Application of human mesenchymal and pluripotent stem cell microcarrier cultures in cellular therapy: Achievements and future direction

Mesenchymal stem cells (MSCs) have recently made significant progress with multiple clinical trials targeting modulation of immune responses, regeneration of bone, cartilage, myocardia, and diseases like Metachromatic leukodystrophy and Hurler syndrome. On the other hand, the use of human embryonic and induced pluripotent stem cells (hPSCs) in clinical trials is rather limited mainly due to safety issues. Only two clinical trials, retinal pigment epithelial transplantation and treatment of spinal cord injury were reported. Cell doses per treatment can range between 50,000 and 6 billion cells. The current 2-dimensional tissue culture platform can be used when low cell doses are needed and it becomes impractical when doses above 50 million are needed. This demand for future cell therapy has reinvigorated interests in the use of the microcarrier platform for generating stem cells in a scalable 3-dimensional manner.     

Clinical trials of CAR-T cells in China

Novel immunotherapeutic agents targeting tumor-site microenvironment are revolutionizing cancer therapy. Chimeric antigen receptor (CAR)-engineered T cells are widely studied for cancer immunotherapy. CD19-specific CAR-T cells, tisagenlecleucel, have been recently approved for clinical application. Ongoing clinical trials are testing CAR designs directed at novel targets involved in hematological and solid malignancies. In addition to trials of single-target CAR-T cells, simultaneous and sequential CAR-T cells are being studied for clinical applications. Multi-target CAR-engineered T cells are also entering clinical trials. T cell receptor-engineered CAR-T and universal CAR-T cells represent new frontiers in CAR-T cell development. In this study, we analyzed the characteristics of CAR constructs and registered clinical trials of CAR-T cells in China and provided a quick glimpse of the landscape of CAR-T studies in China.     

Ex vivo expansion of cord blood mononuclear cells on mesenchymal stem cells

BACKGROUND: Cord blood (CB) cells are being used increasingly as a source of hematopoietic cells to support high dose chemotherapy. However, CB units contain low numbers of cells, including CD34+ cells, and thus their use is associated with significant delays in engraftment of neutrophils and platelets. Exvivo expansion of CB has been proposed to increase the numbers of cells available. We and others have reported the requirement of CD34 selection for optimal expansion of CB products'; however, the selection of frozen CB products in clinical trials results in significant loss of CD34+ cells, with a median recovery of 50, but less than 40% recovery in more than one-third of products. In the present studies we evaluated the potential of mesenchymal stem cells (MSC) to support ex vivo expansion of unselected CB products. METHODS: Mononuclear cells (MNC) from CB products were isolated and cultured on preformed MSC layers in T150 flasks containing 50 mL Stemline II media plus hematopoietic growth factors. Various culture conditions were compared for optimal expansion of the CB MNC. RESULTS: Ex vivo expansion of CB MNC on MSC resulted in 10- to 20-fold expansion of total nucleated cells, seven- to 18-fold expansion of committed progenitor cells, two- to five-fold expansion of primitive progenitor cells and 16- to 37-fold expansion of CD34+ cells. DISCUSSION: These studies demonstrated significant expansion of CB products without CD34 cell selection using culture conditions that are clinically applicable. Our current focus is to initiate clinical trials to evaluate the in vivo potential of CB cells expanded with these conditions.     

异种器官移植：前景如何？

目前异种器官移植在进入临床的过程中存在两大难题——免疫排斥和病原微生物感染.人类为防止免疫排斥而采取的措施使超急性排斥得到了很大程度的避免,但大大增加了器官移植受体感染病原微生物的机会.猪作为人类理想的器官供体,却有多种内源性病毒,其中猪内源性逆转录病毒（PERV）引起了人类较大的关注.PERV的跨种感染存在正、反两方面的证据,使得异种器官移植的前景喜忧掺半.     

