The Biotherapy Section of LMB has focused on four main research areas: toxin-mammalian cell interactions, targeted toxins in a cancer setting,vaccine development and the role of small non-coding RNAs in the gene regulation of Pseudomonas aeruginosa. Characterizing the interactions of Pseudomonas exotoxin with mammalian cells is important because we need to understand the pathway of action of this important virulence factor and component of recombinant immunotoxins. Within this research area, there have been three notable findings: 1) a new receptor termed LRP1B, for PE was discovered, 2) it was determined that cells secreting enzymatically inactive PE were resistant to WT toxin added exogenously and 3) the conversion of dispersed epithelial cells to polarized cells renders cells toxin resistant. The underlying mechanisms for the latter two findings are being investigated. Recombinant immunotoxins are immunogenic in a large percentage of individuals with solid tumors. The Biotherapy Section is investigating the use of human granzyme B as a potential alternative to Pseudomonas exotoxin. Granzyme B is released from cytotoxic T-cells and causes death in the cytosol of target cells by proteolytic activation of the apoptosis pathway. We have cloned, expressed (in E coli) and refolded a recombinant form of CTL granzyme that is enzymatically active against small molecular weight substrates and pro-caspase 3. We will next determine its activity against intact cells. A major problem experienced by many cancer patients is a chronic pain syndrome. A method to permanently relieve this pain by destroying the spinal chord neurons responsible for pain transmission has been investigated in rodents. The instillation of substance P-PE35 has proved active against neurokinin 1 receptor-bearing neurons in the periphery of the dorsal horn of rats and will be developed as a potential agent for relieving the suffering of individuals with intractable pain. Finally, the Biotherapy section has been developing a vaccine that could be used by individuals affected with Cystic Fibrosis to prevent infections with Pseudomonas aeruginosa. The first candidate vaccine is a dual function protein (PE?553-pil) that was constructed to produce antibodies that would prevent bacterial adherence (via an anti-pilin response) and neutralize the cytotoxic activity of exotoxin A. Advanced preclinical development of this vaccine is underway at Trinity Biosystems. Small non-coding RNAs have been shown to play important roles in the regulation of genes in E coli. Functional similar RNAs appear to be active in Pseudomonas. However, they cannot be identified via sequence homomolgy searches. Therefore, we have devised a combination of bioinformatic and wet-lab procedures to identify and characterize these novel regulators.
| Kreitman, Robert J; Stetler-Stevenson, Maryalice; Margulies, Inger et al. (2009) Phase II trial of recombinant immunotoxin RFB4(dsFv)-PE38 (BL22) in patients with hairy cell leukemia. J Clin Oncol 27:2983-90 |
| Pastrana, Diana V; FitzGerald, David J (2006) A nonradioactive, cell-free method for measuring protein synthesis inhibition by Pseudomonas exotoxin. Anal Biochem 353:266-71 |
| Pastan, Ira; Hassan, Raffit; Fitzgerald, David J et al. (2006) Immunotoxin therapy of cancer. Nat Rev Cancer 6:559-65 |
| Hsieh, Jennifer C; Tham, Doris M; Feng, Weijun et al. (2005) Intranasal immunization strategy to impede pilin-mediated binding of Pseudomonas aeruginosa to airway epithelial cells. Infect Immun 73:7705-17 |
| Kreitman, Robert J; Squires, David R; Stetler-Stevenson, Maryalice et al. (2005) Phase I trial of recombinant immunotoxin RFB4(dsFv)-PE38 (BL22) in patients with B-cell malignancies. J Clin Oncol 23:6719-29 |
| Pastrana, Diana V; Hanson, Alison J; Knisely, Jane et al. (2005) LRP 1 B functions as a receptor for Pseudomonas exotoxin. Biochim Biophys Acta 1741:234-9 |
| Wilderman, Paula J; Sowa, Nathaniel A; FitzGerald, David J et al. (2004) Identification of tandem duplicate regulatory small RNAs in Pseudomonas aeruginosa involved in iron homeostasis. Proc Natl Acad Sci U S A 101:9792-7 |
