New Autoimmune Treatment Removes Rogue Immune Cells Without Suppressing the Immune System

New preclinical experiments by scientists at the University of Pennsylvania have established that genetically engineered T-cells can drive severe autoimmune diseases into remission without suppressing the patient’s immune system. If the principles applied in this study also prove to be true in human patients, they can potentially revolutionize the treatment of autoimmune diseases.

Autoimmune diseases result when your immune system recognizes your own cells and tissues as foreign and mounts and immune response against them. Autoimmune diseases like systemic lupus erythematosus (also known as “lupus”), rheumatoid arthritis, scleroderma, multiple sclerosis, celiac disease, Sjögren’s syndrome, polymyalgia rheumatic, or ankylosing spondylitis can deeply affect the health of an individual and can also cause large amounts of tissue damage.

Treatment of autoimmune diseases usually requires high doses of drugs that suppress the immune system, such as corticosteroids, or various types of biological agents that also cause a host of undesirable side effects.

This new study, however, by scientists from the Perelman School of Medicine at the University of Pennsylvania have adapted an already-existing technology to remove the subset of antibody-making cells that cause the autoimmune disease. This strategy removes the rogue immune cells without harming the rest of the immune system.

In these experiments, the University of Pennsylvania team examined an autoimmune disease called pemphigus vulgaris or PV. PV results when the immune system recognizes a protein called desmoglein-3 (Dsg3) as foreign and attacks it. Dsg3 helps form attachment sites called “desmosomes” that normally adhere skin cells together to form tight, tough sheets. Desmosomes are also found between epithelial cells, myocardial cells, and other cell types.


Current therapies for autoimmune diseases like PV use drugs like prednisone and rituximab, which suppress large parts of the immune system. Consequently, prednisone and rituximab can leave patients vulnerable to potentially fatal opportunistic infections and cancers.

To treat PV, University of Pennsylvania researcher Aimee Payne and her colleagues used a mouse version of PV that is fatal in mice. Their experimental treatment, however, successfully treated this otherwise fatal autoimmune disease without causing any unintended side effects, which might harm healthy tissue. The results from these experiments were published in the journal Science.

“This is a powerful strategy for targeting just autoimmune cells and sparing the good immune cells that protect us from infection,” said Dr Payne, who serves as the Albert M. Kligman Associate Professor of Dermatology at the Perelman School of Medicine.

In collaboration with Dr. Michael Milone, assistant professor of Pathology and Laboratory Medicine, Payne and her colleagues adapted the Chimeric Antigen Receptor T-Cell (CART-Cell) technology that is being successfully used to experimentally treat malignant cells in certain leukemias and lymphomas. “Our study effectively opens up the application of this anti-cancer technology to the treatment of a much wider range of diseases, including autoimmunity and transplant rejection,” Milone said.

Aimee Payne, Michael Milone, Christoph Ellebrecht, left to right
Aimee Payne, Michael Milone, Christoph Ellebrecht, left to right

CART-Cells are T-lymphocytes that have been extracted from the peripheral blood of cancer patients and then genetically engineered to express a receptor that specifically recognizes a protein on the surface of tumor cells. These chimeric antigen receptor (CAR)-expressing cytotoxic T-lymphocytes have the ability to recognize and destroy tumor cells, which shrinks the tumor and potentially cures the patient.

CAAR technology

The core concepts behind CAR T-cells were first described in the late 1980s. Unfortunately, technical challenges prevented the development of this technology until later. However, since 2011, experimental CAR T cell treatments for B cell leukemias and lymphomas have been successful in some patients for whom all standard therapies had failed.

Antibody-producing B-lymphocytes or B-cells can also cause autoimmunity. A few years ago, a postdoctoral researcher in Payne’s laboratory named Dr. Christoph T. Ellebrecht came upon CAR T cell technology as a potential strategy for deleting rogue B-cells that make antibodies against a patient’s own tissues. Soon Payne and her team had teamed up with Milone’s, which studies CAR T cell technology. Their goal was to find a new way to treat autoimmune diseases.