Treatment of Parkinson's disease using cell transplantation

The clinical trials with intrastriatal transplantation of human fetal mesencephalic tissue, rich in dopaminergic neurons, in Parkinson''s disease (PD) patients show that cell replacement can work and in some cases induce major, long-lasting improvement. However, owing to poor tissue availability, this approach can only be applied in very few patients, and standardization is difficult, leading to wide variation in functional outcome. Stem cells and reprogrammed cells could potentially be used to produce dopaminergic neurons for transplantation. Importantly, dopaminergic neurons of the correct substantia nigra phenotype can now be generated from human embryonic stem cells in large numbers and standardized preparations, and will soon be ready for application in patients. Also, human induced pluripotent stem cell-derived dopaminergic neurons are being considered for clinical translation. Available data justify moving forward in a responsible way with these dopaminergic neurons, which should be tested, using optimal patient selection, cell preparation and transplantation procedures, in controlled clinical studies.     

Using a dose-finding benchmark to quantify the loss incurred by dichotomization in Phase II dose-ranging studies

While there is recognition that more informative clinical endpoints can support better decision-making in clinical trials, it remains a common practice to categorize endpoints originally measured on a continuous scale. The primary motivation for this categorization (and most commonly dichotomization) is the simplicity of the analysis. There is, however, a long argument that this simplicity can come at a high cost. Specifically, larger sample sizes are needed to achieve the same level of accuracy when using a dichotomized outcome instead of the original continuous endpoint. The degree of “loss of information” has been studied in the contexts of parallel-group designs and two-stage Phase II trials. Limited attention, however, has been given to the quantification of the associated losses in dose-ranging trials. In this work, we propose an approach to estimate the associated losses in Phase II dose-ranging trials that is free of the actual dose-ranging design used and depends on the clinical setting only. The approach uses the notion of a nonparametric optimal benchmark for dose-finding trials, an evaluation tool that facilitates the assessment of a dose-finding design by providing an upper bound on its performance under a given scenario in terms of the probability of the target dose selection. After demonstrating how the benchmark can be applied to Phase II dose-ranging trials, we use it to quantify the dichotomization losses. Using parameters from real clinical trials in various therapeutic areas, it is found that the ratio of sample sizes needed to obtain the same precision using continuous and binary (dichotomized) endpoints varies between 70% and 75% under the majority of scenarios but can drop to 50% in some cases.     

Dendritic cell immunotherapy: mapping the way

Dendritic cells (DCs) are the professional antigen-presenting cells of the immune system, with the potential to either stimulate or inhibit immune responses. Exploiting the immune-regulatory capacities of dendritic cells holds great promise for the treatment of cancer, autoimmune diseases and the prevention of transplant rejection. Although early clinical trials indicate that DC vaccines can induce immune responses in some cancer patients, careful study design and use of standardized clinical and immunological criteria are needed.     

Stem cells for the treatment of musculoskeletal pain

Musculoskeletal-related pain is one of the most disabling health conditions affecting more than one third of the adult population worldwide. Pain from various mechanisms and origins is currently underdiagnosed and undertreated. The complexity of molecular mechanisms correlating pain and the progression of musculoskeletal diseases is not yet fully understood. Molecular biomarkers for objective evaluation and treatment follow-up are needed as a step towards targeted treatment of pain as a symptom or as a disease. Stem cell therapy is already under investigation for the treatment of different types of musculoskeletal-related pain. Mesenchymal stem cell-based therapies are already being tested in various clinical trials that use musculoskeletal system-related pain as the primary or secondary endpoint. Genetically engineered stem cells, as well as induced pluripotent stem cells, offer promising novel perspectives for pain treatment. It is possible that a more focused approach and reassessment of therapeutic goals will contribute to the overall efficacy, as well as to the clinical acceptance of regenerative medicine therapies. This article briefly describes the principal types of musculoskeletal-related pain and reviews the stem cell-based therapies that have been specifically designed for its treatment.     

