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Archive for the ‘IPS Cell Therapy’ Category

A Safeguard System for Induced Pluripotent Stem Cell …

Highlights

iPSC-derived rejuvenated CTLs are effective against EBV-induced tumors invivo

Rejuvenated CTLs are implemented with an inducible caspase-9 (iC9)-based suicide system

Upon induction, the iC9 system efficiently leads to apoptosis in rejuvenated CTLs

The iC9-based system provides a safeguard for future iPSC-mediated cell therapy

The discovery of induced pluripotent stem cells (iPSCs) has created promising new avenues for therapies in regenerative medicine. However, the tumorigenic potential of undifferentiated iPSCs is a major safety concern for clinical translation. To address this issue, we demonstrated the efficacy of suicide gene therapy by introducing inducible caspase-9 (iC9) into iPSCs. Activation of iC9 with a specific chemical inducer of dimerization (CID) initiates a caspase cascade that eliminates iPSCs and tumors originated from iPSCs. We introduced this iC9/CID safeguard system into a previously reported iPSC-derived, rejuvenated cytotoxic T lymphocyte (rejCTL) therapy model and confirmed that we can generate rejCTLs from iPSCs expressing high levels of iC9 without disturbing antigen-specific killingactivity. iC9-expressing rejCTLs exert antitumor effects invivo. The system efficiently and safely induces apoptosis in these rejCTLs. These results unite to suggest that the iC9/CID safeguard system is a promising tool for future iPSC-mediated approaches to clinical therapy.

Human induced pluripotent stem cells (iPSCs) can unlimitedly self-renew and differentiate into various cell types (Takahashi etal., 2007). Their pluripotency makes iPSCs a promising tool for therapy in a wide range of diseases at present refractory to treatment (Inoue etal., 2014). Recent studies, however, reported the tumorigenic potential of contaminated undifferentiated iPSCs and the malignant transformation of differentiated iPSCs (Lee etal., 2013aandNori etal., 2015). The tumorigenic risks of iPSCs could be reduced by several strategies, such as sorting out undifferentiated cells with antibodies targeting surface-displayed biomarkers (Tang etal., 2011), killing undifferentiated cells with cytotoxic antibodies (Choo etal., 2008), or elimination of remaining undifferentiated pluripotent cells with chemical inhibitors (Ben-David etal., 2013andLee etal., 2013b). However, these strategies may not suffice to lower risk to acceptable levels, because the tumorigenic risk of iPSC-based cell therapy arises not just from contamination with undifferentiated iPSCs but also from other unexpected events associated with long-term culture for reprogramming and redifferentiation. There is always a chance of unexpected issues associated with first-in-human clinical studies.

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A Safeguard System for Induced Pluripotent Stem Cell ...

Standards in Cell Therapy

This is a sixth post of the series Not Lost in Translation.

If youre trying to develop a cellular product and just entering the field of cell therapy, you should be aware of existent standards. Why is it important? Knowing standards in your field allows to:

Even though, cell therapy filed relatively new, there are numerous related standards. Unfortunately, many professionals are unaware about organizations and standards in cell therapy field. The purpose of this post is to indicate few leadig organizations, providing standards and types of standards in cell products development. Significant part of this topic was summarized from the recent public FDA workshop Synergizing Efforts in Standards Development for Cellular Therapies and Regenerative Medicine Products.

Type of standards in cell therapy:

Standards-developing organizations and examples: ISO International Organization for Standardization Developing and providing international standards, including medical devices, laboratory testing and some, related to cell therapy and tissue engineered products. Examples: ISO/TC 194/SC 1 Tissue product safety ISO/TC 150/SC 7 Tissue-engineered medical products

ASTM International American Society for Testing and Materials ASTM leading international standards organization. ASTM has Subcommittee F04.43 for developing standards in cell therapy and tissue engineering. Examples: ASTM F2210 Standard Guide for Processing Cells, Tissues, and Organs for Use in Tissue Engineered Medical Products ASTM F2739 Standard Guide for Quantitating Cell Viability Within Biomaterial Scaffolds ASTM F2315 Standard Guide for Immobilization or Encapsulation of Living Cells or Tissue in Alginate Gels ASTM F2944 Standard Test Method for Automated Colony Forming Unit (CFU) Assays

USP U.S. Pharmacopeial Convention Provides standards for use ancillary and raw materials for cellular and tissue products. Examples: Chapter 1046 Cell and Gene Therapies Products Chapter 1047 Gene Therapy Products Chapter 1043 Ancillary Materials for Cell, Gene and Tissue-Engineered Products Chapter 92 Growth Factors and Cytokines Used in Cell Therapy Manufacturing Chapter 90 Fetal Bovine SerumQuality Attributes and Functionality Tests

GBSI Global Biological Standard Institute Developing standards for life sciences, including biomedical research.

ATCC American Type Culture Collection Manufactures and provides reference material (including cells), developing biological standards for basic and translational research. Examples: ATCC Certified reference material ATCC Standards Development Organization

BSI British Standards Institution Has a project for developing regenerative medicine definitions and guidelines for clinical cell products characterization. Examples: PAS 93:2011 Characterization of human cells for clinical applications. Guide PAS 84:2012 Cell therapy and regenerative medicine. Glossary

FACT Foundation for the Accreditation of Cellular Therapy Provides standards for collection and processing cellular products. Accredits clinical stem cell labs, cord blood banks and more than minimal manipulation cell therapy facilities. Examples: FACT-JACIE International Standards for Cellular Therapy Product Collection, Processing and Administration FACT-JACIE Cellular Therapy Accreditation Manual

AABB American Association of Blood Banks Center for Cellular Therapies In cell therapy field, AABB has very similar functions with FACT. Examples: Standards for Cellular Therapy Services

ICCBBA International Council for Commonality in Blood Bank Automation Management of the ISBT-128 Standard the terminology, identification, coding and labeling of medical products of human origin (including blood, cell, tissue, and organ products).

ISCT International Society for Cellular Therapy ISCT leverages expertise of cell therapy professionals to develop guidelines and recommendations for cellular products development, characterization, and quality. Examples: Minimal criteria for defining multipotent mesenchymal stromal cells Potency assay development for cellular therapy products Stromal cells from the adipose tissue-derived stromal vascular fraction and culture expanded adipose tissue-derived stromal/stem cells IFATS/ISCT statement

Coordination and harmonization As you can see, there are many organizations, involved in different aspects of cell therapy standardization. How can we make sure that there are no overlaps between them? How to coordinate and harmonize their activities? There are some good existent examples of such coordination:

*********************** This post is a part of Not Lost in Translation online community project. In this series we will try to bridge the translational gaps between scientific discovery in research labs and clinical cell applications for therapies. We will look at challenges in translation of cell product development and manufacturing in academic and industry settings. If you would like to contribute to this community project, please contact us!

Tagged as: cell therapy, reference material, standard, translation

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Standards in Cell Therapy

What are induced pluripotent stem cells? [Stem Cell …

Induced pluripotent stem cells (iPSCs) are adult cells that have been genetically reprogrammed to an embryonic stem celllike state by being forced to express genes and factors important for maintaining the defining properties of embryonic stem cells. Although these cells meet the defining criteria for pluripotent stem cells, it is not known if iPSCs and embryonic stem cells differ in clinically significant ways. Mouse iPSCs were first reported in 2006, and human iPSCs were first reported in late 2007. Mouse iPSCs demonstrate important characteristics of pluripotent stem cells, including expressing stem cell markers, forming tumors containing cells from all three germ layers, and being able to contribute to many different tissues when injected into mouse embryos at a very early stage in development. Human iPSCs also express stem cell markers and are capable of generating cells characteristic of all three germ layers.

Although additional research is needed, iPSCs are already useful tools for drug development and modeling of diseases, and scientists hope to use them in transplantation medicine. Viruses are currently used to introduce the reprogramming factors into adult cells, and this process must be carefully controlled and tested before the technique can lead to useful treatment for humans. In animal studies, the virus used to introduce the stem cell factors sometimes causes cancers. Researchers are currently investigating non-viral delivery strategies. In any case, this breakthrough discovery has created a powerful new way to "de-differentiate" cells whose developmental fates had been previously assumed to be determined. In addition, tissues derived from iPSCs will be a nearly identical match to the cell donor and thus probably avoid rejection by the immune system. The iPSC strategy creates pluripotent stem cells that, together with studies of other types of pluripotent stem cells, will help researchers learn how to reprogram cells to repair damaged tissues in the human body.

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What are induced pluripotent stem cells? [Stem Cell ...

IPS Cell Therapy IPS Cell Therapy

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Stem Cell Research is an amazing field right now, and promises to be a powerful and potent tool to help us live longer and healthier lives. Just last month, for example, Stem Cell Therapy was used to restore sight in patients with severe retinal deterioration, allowing them to see clearer than they had in years, or even decades.

Now, there is another form of Stem Cell Treatment on the horizonthis one of a very different form. Stem Cells have now been used as a mechanism to deliver medical treatment designed to eliminate cancer cells, even in hard to reach places. One issue with current cancer treatments is that, treatments that are effective at treating tumors on the surface of the brain cannot be performed safely when the tumor is deeper within the brains tissues.

Stem Cells have the fantastic ability to transform into any other kind of cell within the human body, given the appropriate stimulation. As of today, most of these cells come from Embryonic Lines, but researchers are learning how to backwards engineer cells in the human body, reverting them back to their embryonic state. These cells are known as Induced Pluripotent Stem Cells.

How Does This Stem Cell Cancer Treatment Work?

Using genetic engineering, it is possible to create stem cells that are designed to release a chemical known as Pseudomonas Exotoxin, which has the ability to destroy certain tumor cells in the human brain.

What is Pseudomonas Exotoxin?

Pseudomonas Exotoxin is a compound that is naturally released by a form of bacteria known as Pseudomonas Aeruginosa. This chemical is toxic to brain tumor cells because it prevents polypeptides from growing longer, essentially preventing the polypeptides from growing and reproducing. When used in a specific manner, this toxin has the ability to destroy cancerous and malignant tissue without negatively impacting healthy tissue. In addition to its potential as a cancer treatment, there is also evidence that the therapy could be used for the treatment of Hepatitis B.

PE and Similar Toxins Have been Used Therapeutically in the Past

As of now, this chemical, which we will refer to for the rest of the article as PE, has been used as a cancer treatment before, but there are major limitations regarding the use of PE for particular cancers, not because of the risks of the treatment, but because of the lack of an effective method to deliver the medication to where it is needed.

For example, similar chemicals have been highly effective in the treatment of a large number of blood cancers, but havent been nearly as effective in larger, more inaccessible tumors. The chemicals break down or become metabolized before they can fully do their job.

How do Stem Cells Increase the Effectiveness of PE Cancer Treatment

Right now, PE has to be created in a laboratory before it is administered, which is not very effective for these embedded cancers. By using Stem Cells as an intermediary, it is possible to deliver the medication to deeper areas of the brain more effectively, theoretically highly increasing the efficacy of the treatment.

The leader of this Stem Cell Research is Harvard researcher Dr. Khalis Shah. His goal was to find an effective means to treat these deep brain tumors which are not easily treated by methods available today. In utilizing Stem Cells, Dr. Shah has potentially found a means by which the stem cells can constantly deliver this Cancer Toxin to the tumor area. The cells remain active and are fed by the body, which allows them to provide a steady stream of treatment that is impossible to provide via any other known method.

This research is still in its early stages, and has not yet reached human trials, but in mice, the PE Toxin worked exactly as hypothesized and was able to starve out tumors by preventing them from replicating effectively.

Perhaps this might seem a bit less complicated than it actually is. One of the major hurdles that had to be overcome was that this Toxin would normally be strong enough to kill the cell that hosted it. In order for the Stem Cells to release the cancer, they had to be able to withstand the effects of PE, themselves. Using genetic engineering, Dr. Shah and his associates were able to create a cell that is capable of both producing and withstanding the effects of the toxin.

Stem Cell delivered medical therapy is a 21st century form of medical treatment that researchers are just beginning to learn how to effectively utilize. Essentially, this treatment takes a stem cell and converts it into a unique symbiotic tool capable of feeding off of the host for energy in order to perform a potentially life-saving function. Its really quite fascinating.

