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.

Skin Cells Converted into Blood Cells By Direct Reprogramming


Making tissue-specific progenitor cells that possess the ability to survive, but have not passed through the pluripotency state is a highly desirable goal of regenerative medicine. The technique known as “direct reprogramming” uses various genetic tricks to transdifferentiate mature, adult cells into different cell types that can be used for regenerative treatments.

Juan Carlos Izpisua Belmonte and his colleagues from the Salk Institute for Biological Studies in La Jolla, California and his collaborators from Spain have used direct reprogramming to convert human skin cells into a type of white blood cells.

These experiments began with harvesting skin fibroblasts from human volunteers that were then forced to overexpress a gene called “Sox2.” The Sox2 gene is heavily expressed in mice whose bone marrow stem cells are being reconstituted with an infusion of new stem cells. Thus this gene might play a central role is the differentiation of bone marrow stem cells.

Sox2 overexpression in human skin fibroblasts cause the cells express a cell surface protein called CD34. Now this might seem so boring and unimportant, but it is actually really important because CD34 is expressed of the surfaces of hematopoietic stem cells. Hematopoietic stem cells make all the different types of white and red blood cells in our bodies. Therefore, the expression of these protein is not small potatoes.

In addition to the expression of CD34, other genes found in hematopoietic stem cells were also induced, but not strongly. Thus overexpression of SOX2 seems to induce an incipient hematopoietic stem cell‐like status on these fibroblasts. However, could these cells be pushed further?

Gene profiling of hematopoietic stem cells from Umbilical Cord Blood identified a small regulatory RNA known as miR-125b as a factor that pushes SOX2-generated CD34+ cells towards an immature hematopoietic stem cell-like progenitor cell that can be grafted into a laboratory animal.

When SOX2 and miR-125b were overexpressed in combination, the cells transdifferentiated into monocytic lineage progenitor cells.

What are monocytes? They are a type of white blood cells and are, in fact, the largest of all white blood cells. Monocytes compose 2% to 10% of all white blood cells in the human body. They play multiple roles in immune function, including phagocytosis (gobbling up bacteria and other stuff), antigen presentation (identifying and altering other cells to the presence of foreign substances), and cytokine production (small proteins that regulate the immune response).

Monocytes express a molecule on their cell surfaces called CD14, and when human fibroblasts overexpressed Sox2 and miR-125b, they became CD14-expressing cells that looked and acted like monocytes. These cells were able to gobble up bacteria and other foreign material, and when transplanted into a laboratory animal, these directly reprogrammed cells generated cells that established the monocytic/macrophage lineage.

Cancer patients, and other patients with bone marrow diseases can have trouble making sufficient white blood cells. A technique like this can generate transplantable monocytes (at least in laboratory animals) without many of the drawbacks associated with reprogramming human cells into hematopoietic stem cells that possess true clinical potential. Also because this technique skips the pluipotency stage, it is potentially safer.

Stem Cell Treatments to Improve Blood Flow in Angina Patients


Angina pectoris is defined as chest pain or discomfort that results from poor blood flow through the blood vessels in the heart and is usually activated by activity or stress.

In Los Angeles, California, physicians have initiated a double-blind, multicenter Phase III clinical trial that uses a patient’s own blood-derived stem cells to restore circulation to the heart of angina patients.

This procedure utilizes state-of-the-art imaging technology to map the heart and generate a three-dimensional image of the heart. These sophisticated images will guide the physicians as they inject stem cells into targeted sites in the heart.

This is a double-blinded study, which means that neither the patients nor the researcher will know who is receiving stem-cell injections and who is receiving the placebo.

The institution at which this study is being conducted, University of Los Angeles (UCLA), is attempting to establish evidence for a stem cell treatment that might be approved by the US Food and Drug Administration for patients with refractory angina. The subjects in this study had received the standard types of care but did not receive relief. Therefore by enrolling in this trial, these patients had nothing to lose.

Dr. Ali Nasir, assistant professor of cardiology at the David Geffen School of Medicine and co-principal investigator of this study, said: “We’re hoping to offer patients who have no other options a treatment that will alleviate their severe chest pain and improve their quality of life.”

Before injecting the stem cells or the placebo, the team examined the three-dimensional image of the heart and ascertained the health of the heart muscle and voltage it generated. Damaged areas of the heart fail to produce adequate quantities of voltage and show low levels of energy.

Jonathan Tobis, clinical professor of cardiology and director of interventional cardiology research at Geffen School of Medicine, said: “We are able to tell by the voltage levels and motion which area of the [heart] muscle is scarred or abnormal and not getting enough blood and oxygen. We then targeted the injections to the areas just adjacent to the scarred and abnormal heart muscle to try to restore some of the blood flow.”

What did they inject? The UCLA team extracted bone marrow from the pelvic bones and isolated CD34+ cells. CD34 refers to a cell surface protein that is found on bone marrow stem cells and mediates the adhesion of bone marrow stem cells to the bone marrow matrix. It is found on the surfaces of hematopoietic stem cells, placental cells, a subset of mesenchymal stem cells, endothelial progenitor cells, and endothelial cells of blood vessels. These are not the only cells that express this cell surface protein, but it does list the important cells for our purposes. Once the CD34+ cells were isolated, the were injected into the heart through a catheter that was inserted into a vein in the groin.

CD34

The team hopes that these cells (a mixture of mesenchymal stem cells, hematopoietic stem cells, and endothelial progenitor cells) will stimulate the growth of new blood vessels (angiogenesis) in the heart, and improve blood flow and oxygen delivery to the heart muscle.