“We thought we could adapt this technology that’s really good at killing all B cells in the body to target specifically the B cells that make antibodies that cause autoimmune disease,” said Milone.

“Targeting just the cells that cause autoimmunity has been the ultimate goal for therapy in this field,” noted Payne.

Because an excellent mouse model existed for PV, Payne and Milone decided to examine pemphigus vulgaris. Since PV consists of a patient’s antibodies attacking those molecules that normally keep skin cells together, it can cause extensive skin blistering and is almost always fatal. PV is treatable with broadly immunosuppressive drugs such as prednisone, mycophenolate mofetil, and rituximab.

However, to treat PV without causing broad immunosuppression, the Penn team designed an artificial CAR-type receptor that would home the patient’s own genetically engineered T-cells exclusively to those B-cells that produce harmful anti-Dsg3 antibodies.

Payne and Milone and their colleagues developed a “chimeric autoantibody receptor,” or CAAR, that displays fragments of the Dsg3 on their cell surfaces. Since the Dsg3 protein is the target of the PV-causing B-cells, the CAAR acts as a lure for the rogue B cells that target Dsg3. The CAAR effectively brings the cells into fatal contact with the therapeutic T cells.

After testing a battery of different cultured, genetically engineered T-cells, these teams eventually found a CAAR that worked well in cell culture and enabled host T cells to efficiently destroy anti-desmoglein-producing B-cells. These cultured cells worked so well that they even killed B-cells isolated from PV patients. The engineered CAAR T cells also performed successfully in a mouse model of PV. The CAAR T-cell effectively killed desmoglein-specific B cells, prevented blistering, and other manifestations of autoimmunity in the animals. “We were able to show that the treatment killed all the Dsg3-specific B cells, a proof of concept that this approach works,” Payne said.

Not only were these treatments devoid of undesirable side effects in the laboratory mice they studied, but they maintained their potency despite the presence of high levels of anti-Dsg3 antibodies that might have swamped out their CAARs.

Next, Payne plans to test her treatment in dogs, which can also develop PV and often die from it. “If we can use this technology to cure PV safely in dogs, it would be a breakthrough for veterinary medicine, and would hopefully pave the way for trials of this therapy in human pemphigus patients,” Payne said.

Penn scientists would also like to develop applications of CAAR T cell technology for other types of autoimmunity. Organ transplant rejection, which is also related to autoimmunity, complicates organ transplants, and normally requires long-term immunosuppressive drug therapy, may also be treatable with CAAR T cell technology.

“If you can identify a specific marker of a B cell that you want to target, then in principle this strategy can work,” Payne said.

U of Penn Group Releases Hopeful Results of CAR T-Cells Trial

Chimeric Antigen Receptor T-Cells (CART-cells) are a type of genetically engineered type of immune cell that represents one of the most promising avenues of cancer therapy. Such treatments can induce sustained remissions in patients with stubborn disease.

Studies with CART-cells have been tested in patients with relapsed and stubborn chronic lymphocytic leukemia (CLL). Now a new publication by Porter and others reports the results of a clinical trial that examined CART-cells as a treatment for blood-based cancers. This study reports that infused CART-cells were functional up to 4 years after treatment. Patients also achieved completely remission, and no patient who achieved complete remission relapsed, and no minimal residual disease was detected, suggesting that in a subset of patients, CAR T cells may drive disease eradication.

Patients enrolled in this study suffered from CLL and had a poor prognosis. The CART-cells employed in this study targeted the molecule CD19. Porter and others report the mature results of the treatment of 14 patients with relapsed and refractory CLL.

The patient’s own T-Cells were extracted from circulating blood, and genetically engineered to express a CD19-directed receptor. Patients received doses of 0.14 × 10[8] to 11 × 10[8] CTL019 cells. Patients were monitored for toxicity, response, expansion, and persistence of circulating CTL019 T cells.

