How Skeletal Stem Cell form the Blueprint of the Face


A new study from the laboratory of University of Southern California (USC) Stem Cell researcher J Gage Crump, who is at the Eli and Edythe Broad CIRM Center for Regenerative Medicine and Stem Cell Research, has identified the key molecular signals that control the critical timing of the development of the vertebrate face.

Previous work has demonstrated that two molecular signals, in particular the JaggedNotch and Endothelin 1 signaling, are integral for shaping the face. Loss of either of these signals results in facial deformities in zebrafish and humans. This illustrates the essential contribution these signaling pathways make to the development of the face.

Lindsey Barske, a researcher in Crump’s laboratory and her colleagues utilized sophisticated genetic, genomic, and imaging tools to study face formation in zebrafish and showed that the Jagged-Notch and Endothelin 1 pathways work in tandem to control when and where the facial stem cells form face-specific cartilage.

In the lower part of the face, the Endothelin 1 signal accelerates cartilage formation early in development, but in the upper face, the Jagged-Notch signal transduction pathway produces signals that prevent stem cells from making cartilage until later in development.

Barske and her colleagues discovered that these timing differences in facial stem cell activity and facial cartilage production play a major role in making the upper and lower cartilage regions of the face.

The earliest blueprint of the facial skeleton is established by intersecting signals that control when stem cells transform cartilage into bone. It also appears that small tweaks to the timing of these events accounts for the different skull shapes observed in vertebrate animals. Also, small, nuanced changes in facial cartilage production and ossification can also account for the diverse array of facial shapes observed in humans.

This work was published in PLOS Genetics 12(4): e1005967. doi:10.1371/journal.pgen.1005967.

Skin Cell to Eye Transplantation Successful


A presentation at the annual meeting of the Association for Research in Vision and Ophthalmology in Seattle, Washington has reported the safe transplantation of stem cells derived from a patient’s skin to the back of the eye in an effort to restore vision. The subject for this research project suffered from advanced wet age-related macular degeneration that did not respond to current standard treatments.

A small skin biopsy from the patient’s arm was collected and reprogrammed into induced pluripotent stem cells (iPSCs). The iPSCs were then differentiated into retinal pigmented epithelial (RPE) cells, which were transplanted into the patient’s eye. The transplanted cells survived without any adverse events for over a year and resulted in slightly, though significantly, improved vision.

iPSCs are adult cells that have been reprogrammed to an embryonic stem cell-like state, which can then be differentiated into any cell type found in the body.

Abstract Title: #3769: Transplantation of Autologous induced Pluripotent Stem Cell-Derived Retinal Pigment Epithelium Cell Sheets for Exudative Age Related Macular Degeneration: A Pilot Clinical Study by Yasuo Kurimoto and others from the laboratory of Masayo Takahashi’s laboratory at the RIKEN Center for Developmental Biology in Kobe, Japan.

Unfortunately, this clinical trial has been suspended because iPSCs made from other patients proved to possess too many genetic abnormalities.  Therefore, Takahashi and her colleagues have decided that allogeneic iPSCs differentiated into RPEs will probably do a better job than the patient’s own skin cell-derived iPSCs.

Positive Results from Phase 2 Study in Spinal Cord Injury


Stem Cells, Inc., has released the six-month results from cohort I of an ongoing Phase 2 clinical trial of human neural stem cells for the treatment of chronic cervical spinal cord injuries. The data displayed significant improvements in muscle strength had occurred in five of the six patients treated. Of these five patients, four of them also showed improved performance on functional tasks that assesses dexterity and fine motor skills. Furthermore, these four patients improved in the level of spinal cord injury according to the classification system provided by the International Standards for Neurological Classification of Spinal Cord Injury or ISNCSCI.

Stem Cells, Inc., expects to release their detailed final 12-month results on this first open-cohort later this quarter.

Chief medical officer, Stephen Huhn, presented these data at the American Spinal Injury Association annual meeting in Philadelphia, on Friday, April 15.  Dr. Huhn also believes that the interim results are very encouraging and reason to be quite hopeful.