Clinical applications of dendritic cell vaccination in the treatment of cancer

Dendritic cell (DC) immunotherapy has shown significant promise in animal studies as a potential treatment for cancer. Its application in the clinic depends on the results of human trials. Here, we review the published clinical trials of cancer immunotherapy using exogenously antigen-exposed DCs. We begin with a short review of general properties and considerations in the design of such vaccines. We then review trials by disease type. Despite great efforts on the part of individual investigative groups, most trials to date have not yielded data from which firm conclusions can be drawn. The reasons for this include nonstandard DC preparation and vaccination protocols, use of different antigen preparations, variable means of immune assessment, and nonrigorous criteria for defining clinical response. While extensive animal studies have been conducted using DCs, optimal parameters in humans remain to be established. Unanswered questions include optimal cell dose, use of mature versus immature DCs for vaccination, optimal antigen preparation, optimal route, and optimal means of assessing immune response. It is critical that these questions be answered, as DC therapy is labor- and resource-intensive. Cooperation is needed on the part of the many investigators in the field to address these issues. If such cooperation is not forthcoming, the critical studies that will be required to make DC therapy a clinically and commercially viable enterprise will not take place, and this therapy, so promising in preclinical studies, will not be able to compete with the many other new approaches to cancer therapy presently in development. Trials published in print through June 2003 are included. We exclude single case reports, except where relevant, and trials with so many variables as to prevent interpretation about DC therapy effects.     

Gene therapy clinical trials worldwide to 2017: An update

To date, almost 2600 gene therapy clinical trials have been completed, are ongoing or have been approved worldwide. Our database brings together global information on gene therapy clinical activity from trial databases, official agency sources, published literature, conference presentations and posters kindly provided to us by individual investigators or trial sponsors. This review presents our analysis of clinical trials that, to the best of our knowledge, have been or are being performed worldwide. As of our November 2017 update, we have entries on 2597 trials undertaken in 38 countries. We have analysed the geographical distribution of trials, the disease indications (or other reasons) for trials, the proportions to which different vector types are used, and the genes that have been transferred. Details of the analyses presented, and our searchable database are available via The Journal of Gene Medicine Gene Therapy Clinical Trials Worldwide website at: http://www.wiley.co.uk/genmed/clinical . We also provide an overview of the progress being made in gene therapy clinical trials around the world, and discuss key trends since the previous review, namely the use of chimeric antigen receptor T cells for the treatment of cancer and advancements in genome editing technologies, which have the potential to transform the field moving forward.     

Preparation of peptide-loaded dendritic cells for cancer immunotherapy

Dendritic cell-based vaccines are being evaluated in clinical trials to determine their ability to activate clinically relevant tumor antigen-specific immune responses. Although some groups isolate dendritic cells from peripheral blood, most have found it more efficient to generate large numbers from peripheral blood progenitors, particularly plastic adherent or CD14+ monocytes, in media supplemented with GM-CSF and IL-4. These DC may then be matured, if desired, and loaded with antigen, such as tumor-associated peptides, prior to administration. We describe the scheme that we are currently using to generate peptide-loaded dendritic cells for our clinical trials of cancer immunotherapy.     

CpG motif identification for veterinary and laboratory species demonstrates that sequence recognition is highly conserved.

Oligodinucleotides containing CpG motifs stimulate vertebrate immune cells in vitro, have proven efficacy in murine disease models and are currently being tested in human clinical trials as therapies for cancer, allergy, and infectious disease. As there are no known immunostimulatory motifs for veterinary species, the potential of CpG DNA as a veterinary pharmaceutical has not been investigated. Here, optimal CpG motifs for seven veterinary and three laboratory species are described. The preferential recognition of a GTCGTT motif was strongly conserved across two vertebrate phyla, although a GACGTT motif was optimal for inbred strains of mice and rabbits. In a subsequent adjuvanticity trial, the in vitro screening methodology was validated in sheep, representing the first demonstration of CpG DNA efficacy in a veterinary species. These results should provide candidate immunostimulant and therapeutic drugs for veterinary use and enable the testing of CpG DNA in large animal models of human disease.     