The overall response rate in these heavily pretreated CLL patients was 8 of 14 (57%), and there were 4 complete remissions (CR) and 4 partial remissions (PR). The expansion of the CAR T-cells in culture correlated with clinical responses; the better the engineered T-cells grew in culture the better they performed in the Patient’s bodies. Furthermore, the CAR T-cells persisted and remained functional beyond 4 years in the first two patients achieving Complete Remission. None of the patients who experienced Complete Remission have relapsed.

All the patients who responded to the treatment developed “B cell aplastic” (abnormally low B-cell levels) and experienced cytokine release syndrome, which was part and partial of T cell proliferation.

Minimal residual disease was not detectable in patients who achieved Complete Remission, suggesting that disease eradication may be possible in some patients with advanced CLL.

Preclinical Study Results Pave the Way for Newly Opened Clinical Trial of Immune Cells Engineered to Attack Protein Found on Tumors in 30 Percent of Patients with Glioblastoma

Scientists from the University of Pennsylvania have engineered immune cells to seek out and attack a type of deadly brain cancer. In an important preclinical study, these souped-up immune cells were shown to be both safe and effective at controlling tumor growth in mice treated with these modified cells. This work is the result of collaboration between a team from the Perelman School of Medicine at the University of Pennsylvania and the Novartis Institutes for BioMedical Research. These results will hopefully be the impetus for a newly opened clinical trial for glioblastoma patients at Penn.

Marcela Maus, assistant professor of Hematology/Oncology at the Penn Abramson Cancer Center, said: “A series of trials that began in 2010 have found that engineered T cells have an effect in treating some blood cancers, but expanding this approach into solid tumors has posed challenges. A challenging aspect of applying engineered T cell technology is finding the best targets that are found on tumors but not normal tissues. This is the key to making this kind of T cell therapy both effective and safe.”

This new preclinical study, which was conducted with Hideho Okada and his colleagues at the University of Pittsburgh, makes use of T cells engineered to express a chimeric antigen receptor (CAR) that specifically binds to a mutant epidermal growth factor receptor protein called EGFRvIII. EGFRvIII is found on the cell surfaces of approximately 30 percent of glioblastoma tumors. Over 22,000 Americans are diagnosed with glioblastoma each year, and those patients whose glioblastomas express the EGFRvIII mutation tend to be more aggressive and are less likely to respond favorably to standard therapies and more likely to recur after treatment.

“Patients with this type of brain cancer have a very poor prognosis. Many survive less than 18 months following their diagnosis,” said M. Sean Grady, who is the Charles Harrison Frazier Professor and chair of the department of Neurosurgery. “We’ve brought together experts in an array of fields to develop an innovative personalized immunotherapy for certain brain cancers.”

This new trial is being led by Donald O’Rourke, associate professor of Neurosurgery, who heads an interdisciplinary collaboration of neurosurgeons, neuro-oncologists, neuropathologists, immunologists, and transfusion medicine experts.

In order to bring this experiment to fruition, Maus and her colleagues had to characterize the EGFRvIII CAR T cell. They had to develop and tested multiple antibodies that bind to cells expressing EGFRvIII on their surface. The single-chain variable fragments or scFvs that recognized the mutant EGFRvIII protein were then extensively tested in order to confirm that they do not also bind to those normal, EGFR proteins that are widely expressed on cells in the human body.

Maus and her group then generated a panel of humanized scFvs and tested their specificity and function in CAR modified T cells. The humanized scFvs have distinct amino acid sequences that more closely resemble human antibodies. From this huge panel of humanized scFvs, they selected one scFv to explore further based on its binding selectivity for EGFRvIII over normal non-mutated EGFR. They also evaluated the EGFRvIII CAR T cells by testing them against normal EGFR-expressing skin cells in mice grafted with human skin. This test showed that the engineered EGFRvIII CAR T cells did not attack cells with normal EGFR, at least under these conditions.

In order to test the selected scFv for its anti-cancer efficacy, Maus and others used human tumor cells that expressed EGFRvIII and showed that the EGFRvIII CAR T cells could multiply and secrete cytokines in response such to tumor cells. When used inside living animals, it was clear that the EGFRvIII CAR T cells ably controlled tumor growth in several mouse models of glioblastoma. The tumors were measured with magnetic resonance imaging (MRI) and the EGFRvIII CAR T cells caused tumor shrinkage, and were also effective with used in combination with the anticancer drug temozolomide, which is normally used to treat patients with glioblastoma.

