Russell, A. J., & Bertram, T. A. (2007). Moving into the clinic. In R. Lanza, R. Langer & J. P. Vacanti (Eds.), Principles of tissue engineering (3rd ed., pp. 15-32). Burlington, MA: Elsevier Academic Press.
In the early 1930s Charles Lindbergh, who was better known for his aerial activities, went to Rockefeller University and began to study the culture of organs. After the publication of his book about the culturing of organs ex vivo in order to repair or replace damaged or diseased organs, the field lay dormant for many years. Indeed, delivering respite to failing organs with devices or total replacement (transplant) became far more fashionable. Transplantation medicine has been a dramatic success. But in the late 1980s scientists, engineers, and clinicians began to conceptualize how de novo tissue generation might be used to address the tragic shortage of donated organs. The approach they proposed was as simple as it was dramatic. Biodegradable materials would be seeded with cells and cultured outside the body for a period of time before exchanging this artifi cial bioreactor for a natural bioreactor by implanting the seeded material into a patient. These early pioneers believed that the cells would degrade the material, and after implantation the cell-material construct would become a vascularized native tissue. Tissue engineering, as this approach came to be known, can be accomplished once we understand which materials and cells to use, how to culture these together ex vivo, and how to integrate the resulting construct into the body.
Bertram, T. A., & Jayo, M. J. (2008). Tissue engineered products: Preclinical development of neo-organs. In J. Cavagnero (Ed.), Preclinical safety evaluation of biopharmaceuticals: A science-based approach to facilitating clinical trials (pp. 799-826). New York: John Wiley & Sons.
The term tissue engineering was coined at a National Science Foundation workshop in 1987 to mean "the application of principles and methods of engineering and life sciences toward fundamental understanding of structure-function relationships in normal and pathological mammalian tissues and tissue function" (Viola et al., 2003). Tissue engineering draws on specialized expertise from two traditional disciplines: engineering and the life sciences. The combination of these technologies forms a foundation upon which the commercial development of neo-organs is possible.
Preclinical development of neo-organs faces complex scientific questions and regulatory hurdles. The term neo-organ product will be used to indicate any product composed of synthetic or natural biodegradable materials, with or without living cells and/or cellular products, implanted in the body to incorporate, replace, and/or regenerate a damaged tissue or organ. Today ex vivo development of partial and complete neo-organs by the emerging regenerative medical industry fulfills a significant unmet medical need for patients who have partial or complete organ or tissue loss. In this chapter we will consider development challenges and available solution strategies for bringing these transformational technologies to patients in need.
Jayo, M. J., Watson, D. D., Wagner, B. J., & Bertram, T. A. (2008). Tissue engineering and regenerative medicine: role of toxicologic pathologists for an emerging medical technology. Toxicol Pathol, 36(1), 92-96.
Tissue Engineering Regenerative Medical (TERM) products are a new technology currently in human clinical testing for a variety of unmet medical needs involving tissue and organ dysfunction and failure. Safety evaluation of TERM products overlaps 3 established product paradigms: pharmaceuticals (biologically active substances), transplantation (cells or tissue), and devices (biomaterials). As TERM products recapitulate organ or tissue structure and function with unique biological activity and characteristics, they require new preclinical paradigms to bring TERM products through to clinical trials. Establishing TERM-product safety programs requires broad-based knowledge of tissue and organ homeostasis, regenerative biology, and translational medicine to design new preclinical paradigms. Therefore, toxicologic pathologists have a compelling scientific role in evaluating TERM products, characterizing tissue responses, and helping distinguish optimal (regeneration) from deficient or incomplete outcomes indicative of substandard functionality (repair). As new-tissue engineering and regenerative medical technologies develop for tissue and organ regeneration, the toxicologic pathologist will be asked to develop novel testing, reevaluate established toxicologic diagnostic criteria, and reinterpret tissue responses that may extend beyond current standards.
Jayo, M. J., Jain, D., Ludlow, J. W., Payne, R., Wagner, B. J., Seltzer, E., McLorie, G. A., & Bertram, T. A. (2007, October 27). A regenerative neo-bladder construct in trigone-sparing cystectomized dogs: Long-term safety, continence, voiding, and urodynamics. Poster presented at the American Association of Pediatrics Section on Urology National Conference and Exhibition.
Purpose: Trigone-sparing cystectomy was used to study the structural and functional aspects of bladder regeneration in a canine model of an augmentation cystoplasty.
Methods: An autologous neo-bladder augmentation construct composed of a PLGA-based biodegradable mesh scaffold and autologous urothelial and smooth muscle cells (Construct) (n=32) was compared to re-implanted native bladder (Reimplant) (n=32), or PLGA-based biodegradable mesh scaffold alone (Scaffold) (n=8) at 1, 3, 6 and 9 months (mo) post-implantation.