冠状动脉微循环障碍及微血管型心绞痛的治疗现状

10%到30%的胸痛患者并无明显的冠状动脉病变(coronary artery disease,CAD),但50%到65%患者的细小冠状动脉血管舒张功能受损,发生冠状动脉微循环障碍(coronary microvascular dysfunction,CMD),即微血管型心绞痛(microvascular angina,MA),目前尚无针对MA的最佳治疗方案。本文综述了既往7篇微血管型心绞痛的治疗方案的临床研究结果,提示西地那非,喹那普利,雌激素和经皮脊髓电刺激对本病有一定的治疗效果,但并未发现L-精氨酸,多沙唑嗪,普伐他丁和地尔硫卓对本病有明确疗效。然而,这7篇研究对冠状动脉微循环障碍的定义不同、采用的治疗方案和效果评价指标存在较大差异,纳入的样本量较小,这些都严重影响了临床研究结果的可靠性。在将来的相关临床试验中需统一冠脉微循环障碍的定义,评估无阻塞性冠脉病变所致胸痛的冠脉微循环障碍以及相应的治疗效果。     

Challenges to licensure of enterovirus 71 vaccines

Human enteroviruses usually cause self-limited infections except polioviruses and enterovirus 71 (EV71), which frequently involve neurological complications. EV71 vaccines are being evaluated in humans. However, several challenges to licensure of EV71 vaccines need to be addressed. Firstly, EV71 and coxsackievirus A (CA) are frequently found to co-circulate and cause hand-foot-mouth disease (HFMD). A polyvalent vaccine that can provide protection against EV71 and prevalent CA are desirable. Secondly, infants are the target population of HFMD vaccines and it would need multi-national efficacy trials to prove clinical protection and speed up the licensure and usage of HFMD vaccines in children. An international network for enterovirus surveillance and clinical trials is urgently needed. Thirdly, EV71 is found to evolve quickly in the past 15 years. Prospective cohort studies are warranted to clarify clinical and epidemiological significances of the antigenic and genetic variations between different EV71 genogroups, which is critical for vaccine design.     

Current approaches in dendritic cell generation and future implications for cancer immunotherapy

The discovery of tumor-associated antigens, which are either selectively or preferentially expressed by tumors, together with an improved insight in dendritic cell biology illustrating their key function in the immune system, have provided a rationale to initiate dendritic cell-based cancer immunotherapy trials. Nevertheless, dendritic cell vaccination is in an early stage, as methods for preparing tumor antigen presenting dendritic cells and improving their immunostimulatory function are continuously being optimized. In addition, recent improvements in immunomonitoring have emphasized the need for careful design of this part of the trials. Still, valuable proofs-of-principle have been obtained, which favor the use of dendritic cells in subsequent, more standardized clinical trials. Here, we review the recent developments in clinical DC generation, antigen loading methods and immunomonitoring approaches for DC-based trials.     

Recombinant Fv immunotoxins and Fv fragments as novel agents for cancer therapy and diagnosis

Recombinant immunotoxins are new agents being developed for cancer therapy. They are composed of Fv fragments of antibodies that bind to cancer cells fused to a truncated form of a very potent bacterial toxin. The antibody moiety directs the toxin to cancer cells, which are killed, while normal cells are not recognized and thus survive. The excellent preclinical results in vitro and in vivo have led to the initiation of several clinical trials.     

Adjuvant endocrine therapy in postmenopausal hormone-sensitive breast cancer: to start, to switch or to extend?

From the big randomized clinical trials there are evidences that adjuvant endocrine therapy for hormone-sensitive early breast cancer in postmenopausal women should include an aromatase inhibitor (AI). Anastrozole or letrozole should be used upfront for 5 years (ATAC and BIG 1-98), the sequential approach of tamoxifen for 2-3 years, followed by anastrozole or exemestane for 2-3 years is a reasonable alternative (ABCSG8, ARNO 95, IES, ITA), and mostly in patients with node-positive disease completing 5 years of tamoxifen should be offered letrozole up to 4-5 years (MA-17). In each of these trials incorporation of an AI resulted in significant improvement in study endpoints. Further results will be needed to establish the optimal beneficial effect, use, duration and safety of adjuvant AI therapies.     