On the strength of these preclinical successes, this team designed a phase 1 clinical study of CAR T cells transduced with humanized scFv directed to EGFRvIII for both newly diagnosed and recurrent glioblastoma patients who carry the EGFRvIII mutation. “There are unique aspects about the immune system that we’re now able to utilize to study a completely new type of therapy,” said O’Rourke.

For these glioblastoma patients, their T cells were removed by means of apheresis (a process similar to dialysis), and then the T cells were genetically engineered using a viral vector that programs them to find EGFRvIII-expressing cancer cells. The patient’s own engineered cells are infused back into their body, and when the T cells find the EGFRvIII-expressing cells, a signaling domain built into the CAR promotes proliferation of these “hunter” T-cells. This procedure is distinct from other T cell-based therapies that also target some healthy cells, since EGFRvIII seems to only be found on tumor tissue, which the study’s leaders hope will minimize side effects.

This new phase I clinical trial will enroll 12 adult patients whose tumors express EGFRvIII, in two groups: One arm of 6 patients whose cancers have returned after receiving other therapies, and one arm of 6 patients who are newly diagnosed with the disease and still have 1 cm or more of tumor tissue remaining after undergoing surgery to remove it.

The clinical trial is sponsored by the biotech company Novartis. In 2012, the University of Pennsylvania and Novartis announced an exclusive global research and licensing agreement to further study and commercialize novel cellular immunotherapies using CAR technologies. This STM study is the first pre-clinical paper developed within the Penn-Novartis alliance, with Penn and Novartis scientists working collaboratively. Ongoing clinical trials evaluating a different type of Penn-developed CAR therapy known as CTL019 have yielded promising results among some patients with certain blood cancers. In July 2014, the FDA granted CTL019 its Breakthrough Therapy designation for the treatment of relapsed and refractory acute lymphoblastic leukemia in both children and adults.

CAR Immune Cells to Treat Childhood Cancers

In clinical trials, cancer treatments that use genetically modified versions of a patient’s own cells to specifically target the disease have remarkable results. The next step for these companies that spent enormous amounts of time, capital, and intellectual energy inventing and designing these treatments is to get them into hospitals despite their enormous price tags.

Novartic CAR T-Cell therapy

In two separate clinical trials, one sponsored by the Swiss company Novalis AG and another by the Seattle-based biotech company Juno Therapeutics Inc., close to 90% of all patients saw their leukemia completely disappear after being given experimental “CAR” or “chimeric antigen receptor” T-cell therapies.

Both trials examined small numbers of patients (22 children in the Novartis trial and 16 adults in the Juno trial). These patients had acute lymphoblastic leukemia, which is the most common childhood cancer. All of them had also not responded to the available standard treatments. Consequently, both companies are now conducting larger trials.

“CAR T cells are probably one of the most exciting concepts and fields to come out in cancer in a very, very long time,” says Dr. Daniel DeAngelo, a Boston-based hematologist and associate professor of medicine at Harvard Medical School, who wasn’t involved in either study.

Usman Azam, head of cell and gene therapies at Novartis, calls the therapies “critically important” for Novartis. “I think that a cure for cancers such as leukemia and lymphoma through a CAR technology is plausible,” said Dr. Azam in an interview with The Wall Street Journal. “Our job is to get this into patients as soon as we feasibly can.”

Novatis created a new research unit headed by Dr. Azam. Novartis’ rationale is to accelerate the advent of CAR T-Cell Therapy to medical markets. The U.S. Food and Drug Administration (US FDA) granted Novartis’ leading CAR therapy “breakthrough” designation in July of 2014. Presently Novartis wants to file it with regulators in 2016.

CAR therapies use the patient’s own immune system to fight the cancer, but with a genetic-engineering twist. “Immunotherapies,” culture immune cells from the patient and manipulate them in culture to sensitize them to the cancer. CAR therapies extract T-cells, which are disease-fighting white blood cells, from a patient’s blood. These T-cells are then genetically engineered and grown in a laboratory for around 10 days and reintroduced into the patient.