“The emerging data continue to be very encouraging,” said Dr. Huhn. “We believe that these types of motor changes will improve the independence and quality of life of patients and are the first demonstration that a cellular therapy has the ability to impact recovery in chronic spinal cord injury. We currently have thirteen sites in the United States and Canada that are actively recruiting patients. We have enrolled and randomized 19 of the 40 total patients in the statistically powered, single-blind, randomized controlled, Cohort II. We are projecting to complete enrollment by the end of September so that we can have final results in 2017.”

The present Phase 2 clinical trial is a multi-center enterprise that includes physicians and scientists at 13 different sites in the united States and Canada. Incidentally, these sites are presently actively recruiting patients.

Stem Cells, Inc., has enrolled and randomized 19 of the 40 total patients in this statistically powered, single-blind, randomized controlled, cohort II.

The Phase 2 study, “Study of Human Central Nervous System (CNS) Stem Cell Transplantation in Cervical Spinal Cord Injury,” will determine the safety and efficacy of transplanting the company’s proprietary human neural stem cells (HuCNS-SC cells) into patients with traumatic injury of the cervical region of the spinal cord.

Cohort I is an open label dose-ranging cohort in six AIS-A or AIS-B subjects. For those of you not familiar with the American Spinal Injury Impairment Scale (ASI A-E scale), here is a summary of the classification scheme:

ASI – A = Complete paralysis; No sensory or motor function is preserved in the sacral segments S4-5.
ASI – B = Sensory Incomplete; Sensory but not motor function is preserved below the neurological level and includes the sacral segments S4-5 (light touch or pin prick at S4-5 or deep anal pressure) AND no motor function is preserved more than three levels below the motor level on either side of the body.
ASI – C = Motor Incomplete; Motor function is preserved below the neurological level**, and more than half of key muscle functions below the neurological level of injury (NLI) have a muscle grade less than 3 (Grades 0-2).
ASI – D = Motor Incomplete; Motor function is preserved below
the neurological level**, and at least half (half or more) of key muscle functions below the NLI have a muscle grade > 3.
ASI – E = Normal; If sensation and motor function as tested with the ISNCSCI are graded as normal in all segments, and the patient had prior deficits, then the AIS grade is E. Someone without an initial SCI does not receive an AIS grade.
Cohort II is a randomized, controlled, single-blinded cohort in forty AIS-B subjects. Cohort III, which will only be conducted at the discretion of the sponsor, is an open-label arm that involves six AIS-C subjects.
The primary efficacy outcome will focus on changes in the upper extremity strength as measured in the hands, arms, and shoulders.  This trial will enroll up to 52 subjects.
StemCells, Inc. has demonstrated the safety and efficacy of their HuCNS-SC cell in preclinical studies in laboratory rodents.  Additional Phase I studies yielded positive human safety data.  Furthermore, completed and ongoing clinical studies in which its proprietary HuCNS-SC cells have been transplanted directly into all three components of the central nervous system: the brain, the spinal cord and the retina of the eye, have further demonstrated the safety of HuCNS SC cells in human patients.
StemCells, Inc. clinicians and scientists believe that HuCNS-SC cells may have broad therapeutic application for many diseases and disorders of the CNS. Because the transplanted HuCNS-SC cells have been shown to engraft and survive long-term, there is the possibility of a durable clinical effect following a single transplantation.
The HuCNS-SC platform technology is a highly purified composition of human neural stem cells (tissue-derived or “adult” stem cells). Manufactured under cGMP standards, the Company’s HuCNS-SC cells are purified, expanded in culture, cryopreserved, and then stored as banks of cells, ready to be made into individual patient doses when needed.

SEPCELL Trial Tests Fat-Derived Stem Cells as a Treatment for Sepsis


The Belgium-based biotechnology company, TiGenix, has launched a clinical trial entitled SEPCELL that uses fat-derived stem cells (called Cx611) to treat severe sepsis secondary to acquired pneumonia (also known as sCAP). SEPCELL is a randomized, double-blind, placebo-controlled, Phase 1b/2a study of sCAP patients who require mechanical ventilation and/or vasopressors.

SEPCELL will, hopefully, enroll 180 patients and will be conducted at approximately 50 centers throughout Europe. Subjects who participate in this trial will be randomly assigned to receive either an investigational product or placebo on days 1 and 3. All patients will be treated with standard care, which usually includes broad-spectrum antibiotics and anti-inflammatory drugs.