How Does PE Not Damage or Kill Brain Cells Indiscriminately?

You might be concerned about the idea of a patient having a toxin injected into the brain to cure a disease. It sounds almost like a dangerous, tribal, homeopathic remedy. In reality, the researchers have been able to harness the destructive power of the toxin and re-engineer it so that it directly targets cancer cells while having limited negative effects on healthy, non-cancerous tissue.

The toxin does its damage after it has been absorbed by a cell. By retooling the toxin so that it does not readily absorb into healthy cells, the dangers associated with having such a potentially dangerous toxin in the brain are seriously and significantly mitigated.

Beyond that, Dr. Shah and his associates have been able to take steps to effectively turn off PE while it is inside the host stem cell, and only activates when it has entered the cancerous tissue. Dr. Shah explains that, although this research has only been conducted in animal subjects, there is no known reason why the effectiveness and safety of the treatment would not be applicable to human patients.

In this treatment, surgeons remove as much of the tumor as possible from the brain, and insert the engineered Stem Cells submerged in a sterile gel in the area where the tumor was removed or partially still exists. Researchers found that, when they used this treatment on laboratory rats, they could tell through imaging and analysis that the modified PE toxin effectively killed the cancer cells, and that this cancer treatment effectively lengthened the life of the rat, as compared to control subjects.

Whats the Next Step?

Of course, cancer treatment is far more complex than a single treatment, no matter how effective that treatment may be. Because human cancer treatment is a comprehensive therapy approach, the end goal of this research is to create a form of therapy in which the method used in animal subjects is combined with other existing approaches, increasing and maximizing the effectiveness of the comprehensive treatment.

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A recent change in how well we understand stem cells may make it easier for scientists and researchers to gather stem cells for use in scientific research as well as medical application. A new study was released in the research publication, Cell, which was performed by representatives from the University of California San Francisco.

One of the issues which hinder the use of stem cells as a more widespread treatment or field of research is that researchers and patients have a bottleneck of available healthy stem cell lines which can be used for research. Researchers hope that this new discovery will allow future scientific discoveries and applications in the areas of creating new and healthy tissue for patients with kidney failure or any other form of organ tissue failure. The future of medical therapy lies with Stem Cell Research, but many other forms of treatment, including Hormone Replacement Therapy, are already in practice today.

Researchers have discovered that it is possible to essentially flip a switch in an adult cell, reverting it back to the preliminary state at which cells existed in one of the earliest stages of developmentthe embryonic stem cell. Medical researchers hypothesize that Stem Cell treatments could be used for a variety of medical health issues which plague the world today, including kidney failure, liver disease, and Type-1 and Type-2 Diabetes.

Use of Embryonic Stem Cells Contentious

There is an ethical issue in Stem Cell Research today. Many Pro-Life Advocates are vociferously against the use of Embryonic Stem Cells harvested from procedures such as fertility treatments designed for conception. They believe that the use of embryonic stem cells harvested from donors and couples looking to conceive is unethical.

Using current research, it may be possible to bypass this ethical quandary completely by using adult cells and converting them into embryonic stem cells. Furthermore, because these stem cells are genetic derivatives of the patient from which the adult cells were harvested, this potentially paves the way for patient-specific medical treatments using stem cells.

After adult cells have been converted back into Embryonic Stem Cells, it will be possible to convert them into any possible cell that the patient needs or would benefit from.

Hijacking the Blueprint of the Cell Allows Scientists to Revert Adult Cells to their Earliest State

Researchers have increased the capacity to produce Embryonic Stem Cells by identifying previously unrecognized biochemical processes which tell human cells how to develop. In essence, researchers have discovered how the body blueprints cells, and can change the blueprints so that a new cell is made.

By utilizing these newly recognized pathways, it is possible to create new stem cells more quickly than ever before. One of the researchers explains the implications of this research. Dr. Miguel Ramalho-Santos is an associate professor of obstetrics, medicine, and cancer research at the University of California San Francisco. Dr. Ramalho-Santos is also a member of the Broad Center of Regenerative Medicine and Stem Cell Research.

He explains that these stem cell discoveries have the ability to alter the way that the medical sciences can take advantage of stem cells with regard to both cancer research and regenerative medicine. Dr. Ramalho-Santos was the lead researcher for this study, and the research was largely funded by the Director of the National Institutes of Health New Innovator Award, granted to promising young researchers which are leading highly innovative and promising medical research studies.

Dr. Ramalho-Santos research builds off of earlier research which discovered that it was possible to take adult cells and turn them back into embryonic stem cells. These stem cells dont have any inherent aging processes, and they can be turned into any other kind of tissue. In the process of this conversion, the adult cells lose all of their unique characteristics, leaving them in an ultimately immature and malleable state.

This earlier research was conducted by researchers from UC San Francisco in partnership with Dr. Shinya Yamanaka from Kyoto University and Gladstone Institutes. These entities all gained a piece of the Nobel Prize in Physiology or Medicine from their part in the study.

Pluripotent Stem Cells vs. Embryonic Stem Cells

Thus far, weve described these cells as Embryonic Stem Cells, but in fact, the more accurate term for these cells are Induced Pluripotent Stem Cells (IPS). These cells are biologically and functionally similar to Embryonic Stem Cells, but have a different name because they are sourced from adult cells. The difference between Induced Pluripotent Stem Cells and Embryonic Stem Cells is that Induced Pluripotent Stem Cells do seem to retain some of the characteristics of their previous state, which appears to limit their ability to convert into any other type of cell. This new research identifies new pathways by which it may be possible to increase the number of cells that an individual IPS Cell can turn into, perhaps allowing them to convert into any other kind of human cell.

Induced Pluripotent Stem Cells are not explicitly considered an alternative to Embryonic Stem Cells, but are considered a different approach to produce similar cells. If researchers fully uncover the mechanisms of how to reprogram these cells, it will lower many barriers to stem cell research and the availability of stem cell treatments.

As of today, researchers have figured out how to make these Induced Pluripotent Stem Cells, but the percentage of adult cells which are reverted successfully is quite low, and frequently, these cells still show some aspects of specialization, which limits their use.

How Do Scientists Make Stem Cells From Adult Cells?

There are genes within every cell which have the ability to induce pluripotency, reverting the cell to an earlier stage of specialization. The initial stage of this process is the result of activating Yamanaka Factors, specific genes that initiate this reversion process.

As of today, this process of de-maturation is not completely understood, and researchers realized from the start that the cells they created were not truly identical to Embryonic Stem Cells, because they still showed signs of their former lives, which often prevented them from being successfully reprogrammed.

The new research conducted by Dr. Ramalho-Santos appears to increase our knowledge regarding how these cells work, and how to program them more effectively. Dr. Ramalho-Santos and his team discovered more genes associated with these programming/reprogramming processes, and by manipulating them, they have increased the viability and range of particular stem cells.

It appears that these genetic impulses are constantly at play to maintain the structure and function of a cell, and that by systematically removing these safeguards, it is possible to increase the ability to alter these cells.

This research increases researchers ability to produce these stem cells, by increasing the ability of medical scientists to produce adequate numbers of stem cells, while also increasing the range of potential treatment options by more effectively inducing the total pluripotency which is available in Embryonic Stem Cells. This research may also help scientists treat certain forms of cancer which are the result of malfunctions of these genes.

Introduction

[Note: Many of the medical and scientific terms used in this summary are found in the NCI Dictionary of Genetics Terms. When a linked term is clicked, the definition will appear in a separate window.]

[Note: Many of the genes described in this summary are found in the Online Mendelian Inheritance in Man (OMIM) database. When OMIM appears after a gene name or the name of a condition, click on OMIM for a link to more information.]

The genetics of skin cancer is an extremely broad topic. There are more than 100 types of tumors that are clinically apparent on the skin; many of these are known to have familial components, either in isolation or as part of a syndrome with other features. This is, in part, because the skin itself is a complex organ made up of multiple cell types. Furthermore, many of these cell types can undergo malignant transformation at various points in their differentiation, leading to tumors with distinct histology and dramatically different biological behaviors, such as squamous cell carcinoma (SCC) and basal cell cancer (BCC). These have been called nonmelanoma skin cancers or keratinocytic cancers.

Figure 1 is a simple diagram of normal skin structure. It also indicates the major cell types that are normally found in each compartment. Broadly speaking, there are two large compartmentsthe avascular cellular epidermis and the vascular dermiswith many cell types distributed in a largely acellular matrix.[1]

Figure 1. Schematic representation of normal skin. The relatively avascular epidermis houses basal cell keratinocytes and squamous epithelial keratinocytes, the source cells for BCC and SCC, respectively. Melanocytes are also present in normal skin and serve as the source cell for melanoma. The separation between epidermis and dermis occurs at the basement membrane zone, located just inferior to the basal cell keratinocytes.

The outer layer or epidermis is made primarily of keratinocytes but has several other minor cell populations. The bottom layer is formed of basal keratinocytes abutting the basement membrane. The basement membrane is formed from products of keratinocytes and dermal fibroblasts, such as collagen and laminin, and is an important anatomical and functional structure. As the basal keratinocytes divide and differentiate, they lose contact with the basement membrane and form the spinous cell layer, the granular cell layer, and the keratinized outer layer or stratum corneum.

The true cytologic origin of BCC remains in question. BCC and basal cell keratinocytes share many histologic similarities, as is reflected in the name. Alternatively, the outer root sheath cells of the hair follicle have also been proposed as the cell of origin for BCC.[2] This is suggested by the fact that BCCs occur predominantly on hair-bearing skin. BCCs rarely metastasize but can invade tissue locally or regionally, sometimes following along nerves. A tendency for superficial necrosis has resulted in the name rodent ulcer.[3]

Some debate remains about the origin of SCC; however, these cancers are likely derived from epidermal stem cells associated with the hair follicle.[4] A variety of tissues, such as lung and uterine cervix, can give rise to SCC, and this cancer has somewhat differing behavior depending on its source. Even in cancer derived from the skin, SCC from different anatomic locations can have moderately differing aggressiveness; for example, SCC from glabrous (smooth, hairless) skin has a lower metastatic rate than SCC arising from the vermillion border of the lip or from scars.[3]

Additionally, in the epidermal compartment, melanocytes distribute singly along the basement membrane and can transform into melanoma. Melanocytes are derived from neural crest cells and migrate to the epidermal compartment near the eighth week of gestational age. Langerhans cells, or dendritic cells, are a third cell type in the epidermis and have a primary function of antigen presentation. These cells reside in the skin for an extended time and respond to different stimuli, such as ultraviolet radiation or topical steroids, which cause them to migrate out of the skin.[5]

The dermis is largely composed of an extracellular matrix. Prominent cell types in this compartment are fibroblasts, endothelial cells, and transient immune system cells. When transformed, fibroblasts form fibrosarcomas and endothelial cells form angiosarcomas, Kaposi sarcoma, and other vascular tumors. There are a number of immune cell types that move in and out of the skin to blood vessels and lymphatics; these include mast cells, lymphocytes, mononuclear cells, histiocytes, and granulocytes. These cells can increase in number in inflammatory diseases and can form tumors within the skin. For example, urticaria pigmentosa is a condition that arises from mast cells and is occasionally associated with mast cell leukemia; cutaneous T-cell lymphoma is often confined to the skin throughout its course. Overall, 10% of leukemias and lymphomas have prominent expression in the skin.[6]

Epidermal appendages are also found in the dermal compartment. These are derivatives of the epidermal keratinocytes, such as hair follicles, sweat glands, and the sebaceous glands associated with the hair follicles. These structures are generally formed in the first and second trimesters of fetal development. These can form a large variety of benign or malignant tumors with diverse biological behaviors. Several of these tumors are associated with familial syndromes. Overall, there are dozens of different histological subtypes of these tumors associated with individual components of the adnexal structures.[7]

Finally, the subcutis is a layer that extends below the dermis with varying depth, depending on the anatomic location. This deeper boundary can include muscle, fascia, bone, or cartilage. The subcutis can be affected by inflammatory conditions such as panniculitis and malignancies such as liposarcoma.[8]

These compartments give rise to their own malignancies but are also the region of immediate adjacent spread of localized skin cancers from other compartments. The boundaries of each skin compartment are used to define the staging of skin cancers. For example, an in situ melanoma is confined to the epidermis. Once the cancer crosses the basement membrane into the dermis, it is invasive. Internal malignancies also commonly metastasize to the skin. The dermis and subcutis are the most common locations, but the epidermis can also be involved in conditions such as Pagetoid breast cancer.

