Teresa L. K. Low

Current Status of Thymosin and Other Hormones of the Thymus Gland

Publisher Summary This chapter discusses the current status of the chemistry, biology, and clinical applications of the well-defined thymic hormones. The first biologically active polypeptide to be isolated from among the highly acidic components of bovine thymosin fraction 5 has been termed thymosin α1. This peptide is highly active in amplifying T-cell immunity and is active in modulating the expression of terminal deoxynucleotidyl transferase. The ultimate application of the thymosins and other thymic hormones and factors in cancer treatment should be in providing a means of safely augmenting specific T lymphocyte functions in patients with diminished thymic-dependent immunity. In anergic cancer patients, thymic hormones may be of importance as an adjunct to conventional treatments by increasing T-cell function in response not only to tumor cells but also to pathogens, thus reducing the high incidence of infection that often accompanies cancer treatment.

Current status of thymosin research: evidence for the existence of a family of thymic factors that control T-cell maturation*..

Thymosin fraction 5 contains several distinct hormonal-like factors which are effective in partially or fully inducing and maintaining immune function. Several of the peptide components of fraction 5 have been purified, sequenced and studied in assay systems designed to measure T-cell differentiation and function. These studied indicate that a number of the purified peptides act on different subpopulations of T-cells (see Figure 1). Thymosin beta 3 and beta 4 peptides act on terminal deoxynucleotidyl transferase (TdT) negative precursor T-cells to induce TdT positive cells. Thymosin alpha 1 induces the formation of functional helper cells and conversion of Lyt- cells to Lyt 1+, 2+, 3+ cells. Thymosin alpha 7 induces the formation of functional suppressor T-cells and also converts Lyt- cells to Lyt 1+, 2+, 3+ cells. These studies have provided further evidence that the thymus secretes a family of distinct peptides which act at various sites of the maturation sequence of T-cells to induce and maintain immune function. Phase I and Phase II clinical studied with thymosin in the treatment of primary immunodeficiency diseases, autoimmune diseases, and cancer point to a major role of the endocrine thymus in the maintenance of immune balance and in the treatment of diseases characterized by thymic malfunction. It is becoming increasingly clear that immunological maturation is a process involving a complex number of steps and that a single factor initiating a single cellular event might not be reflected in any meaningful immune reconstitution unless it is the only peptide lacking. Given the complexity of the maturation sequence of T-cells and the increasing numbers of T-cell subpopulations that are being identified, it would be surprising if a single thymic factor could control all of the steps and populations involved. Rather, it would appear that the control of T-cell maturation and function involves a complex number of thymic-specific factors and other molecules that rigidly control the intermediary steps in the differentiation process.

Immunochemical studies on thymosin: Radioimmunoassay of thymosin β4

Thymosin beta 4, a peptide with hormonal-like properties first isolated from the thymus gland, can be measured in serum using a newly described radioimmunoassay. The radioimmunoassay utilizes an antibody raised in rabbits against synthetic thymosin beta 4 conjugated by glutaraldehyde to keyhole limpet hemocyanin. A 125I-tyrosine-C13 analogue of the biologically active C-terminal fragment is used as the radioactive tracer. The radioimmunoassay is sensitive in the nanogram range and no cross-reactivity with common serum proteins is demonstrable. High performance liquid chromatography of serum samples indicates that two thymosin beta 4 cross-reactive species are present in human serum. Levels in serum range from 450 to 1100 ng/ml and decline with age.

[13] Thymic hormones: An overview

Publisher Summary This chapter provides an overview of thymic hormones. The thymus gland is a bilobed lymphoid organ located in the chest immediately beneath the breastbone. The involution of the thymus is among the first noticed indications of aging. A vital part of the process by which the thymus works occurs via a hormonal mechanism. A partially purified thymic extract termed thymosin fraction is been demonstrated to be effective in reconstituting immune functions in thymic-deprived or immuno deprived animals as well as in humans with primary immunodeficiency diseases and in immunosuppressed cancer patients. In addition to thymosin fraction 5, a number of preparations with thymic hormone-like activity are isolated from thymus tissue, blood and thymic epithelial supernatants. Several of the newly described thymosin peptides have been found to bear high degree of homology with either thymosin a α 1 and β 4 . The most important contribution of thymic hormone research resides in its application to the clinical management of thymus-dependent diseases.