The primary endpoint of this clinical trial will examine the number, frequency, and type of adverse reactions during the 90-day period of the trial. The secondary endpoints of the SEPCELL trial include reduction in the duration of mechanical ventilation and/or vasopressors, overall survival, clinical cure of sCAP, and other infection-related endpoints. SEPCELL will also assess the safety and efficacy of the expanded allogeneic adipose stem cells (eASCs) that will be intravenously delivered to some of the patients in this study.

The SEPCELL trial will be managed by TFS International, a company based in Lund, Sweden. TFS has extensive experience in running sepsis trials and hospital-based trials.

Sepsis is a potentially life-threatening complication of infection that occurs when inflammatory molecules (cytokines and chemokines) released into the bloodstream to fight the infection trigger systemic inflammation.  This body-wide inflammation has the ability to trigger a cascade of detrimental changes that damage multiple organ systems and cause them to fail. If sepsis progresses to “septic shock,” blood pressure drops dramatically, which may lead to death. Patients with “severe sepsis” require close monitoring and treatment in a hospital intensive care unit. Drug therapy is likely to include broad-spectrum antibiotics, corticosteroids, vasopressor drugs to increase blood pressure, as well as oxygen and large amounts of intravenous fluids. Supportive therapy may be needed to stabilize breathing and heart function and to replace kidney function. Patients with severe sepsis have a low survival rate so there is a critical need to improve the effectiveness of current therapy. Only a small number of new molecular entities are currently in development for severe sepsis.

Severe sepsis and septic shock significantly affect public health and these event also are leading causes of mortality in intensive care units.

Severe sepsis and septic shock have an incidence of about 3 cases per 1,000, but due to the aging of the population and an increase in drug resistant bacteria.

Cx611 is an intravenously-administered concoction that consists of allogeneic eASCs. These cells are largely mesenchymal stem cells that secrete an impressive array of molecules that suppress the type of immune responses that damage organs during events like septic shock.  eASCs have a higher proliferation rate in culture and faster attachment than bone marrow-based mesenchymal stem cells in cell culture.  ASCs are also less prone to senescence and differentiation.  Their differentiation capacity decreases with expansion time without losing immunomodulatory properties.  These eASCs also have superior inflammation targeting capacities than bone marrow-based mesenchymal stem cells, and are safe, since they do not express ligands for receptors on Natural Killer cells that, and therefore, are unlikely to elicit an immune rejection.

In May 2015, TiGenix completed a Phase 1 sepsis challenge that demonstrated that Cx611 is safe and well tolerated. That trial began in December 2014, and was a placebo-controlled dose-ranging study (3 doses of eASC’s) in which 32 healthy male volunteers were randomized to receive Cx611 or placebo in a ratio of 3:1. Primary endpoints were vital signs and symptoms, laboratory measures and functional assays of innate immunity. All 32 volunteer subjects were recruited and dosed by March 2015. By May, 2015, the phase I trial data essentially demonstrated the safety and tolerability of Cx611.  On the strength of that phase I trial, TiGenix designed a Phase 1b/2a trial in severe sepsis secondary to sCAP in which they expecet to enroll 180 subjects across Europe.

SEPCELL was funded by a €5.4 million grant ($6.14 million) from the European Union.

CardioCell LLC Clincal Trial Tests Ischemia-Resistant Mesenchymal Stem Cells in Heart Failure


The cell therapy company CardioCell LLC has completed enrolling 23 patients for its Phase 2a chronic heart failure trial. These subjects were enrolled at Emory University in Atlanta, GA, MedStar Washington Hospital Center in Washington DC, and three other hospitals.

This study has the ponderous title of “Single-blind, Placebo-controlled, Crossover, Multicenter, Randomized Study to Assess the Safety, Tolerability and Preliminary Efficacy of Single Intravenous Dose of Ischemia-tolerant of Allogeneic Mesenchymal Bone Marrow Cells to Subjects With Heart Failure of Non-ischemic Etiology.”

This clinical trial will examine the safety of CardioCell’s proprietary ischemic-tolerant mesenchymal stem cells in heart failure patients. The trial will also test the ability of these cells to improve the heart function of these safe patients.

Ischemia-resistant mesenchymal stem cells have are extracted from bone marrow and then subjected to harsh cell culture conditions that toughen them up and improves their therapeutic capacities.