The skin has a wide variety of functions. First, the skin is an important barrier preventing extensive water and temperature loss and providing protection against minor abrasions. These functions can be aberrantly regulated in cancer. For example, in the erythroderma associated with advanced cutaneous T-cell lymphoma, alterations in the regulations of body temperature can result in profound heat loss. Second, the skin has important adaptive and innate immunity functions. In adaptive immunity, antigen-presenting cells engender a TH1, TH2, and TH17 response.[9] In innate immunity, the immune system produces numerous peptides with antibacterial and antifungal capacity. Consequently, even small breaks in the skin can lead to infection. The skin-associated lymphoid tissue is one of the largest arms of the immune system. It may also be important in immune surveillance against cancer. Immunosuppression, which occurs during organ transplant, is a significant risk factor for skin cancer. The skin is significant for communication through facial expression and hand movements. Unfortunately, areas of specialized function, such as the area around the eyes and ears, are common places for cancer to occur. Even small cancers in these areas can lead to reconstructive challenges and have significant cosmetic and social ramifications.[1]

While the appearance of any one skin cancer can vary, there are general physical presentations that can be used in screening. BCCs most commonly have a pearly rim (see Figure 3) or can appear somewhat eczematous. They often ulcerate (see Figure 3). SCCs frequently have a thick keratin top layer (see Figure 4). Both BCCs and SCCs are associated with a history of sun-damaged skin. Melanomas are characterized by asymmetry, border irregularity, color variation, a diameter of more than 6 mm, and evolution (ABCDE criteria). (Refer to What Does Melanoma Look Like? on NCIs website for more information about the ABCDE criteria.) Photographs representing typical clinical presentations of these cancers are shown below.

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Figure 2. Superficial basal cell carcinoma (left panel) and nodular basal cell carcinoma (right panel).

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Figure 3. Ulcerated basal cell carcinoma (left panel) and ulcerated basal cell carcinoma with characteristic pearly rim (right panel).

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Figure 4. Squamous cell carcinoma on the face with thick keratin top layer (left panel) and squamous cell carcinoma on the leg (right panel).

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Figure 5. Melanomas with characteristic asymmetry, border irregularity, color variation, and large diameter.

Basal cell carcinoma (BCC) is the most common malignancy in people of European descent, with an associated lifetime risk of 30%.[1] While exposure to ultraviolet (UV) radiation is the risk factor most closely linked to the development of BCC, other environmental factors (such as ionizing radiation, chronic arsenic ingestion, and immunosuppression) and genetic factors (such as family history, skin type, and genetic syndromes) also potentially contribute to carcinogenesis. In contrast to melanoma, metastatic spread of BCC is very rare and typically arises from large tumors that have evaded medical treatment for extended periods of time. BCCs can invade tissue locally or regionally, sometimes following along nerves. A tendency for superficial necrosis has resulted in the name rodent ulcer. With early detection, the prognosis for BCC is excellent.

Sun exposure is the major known environmental factor associated with the development of skin cancer of all types. There are different patterns of sun exposure associated with each major type of skin cancer (BCC, squamous cell carcinoma [SCC], and melanoma).

While there is no standard measure, sun exposure can be generally classified as intermittent or chronic, and the effects may be considered acute or cumulative. Intermittent sun exposure is obtained sporadically, usually during recreational activities, and particularly by indoor workers who have only weekends or vacations to be outdoors and whose skin has not adapted to the sun. Chronic sun exposure is incurred by consistent, repetitive sun exposure, during outdoor work or recreation. Acute sun exposure is obtained over a short time period on skin that has not adapted to the sun. Depending on the time of day and a persons skin type, acute sun exposure may result in sunburn. In epidemiology studies, sunburn is usually defined as burn with pain and/or blistering that lasts for 2 or more days. Cumulative sun exposure is the additive amount of sun exposure that one receives over a lifetime. Cumulative sun exposure may reflect the additive effects of intermittent sun exposure, chronic sun exposure, or both.

Specific patterns of sun exposure appear to lead to different types of skin cancer among susceptible individuals. Intense intermittent recreational sun exposure has been associated with melanoma and BCC,[2,3] while chronic occupational sun exposure has been associated with SCC. Given these data, dermatologists routinely counsel patients to protect their skin from the sun by avoiding mid-day sun exposure, seeking shade, and wearing sun-protective clothing, although evidence-based data for these practices are lacking. The data regarding skin cancer risk reduction by regular sunscreen use are variable. One randomized trial of sunscreen efficacy demonstrated statistically significant protection for the development of SCC but no protection for BCC,[4] while another randomized study demonstrated a trend for reduction in multiple occurrences of BCC among sunscreen users [5] but no significant reduction in BCC or SCC incidence.[6]

Level of evidence (sun-protective clothing, avoidance of sun exposure): 4aii

Level of evidence (sunscreen): 1aii

Tanning bed use has also been associated with an increased risk of BCC. A study of 376 individuals with BCC and 390 control subjects found a 69% increased risk of BCC in individuals who had ever used indoor tanning.[7] The risk of BCC was more pronounced in females and individuals with higher use of indoor tanning.[8]

Environmental factors other than sun exposure may also contribute to the formation of BCC and SCC. Petroleum byproducts (e.g., asphalt, tar, soot, paraffin, and pitch), organophosphate compounds, and arsenic are all occupational exposures associated with cutaneous nonmelanoma cancers.[9-11]

Arsenic exposure may occur through contact with contaminated food, water, or air. While arsenic is ubiquitous in the environment, its ambient concentration in both food and water may be increased near smelting, mining, or coal-burning establishments. Arsenic levels in the U.S. municipal water supply are tightly regulated; however, control is lacking for potable water obtained through private wells. As it percolates through rock formations with naturally occurring arsenic, well water may acquire hazardous concentrations of this material. In many parts of the world, wells providing drinking water are contaminated by high levels of arsenic in the ground water. The populations in Bangladesh, Taiwan, and many other locations have high levels of skin cancer associated with elevated levels of arsenic in the drinking water.[12-16] Medicinal arsenical solutions (e.g., Fowlers solution and Bells asthma medication) were once used to treat common chronic conditions such as psoriasis, syphilis, and asthma, resulting in associated late-onset cutaneous malignancies.[17,18] Current potential iatrogenic sources of arsenic exposure include poorly regulated Chinese traditional/herbal medications and intravenous arsenic trioxide utilized to induce remission in acute promyelocytic leukemia.[19,20]

Aerosolized particulate matter produced by combustion of arsenic-containing materials is another source of environmental exposure. Arsenic-rich coal, animal dung from arsenic-rich regions, and chromated copper arsenatetreated wood produce airborne arsenical particles when burned.[21-23] Burning of these products in enclosed unventilated settings (such as for heat generation) is particularly hazardous.[24]

Clinically, arsenic-induced skin cancers are characterized by multiple recurring SCCs and BCCs occurring in areas of the skin that are usually protected from the sun. A range of cutaneous findings are associated with chronic or severe arsenic exposure, including pigmentary variation (poikiloderma of the skin) and Bowen disease (SCC in situ).[25]

However, the effect of arsenic on skin cancer risk may be more complex than previously thought. Evidence from in vivo models indicate that arsenic, alone or in combination with itraconazole, can inhibit the hedgehog pathway in cells with wild-type or mutated Smoothened by binding to GLI2 proteins; in this way, these drugs demonstrated inhibition of BCC growth in these animal models.[26,27] Additionally, the effect of arsenic on skin cancer risk may be modified by certain variants in nucleotide excision repair genes (xeroderma pigmentosum [XP] types A and D).[28]

The high-risk phenotype consists of individuals with the following physical characteristics:

Specifically, people with more highly pigmented skin demonstrate lower incidence of BCC than do people with lighter pigmented skin. Individuals with Fitzpatrick skin types I or II were shown to have a twofold increased risk of BCC in a small case-control study.[29] (Refer to the Pigmentary characteristics section in the Melanoma section of this summary for a more detailed discussion of skin phenotypes based upon pigmentation.) Blond or red hair color was associated with increased risk of BCC in two large cohorts: the Nurses Health Study and the Health Professionals Follow-Up Study.[30]

Immunosuppression also contributes to the formation of nonmelanoma (keratinocyte) skin cancers. Among solid-organ transplant recipients, the risk of SCC is 65 to 250 times higher, and the risk of BCC is 10 times higher than in the general population.[31-33] Nonmelanoma skin cancers in high-risk patients (i.e., solid-organ transplant recipients and chronic lymphocytic leukemia patients) occur at a younger age and are more common, more aggressive, and have a higher risk of recurrence and metastatic spread than nonmelanoma skin cancers in the general population.[34,35] Among patients with an intact immune system, BCCs outnumber SCCs by a 4:1 ratio; in transplant patients, SCCs outnumber BCCs by a 2:1 ratio.

This increased risk has been linked to the level of immunosuppression and UV exposure. As the duration and dosage of immunosuppressive agents increases, so does the risk of cutaneous malignancy; this effect is reversed with decreasing the dosage of, or taking a break from, immunosuppressive agents. Heart transplant recipients, requiring the highest rates of immunosuppression, are at much higher risk of cutaneous malignancy than liver transplant recipients, in whom much lower levels of immunosuppression are needed to avoid rejection.[31,36] The risk appears to be highest in geographic areas of high UV radiation exposure: when comparing Australian and Dutch organ transplant populations, the Australian patients carried a fourfold increased risk of developing SCC and a fivefold increased risk of developing BCC.[37] This speaks to the importance of rigorous sun avoidance among high-risk immunosuppressed individuals.

Individuals with BCCs and/or SCCs report a higher frequency of these cancers in their family members than do controls. The importance of this finding is unclear. Apart from defined genetic disorders with an increased risk of BCC, a positive family history of any skin cancer is a strong predictor of the development of BCC.

A personal history of BCC or SCC is strongly associated with subsequent BCC or SCC. There is an approximate 20% increased risk of a subsequent lesion within the first year after a skin cancer has been diagnosed. The mean age of occurrence for these nonmelanoma skin cancers is the mid-60s.[38-43] In addition, several studies have found that individuals with a history of skin cancer have an increased risk of a subsequent diagnosis of a noncutaneous cancer;[44-47] however, other studies have contradicted this finding.[48-51] In the absence of other risk factors or evidence of a defined cancer susceptibility syndrome, as discussed below, skin cancer patients are encouraged to follow screening recommendations for the general population for sites other than the skin.

Mutations in the gene coding for the transmembrane receptor protein PTCH1, or PTCH, are associated with basal cell nevus syndrome (BCNS) and sporadic cutaneous BCCs. PTCH1, the human homolog of the Drosophila segment polarity gene patched (ptc), is an integral component of the hedgehog signaling pathway, which serves many developmental (appendage development, embryonic segmentation, neural tube differentiation) and regulatory (maintenance of stem cells) roles.

In the resting state, the transmembrane receptor protein PTCH1 acts catalytically to suppress the seven-transmembrane protein Smoothened (Smo), preventing further downstream signal transduction.[52] Stoichiometric binding of the hedgehog ligand to PTCH1 releases inhibition of Smo, with resultant activation of transcription factors (GLI1, GLI2), cell proliferation genes (cyclin D, cyclin E, myc), and regulators of angiogenesis.[53,54] Thus, the balance of PTCH1 (inhibition) and Smo (activation) manages the essential regulatory downstream hedgehog signal transduction pathway. Loss-of-function mutations of PTCH1 or gain-of-function mutations of Smo tip this balance toward constitutive activation, a key event in potential neoplastic transformation.