Cartilage repair: past and future – lessons for regenerative medicine

Since the first cell therapeutic study to repair articular cartilage defects in the knee in 1994, several clinical studies have been reported. An overview of the results of clinical studies did not conclusively show improvement over conventional methods, mainly because few studies reach level I of evidence for effects on middle or long term. However, these explorative trials have provided valuable information about study design, mechanisms of repair and clinical outcome and have revealed that much is still unknown and further improvements are required. Furthermore, cellular and molecular studies using new technologies such as cell tracking, gene arrays and proteomics have provided more insight in the cell biology and mechanisms of joint surface regeneration. Besides articular cartilage, cartilage of other anatomical locations as well as progenitor cells are now considered as alternative cell sources. Growth Factor research has revealed some information on optimal conditions to support cartilage repair. Thus, there is hope for improvement. In order to obtain more robust and reproducible results, more detailed information is needed on many aspects including the fate of the cells, choice of cell type and culture parameters. As for the clinical aspects, it becomes clear that careful selection of patient groups is an important input parameter that should be optimized for each application. In addition, the study outcome parameters should be improved. Although reduced pain and improved function are, from the patient's perspective, the most important outcomes, there is a need for more structure/tissue-related outcome measures. Ideally, criteria and/or markers to identify patients at risk and responders to treatment are the ultimate goal for these more sophisticated regenerative approaches in joint surface repair in particular, and regenerative medicine in general.     

Cancer therapy by biological response modifiers

Biological response modifiers (BRMs) are agents or approaches which can modify the biological response of the host to tumors and thereby are expected to augment the resistance to development or progression of cancer. Recent advances in the technology of genetic engineering and monoclonal antibodies have led to rapid progress in this field. There is an increasing number of genetically engineered cytokines, which appear promising for cancer treatment and are becoming available for clinical trials. These include the interferons, leukin-2, tumor necrosis factor, and colony-stimulating factors. For development of optimal therapeutic protocols with these and a variety of immunomodulatory agents, it appears necessary to develop a detailed understanding of the possible mechanisms of their antitumor effects, and to determine the optimal dose and schedule for altering the antitumor effector mechanisms. BRMs would also be expected to be quite useful in the prevention of cancer. Indeed, in a variety of animal tumor model systems, potent protective effects can be demonstrated. However, the strategies for clinical application of such information have yet to be adequately worked out. One more immediate application of this approach is for the prevention of metastatic spread of tumor cells. BRMs, which stimulate natural killer cell activity, have been shown to strongly protect against dissemination of tumors, and clinical strategies for this important aspect of cancer treatment are being developed.     

Human adipose stem cells: current clinical applications

Adipose-derived stem cells are multipotent cells that can easily be extracted from adipose tissue, are capable of expansion in vitro, and have the capacity to differentiate into multiple cell lineages, which have the potential for use in regenerative medicine. However, several issues need to be studied to determine safe human use. For example, there are questions related to isolation and purification of adipose-derived stem cells, their effect on tumor growth, and the enforcement of U.S. Food and Drug Administration regulations. Numerous studies have been published, with the interest in the potential for regenerative medicine continually growing. Several clinical trials using human adipose stem cell therapy are currently being performed around the world, and there has been a rapid evolution and expansion of their number. The purpose of this article was to review the current published basic science evidence and ongoing clinical trials involving the use of adipose-derived stem cells in plastic surgery and in regenerative medicine in general. The results of the studies and clinical trials using adipose-derived stem cells reported in this review seem to be promising not only in plastic surgery but also in a wide variety of other specialties. Nevertheless, those reported showed disparity in the way adipose-derived stem cells were used. Further basic science experimental studies with standardized protocols and larger randomized trials need to be performed to ensure safety and efficacy of adipose-derived stem cells use in accordance with U.S. Food and Drug Administration guidelines.