‘One-Two Punch’ Targets Resistance Mechanism in Aggressive Triple-Negative Breast Cancer

Triple-negative breast cancer (TNBC) may initially respond well to chemotherapy but will often develop resistance to treatment. Researchers at Medical University of South Carolina Hollings Cancer Center have now uncovered one of the mechanisms behind that resistance and a potential approach to its reversal.

The team, headed by Ozgur Sahin, PhD, co-leader of the Hollings Cancer Biology and Immunology Research Program, discovered an unexpected role for a protein called lysyl oxidase, or LOX, inside TNBC cells. They found that blocking LOX disrupted several processes on which the cancer depends, creating a weakness that can then be exploited using a second drug.

Experiments showed that the combination strategy significantly blocked tumor growth in multiple preclinical models of TNBC, without chemotherapy. This is important as chemotherapy treatment is often associated with adverse effects and persistent tumor growth in the clinic. The new strategy offers a safe way to target chemoresistant tumors by first creating a weakness and then exploiting it to close the escape route the tumor cells use to survive.

“It’s a one-two-punch approach,” said Sahin, also program director of science translation for the Hollings Advisory for Rapid Translation, and senior and corresponding author of the team’s published paper in Cell Reports Medicine. “First, we block LOX, which weakens the cancer cells. As they adapt and become dependent on a backup survival pathway, we deliver the second punch by blocking that pathway, too.” The researchers’ paper is titled “Lysyl oxidase inhibition disrupts mitochondrial homeostasis to create vulnerability to ferroptosis in TNBC.”

TNBC gets its name because the cancer cells lack three common targets used to treat other forms of breast cancer, leaving patients with fewer treatment options. Chemotherapy remains a primary treatment for TNBC, but it comes with significant side effects. Even when tumors initially respond well, they often develop resistance. “TNBCs are characterized by high metabolic heterogeneity and plasticity, contributing to their aggressiveness and therapy resistance,” the authors noted. “Thus, identification of therapeutic vulnerabilities to improve clinical outcomes is urgently needed.”

Sahin added, “Triple-negative breast cancer is one of the most aggressive, deadliest versions of breast cancer. Chemotherapy is really the mainstay, and interestingly, this subtype is sensitive to chemotherapy compared to others, but resistance develops quite quickly.”

Sahin’s laboratory has spent years studying LOX. “High LOX expression has been associated with poor prognosis, chemoresistance, and metastasis across multiple cancer types the researchers noted.

Traditionally, scientists have focused on what the protein does outside cancer cells, where it reshapes the tissue surrounding tumors. As that tissue becomes denser and stiffer, cancer can spread more easily, and drugs have a harder time reaching the tumor. “LOX promotes collagen crosslinking and matrix stiffening, which contributes to tumor progression, invasion, and the establishment of pre-metastatic niche,” the researchers continued. But despite it’s well-established “extracellular functions,” they noted, “… whether LOX regulates energy metabolism and mitochondrial homeostasis in cancer cells remains poorly understood.

When Sahin and team looked inside cancer cells, they discovered something unexpected. For the first time, they showed that LOX also helps TNBC cells to survive from the inside by supporting energy production, maintaining healthy mitochondria and protecting the cells from stress.

Mitochondria produce the energy on which cells rely, and also help cells to respond to stress and maintain the balance that keeps them alive. The researchers found that LOX plays a key role in both. When they blocked LOX, cancer cells struggled to produce energy, cope with stress and ultimately survive.

“LOX helps cancer cells keep multiple survival systems running,” explained postdoctoral fellow and co-first author Burge Ulukan, PhD. “When we blocked LOX, the cancer cells lost that advantage. When we inhibit it, we are inhibiting multiple arms. We’re disrupting cells’ energy production and making them much more vulnerable to treatment.” By disrupting multiple systems that cancer cells depend on, the researchers had left them vulnerable. The next question was how to capitalize on that weakness. The researchers realized they could do that by triggering ferroptosis, a form of cell death caused by toxic damage inside the cell. Scientists are increasingly interested in harnessing ferroptosis to destroy cancer cells that resist other treatments.