“We will be tracking patients to see how they’re doing,” said William Suh MD, assistant clinical professor of medicine in the division of cardiology at Geffen School of Medicine.

The goal of this study is to enroll 444 patients nation-wide, of which 222 will receive the stem cell treatment, 111 will receive the placebo, and 111 who will be given standard heart care.

Producing blood cells from stem cells could yield a purer, safer cell therapy


The journal Stem Cells Translational Medicine has published a new protocol for reprogramming induced pluripotent stem cells (iPSCs) into mature blood cells. This protocol uses only a small amount of the patient’s own blood and a readily available cell type. This novel method skips the generally accepted process of mixing iPSCs with either mouse or human stromal cells. Therefore, is ensures that no outside viruses or exogenous DNA contaminates the reprogrammed cells. Such a protocol could lead to a purer, safer therapeutic grade of stem cells for use in regenerative medicine.

The potential for the field of regenerative medicine has been greatly advanced by the discovery of iPSCs. These cells allow for the production of patient-specific iPSCs from the individual for potential autologous treatment, or treatment that uses the patient’s own cells. Such a strategy avoids the possibility of rejection and numerous other harmful side effects.

CD34+ cells are found in bone marrow and are involved with the production of new red and white blood cells. However, collecting enough CD34+ cells from a patient to produce enough blood for therapeutic purposes usually requires a large volume of blood from the patient. However, a new study outlined But scientists found a way around this, as outlined by Yuet Wai Kan, M.D., FRS, and Lin Ye, Ph.D. from the Department of Medicine and Institute for Human Genetic, University of California-San Francisco has devised a way around this impasse.

“We used Sendai viral vectors to generate iPSCs efficiently from adult mobilized CD34+ and peripheral blood mononuclear cells (MNCs),” Dr. Kan explained. “Sendai virus is an RNA virus that carries no risk of altering the host genome, so is considered an efficient solution for generating safe iPSC.”

“Just 2 milliliters of blood yielded iPS cells from which hematopoietic stem and progenitor cells could be generated. These cells could contain up to 40 percent CD34+ cells, of which approximately 25 percent were the type of precursors that could be differentiated into mature blood cells. These interesting findings reveal a protocol for the generation iPSCs using a readily available cell type,” Dr. Ye added. “We also found that MNCs can be efficiently reprogrammed into iPSCs as readily as CD34+ cells. Furthermore, these MNCs derived iPSCs can be terminally differentiated into mature blood cells.”

“This method, which uses only a small blood sample, may represent an option for generating iPSCs that maintains their genomic integrity,” said Anthony Atala, MD, Editor of STEM CELLS Translational Medicine and director of the Wake Forest Institute for Regenerative Medicine. “The fact that these cells were differentiated into mature blood cells suggests their use in blood diseases.”

Directly Programming Skin Cells to Become Blood-Making Stem Cells


Within our bones lies a spongy, ribbon-like material called bone marrow.  Bone marrow is home to several different populations of stem cells, but the star of the stem cell show in the bone marrow are the hematopoietic stem cells or blood-making stem cells.   When a patient receives a bone marrow transplant these are the stem cells that are transferred, take up residence in the new bone marrow, and begin making new red and white blood cells for the patient.  Because bone marrow is such a precious commodity from a clinical standpoint, finding a way to make more of it is essential.

Hematopoiesis from Pluripotent Stem Cell

A new report from scientists at Mt Sinai Hospital in New York suggest that the transfer of specific genes into skin fibroblasts can reprogram mature, adult cells into hematopoietic stem cells that look and function exactly like the ones normally found within our bone marrow.

A research team at the Icahn School of Medicine at Mount Sinai led by Kateri Moore screen a panel of 18 different genes for their ability to induce blood-forming activity when transfected into fibroblasts. Kateri and others discovered that a combination four different genes (GATA2, GFI1B, cFOS, and ETV6) is sufficient to generate blood vessel precursors with the subsequent appearance of hematopoietic stem cells. These cells expressed several known hematopoietic stem cell surface proteins (CD34, Sca1 and Prominin1/CD133).

Reprogramming of fibroblasts to HSCs

“The cells that we grew in a Petri dish are identical in gene expression to those found in the mouse embryo and could eventually generate colonies of mature blood cells,” said Carlos Filipe Pereira, first author of this paper and a postdoctoral research fellow in Moore’s laboratory.

The combination of gene factors that we used was not composed of the most obvious or expected proteins,” said Ihor Lemischka, a colleague of Dr. Moore at Mt. Sinai Hospital.  “Many investigators have been trying to grow hematopoietic stem cells from embryonic stem cells, but this process has been problematic.  Instead, we used mature mouse fibroblasts, pick the right combination of proteins, and it worked.”

According to Pereira, there is a rather critical shortage of suitable donors for blood stem cells transplants.  Bone marrow donors are currently necessary to meet the needs of patients suffering from blood diseases such as leukemia, aplastic anemia, lymphomas, multiple myeloma and immune deficiency disorders.  “Programming of hematopoietic stem cells represents an exciting alternative,” said Pereira.

“Dr. Lemischka and I have been working together for over 20 years in the fields of hematopoiesis and stem cell biology,” said Kateri Moore.  “It is truly exciting to be able to grow these blood forming cells in a culture dish and learn so much from them.  We have already started applying this new approach to human cells and anticipate similar success.”