Results:Within 14 days, all 72 dogs were continent. Within 1 mo, acute phase responses, hematological and urinalysis parameters returned to baseline. Treatment-related morbidity was only observed in Scaffod and Reimplant dogs. Only Construct dogs achieved functional recovery (i.e., urodynamics) and a regenerative tissue response. Construct dogs regained baseline bladder capacity by 4 mo and compliance by 6 mo and were sustained throughout the study. Urodynamic parameters in Reimplant animals were initially comparable to Construct dogs but were unstable and significantly lower than baseline by 9 mo (60-75% decrease from baseline). Histologically, decreased compliance in Reimplant and Scaffold groups at 9 mo correlated with limited healing and incomplete bladder wall regeneration.
Conclusions: Construct implants were safe and able to restore urodynamic, continence, and voiding functions by 6 mo and retained these functions to study termination. Bladder wall regeneration was obtained only in the Construct implanted animals. Scaffolds lacking cells (Scaffold) elicit repair of bladder wall with incompletely developed components reduced organ capacity and restricted compliance.
Bertram, T.A. (2007). Tengion Autologous Neo-bladder Construct. In a presentation to the FDA/NIST Sponsored Workshop In Vitro Analyses of Cell/Scaffold Products. December 2007.
Bertram, T. A., Christ, G. J., Wagner, B. J., Jain, D., Aboushwareb, T., Ludlow, J. W., Payne, R., Jarapu, Y., Turner, C., & Jayo, M. J. (2008, April 6). Total urinary bladder regeneration with restoration of native structure and pharmacological response. Poster presented at the 2008 Experimental Biology Meeting, San Diego, CA.
Neo-bladder histology was investigated in 21 canines at 30-79 days (n=9) and 80-180 days (n=9) after radical cystectomy and implantation of a Neo-Bladder Replacement Construct (Construct). Pharmacological responses of neo-bladder tissue strips from 9 retrieved neo-bladders (n=1, 30-79 days; n=8, 80-180 days) were compared to age-matched native bladder tissue (n=17).
In both groups, regenerated bladder was histologically consistent with native bladder including mucosal and serosal linings, detrusor muscle, vascular, and nerve composition. Logistic analysis revealed similar EC50 and slope factor values for contraction of bladder tissue strips derived from both groups induced by carbachol (Car) and electrical field stimulation (EFS). A progressive increase in the mean Emax values occurred in response to both Car and EFS over time. While contractile responses to Car and EFS increased over time in the neo-bladder tissue, they were lower than native bladder responses in the 30-79 day group. Car-induced contractions, but not EFS-induced contractions, became equivalent to native tissue in 80-180 days post-implantation (p.i.).
The Neo-Bladder Replacement Construct is capable of regenerating urinary bladder in vivo with histology similar to native bladder in 30-79 days p.i. and pharmacological responses became similar to native bladder in 80-180 days p.i. with no evidence of abnormal cell growth, immune response, or adverse systemic effects.
Tengion is currently conducting a GLP study to support clinical trial studies in 2009.
Jayo, M. J., Jain, D., Wagner, B. J., & Bertram, T. A. (2008). Early Cellular and Stromal Responses in Regeneration Versus Repair of a Mammalian Bladder Using Autologous Cell and Biodegradable Scaffold Technologies. J Urol, Volume 180, Issue 1, Pages 392-397 (July 2008).
Purpose: Internal organ regeneration holds promise for changing medical technology and reducing organ shortages. Current medical treatment for internal organ failure is largely limited to organ transplantation. A synthetic biopolymer with autologous cells (Construct) has exhibited long-term clinical benefit in patients undergoing augmentation cystectomy; however, early cellular and stromal events during bladder regeneration have not been elucidated.
Materials and Methods: In situ cellular responses to two biopolymer implants; one with autologous cells (Construct) and one without cells (Scaffold) were compared in a canine model of augmentation cystoplasty. Healing events were correlated with urodynamic assessments.
Results: Construct implants regenerated baseline urodynamics as early as 4 months post-implantation. Urodynamics following Scaffold implantation failed to return to baseline by study termination at 9 months. Functional differences elicited by Construct and Scaffold implants correlated with structural differences in the neo- tissues. Construct stroma had greater vascularization with gently folded interwoven connective tissue elements. Scaffold stroma was dense, haphazardly-organized connective tissue. Urothelium regenerated in response to both Construct and Scaffold implantation; however, only Construct had normal stroma, well developed detrusor, and abundant aSMA-staining cells at early time points leading to a structurally and functionally complete, regenerated bladder wall at 9 months.