Cancer cells have built-in defenses that protect them from ferroptosis. Blocking LOX weakened two of those defenses. But the cancer cells did not give up. Instead, they relied more heavily on a backup defense controlled by a protein called DHODH. That helped the cells to survive after LOX was blocked—but it also created a new weakness as the cancer cells became increasingly dependent on DHODH.

The researchers saw an opportunity. If blocking LOX was the first punch, blocking DHODH could provide the second. In laboratory experiments, blocking both pathways overwhelmed the cancer cells and caused damaging molecules to build up until the cells underwent ferroptosis.

The team then tested the strategy in several preclinical models, including models developed from patients whose tumors had become resistant to chemotherapy. They paired an experimental drug they developed to block LOX with the FDA-approved drug leflunomide, works by blocking the backup defense on which the cancer cells had become dependent.

The combination significantly blocked tumor growth across multiple patient-derived models. It did so without causing major weight loss or signs of kidney or liver toxicity. It also outperformed a combination of the LOX inhibitor and a standard chemotherapy drug. “Here, we identify non-canonical functions of LOX in promoting glucose metabolism, inhibiting mitophagy, and sustaining redox homeostasis and show that inhibiting LOX generates a targetable vulnerability to DHODH-inhibition-mediated ferroptosis,” the authors wrote in summary.

“One of the most exciting aspects of this work is that we uncovered an entirely new role for LOX inside cancer cells,” said postdoctoral fellow and co-first author Ozge Saatci, PhD. “That discovery revealed a weakness we could exploit. Rather than attacking cancer cells from just one direction, we first weaken the cells and then target the backup system they rely on to survive. That opens the door to a new treatment strategy.”

In their paper the team further commented, “In summary, our study identified non-canonical functions of LOX orchestrating glucose metabolism, mitophagy, and ferroptosis in TNBC. We demonstrated the therapeutic potential of targeting LOX in combination with DHODH inhibitors to eradicate highly aggressive TNBC tumors that could pave the way for clinical testing of LOX inhibitors in combination with DHODH inhibitors.”

The findings are still preclinical and do not mean the combination is ready for patients yet. But use of an existing drug could potentially make that path toward clinical testing far easier. “The good thing is when the drug is FDA approved, you know the side effect profile,” Sahin said. “It makes it faster and potentially safer to repurpose it, in other words, adapt it for a different disease condition.”

For patients, one of the most promising aspects of the research is the possibility of one day reducing reliance on chemotherapy. Although chemotherapy can be highly effective, it can also cause serious side effects, including heart damage and nerve damage that leads to numbness, tingling or pain in hands and feet. A successful nonchemotherapy approach could potentially avoid some of those toxicities. However, more research is needed to determine whether the new strategy is safe and effective in people.

Importantly, the researchers found evidence that the same biology they observed in the laboratory may also be at work in patients. In tumor samples from people with TNBC, higher levels of LOX were linked to increased activity in the same energy and survival pathways identified in the study. “High levels of both LOX and DHODH is associated with worse overall survival in TNBC patients,” the team further pointed out.

Together, the findings raise the possibility that these proteins could eventually serve as biomarkers, helping to identify patients most likely to benefit from treatments targeting this vulnerability. “LOX may act as a biomarker of response or resistance to the metabolic targeting of the tumors,” Ulukan said. “If we can identify patients whose tumors depend on this pathway, those may be the patients who benefit most from this type of treatment.”

The team is already developing a newer version of its LOX-blocking drug in collaboration with the University of South Carolina. The next step is completing the studies needed to test the drug safely in humans. Sahin hopes that process can be completed within the next few years. An earlier goal is to determine whether the strategy can help patients whose cancers have stopped responding to current treatments.

For the researchers, the study represents more than the discovery of a new drug target. It offers a new way to think about how to outsmart one of cancer’s greatest strengths, which is the ability to adapt. That approach could prove especially valuable for aggressive cancers such as TNBC, where treatment resistance remains one of the biggest barriers to long-term success.