Hiromu Takematsu

Abstract Coley’s Toxin comprised a mixture of cell-free, heat-treated culture media from Streptococcus pyogenes (originally Streptococus erysipelatos) and Serratia marcescens (originally Bacillus prodigiosus). A 250 kDa tumor hemorrhage-inducing polysaccharide “PS1” is reported here secreted into culture medium by S. marcescens. Four h after PS1 is injected at 32 μg/kg (10pM) into the tail vein of Balb/C mice bearing C26 subcutaneous colon-derived tumors, tumor-specific capillary hemorrhage is exhibited in 90% of tumors. As a positive control, CM101, a similar tumor hemorrhagic polysaccharide from Streptococcus agalactica caused tumor hemorrhage in 75% of tumors in the Balb/C-C26 model at 7.5 μg/kg(2.5pM). CM101 has previously been safety tested in a Phase I clinical trial. These two polysaccharides have merit to be identified as the active principal ingredients (API’s) of Coley’sToxin. Additional approaches to cancer therapy are a global need. No matter the level of wealth of victims, some cancers are still incurable. Recall in recent years the tragic early cancer deaths of Steve Jobs and Paul Allen among other luminaries. Streptococcal and Serratia bacterial extracts have unique tumor specific capillary destructive activity, with observations originating with sarcomas cured by nosocomial  erysipelas  infections in the 1860’s. The active pharmaceutical ingredients (API’s) in these extracts and Coley’s Toxins are proposed to be polysaccharides.

CD22, one of the sialic acid-binding immunoglobulin-like lectins (Siglecs), regulates B lymphocyte signaling via its interaction with glycan ligands bearing the sequence Neu5Ac/Gcα(2→6)Gal. We have developed the synthetic sialoside GSC-718 as a ligand mimic for CD22 and identified it as a potent CD22 inhibitor. Although the synthesis of CD22-binding sialosides including GSC-718 has been reported by our group, the synthetic route was unfortunately not suitable for large-scale synthesis. In this study, we developed an improved scalable synthetic procedure for sialosides which utilized 1,5-lactam formation as a key step. The improved procedure yielded sialosides incorporating a series of aglycones at the C2 position. Several derivatives with substituted benzyl residues as aglycones were found to bind to mouse CD22 with affinity comparable to that of GSC-718. The new procedure developed in this study affords sialosides in sufficient quantities for cell-based assays, and will facilitate the search for promising CD22 inhibitors that have therapeutic potential.

Spermatid production is a complex regulatory process in which coordination between hormonal control and apoptosis plays a pivotal role in maintaining a balanced number of sperm cells. Apoptosis in spermatogenesis is controlled by pro-apoptotic and anti-apoptotic molecules. Hormones involved in the apoptotic process during spermatogenesis include gonadotrophins, sex hormones, and glucocorticoid (GC). GC acts broadly as an apoptosis inducer by binding to its receptor (glucocorticoid receptor: GR) during organ development processes, such as spermatogenesis. However, the downstream pathway induced in GC-GR signaling and the apoptotic process during spermatogenesis remains poorly understood. We reported previously that GC induces full-length glucocorticoid-induced transcript 1 (GLCCI1-long), which functions as an anti-apoptotic mediator in thymic T cell development. Here, we demonstrate that mature murine testis expresses a novel isoform of GLCCI1 protein (GLCCI1-short) in addition to GLCCI1-long. We demonstrate that GLCCI1-long is expressed in spermatocytes along with GR. In contrast, GLCCI1-short is primarily expressed in spermatids where GR is absent; instead, the estrogen receptor is expressed. GLCCI1-short also binds to LC8, which is a known mediator of the anti-apoptotic effect of GLCCI1-long. A luciferase reporter assay revealed that β-estradiol treatment synergistically increased Glcci1-short promotor-driven luciferase activity in Erα-overexpressing cells. Together with the evidence that the conversion of testosterone to estrogen is preceded by aromatase expression in spermatids, we hypothesize that estrogen induces GLCCI1-short, which, in turn, may function as a novel anti-apoptotic mediator in mature murine testis.