Demonstration of allelic loss on chromosome 9q22 in both sporadic and familial BCCs suggested the potential presence of an associated tumor suppressor gene.[55,56] Further investigation identified a mutation in PTCH1 that localized to the area of allelic loss.[57] Up to 30% of sporadic BCCs demonstrate PTCH1 mutations.[58] In addition to BCC, medulloblastoma and rhabdomyosarcoma, along with other tumors, have been associated with PTCH1 mutations. All three malignancies are associated with BCNS, and most people with clinical features of BCNS demonstrate PTCH1 mutations, predominantly truncation in type.[59]

Truncating mutations in PTCH2, a homolog of PTCH1 mapping to chromosome 1p32.1-32.3, have been demonstrated in both BCC and medulloblastoma.[60,61] PTCH2 displays 57% homology to PTCH1, differing in the conformation of the hydrophilic region between transmembrane portions 6 and 7, and the absence of C-terminal extension.[62] While the exact role of PTCH2 remains unclear, there is evidence to support its involvement in the hedgehog signaling pathway.[60,63]

BCNS, also known as Gorlin Syndrome, Gorlin-Goltz syndrome, and nevoid basal cell carcinoma syndrome, is an autosomal dominant disorder with an estimated prevalence of 1 in 57,000 individuals.[64] The syndrome is notable for complete penetrance and extremely variable expressivity, as evidenced by evaluation of individuals with identical genotypes but widely varying phenotypes.[59,65] The clinical features of BCNS differ more among families than within families.[66] BCNS is primarily associated with germline mutations in PTCH1, but families with this phenotype have also been associated with alterations in PTCH2 and SUFU.[67-69]

As detailed above, PTCH1 provides both developmental and regulatory guidance; spontaneous or inherited germline mutations of PTCH1 in BCNS may result in a wide spectrum of potentially diagnostic physical findings. The BCNS mutation has been localized to chromosome 9q22.3-q31, with a maximum logarithm of the odd (LOD) score of 3.597 and 6.457 at markers D9S12 and D9S53.[64] The resulting haploinsufficiency of PTCH1 in BCNS has been associated with structural anomalies such as odontogenic keratocysts, with evaluation of the cyst lining revealing heterozygosity for PTCH1.[70] The development of BCC and other BCNS-associated malignancies is thought to arise from the classic two-hit suppressor gene model: baseline heterozygosity secondary to germline PTCH1 mutation as the first hit, with the second hit due to mutagen exposure such as UV or ionizing radiation.[71-75] However, haploinsufficiency or dominant negative isoforms have also been implicated for the inactivation of PTCH1.[76]

The diagnosis of BCNS is typically based upon characteristic clinical and radiologic examination findings. Several sets of clinical diagnostic criteria for BCNS are in use (refer to Table 1 for a comparison of these criteria).[77-80] Although each set of criteria has advantages and disadvantages, none of the sets have a clearly superior balance of sensitivity and specificity for identifying mutation carriers. The BCNS Colloquium Group proposed criteria in 2011 that required 1 major criterion with molecular diagnosis, two major criteria without molecular diagnosis, or one major and two minor criteria without molecular diagnosis.[80] PTCH1 mutations are found in 60% to 85% of patients who meet clinical criteria.[81,82] Most notably, BCNS is associated with the formation of both benign and malignant neoplasms. The strongest benign neoplasm association is with ovarian fibromas, diagnosed in 14% to 24% of females affected by BCNS.[74,78,83] BCNS-associated ovarian fibromas are more likely to be bilateral and calcified than sporadic ovarian fibromas.[84] Ameloblastomas, aggressive tumors of the odontogenic epithelium, have also been proposed as a diagnostic criterion for BCNS, but most groups do not include it at this time.[85]

Originally posted here:
IPS Cell Therapy IPS Cell Therapy

Patients guide to treatments | Knoepfler Lab Stem Cell Blog

Top 10 list of important, easy to understand facts for patients about stem cell treatments

For better or worse, I am in the unique position of being a stem cell scientist and also a patient. Looking on the bright side this gives me a unique perspective on things.

I know there are thousands of people out there looking for more practical information about stem cell therapies and treatments. These folks understandably are using the Internet to look for some clear, good info on stem cell treatments either for themselves or their loved ones. Too often the info that is out there is either wrong, misleading, or overly complex.

So in this post I want to address this need speaking as a scientist, patient advocate and cancer survivor in the form of 10 key facts to help you guide your way through the jungle of stuff out there about stem cells.

1) Stem cells are essentially a type of drug or biological and possibly permanent in nature. Yeah, they are extremely unusual drugs, but they are drugs. The FDA considers them drugs. Unlike other drugs, once a patient receives a stem cell drug, it will not necessarily simply go away like other drugs because a stem cell drug consists of living cells that often behave in unpredictable ways. What this means is if the stem cells are doing bad things your doctor has no way to stop it.

2) Like any medical product, even aspirin, stem cells treatments will have side effects. Not maybe. Definitely. Our hope is the side effects will be relatively mild.

3) The only stem cell treatment explicitly approved by the FDA for use in the U.S. is bone marrow transplantation. What this means is that any other stem cell treatment you see advertised on Facebook or Google or elsewhere that indicates it will be given to you inside the U.S. may in fact be illegal and unsafe. The exception to this is if it is part of an FDA-approved clinical trial.

4) If you venture outside the U.S. for a stem cell treatment, use extra caution and have a knowledgeable physician inside the U.S. guiding you. We have to avoid the trap of thinking that only the U.S. can offer advanced medical treatments, but on the other hand within the U.S. you have the added safety of the FDA, which is trying to protect you. In the vast majority of other countries regulatory agencies are practically non-existent or are far less strict than the FDA.

5) Stem cells are not a cure all. I am as excited as anybody about the potential of stem cells to treat a whole bunch of diseases and injuries, but they are not some kind of miracle cure for everything. When a doctor offers to inject some kind of stem cells or a stem cell-derived product into a patient either into the bloodstream or into a specific place that is injured such as a shoulder, we just do not know at this point if it will do any good with the exception of bone marrow transplant.

6) Dont let celebrities be your guide to medical care. The number of famous people getting stem cell treatments is increasing including sports stars and politicians. Dont let what these folks do influence what you decide to do about your health. Just because they are famous do not believe for one minute that they are any more informed than you or your personal doctor about medical treatments or stem cells. If anything I think sometimes famous people are more reckless with their health than average people like you and me.

7) Reach out to scientists as a source of info. As a scientist I am always happy to hear from people outside the scientific community with questions about stem cells and other research. I cant speak for all stem cell scientists but you might be surprised at how likely it is that if you send them a very short, clear email with one or two questions that they will respond and be helpful. We cant or shouldnt offer medical advice, but we can give our perspectives on stem cell research and its clinical potential, etc. Just do not cold call scientists as you are unlikely to find them that way and even if you do, they may be cranky. Email.

8 )The people selling you non-FDA approved stem cell treatments want your money. Unlike stem cell researchers, the people out there advertising stem cell treatments that are not FDA approved are only really after one thing: your money. As such they will do their best to convince you that their treatment is safe and effective. They may offer patient testimonials either from patients who truly believe they were helped or from people who are paid to say the treatment helped them. The bottom line is that the sellers of dubious stem cell treatments simply want your money.

9) There is no such thing as completely proven safe and if something sounds too good to be true, it probably is. I am contacted fairly regularly by patients or their families and they often mention that the doctors offering stem cell treatments told them that the treatments are proven safeor that umbilical cord blood cannot harm you.or that your own stem cells cannot harm you..or that adult stem cells are harmless. Ill believe it when the FDA says it is so and you should be skeptical too.

10) The most important thing is data and you have a right to see it before treatment.Before you or a loved one get a stem cell treatment, ask two key questions. First, is the treatment FDA approved and if not, why not? Second, can you please show me the data that proves your treatment is safe and effective. See what kind of answer you get. If they criticize the FDA then that is a warning flag. If they refuse to show you data, then that is a big red warning flag. They may say it is confidential or that it is not published yet, but as a patient you have a right to see the data, assuming they have any data at all.

These facts will hopefully change over the coming years, but right now I think they represent reality. I know as patients we need hope, but these unapproved stem cell treatments will at best take your money for nothing, and at worst will endanger you or your loved ones.

The post above is for information only and is not medical advice. All medical decisions should be made by patients in consultation with their personal physicians.

See original here:
Patients guide to treatments | Knoepfler Lab Stem Cell Blog

Patients guide to treatments | Knoepfler Lab Stem Cell Blog

Top 10 list of important, easy to understand facts for patients about stem cell treatments

For better or worse, I am in the unique position of being a stem cell scientist and also a patient. Looking on the bright side this gives me a unique perspective on things.

I know there are thousands of people out there looking for more practical information about stem cell therapies and treatments. These folks understandably are using the Internet to look for some clear, good info on stem cell treatments either for themselves or their loved ones. Too often the info that is out there is either wrong, misleading, or overly complex.

So in this post I want to address this need speaking as a scientist, patient advocate and cancer survivor in the form of 10 key facts to help you guide your way through the jungle of stuff out there about stem cells.

1) Stem cells are essentially a type of drug or biological and possibly permanent in nature. Yeah, they are extremely unusual drugs, but they are drugs. The FDA considers them drugs. Unlike other drugs, once a patient receives a stem cell drug, it will not necessarily simply go away like other drugs because a stem cell drug consists of living cells that often behave in unpredictable ways. What this means is if the stem cells are doing bad things your doctor has no way to stop it.

2) Like any medical product, even aspirin, stem cells treatments will have side effects. Not maybe. Definitely. Our hope is the side effects will be relatively mild.

3) The only stem cell treatment explicitly approved by the FDA for use in the U.S. is bone marrow transplantation. What this means is that any other stem cell treatment you see advertised on Facebook or Google or elsewhere that indicates it will be given to you inside the U.S. may in fact be illegal and unsafe. The exception to this is if it is part of an FDA-approved clinical trial.

4) If you venture outside the U.S. for a stem cell treatment, use extra caution and have a knowledgeable physician inside the U.S. guiding you. We have to avoid the trap of thinking that only the U.S. can offer advanced medical treatments, but on the other hand within the U.S. you have the added safety of the FDA, which is trying to protect you. In the vast majority of other countries regulatory agencies are practically non-existent or are far less strict than the FDA.

5) Stem cells are not a cure all. I am as excited as anybody about the potential of stem cells to treat a whole bunch of diseases and injuries, but they are not some kind of miracle cure for everything. When a doctor offers to inject some kind of stem cells or a stem cell-derived product into a patient either into the bloodstream or into a specific place that is injured such as a shoulder, we just do not know at this point if it will do any good with the exception of bone marrow transplant.

6) Dont let celebrities be your guide to medical care. The number of famous people getting stem cell treatments is increasing including sports stars and politicians. Dont let what these folks do influence what you decide to do about your health. Just because they are famous do not believe for one minute that they are any more informed than you or your personal doctor about medical treatments or stem cells. If anything I think sometimes famous people are more reckless with their health than average people like you and me.

7) Reach out to scientists as a source of info. As a scientist I am always happy to hear from people outside the scientific community with questions about stem cells and other research. I cant speak for all stem cell scientists but you might be surprised at how likely it is that if you send them a very short, clear email with one or two questions that they will respond and be helpful. We cant or shouldnt offer medical advice, but we can give our perspectives on stem cell research and its clinical potential, etc. Just do not cold call scientists as you are unlikely to find them that way and even if you do, they may be cranky. Email.

8 )The people selling you non-FDA approved stem cell treatments want your money. Unlike stem cell researchers, the people out there advertising stem cell treatments that are not FDA approved are only really after one thing: your money. As such they will do their best to convince you that their treatment is safe and effective. They may offer patient testimonials either from patients who truly believe they were helped or from people who are paid to say the treatment helped them. The bottom line is that the sellers of dubious stem cell treatments simply want your money.

9) There is no such thing as completely proven safe and if something sounds too good to be true, it probably is. I am contacted fairly regularly by patients or their families and they often mention that the doctors offering stem cell treatments told them that the treatments are proven safeor that umbilical cord blood cannot harm you.or that your own stem cells cannot harm you..or that adult stem cells are harmless. Ill believe it when the FDA says it is so and you should be skeptical too.

10) The most important thing is data and you have a right to see it before treatment.Before you or a loved one get a stem cell treatment, ask two key questions. First, is the treatment FDA approved and if not, why not? Second, can you please show me the data that proves your treatment is safe and effective. See what kind of answer you get. If they criticize the FDA then that is a warning flag. If they refuse to show you data, then that is a big red warning flag. They may say it is confidential or that it is not published yet, but as a patient you have a right to see the data, assuming they have any data at all.

