., Ting, J. G. W., Tan, J., Shekeran, S. G., Carling, D., … & Cook, S. A. (2022). IL11 stimulates ERK/P90RSK to inhibit LKB1/AMPK and activate mTOR initiating a mesenchymal program in stromal, epithelial, and cancer cells. IScience, 25(8).

[3] Harrison, D. E., Strong, R., Sharp, Z. D., Nelson, J. F., Astle, C. M., Flurkey, K., … & Miller, R. A. (2009). Rapamycin fed late in life extends lifespan in genetically heterogeneous mice. nature, 460(7253), 392-395.