Conclusion: Early cellular and stromal events distinguish healing processes that lead to bladder wall regeneration or repair. Construct implants containing cells elicit early healing processes that culminate with regeneration of complete mucosal and muscular components whereas the response to Scaffold implantation is consistent with reparative healing; with mucosal growth but incomplete tissue layer development.
Bertram, T.A., Christ, G.J., Aboushwareb, T.A., Wagner, B.J., Jain, D., Ludlow, J. W., Payne, R., Jarajapu, Y.R., Turner, C., Jayo, M.J. (2008, May 18). Total regenerated urinary bladders are structurally and pharmacologically similar to native tissue. Presented at the American Urological Association’s 2008 Meeting, held in Orlando, FL, May 17-22, 2008.
Neo-bladder structure and function was investigated in 23 canines at 30- 79 days (n=9) and 80-188 days (n=14) after radical cystectomy and implantation of a Neo-Bladder Replacement Construct (Construct). Pharmacological responses of neo-bladder tissue strips from 11 retrieved neo-bladders (n=1, 30-79 days; n=10, 80-188 days) were compared to age- matched native bladder tissue (n=17).
Regenerated bladder was histologically consistent with native bladder including mucosal and serosal linings, detrusor muscle, vascular, and nerve composition. Logistic analysis revealed similar EC50 and slope factor values for contraction of bladder tissue strips derived from both groups induced by carbachol (Car) and electrical field stimulation (EFS). A progressive increase in the mean Emax values occurred in response to both Car and EFS over time. While contractile responses to Car and EFS increased over time in the neo-bladder tissue, they were lower than native bladder responses in the 30-79 day group. In contrast, strips from 80-188 day post-implantation neo-bladders reached the 95% confidence interval of native tissue in the linear range of the response.
The Neo-Bladder Replacement Construct is capable of regenerating urinary bladder in vivo with histology similar to native bladder in 30-79 days p.i. and pharmacological and electrical field responses became similar to native bladder in 80-188 days p.i. with no evidence of abnormal cell growth, immune response, or adverse systemic effects.
Tengion is currently conducting a GLP study to support clinical trial studies in 2009.
Jayo MJ, Jain D, Ludlow JW, Payne R, Wagner BJ, McLorie G, Bertram TA (2008, September). Long-term durability, tissue regeneration and neo-organ growth during skeletal maturation with a neo-bladder augmentation construct. Regen Med. 2008 Sep;3(5):671-82
AIMS: To comparatively evaluate bladder regeneration following 80% cystectomy and augmentation using a synthetic biopolymer with autologous urothelial and smooth muscle cells (autologous neo-bladder augmentation construct [construct]) or autotransplantation of native bladder (reimplanted native urinary bladder [reimplant]) in canines.
MATERIALS & METHODS: Voiding function, urodynamic assessment and neo-organ capacity-to-body-weight ratio (C:BW) were assessed longitudinally for a total of 24 months following trigone-sparing augmentation cystoplasty in juvenile canines.
RESULTS: Within 30 days postimplantation, hematology and urinalysis returned to baseline. Constructs and reimplants yielded neo-organs with statistically equivalent urodynamics and histology. Linear regression analysis of C:BW showed that constructs regained baseline slope and continued to adapt with animal growth.
CONCLUSIONS: Constructs and reimplants regained and maintained native bladder histology by 3 months, capacity at 3-6 months and compliance by 12-24 months. Furthermore, construct C:BW demonstrated the ability of regenerated bladder to respond to growth regulation.
Kelley, R., Bruce, A., Wallace, S., Choudhury, S., Tatsumi, P., Das, A., Riveras, E., & Presnell, S. (2008, September 17-19). Enhanced renal cell function in dynamic 3D culture system. Poster presented at the KIDSTEM International Conference, Liverpool, England.
Traditional primary culture of renal cells involves enzymatic dissociation of kidney tissue followed by propagation of the cells on tissue-culture treated plastic with or without extracellular matrix coatings. In the present study, we examined the effects of three-dimensional (3D) architecture and perfusion on metabolism, phenotype, and tubular function of primary kidney cells in a variety of culture configurations. 3D architecture promoted cell-cell interaction and organization, as determined by scanning electron microscopy, histology, and confocal immunochemistry. The addition of perfusion to the culture system resulted in enhanced metabolic activity and a significant and sustained upregulation of genes associated with tubular function. Importantly, the tubular function of renal cells was confirmed in culture systems via the demonstration of megalin/cubilin-mediated uptake of albumin. In summary, dynamic 3D culture systems provide a means to examine tubular cell phenotype and function in an environment that better recapitulates in vivo biology.