These facts will hopefully change over the coming years, but right now I think they represent reality. I know as patients we need hope, but these unapproved stem cell treatments will at best take your money for nothing, and at worst will endanger you or your loved ones.

The post above is for information only and is not medical advice. All medical decisions should be made by patients in consultation with their personal physicians.

More here:
Patients guide to treatments | Knoepfler Lab Stem Cell Blog

Stem Cell Biology & Gene Therapy: Mostoslavsky Lab BUMC

Welcome to the Mostoslavsky Lab!

Mission Statement

The Mostoslavsky Lab is a basic science laboratory in the Section of Gastroenterology in the Department of Medicine at Boston University, affiliated with the Boston University Center for Regenerative Medicine (CReM). Our goal is to advance our understanding of stem cell biology with a focus on their genetic manipulation via gene transfer and their potential use for stem cell-based therapy. We believe that by discovering the mechanisms involved in stem cell self-renewal and differentiation we will be able to manipulate stem cell fate and use it as the basis for the correction of several diseases. Project areas in the lab focuses on the use of different stem cell populations, including embryonic stem cells, induced Pluripotent Stem (iPS) cells, hematopoietic stem cells and intestinal stem cells and their genetic manipulation by lentiviral vectors.

Specific Areas of Research

Embryonic Stem Cell Modeling of Intestinal Differentiation Embryonic Stem Cells (ESC) are pluripotent undifferentiated cells capable of giving rise to cells from all three germ layers. This unique ability makes them ideal candidates to model early development allowing us to study the basic signaling mechanisms involved in stem cell fate determination. At the same time, manipulating ESC differentiation toward a specific developmental pathway holds a great promise for their use in regenerative medicine. One focus of our lab is differentiating mouse ESC into intestinal epithelial cells in order to understand the complex signaling pathways involved in intestinal commitment from endodermal progenitors and undifferentiated stem cells.

iPS cells Our lab has a major interest in the study of induced Pluripotent Stem cells or iPS cells and the development of tools for their generation and characterization. Pioneering work by the laboratory of Dr. Yamanaka showed that fibroblasts transduced with retroviral vectors expressing four transcription factors, Oct4, Klf4, Sox2 and cMyc can be reprogrammed to become pluripotent stem cells that appear almost indistinguishable from ESC. In contrast to ESC, iPS cells are genetically identical to the individual from whom they are derived, raising the prospect of utilizing iPS cells for autologous cell based therapies without risk of rejection. We have previously developed a single lentiviral vector expressing a stem cell cassette, named STEMCCA, capable of generating iPS cells from post-natal fibroblasts with the highest efficiency reported to date. We have recently modified it to make it excisable and have used it to generate mouse and human iPS cells free of exogenous transgenes. We aimed at using iPS cells in parallel to ESC for the study of endoderm/intestinal lineage specification, as well as for disease modeling and their potential for regenerative medicine. We are currently establishing and characterizing iPSC lines from several GI tract related diseases, including Familial Adenomatous Polyposis (FAP), Crohns disease and Hemochromatosis.

Hematopoietic Stem Cell Manipulation for the Study of Stem Cell Self-Renewal and Differentiation Hematopoietic Stem Cells (HSCs) are the most thoroughly characterized stem cell population in the body and their study has resulted in well established methods for their isolation, purification and reliable assays of HSC function. During the last few years we have substantially improved our ability to genetically manipulate HSCs using viral vectors for gene transfer. Despite these efforts, few genes are known to play a role in the processes of stem cell self-renewal and differentiation. Understanding the molecular mechanisms that govern those unique functions are crucial for developing the promise that stem cells hold for developmental biology and regenerative medicine. In our lab, we use lentiviral viral gene transfer to study the role of several molecules in long-term HSC self-renewal and differentiation.

Read more from the original source:
Stem Cell Biology & Gene Therapy: Mostoslavsky Lab BUMC

Potential use of iPS cells to combat acute kidney disease …

Whilst transplantation often remains the only effective treatment for acute kidney disease, a new study from Kyoto University points to a future where renal progenitor cells derived from iPSCs could be transplanted into affected kidneys to combat these debilitating conditions.

In recent years, a popular avenue of investigation for treating kidney disease and damage has been transplantation of renal progenitor cells (RPCs), which can develop into the variety of cells required for organ repair. One problem with this line of study has been growing the number of RPCs required for effective treatment. This investigation, lead by Professor Kenji Osafune and published in Stem Cells Translational Medicine, shows iPSCs can be expanded and differentiated into RPCs at high enough levels to make them a strong candidate for the therapy.

One issue outstanding with this potential therapy is the difficulty associated with transplanting the RPCs directly into kidney parenchyma, with few studies managing to introduce sufficient cell numbers. The kidney is a very solid organ, which makes it very difficult to bring enough number of cells upon transplantation, Osafune explained.

To circumvent this problem, the team transplanted RPCs derived from iPSCs into the kidney subcapsule at the kidney surface. These cells never integrated into the host organ, but the mice receiving the treatment showed better recovery from their acute kidney injury nevertheless. Compared to control experiments, introduction of RPCs was concomitant with reduced necrosis and fibrosis of the damaged kidneys. Osafune has suggested that these improvements may be due to the RPCs expressing two known renal progenitor marker proteins, Osr1 and Six2, which have not been tested together until now.

As the cells did not integrate into the host kidney, another mode of action must have caused the benefits observed. The study concluded that paracrine secretions of renal protective factors from the RPCs caused the improvements seen in the treated mice. As kidney fibrosis marks progression towards chronic disease, Osafune hinted the paracrine secretions could be utilised as a preventative therapy for other diseases, or give clues for drug discovery. There is no medication for acute kidney injury. If we can identify the paracrine factor, maybe it will lead to a drug.

Sources: Toyohara T, Mae SU, Sueta SU et al. Cell Therapy Using Human Induced Pluripotent Stem Cell-Derived Renal Progenitors Ameliorates Acute Kidney Injury In Mice. Stem Cells Translational Medicine. doi: 10.5966/sctm.2014-0219

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Potential use of iPS cells to combat acute kidney disease ...

Combining Stem Cell Therapy with Gene Therapy | Boston …

When pluripotent stem cells are made from a patients own cells, it may be also be possible to replace the faulty gene that caused their disease with a normal, healthy copy. The repaired stem cells could then be directed to form the tissue type needed, introduced into the body, allowed to divide, and used to reconstitute the diseased tissue. It's a treatment that should last a lifetime.

Boston Childrens Hospital researcher George Q. Daley, MD, PhD, then at the Whitehead Institute, was the first to demonstrate, in 2002, that pluripotent stem cells could successfully treat a disease. Working with mice that possess a genetic defect caused by an immune deficiency, the research team created genetically-matched embryonic stem cells through nuclear transfer, introduced corrective genes, then derived healthy blood stem cells and infused them into the mice, partially restoring their immune function. Daley, Director of Stem Cell Transplantation at Childrens, would like to do the same for his patients with blood diseases.

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Combining Stem Cell Therapy with Gene Therapy | Boston ...

Clinical GMP-grade iPS cell production – Stem Cell Assays

Recently, Ive written about transition from iPS cell research to iPS cell large-scale manufacturing and automation. Ive described iPS cell process development in Cellular Dynamics International and New York Stem Cell Foundation Research Institute. Today, Id like to share presentations of 2 more players in the field Lonza and Roslin Cells. Both presentations were recorded at Stem Cell Meeting on the Mesa, held on October 14-16, 2013.

What was especially interesting to see a cost comparison between research and clinical-grade GMP-produced iPS cell lines:

(Screenshot from Lonza presentation at Stem Cell Meeting on the Mesa, 2013)

Interestingly, the major cost contributor in GMP-grade iPS cell production is a facility cost. I think, this is a first estimation of cost difference, presented for public.

The framework for establishing clinical-grade iPS cell manufacturing, nicely outlined in the recent article. Id also recommend you to read the following open access articles:

Tagged as: cost, iPS, manufacturing

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Clinical GMP-grade iPS cell production - Stem Cell Assays

Cell Therapy Blog

Under agreement withStreetWise Reports, I'm pleased to share their recent coverage of the regenerative medicine sector.

Original Source: George S. Mack of The Life Sciences Report (01/18/2015) http://www.thelifesciencesreport.com/pub/na/where-do-cures-reside-morrie-ruffin-and-michael-werner-of-the-alliance-for-regenerative-medicine-think-the-answer-is-in-regenerative-medicine

Where Do Cures Reside? Morrie Ruffin and Michael Werner of the Alliance for Regenerative Medicine Think the Answer Is in Regenerative Medicine

The practice of medicine is being transformed. The journey has been arduous, but revolutionary stem cell, gene and immunocellular therapies are rapidly moving toward pivotal milestonesand investors in the space should strap in for a rewarding joy ride. In this interview with The Life Sciences Report, Alliance for Regenerative Medicine cofounders Morrie Ruffin and Michael Werner consider how new federal guidance might enable new standards of care in cardiovascular, neurologic and oncologic medicine, and offer a preview of next week's Biotech Showcase in San Francisco.

The Life Sciences Report: On Dec. 22, the stem cell and regenerative medicine space saw a very significant development. The U.S. Food and Drug Administration (FDA) published a new draft guidance paperon what constitutes minimal manipulation of human tissues and cells. Although this document is not binding, it does tell us what the agency is thinking. In effect, tissues and cells that have been processed in some way will be treated as drugs, and will come under the regulatory umbrella of the FDA. What does this mean for the industry?

Michael Werner: As sometimes happens with government agencies, major regulatory documents are released right before the holidays. As far as the Alliance for Regenerative Medicine (ARM) is concerned, our member companies are currently looking at the paper and trying to digest it. This draft guidance is open for comment for 60 days after its publication in the Federal Register. We will go through it with a fine-tooth comb, and we will prepare formal comments, which we will submit to the FDA early this year.

Generally speaking, I think the most significant aspect of the guidance is that it provides greater clarity on the FDA's definition of minimal manipulation and, therefore, which products can qualify as minimally manipulated, and which will be regulated as biologics or drugs.

What ARM has said, from its inception many years ago, is that this industry needs a clear and predictable regulatory pathway. The regulations for minimal manipulation have been around for a while, but as this field has expanded, and as the technology has changed, questions have arisen about how the FDA is applying its regulations, and what minimally manipulated means in the context of new, tissue-engineered products.

At the very least, it's important that the FDA has, through this document, been transparent about its views.

TLSR: Michael, will you explain further why this transparency is important?

See the original post here:
Cell Therapy Blog

japanese | StemCell Therapy MD

SAN DIEGO(BUSINESS WIRE)Cytori Therapeutics, Inc. (NASDAQ: CYTX) today confirmed that two Japanese regenerative medicine laws, which went into effect on November 25, 2014, remove regulatory uncertainties and provide a clear path for the Company to commercialize and market Cytori Cell Therapy and its Celution System under the Companys existing and planned regulatory approvals.

Japans new regenerative medicine laws substantially clarify regulatory ambiguities of pre-existing guidelines and this news represents a significant event for Cytori, said Dr. Marc Hedrick, President & CEO of Cytori. We have a decade of operating experience in Japan and Cytori is nicely positioned to see an impact both on existing commercial efforts and on our longer-term efforts to obtain therapeutic claims and reimbursement for our products.

Under the two new laws, Cytori believes its Celution System and autologous adipose-derived regenerative cells (ADRCs) can be provided by physicians under current Class I device regulations and used under the lowest risk category (Tier 3) for many procedures with only the approval by accredited regenerative medicine committees and local agencies of the Ministry of Health, Labour and Welfare (MHLW). This regulatory framework is expected to streamline the approval and regulatory process and increase clinical use of Cytori Cell Therapy and the Celution System over the former regulations.

Before these new laws were enacted, the regulatory pathway for clinical use of regenerative cell therapy was one-size-fits-all, irrespective of the risk posed by certain cell types and approaches, said Dr. Hedrick. Now, Cytoris point-of-care Celution System can be transparently integrated into clinical use by providers under our Class I device status and the streamlined approval process granted to cell therapies that pose the lowest risk. Our technology is unique in that respect.