Cardiologist Javed Butler said that this clinical trial has been designed to use this novel intervention in a carefully selected group of patients who met rigorous inclusion and exclusion criteria.

This trial will deliver ischemia-tolerant mesenchymal stem cells (itMSCs) by means of intravenous infusion into heart failure patients and then monitor these patients to determine if the itMSC-treated patients show signs of improvement in heat function.

These itMSCs are licensed under the parent company Stemedica and these are allogeneic cells that were isolated from young, healthy donors and grown under hypoxic conditions. Once grown under these harsh culture conditions, the itMSCs increase their ability to home to damaged tissues and engraft into those tissues. itMSCs also secrete increased levels of growth and trophic factors that promote neurogenesis and tissue healing.

Insulin-Secreting Beta Cells from Human Fat


In a study led by Martin Fussenegger of ETH Zurich, stem cells extracted from the fat of a 50-year-old test subject were transformed into mature, insulin-secreting pancreatic beta cells.

Fussenegger and his colleagues isolated stem cells from the fat of a 50-year-old man and used these cells to make induced pluripotent stem cells (iPSCs). These iPSCs were then differentiated into pancreatic progenitor cells and then into insulin-secreting beta cells but means of a “genetic software” approach.

Genetic software refers to the complex synthetic network of genes required to differentiate pancreatic progenitor cells into insulin-secreting beta cells. In particular, three genes, all of which expression transcription factors, Ngn3, Pdx1, and MafA, are particularly crucial for beta cell differentiation.

Fussenegger and his team designed a a protocol that would express within these fat-based stem cells the precise concentration and combination of these transcription factors. This feature is quite important because the concentration of these factors changes during the differentiation process. For example, MafA is not present at the start of beta cells maturation, but appears on day four on the final data of maturation when its concentration rises precipitously. The concentration of Ngn3 rises and then falls and the levels of Pdx1 rise at the beginning and towards the end of maturation.

The Zurich team used ingenious genetic tools to reproduce these vicissitudes of gene expression as precisely as possible. By doing so, they were able to differentiate the iPSC-derived pancreatic progenitor cells into insulin-secreting beta cells.

This work was published in Nature Communications 7, doi:10.1038/ncomms11247.

The fact that Fussenegger’s team was able to use a synthetic gene network to form mature beta cells from adult stem cells is a genuine breakthrough. The genetic network approach also seems to work better than the traditional technique of adding various chemicals and growth factors to cultures cells. “It’s not only really hard to add just the right quantities of these components (growth factors) at just the right time, it’s also inefficient and impossible to scale up,” said Fussenegger.

This new process can successfully transform three out of four fat stem cells into beta cells. Also the beta cells made with this method have the same microscopic appearance of natural beta cells in that they contain internal granules full of insulin. They also secrete insulin in response to increased blood glucose concentrations. Unfortunately the amount of insulin made by these cells is lower than that made by natural beta cells.

Pancreatic islet transplants have been performed in diabetic patients, but such transplantations also require treatment with potent antirejection drugs that have potent side effects.

“With our beta cells, there would likely be no need for this action (administering antitransplantation drugs), since we can make them using endogenous cell material taken from the patient’s own body. This is why our work is of such interest in the treatment of diabetes,” said Fussenegger.

Fussenegger and his group have made these beta cells in the laboratory, but they have yet to transplant them into a diabetic patient. However, the success of this synthetic genetic software technology might also be useful in the reprogramming of adult cells into other types of cells that are useful for therapeutic purposes.

RENEW Trial Shows Stem Cell Mobilization Has Some Potential for Refractory Angina


The RENEW clinical trial has examined the ability of “CD34+” stem cells from bone marrow to alleviate the symptoms of refractory angina.

Angina pectoris is a crushing chest pain that afflicts people when the heart receives too little oxygen to support it for the workload placed upon it. Angina pectoris typically results from the blockage of coronary arteries as a result of atherosclerosis. Treatment of angina pectoris usually includes PCI or percutaneous coronary intervention, which involves the placement of a stent in the narrowed coronary artery, in combination with drug treatments like beta blockers, and/or cardiac nitrate (e.g., nitroglycerine).