Isolation of thymosin α1 from thymosin fraction 5 of different species by high-performance liquid chromatography

High-performance liquid chromatography (muBondapak C18 column with 0.05% trifluoroacetic acid in acetonitrile as solvent system) was used to isolate thymosin alpha 1 (alpha 1) from thymosin fraction 5 (f5) of various species (calf, pig, sheep and mouse). Each of the f5 preparations gave a protein peak similar in retention time to bovine thymosin alpha 1. This peak coincided with the immunoreactive peak determined by a radioimmunoassay for alpha 1. Chromatographic analysis of fresh thymus tissue extracts using a high-performance liquid chromatographic similar system did not reveal a detectable protein peak or immunoreactive peak at the alpha 1 position. Our results suggest that alpha 1 may be synthesized in a precursor form in animal tissues.

Thymosin and Other Thymic Hormones and Their Synthetic Analogues

The central role of the thymus gland in the development, growth, and function of lymphoid tissue and in the maintenance of immune balance has now been fully recognized [30]. Investigations on in vivo and in vitro effects of thymic extracts [43] and the effect of thymus grafts in cell-impermeable millipore diffusion chambers [5, 40, 41, 46, 51, 56] have helped to establish the endocrine function of the thymus. As shown in Fig. 1, ongoing studies by us and by others have suggested that the lack of adequate production and utilization of thymic factors causes an immune imbalance and may contribute to the etiology of many diseases.

[16] Thymosin β4

Publisher Summary This chapter provides an overview of β4. Thymosin β4 is a polypeptide isolated from thymosin fraction 5 (F5) and fraction 5A (5A). This peptide exhibits important activities in the regulation and differentiation of thymus-dependent lymphocytes. It induces expression of TdT activity in vivo 2 and in vitro. 3 It also inhibits the migration of macrophages, and exerts biological effects on the hypothalamus and pituitary. The chapter further provides purification procedure, thymosin β4 is present in both thymosin F5 and F5A. However, its relative concentration is higher in F5A than in F5. This peptide can be purified either by conventional column technique or by HPLC. Conventional column techniques are used for large-scale purification, while the HPLC procedure is used when the starting material available is limited. In addition, the chapter also explains purification by HPLC from thymosin F5A or tissue extracts.

Thymosin: Basic Properties and Clinical Application in the Treatment of Immunodeficiency Diseases and Cancer

Thymosin fraction 5 contains a family of polypeptides with varying biological activities. Current efforts in the thymosin research program are involved in further chemical characterization of thymosin peptides and evaluation of clinical immunotherapeutic protocols. Recent clinical studies with thymosin fraction 5 have shown therapeutic potentials for treatment of patients with primary immunodeficiency diseases and cancer.

Isolation and structural studies of porcine, ovine and murine thymosin β4 by high-performance liquid chromatography

Abstract Rapid high-performance liquid chromatographic (HPLC) procedures have been used to isolate and characterize thymosin β 4 from different species. Crude extracts termed thymosin fraction 5A were prepared from porcine and ovine thymus glands as well as murine spleen. Each fraction 5A preparation was then fractionated by HPLC on a μBondapak C 18 reversed-phase column. Porcine and ovine thymus fraction 5A, and murine spleen 5A, each yields a predominant peak at a retention time similar to that of bovine thymosin β 4 . Amino acid analysis as well as HPLC tryptic peptide mapping of these peaks indicate that they have homologous sequences to bovine thymosin β 4 . Chromatographic analysis of fresh murine thymus and spleen tissues also revealed protein peaks at the position of bovine β 4 , suggesting that thymosin β 4 is the native protein present in the animal tissues.

POTENTIAL ROLE OF THYMOSIN IN THE TREATMENT OF AUTOIMMUNE DISEASES

Fifteen years ago, at the first Myasthenia Gravis Symposium sponsored by the New York Academy of Sciences, we presented our first report on the isolation of a soluble lymphocytopoietic factor prepared from calf thymus h0mogenates.l Shortly thereafter, we were able to partially purify the active principle from calf thymus homogenates, which we named thymosin.? These early studies supported the theory that the thymus gland has important endocrine, as well as lymphopoietic, functions. Ongoing studies in immunobiology continue to support the concept of an important regulatory role of the endocrine thymus in the development and maintenance of the immune system. Diminished secretion of thymosin and perhaps other thymic factors appears to underlie many of the immunological deficiencies observed in animals after experimental thymectomy or in humans with thymic dysplasia. A t the last Myasthenia Gravis Symposium in 1976, we summarized the aiiimal and clinical data supporting our hypothesis that thymosin imbalances are involved in the etiology of autoimmune d i ~ e a s e . ~ We proposed that a thymic endocrine malfunction could lead to autoimmune disease and suggested that new strategies for using thymosin in the therapeutic manipulation of the immune system should be developed. In this paper, we will summarize the current status of the chemical and biological characterization of the thymosin peptides and the initial clinical applications of thymosin in the treatment of autoimmune diseases.