Cytoris Celution System Is in Lowest of Three Risk Categories

The Act on the Safety of Regenerative Medicines and an amendment of the 2013 Pharmaceutical Affairs Act (the PMD Act), collectively termed the Regenerative Medicine Laws, replace the Human Stem Cell Guidelines. Under the new laws, the cell types used in cell therapy and regenerative medicine are classified based on risk. Cell therapies using cells derived from embryonic, induced pluripotent, cultured, genetically altered, animal and allogeneic cells are considered higher risk (Tiers 1 and 2) and will undergo an approval pathway with greater and more stringent oversight due to the presumed higher risk to patients. Cytoris Celution System, which uses the patients own cells at the point-of-care, will be considered in the lowest risk category (Tier 3) for most cases, and will be considered in Tier 2 if used as a non-homologous therapy.

Streamlined Regulatory Approval for Certain Medical Devices

In the near future, Cytori intends to pursue disease-specific or therapeutic claims and reimbursement for Cytoris Celution System and the Company would, at that point, sponsor a clinical trial to obtain Class III device-based approval and reimbursement. The new laws include changes to streamline regulation of Class II and some Class III devices, which will now require the approval of certification bodies rather than the PMDA, similar to the European notified body model. To date, certification bodies have only been used for some Class II devices.

Conditional Regulatory Approval and Reimbursement Potential

As a supplementary benefit to Cytori, the Company may also choose to take advantage of the new conditional approval opportunities granted under the new laws. Once clinical safety and an indication of efficacy are shown, sponsors may apply for their cell product to receive conditional approval for up to seven years and may be eligible for reimbursement under Japans national insurance coverage. Under the conditional approval, the sponsor can then generate post-marketing data to demonstrate further efficacy and cost effectiveness.

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japanese | StemCell Therapy MD

japanese | StemCell Therapy MD

SAN DIEGO(BUSINESS WIRE)Cytori Therapeutics, Inc. (NASDAQ: CYTX) today confirmed that two Japanese regenerative medicine laws, which went into effect on November 25, 2014, remove regulatory uncertainties and provide a clear path for the Company to commercialize and market Cytori Cell Therapy and its Celution System under the Companys existing and planned regulatory approvals.

Japans new regenerative medicine laws substantially clarify regulatory ambiguities of pre-existing guidelines and this news represents a significant event for Cytori, said Dr. Marc Hedrick, President & CEO of Cytori. We have a decade of operating experience in Japan and Cytori is nicely positioned to see an impact both on existing commercial efforts and on our longer-term efforts to obtain therapeutic claims and reimbursement for our products.

Under the two new laws, Cytori believes its Celution System and autologous adipose-derived regenerative cells (ADRCs) can be provided by physicians under current Class I device regulations and used under the lowest risk category (Tier 3) for many procedures with only the approval by accredited regenerative medicine committees and local agencies of the Ministry of Health, Labour and Welfare (MHLW). This regulatory framework is expected to streamline the approval and regulatory process and increase clinical use of Cytori Cell Therapy and the Celution System over the former regulations.

Before these new laws were enacted, the regulatory pathway for clinical use of regenerative cell therapy was one-size-fits-all, irrespective of the risk posed by certain cell types and approaches, said Dr. Hedrick. Now, Cytoris point-of-care Celution System can be transparently integrated into clinical use by providers under our Class I device status and the streamlined approval process granted to cell therapies that pose the lowest risk. Our technology is unique in that respect.

Cytoris Celution System Is in Lowest of Three Risk Categories

The Act on the Safety of Regenerative Medicines and an amendment of the 2013 Pharmaceutical Affairs Act (the PMD Act), collectively termed the Regenerative Medicine Laws, replace the Human Stem Cell Guidelines. Under the new laws, the cell types used in cell therapy and regenerative medicine are classified based on risk. Cell therapies using cells derived from embryonic, induced pluripotent, cultured, genetically altered, animal and allogeneic cells are considered higher risk (Tiers 1 and 2) and will undergo an approval pathway with greater and more stringent oversight due to the presumed higher risk to patients. Cytoris Celution System, which uses the patients own cells at the point-of-care, will be considered in the lowest risk category (Tier 3) for most cases, and will be considered in Tier 2 if used as a non-homologous therapy.

Streamlined Regulatory Approval for Certain Medical Devices

In the near future, Cytori intends to pursue disease-specific or therapeutic claims and reimbursement for Cytoris Celution System and the Company would, at that point, sponsor a clinical trial to obtain Class III device-based approval and reimbursement. The new laws include changes to streamline regulation of Class II and some Class III devices, which will now require the approval of certification bodies rather than the PMDA, similar to the European notified body model. To date, certification bodies have only been used for some Class II devices.

Conditional Regulatory Approval and Reimbursement Potential

As a supplementary benefit to Cytori, the Company may also choose to take advantage of the new conditional approval opportunities granted under the new laws. Once clinical safety and an indication of efficacy are shown, sponsors may apply for their cell product to receive conditional approval for up to seven years and may be eligible for reimbursement under Japans national insurance coverage. Under the conditional approval, the sponsor can then generate post-marketing data to demonstrate further efficacy and cost effectiveness.

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japanese | StemCell Therapy MD

Mobilephone | IPSCELLTHERAPY

Most know that Apple is always a bit behind when delivering latest technology in their iOS gadgets, even though the iOS realistic would no doubt say or else. But as far as I am aware Apple doesnt have any 3D plans for their device. 3D appears to be the next big thing in the mobile space and no doubt the iOS faithful wouldnt want to lose out, but if Apple doesnt bring 3D to their gadgets it appears someone else will.

Actually this resolution involves placing a film over your iOS display and combining with softwareto deliver 3D and actually the film doesnt hamper with your multi-touch gestures. So just so you can check out what this new 3D film and software does, we have a video of 3D in action on the Apple iPad for your viewing contentment below which lasts just 46 seconds by does look relatively remarkable. More information about sell my mobile phone can be found at this http://www.onrecycle.co.uk.

PC provider Acer has provided the Liquid Metal smartphone, a device which it hopes will allow it to compete with the major rivals in what is becoming an increasingly crowded market.

The Acer Liquid Metals specifications make interesting reading, with version 2.2 of Android onboard accompanied by the specialised Breeze user interface.

Its 5MP camera with HD video capture lurks on the rear, whilst a 3.6 inch display using capacitive touch technology makes an appearance on the front, making it a hair larger than the iPhone 4. Sadly the displays resolution is unlikely to match that of Apples smartphone king.

An 800MHz processor will give life to Android, but it is slightly strange to see a new mobile emerging with anything less than 1GHz of processing power under the hood, so it will be interesting to see how the Acer Liquid Metal has been optimised to squeeze the most from this chip.

Officially announced less than two months ago, the Garmin-Asus M10 has just been launched in India, being the countrys first Windows Mobile 6.5.3 smartphone. The M10 offers maps for 62 major Indian cities, Garmin turn-by-turn navigation, lane assistance, and a Ciao feature that keeps you informed on the roads your friends are traveling on.

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Mobilephone | IPSCELLTHERAPY

Patient-specific stem cells and personalized gene therapy …

These are images of normal (above) and diseased retinas. Patients with MFRP mutations, a cause of retinitis pigmentosa, lose the function of most retinal cells, particularly at the periphery of the retina, leaving them with drastically reduced vision. Personalized gene therapy, using iPS cells, may offer a way to correct this genetic disorder.

Vision loss patients own cells transformed into model for studying disease and developing potential treatment

Columbia University Medical Center (CUMC) researchers have created a way to develop personalized gene therapies for patients with retinitis pigmentosa (RP), a leading cause of vision loss. The approach, the first of its kind, takes advantage of induced pluripotent stem (iPS) cell technology to transform skin cells into retinal cells, which are then used as a patient-specific model for disease study and preclinical testing.

Using this approach, researchers led by Stephen H. Tsang, MD, PhD, showed that a form of RP caused by mutations to the gene MFRP (membrane frizzled-related protein) disrupts the protein that gives retinal cells their structural integrity. They also showed that the effects of these mutations can be reversed with gene therapy. The approach could potentially be used to create personalized therapies for other forms of RP, as well as other genetic diseases. The paper was published recently in the online edition of Molecular Therapy, the official journal of the American Society for Gene & Cell Therapy.

In normal, or wild-type, retinal cells (left), the protein actin forms the cells cytoskeleton, creating an internal support structure that looks like a series of connected hexagons. In cells with MFRP mutations (center), this structure fails to form, compromising cellular function. When diseased retinal cells are treated with gene therapy to insert normal copies of MFRP (right), the cells cytoskeleton and function are restored. (Image credit: Lab of Stephen H. Tsang, MD, PhD/Columbia University Medical Center.)

The use of patient-specific cell lines for testing the efficacy of gene therapy to precisely correct a patients genetic deficiency provides yet another tool for advancing the field of personalized medicine, said Dr. Tsang, the Laszlo Z. Bito Associate Professor of Ophthalmology and associate professor of pathology and cell biology.

While RP can begin during infancy, the first symptoms typically emerge in early adulthood, starting with night blindness. As the disease progresses, affected individuals lose peripheral vision. In later stages, RP destroys photoreceptors in the macula, which is responsible for fine central vision. RP is estimated to affect at least 75,000 people in the United States and 1.5 million worldwide.

More than 60 different genes have been linked to RP, making it difficult to develop models to study the disease. Animal models, though useful, have significant limitations because of interspecies differences. Researchers also use human retinal cells from eye banks to study RP. As these cells reflect the end stage of the disease process, however, they reveal little about how the disease develops. There are no human tissue culture models of RP, as it would dangerous to harvest retinal cells from patients. Finally, human embryonic stem cells could be useful in RP research, but they are fraught with ethical, legal, and technical issues.

The use of iPS technology offers a way around these limitations and concerns. Researchers can induce the patients own skin cells to revert to a more basic, embryonic stem celllike state. Such cells are pluripotent, meaning that they can be transformed into specialized cells of various types.

In the current study, the CUMC team used iPS technology to transform skin cells taken from two RP patientseach with a different MFRP mutationinto retinal cells, creating patient-specific models for studying the disease and testing potential therapies.

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Patient-specific stem cells and personalized gene therapy ...

IPS Stem Cells: New Ethical Quandaries

IPS Stem Cells: New Ethical Quandaries By Sally Lehrman

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When scientists learned how to turn back the clock in a young skin cell, to bring it back to an early-stage cell that could become any other type in the body, both they and ethicists rejoiced. The reprogrammed cell was pluripotent, much like an embryonic stem cell. Maybe even better, it also might be prompted to jump from one cell type to another.

One day, these induced pluripotent stem cells -- iPS cells for short -- might be able to correct any number of life-threatening and disabling conditions. Much sooner, these cells will almost certainly serve as extremely useful models for studying disease.

The researchers used viruses to deliver three to four new genes into the cell nucleus. And with the new information, the skin cells reprogrammed themselves. They behaved almost exactly like embryonic stem cells, which are derived from fertilized eggs. But with these reprogrammed cells, people thought, there would be no moral and political controversy. No embryo would be destroyed.

Recently, new studies have taken the work a step further. Researchers used synthetic RNA instead of viruses to get new instructions into the cell nucleus. This sped up the process and lessened the possibility of side effects such as cancer when the cells finally become a treatment for patients. (They're called RNA-induced pluripotent cells.)

But as researchers and ethicists take a closer look at these iPS cells, they are realizing that the issues posed are as thorny as ever. In fact, the issues may be even more urgent because the new techniques are so much easier and cheaper. The concerns fall into three main areas.

First, the possibility of human cloning from one person's skin cells or human reproduction from cells made into sperm and egg. The possibility is far-off, but real. Scientists already have reported progress that could lead to either. One could become a parent at any age, using tissue from someone either living or dead.

More immediate concerns have to do with control of the original tissue donation and the purposes to which it is applied.

For instance, privacy. Because of the desire to use these cells to study or treat diseases such as Parkinson's, juvenile diabetes or Alzheimer's, it will be important to know the donor's health history. The donor's personal information and health history must always be linked to the cells. It may be impossible to maintain donor privacy.

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IPS Stem Cells: New Ethical Quandaries

CDI | iPS Cells – Cellular Dynamics | Home

How does CDI's technology work? A human biological sample, for example blood or skin, is obtained, and the cells within the sample are grown under appropriate cell culture conditions. In the episomal reprogramming method, vectors containing multiple reprogramming genes are introduced into the cells.