Angina pectoris is also classified according to the severity of the disease. The Canadian Cardiovascular Society grading of angina pectoris (which is very similar to the New York Heart Association classification) uses four classes (I-IV) to classify the disease. Patients with Class I angina only experience pain during strenuous or prolonged physical activity. Those with Class II angina have a slight limitation in physical activity and experience pain during vigorous physical activity (climbing several flights of stairs). Class III angina manifests as pain during everyday living activities, such as climbing one flight of stairs. These patients experience moderate limitation of their physical activity. Those with Class IV angina experience pain at rest and are unable to perform any activity without angina, and therefore, suffer from severe limitations on their activity.

Refractory angina pectoris (also known as chronic symptomatic coronary artery disease) stubbornly resists medical therapy and is unamenable to conventional revascularization procedures. Patients with refractory angina pectoris have reproducible lifestyle-limiting symptoms of chest pain, shortness of breath, and easy fatigability.

The results of the RENEW clinical trial were presented at the Society for Cardiovascular Angiography and Interventions 2016 sessions. Even though the trial was prematurely ended for financial reasons, the results that were collected suggest that cell-based therapies might provide relief for suffers of refractory angina pectoris.

RENEW tested the effectiveness of the intravenous infusion of the protein called granulocyte-colony simulating factor (G-CSF), which mobilizes CD34+ stem cells from the bone marrow. Once summoned from the bone marrow, CD34+ stem cells can help establish new blood vessels and increase blood flow throughout the heart. CD34+ stem cells also seem to have some ability to home to sites of damage. Therefore, G-CSF infusions might provide some relief to patients with refractory angina pectoris.

Dr. Timothy D. Henry of the Cedars-Sinai Heart Institute in Los Angles, CA, said: “Clinicians are seeing more RA (refractory angina) patients because people are living longer. Unfortunately, despite better medical care, these people are still confronting ongoing symptoms that affect their daily lives.”

Patients enrolled in the RENEW trial had either class III or IV angina and experiences ~7 chest pain episodes each week. These patients were also not candidates for revascularization (PCI) and their treadmill exercise times were between 3-10 minutes.

112 RA patients were randomly broken into three groups. Group 1 received standard care (28), group 2 received placebo injections (27), and group 3 received treatment with CD34+ cells. The trial was double-blinded and placebo controlled. The original aim was to test 444 RA patients, but financial concerns truncated the study at 112 patients.

All patients were assessed at three, six, 12, and 24 months after treatment by means of exercise tolerance, anginal attacks, and major adverse cardiovascular events (MACEs).

The cell-treated patients increased their exercise times by more than two minutes at three (average 122-second increase), six (average 142-second increase), and twelve (average 124-second increase) months. This is significant, since the other two groups showed no significant increase in their exercise times.

Patients in the cell-treated group also experienced 40 percent fewer anginal attacks at six months relative to the placebo-treated group.

At two years after the treatment, the CD34+-treated group have lower mortality rates (3.7 percent) compared to those who received standard care (7.1 percent) and those who received the placebo (10 percent).

Finally, after two years, the cell-treated group had lower MACE rates (46 percent) than the standard care group (68 percent). The MACE rate for the placebo-treated group was 43 percent.

On the strength of these results, Dr. Henry said, “Cell therapy appears to be a promising approach for these patients who have few options. Our results were consistent with phase 2 results from the ACT34 trial (author’s note: which gave patients infusions of cells and not G-CSF).”

Tom Povsic of the Duke Clinical Research Institute said of the RENEW trial, “It is unfortunate the early termination of this study precludes a full evaluation of the efficacy of this therapy for these patients with very few options.  Studies like RENEW are critical to developing reliable and effective therapies for heart patients, and continued cellular therapies for heart patients, and continued funding is essential to advancing the work that this study began.  We need to find a way to bring these therapies as quickly as safely as possible.”

Dr. Povsic’s words certainly ring true.  Even though the results of the RENEW study are essentially positive, RENEW was planed to be almost three times the size of Douglas Losordo’s earlier, successful ACT34 study.  The results of both the ACT34 and RENEW studies are largely positive.  Perhaps more importantly, both studies have also established that cell-based treatments for RA patients are safe.  However, given the voracity of the FDA for clinical data before it will approve a treatment, even for patients with few current options, it is unlikely that these studies will prove large enough to satisfy the agency.  Until a very large study shows cell-based treatments to be not only safe but efficacious, only then will the mighty turtle known as the FDA approve such treatments for RA patients.