The T-cells are usually infected with a hamstrung virus that can introduce genes into cells but cannot productively infect them. These recombinant viruses endows the T-cells with genes that encode chimeric antigen receptors, or CARs. CARS bind specifically to proteins on the surface of malignant cancer cells. Once attached to the cancer cells, the T-cells can kill them very effectively.

Both Novartis and Juno are tapping academic scientists to develop their treatments. For example, Novartis has teamed with the University of Pennsylvania and Juno has formed a formal relationships with scientists at Memorial Sloan-Kettering Cancer Center in New York, Seattle Children’s Hospital and the Fred Hutchinson Cancer Research Center, which is also in Seattle.

Even though Novartis and Juno will probably be the first to bring their immunotherapies to the market, other companies are also in the hunt to bring similar therapies to medical markets. Pfizer Inc., Kite Pharma Inc., and Celgene Corp., which is working in collaboration with Bluebird Bio Inc. all are developing competing strategies.

“Competition will keep all of the companies involved on their toes,” said Hans Bishop, Juno’s chief executive.

Unfortunately, CAR therapies still have a few unanswered questions surrounding them. For example: “How long do they last?” Given the small numbers of patients who have been treated with these treatments to date, it is very hard to tell with the available data. Another confounding factor is that those patients in the previous clinical trials whose cancer went into remission after the CAR therapies then became eligible for stem-cell transplants, which can also prolong survival.

Secondly, a potentially dangerous side effect called “cytokine-release syndrome,” shows the therapy is working, but can cause a sharp drop in blood pressure and a surge in the heart rate. The deaths of two patients in a Juno-backed Sloan-Kettering trial in March caused a temporary halt in the study because of worries over these particular adverse reactions.  “Patients need to be healthy enough to combat that side effect,” says Mr. Bishop, who thinks it is now manageable. Patients are once again being recruited for this trial, and patients with a risk of heart failure are excluded, and the modified cell dose for patients with very advanced leukemia also has been lowered.

But largest hurdle of all will probably be the cost of these therapies. Since they are a genetically engineered product, CAR T-cells are very complex to manufacture; each batch is composed of unique, personalized T-cells that were made from a patient’s own blood cells. The inability to mass-produce CAR T-cells will definitely increase the price companies charge for them.

“What we’re talking about here is a single, very expensive therapy that’s used once for a specific patient and is not generalizable,” says Dr. Malcolm Brenner, director of the Center for Cell and Gene Therapy at the Texas Children’s Hospital in Houston, who, in MArch, signed an agreement to commercialize his own CAR research with Celgene.

Novartis and Juno both insist that it is too early to speculate on the price of the treatment, but Dr. Usman agrees the challenge is getting the manufacturing process to “a viable level where it’s both affordable and attractive.”

Citigroup believes CAR therapies could cost in excess of $500,000 per patient, which it notes is roughly in line with the cost of a stem cell transplant, even though most analysts think it is too early to estimate potential revenue or price.

“This technology needs to be widely developed and accessible to patients,” says Dr. DeAngelo. “If the cost is going to be a hindrance, it’s going to be a really sad day.”

Scalability and cost are one reason Pfizer is taking a different approach to this field. “We would like to take it to the next level, where CAR therapies become a more standardized, highly controlled treatment,” said Mikael Dolsten, Pfizer’s head of global research and development.

Working with French biotech Cellectis SA, Pfizer wants to develop a generic CAR therapy for use in any patient. While this will certainly lower the cost of the treatment, since it is the result of a mass-produced, off-the-shelf-product, this work is still at the preclinical stages and may not work in humans.

Global head of health-care research at Société Générale, Stephen McGarry, thinks that the revolutionary treatments being developed by Novartis and Juno could justify “astronomical” prices, he believes health-care payers and patients will probably protest such high prices. “When you look at the initial data with the Novartis therapy, you’re getting cures in some kids—what do you charge for that?” he asks.