While the vectors turn genes in the cell on and off, reprogramming them to a stem cell state, they do not integrate into the genome itself. This method alleviates concerns arising over the potential risks associated with the insertion of foreign DNA to induce reprogramming, which other prior iPS methods use (bottom row in illustration above).

iPS cells are somatic cells (e.g., skin or blood) that have been genetically reprogrammed to a pluripotent stem cell state through forced expression of pluripotency genes.By definition, iPS cells replicate indefinitely and have the potential to differentiate into any cell type in the human body.

Reprogramming factors are the genes introduced into somatic cells that induce a pluripotent stem cell state. Initial reports describing the creation of human iPS cells utilized four reprogramming factors: OCT4, SOX2, KLF4 and MYC (OSKM) (Takahashi, et al. 2007) or OCT4, SOX2, NANOG and LIN28 (OSNL) (Yu, et al. 2007). Subsequent studies revealed that reprogramming using a specific combination of all 6 of these factors combined with SV40LT and a cocktail of small molecules yields iPS cells at much higher efficiency (Yu, et al. 2009; Yu, et al. 2011).

iPS cells are genetically reprogrammed through forced expression of pluripotency genes into somatic cells.The expression of these genes can be accomplished using a variety of different methods.The episomal reprogramming method introduces pluripotency genes into a target cell using circular DNA plasmid vectors (i.e. episomes) that replicate autonomously within the cell cytoplasm and do not integrate into the host cell genome.

Initial methods of iPS cell reprogramming utilized retroviral and lentiviral vectors to introduce pluripotency genes into somatic cells. While these methods generally work well, the viral DNA integrates into the genome of the target cell, and the resulting iPS cells (and cells differentiated from them) will contain foreign DNA, which may result in defects and errors. By contrast, episomal vectors replicate autonomously within the cell cytoplasm and do not integrate into the host genome. In addition, the episomal vectors are released from the target cell at a rate of ~5% per cell cycle resulting in transgene-free or footprint-free iPS cells.These features, combined with recent advancements in episomal reprogramming efficiency, have led to a strong preference for this method to alleviate concerns about genome integrity for drug discovery and cell therapy applications.

Episomal reprogramming has been reported successful from a variety of somatic cells, including fibroblasts, lymphoblastoid cells, and peripheral blood mononuclear cells. Importantly, CDI has optimized its episomal reprogramming method to achieve high efficiency iPS cell generation from small amounts of human peripheral blood. Not only does this enable more streamlined and less invasive collection of donor samples, but ensures increased sterility and lower cost production of iPS cells. In addition, efficient iPS cell production from peripheral blood enables access to large banks of normal and disease-associated clinical samples for disease research and drug screening.

CDIs suite of MyCell Products includes episomal reprogramming of customer-provided donor samples and subsequent genetic engineering and/or differentiation of the iPS cells. In addition, for researchers who would like to generate their own iPS cells, CDIs episomal reprogramming technology is available as a kit from Life Technologies, including Episomal iPSC Reprogramming Vectors, Vitronectin, and Essential 8 Medium. Customer-generated iPS cells using this kit may then be transferred to CDI for genetic engineering and/or differentiation through MyCell Products.

Integration-free iPS cells have been generated using a variety of methods including adenovirus, Sendai virus, piggyBac, minicircle vectors, and direct introduction of protein or synthesized mRNA. The efficiency and success rate of these methods varies depending on the source of somatic cells and experimental conditions, but in general these approaches are limited by impractically low reprogramming efficiency, requirement for higher biosafety containment, and/or labor- and cost-intensive protocols that require repeated transfection/infection.Compared to these methods, episomal reprogramming is virus-free, safe to use, stable, and inexpensive.

A variety of small molecules have been identified that can functionally substitute for one or more reprogramming factors and/or improve the efficiency of iPS cell reprogramming. However, no combination of small molecules has been shown to functionally substitute for all four reprogramming factors. The use of small molecules in iPS cell reprogramming offers some practical advantages including the ability to optimize the chemical structure, fine-tune dose and concentration, and simplify handling and application protocols. However, the use of small molecules presents a number of scientific challenges. Most notably, small molecules may have more than one target, which may or may not be known. In addition, unexpected toxicity and other side effects in vivo may interfere with the clinical application of small molecules.

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CDI | iPS Cells - Cellular Dynamics | Home

Stem Cell Therapy for Neuromuscular Diseases | InTechOpen

Invest in iPS @ TDI | ALS Therapy Development Institute

Steve Perrin, Ph.D., CEO and CSO, discusses why iPS technology is ready for drug discovery for today's ALS patients. Click here to learn why Steve believes TDI is uniquely suited to implement this technology in ALS research.

Fernando Vieira, M.D., director of research operations, discusses how iPS technology can be used to model sporadic ALS, help to identify sub-types of ALS patients and accelerate drug development as part of a comprehensive translational research program at ALS TDI.

Jessie St. Martin, associate scientist, talks about induced pluripotent stem cells (iPS cells) and their importance in ALS research. Jessie, a recent addition to the translational research team, will play an integral part in developing this program at ALS TDI. Click here to learn more about iPS cells.

Jenny Dwyer, board member, explains why your support of the iPS program at ALS TDI may have the ability to rapidly accelerate treatments for today's patients. Jenny was a longtime ALS caregiver of her husband, Pat. Together, they were advocates for ALS research. Click here to listen to her message.

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Invest in iPS @ TDI | ALS Therapy Development Institute

Induced pluripotent stem cell therapy – Wikipedia, the …

In 2006, Shinya Yamanaka of Kyoto University in Japan was the first to disprove the previous notion that reversible cell differentiation of mammals was impossible. He reprogrammed a fully differentiated mouse cell into a pluripotent stem cell by introducing four genes, Oct-4, SOX2, KLF4, and Myc, into the mouse fibroblast through gene-carrying viruses. With this method, he and his coworkers created induced pluripotent stem cells (iPS cells), the key component in this experiment.[1] Scientists have been able to conduct experiments that show the ability of iPS cells to treat and even cure diseases. In this experiment, tests were run on mice with inherited sickle cell anemia.Skin cells were turned into cells containing genes that transformed the cells into iPS cells. These replaced the diseased sickled cells, curing the test mice. The reprogramming of the pluripotent stem cells in mice was successfully duplicated with human pluripotent stem cells within about a year of the experiment on the mice.

Sickle cell anemia is a disease in which the body produces abnormally shaped red blood cells. Red blood cells are flexible and round, moving easily through the blood vessels. Infected cells are shaped like a crescent or sickle (the namesake of the disease). As a result of this disorder the hemoglobin protein in red blood cells is faulty. Normal hemoglobin bonds to oxygen, then releases it into cells that need it. The blood cell retains its original form and is cycled back to the lungs and re-oxygenated.

Sickle cell hemoglobin, however, after giving up oxygen, cling together and make the red blood cell stiff. The sickle shape also makes it difficult for the red blood cell to navigate arteries and causes blockages.[2] This can cause intense pain and organ damage. The sickled red blood cells are fragile and prone to rupture. When the number of red blood cells decreases from rupture (hemolysis), anemia is the result. Sickle cells also die in 1020 days as opposed to the traditional 120-day lifespan of a normal red blood cell.

Sickle cell anemia is inherited as an autosomal (meaning that the gene is not linked to a sex chromosome) recessive condition.[2] This means that the gene can be passed on from a carrier to his or her children. In order for sickle cell anemia to affect a person, the gene must be inherited from both the mother and the father, so that the child has two recessive sickle cell genes (a homozygous inheritance). People who inherit one sickle cell gene from one parent and one normal gene from the other parent, i.e. heterozygous patients, have a condition called sickle cell trait. Their bodies make both sickle hemoglobin and normal hemoglobin. They may pass the trait on to their children.

The effects of sickle cell anemia vary from person to person. People who have the disease suffer from varying degrees of chronic pain and fatigue. With proper care and treatment, the quality of health of most patients will improve. Doctors have learned a great deal about sickle cell anemia since its discovery in 1979. They know its causes, its effects on the body, and possible treatments for complications. Sickle cell anemia has no widely available cure. A bone marrow transplant is the only treatment method currently recognized to be able to cure the disease, though it does not work for every patient. Finding a donor is difficult and the procedure could potentially do more harm than good. Treatments for sickle cell anemia are generally aimed at avoiding crises, relieving symptoms, and preventing complications. Such treatments may include medications, blood transfusions, and supplemental oxygen.

During the first step of the experiment, skin cells (also known as fibroblasts) were collected from infected test mice and put in a culture. The fibroblasts were reprogrammed by infecting them with retroviruses that contained genes common to embryonic stem cells. These genes were the same four used by Yamanaka (Oct-4, SOX2, KLF4, and Myc) in his earlier study. The investigators were trying to produce cells with the potential to differentiate into any type of cell needed (i.e. pluripotent stem cells). As the experiment continued, the fibroblasts multiplied into identical copies of iPS cells. The cells were then treated to form the mutation needed to reverse the anemia in the mice. This was accomplished by restructuring the DNA containing the defective globin gene into DNA with the normal gene through the process of homologous recombination. The iPS cells then differentiated into blood stem cells, or hematopoietic stem cells. The hematopoietic cells were injected back into the infected mice, where they proliferate and differentiate into normal blood cells, curing the mice of the disease.[3][4][verification needed]

To determine whether the mice were cured from the disease, the scientists checked for the usual symptoms of sickle cell disease. They examined the blood for mean corpuscular volume (MCV) and red cell distribution width (RDW) and urine concentration defects. They also checked for sickled red blood cells. They examined the DNA through gel electrophoresis, checking for bands that display an allele that causes sickling. Compared to the untreated mice with the disease, which they used as a control, the treated animals had marked increases in RBC counts, healthy hemoglobin, and packed cell volume levels.[5]

Researchers examined the urine concentration defect, which results from RBC sickling in renal tubules and consequent reduction in renal medullary blood flow, and the general deteriorated systemic condition reflected by lower body weight and increased breathing.[5] They were able to see that these parts of the body of the mice had healed or improved. This indicated that all hematological and systemic parameters of sickle cell anemia improved substantially and were comparable to those in control mice.[5] They cannot say if this will work in humans because a safe way to inject the genes for the induced pluripotent cells is still needed.[citation needed]

The reprogramming of the induced pluripotent stem cells in mice was successfully duplicated in humans within a year of the successful experiment on the mice. This reprogramming was done in several labs and it was shown that the iPS cells in humans were almost identical to original embryonic stem cells (ES cells) that are responsible for the creation of all structures in a fetus.[1] An important feature of iPS cells is that they can be generated with cells taken from an adult, which would circumvent many of the ethical problems associated with working with ES cells. These iPS cells also have potential in creating and examining new disease models and developing more efficient drug treatments.[6] Another feature of these cells is that they provide researchers with a human cell sample, as opposed to simply using an animal with similar DNA, for drug testing.

One major problem with iPS cells is the way in which the cells are reprogrammed. Using gene-carrying viruses has the potential to cause iPS cells to develop into cancerous cells.[1] Also, an implant made using undifferentiated iPS cells, could cause a teratoma to form. Any implant that is generated from using these iPS cells would only be viable for transplant into the original subject that the cells were taken from. In order for these iPS cells to become viable in therapeutic use, there are still many steps that must be taken.[5][7]

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Induced pluripotent stem cell therapy - Wikipedia, the ...

MDC researchers greatly increase precision of new genome editing tool

CRISPR-Cas9 is a powerful new tool for editing the genome. For researchers around the world, the CRISPR-Cas9 technique is an exciting innovation because it is faster and cheaper than previous methods. Now, using a molecular trick, Dr. Van Trung Chu and Professor Klaus Rajewsky of the Max Delbrck Center for Molecular Medicine (MDC) Berlin-Buch and Dr. Ralf Khn, MDC and Berlin Institute of Health (BIH), have found a solution to considerably increase the efficiency of precise genetic modifications by up to eightfold (Nature Biotechnology: doi:10.1038/nbt.3198)**.

"What we used to do in years, we can now achieve in months," said gene researcher and immunologist Klaus Rajewsky, indicating the power of this new genome-editing technology. CRISPR-Cas9 not only speeds up research considerably - at the same time it is much more efficient, cheaper and also easier to handle than the methods used so far.

The CRISPR-Cas9 technology allows researchers to transiently introduce DNA double-strand breaks into the genome of cells or model organisms at genes of choice. In these artificially produced strand breaks, they can insert or cut out genes and change the genetic coding according to their needs.

Mammalian cells are able to repair DNA damage in their cells using two different repair mechanisms. The homology-directed repair (HDR) pathway enables the insertion of preplanned genetic modifications using engineered DNA molecules that share identical sequence regions with the targeted gene and which are recognized as a repair template. Thus, HDR repair is very precise but occurs only at low frequency in mammalian cells.

The other repair system, called non-homologous end-joining (NHEJ) is more efficient in nature but less precise, since it readily reconnects free DNA ends without repair template, thereby frequently deleting short sequences from the genome. Therefore, NHEJ repair can only be used to create short genomic deletions, but does not support precise gene modification or the insertion and replacement of gene segments.

Many researchers, including Van Trung Chu, Klaus Rajewsky and Ralf Khn, are seeking to promote the HDR repair pathway to make gene modification in the laboratory more precise in order to avoid editing errors and to increase efficiency. The MDC researchers succeeded in increasing the efficiency of the more precisely working HDR repair system by temporarily inhibiting the most dominant repair protein of NHEJ, the enzyme DNA Ligase IV. In their approach they used various inhibitors such as proteins and small molecules.

"But we also used a trick of nature and blocked Ligase IV with the proteins of adeno viruses. Thus we were able to increase the efficiency of the CRISPR-Cas9 technology up to eightfold," Ralf Khn explained. For example, they succeeded in inserting a gene into a predefined position in the genome (knock-in) in more than 60 per cent of all manipulated mouse cells. Khn has just recently joined the MDC and is head of the research group for "iPS cell based disease modeling". Before coming to the MDC, he was on the research staff of Helmholtz Zentrum Mnchen. "The expertise of Ralf Khn is very important for gene research at MDC and especially for my research group," Klaus Rajewsky said.

Concurrent with the publication of the article by the MDC researchers, Nature Biotechnology published another, related paper on CRISPR-Cas9 technology. It comes from the laboratory of Hidde Ploegh of the Whitehead Institute in Cambridge, MA, USA.

Somatic gene therapy with CRISPR-Cas9 is a goal

The new CRISPR-Cas9 technology, developed in 2012, is already used in the laboratory to correct genetic defects in mice. Researchers also plan to modify the genetic set up of induced pluripotent stem cells (iPS), which can be differentiated into specialized cell types or tissues. That is, researchers are able to use the new tool to introduce patient-derived mutations into the genome of iPS cells for studying the onset of human diseases. "Another future goal, however, is to use CRISPR-Cas9 for somatic gene therapy in humans with severe diseases," Klaus Rajewsky pointed out.

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MDC researchers greatly increase precision of new genome editing tool

Researchers greatly increase precision of new genome editing tool

CRISPR-Cas9 is a powerful new tool for editing the genome. For researchers around the world, the CRISPR-Cas9 technique is an exciting innovation because it is faster and cheaper than previous methods. Now, using a molecular trick, Dr. Van Trung Chu and Professor Klaus Rajewsky of the Max Delbrck Center for Molecular Medicine (MDC) Berlin-Buch and Dr. Ralf Khn, MDC and Berlin Institute of Health (BIH), have found a solution to considerably increase the efficiency of precise genetic modifications by up to eightfold.

"What we used to do in years, we can now achieve in months," said gene researcher and immunologist Klaus Rajewsky, indicating the power of this new genome-editing technology. CRISPR-Cas9 not only speeds up research considerably - at the same time it is much more efficient, cheaper and also easier to handle than the methods used so far.

The CRISPR-Cas9 technology allows researchers to transiently introduce DNA double-strand breaks into the genome of cells or model organisms at genes of choice. In these artificially produced strand breaks, they can insert or cut out genes and change the genetic coding according to their needs.

Mammalian cells are able to repair DNA damage in their cells using two different repair mechanisms. The homology-directed repair (HDR) pathway enables the insertion of preplanned genetic modifications using engineered DNA molecules that share identical sequence regions with the targeted gene and which are recognized as a repair template. Thus, HDR repair is very precise but occurs only at low frequency in mammalian cells.

The other repair system, called non-homologous end-joining (NHEJ) is more efficient in nature but less precise, since it readily reconnects free DNA ends without repair template, thereby frequently deleting short sequences from the genome. Therefore, NHEJ repair can only be used to create short genomic deletions, but does not support precise gene modification or the insertion and replacement of gene segments.

Many researchers, including Van Trung Chu, Klaus Rajewsky and Ralf Khn, are seeking to promote the HDR repair pathway to make gene modification in the laboratory more precise in order to avoid editing errors and to increase efficiency. The MDC researchers succeeded in increasing the efficiency of the more precisely working HDR repair system by temporarily inhibiting the most dominant repair protein of NHEJ, the enzyme DNA Ligase IV. In their approach they used various inhibitors such as proteins and small molecules.

"But we also used a trick of nature and blocked Ligase IV with the proteins of adeno viruses. Thus we were able to increase the efficiency of the CRISPR-Cas9 technology up to eightfold," Ralf Khn explained. For example, they succeeded in inserting a gene into a predefined position in the genome (knock-in) in more than 60 per cent of all manipulated mouse cells. Khn has just recently joined the MDC and is head of the research group for "iPS cell based disease modeling." Before coming to the MDC, he was on the research staff of Helmholtz Zentrum Mnchen. "The expertise of Ralf Khn is very important for gene research at MDC and especially for my research group," Klaus Rajewsky said.

Concurrent with the publication of the article by the MDC researchers, Nature Biotechnology published another, related paper on CRISPR-Cas9 technology. It comes from the laboratory of Hidde Ploegh of the Whitehead Institute in Cambridge, MA, USA.

Somatic gene therapy with CRISPR-Cas9 is a goal

The new CRISPR-Cas9 technology, developed in 2012, is already used in the laboratory to correct genetic defects in mice. Researchers also plan to modify the genetic set up of induced pluripotent stem cells (iPS), which can be differentiated into specialized cell types or tissues. That is, researchers are able to use the new tool to introduce patient-derived mutations into the genome of iPS cells for studying the onset of human diseases. "Another future goal, however, is to use CRISPR-Cas9 for somatic gene therapy in humans with severe diseases," Klaus Rajewsky pointed out.

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Researchers greatly increase precision of new genome editing tool

A single-cell breakthrough

Mar 18, 2015 by Marla Vacek Broadfoot A jelly fish-green fluorescent gene marks stem cells and other proliferating primitive cells of an intestine-like structure. The central lumen hollow space is stained red. Credit: Magness Lab

The human gut is a remarkable thing. Every week the intestines regenerate a new lining, sloughing off the equivalent surface area of a studio apartment and refurbishing it with new cells. For decades, researchers have known that the party responsible for this extreme makeover were intestinal stem cells, but it wasn't until this year that Scott Magness, PhD, associate professor of medicine, cell biology and physiology, and biomedical engineering, figured out a way to isolate and grow thousands of these elusive cells in the laboratory at one time. This high throughput technological advance now promises to give scientists the ability to study stem cell biology and explore the origins of inflammatory bowel disease, intestinal cancers, and other gastrointestinal disorders.

But it didn't come easy.

One step forward

When Magness and his team first began working with intestinal stem cells some years ago, they quickly found themselves behind the eight ball. Their first technique involved using a specific molecule or marker on the surface of stem cells to make sure they could distinguish stem cells from other intestinal cells.

Then Magness's team would fish out only the stem cells from intestinal tissues and grow the cells in Petri dishes. But there was a problem. Even though all of the isolated cells had the same stem cell marker, only one out of every 100 could "self-renew" and differentiate into specialized cells like a typical stem cell should. (Stem cells spawn cells that have specialized functions necessary for any organ to work properly.)

"The question was: why didn't the 99 others behave like stem cells?" Magness said. "We thought it was probably because they're not all the same, just like everybody named Judy doesn't look the same. There are all kinds of differences, and we've been presuming that these cells are all the same based on this one name, this one molecular marker. That's been a problem. But the only way to solve it so we could study these cells was to look at intestinal stem cells at the single cell level, which had never been done before."

Magness is among a growing contingent of researchers who recognize that many of the biological processes underlying health and disease are driven by a tiny fraction of the 37 trillion cells that make up the human body. Individual cells can replenish aging tissues, develop drug resistance, and become vehicles for viral infections. And yet the effects of these singular actors are often missed in biological studies that focus on pooled populations of thousands of seemingly "identical" cells.

Distinguishing between the true intestinal stem cells and their cellular look-a-likes would require isolating tens of thousands of stem cells and tracking the behavior of each individual cell over time. But Magness had no idea how to accomplish that feat. Enter Nancy Allbritton, PhD, chair of the UNC/NCSU Joint Department of Biomedical Engineering. The two professors met one day to discuss Magness joining the biomedical engineering department as an adjunct faculty member. And they did discuss it. And Magness did join. But the meeting quickly turned into collaboration.

One of Allbritton's areas of expertise is microfabrication the ability to squeeze large devices into very small footprints. During their meeting, Allbritton showed Magness her latest creation, a device smaller than a credit card dotted with 15,000 tiny wells for culturing cells.

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A single-cell breakthrough

Invest in iPS @ TDI | ALS Therapy Development Institute

Steve Perrin, Ph.D., CEO and CSO, discusses why iPS technology is ready for drug discovery for today's ALS patients. Click here to learn why Steve believes TDI is uniquely suited to implement this technology in ALS research.

Fernando Vieira, M.D., director of research operations, discusses how iPS technology can be used to model sporadic ALS, help to identify sub-types of ALS patients and accelerate drug development as part of a comprehensive translational research program at ALS TDI.

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JCM | Free Full-Text | Myogenic Precursors from iPS Cells …

1 Control of Pluripotency Laboratory, Department of Physiological Sciences I, Faculty of Medicine, University of Barcelona, Hospital Clinic, Casanova 143, 08036, Barcelona, Spain 2 Faculty of Medicine, University of Sydney Medical School, Division of Pediatrics and Child Health, Westmead Children's Hospital, Locked Bag 4001, Westmead NSW 2145, Sydney, Australia 3 School of Anatomy Physiology & Human Biology and The Harry Perkins Institute for Medical Research (CCTRM), the University of Western Australia, 6 Verdun St, Nedlands WA 6009, Perth, Australia

* Author to whom correspondence should be addressed.

Received: 1 October 2014 / Accepted: 3 December 2014 / Published: 29 January 2015

Abstract: The use of adult myogenic stem cells as a cell therapy for skeletal muscle regeneration has been attempted for decades, with only moderate success. Myogenic progenitors (MP) made from induced pluripotent stem cells (iPSCs) are promising candidates for stem cell therapy to regenerate skeletal muscle since they allow allogenic transplantation, can be produced in large quantities, and, as compared to adult myoblasts, present more embryonic-like features and more proliferative capacity in vitro, which indicates a potential for more self-renewal and regenerative capacity in vivo. Different approaches have been described to make myogenic progenitors either by gene overexpression or by directed differentiation through culture conditions, and several myopathies have already been modeled using iPSC-MP. However, even though results in animal models have shown improvement from previous work with isolated adult myoblasts, major challenges regarding host response have to be addressed and clinically relevant transplantation protocols are lacking. Despite these challenges we are closer than we think to bringing iPSC-MP towards clinical use for treating human muscle disease and sporting injuries.

Roca, I.; Requena, J.; Edel, M.J.; Alvarez-Palomo, A.B. Myogenic Precursors from iPS Cells for Skeletal Muscle Cell Replacement Therapy. J. Clin. Med. 2015, 4, 243-259.

Roca I, Requena J, Edel MJ, Alvarez-Palomo AB. Myogenic Precursors from iPS Cells for Skeletal Muscle Cell Replacement Therapy. Journal of Clinical Medicine. 2015; 4(2):243-259.

Roca, Isart; Requena, Jordi; Edel, Michael J.; Alvarez-Palomo, Ana B. 2015. "Myogenic Precursors from iPS Cells for Skeletal Muscle Cell Replacement Therapy." J. Clin. Med. 4, no. 2: 243-259.

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