Mathematical Model Studies on the Optimal Scheduling of the Treatment of Systemic Malignant Disease by Radiation

The work reported in this thesis deals with mathematical model studies on the optimal scheduling of treatment of systemic malignant disease by radiation. To provide the necessary background to the original aspects of the work reviews of several fields are required. Chapter 1 is a general review of normal tissue radiobiology. Chapter 2 is a review of human tumour radiobiology. Chapter 3 focusses on one particular isoeffect model, the linear-quadratic or LQ model, which is employed throughout this thesis to describe the effects of radiation on normal tissues. These basic radiobiological principles are applied to the clinical modalities of total body irradiation (TBI) and biologically targeted radiotherapy (BTR). Chapter 4 reviews the principles of TBI. Chapter 5 is a review of published data on the in-vitro radiosensitivities of human leukaemia/lymphoma and neuroblastoma, two conditions which require a systemic approach to treatment. Chapter 8 is a review of the principles of BTR. The original work is contained in the appendix to chapter 3, which examines the correspondence between the LQ model and CRE models for continuous radiation exposures with constant and exponentially decaying dose-rates; chapter 6, Which examines the question of whether fractionated or low dose-rate TBI is the superior method of treatment; chapter 7, where the optimal scheduling of fractionated TBI is investigated; chapter 9, where the LQ isoeffect model and a dosimetric approach is used for the evaluation of alternative therapeutic strategies for the treatment of widespread micrornetastatic disease by BTR. Finally, in chapter 10 a simple model is used to investigate optimal scheduling of BTR, TBI and marrow rescue. CONCLUSIONS 1/ Comparison of the LQ model and the CRE model for continuous radiation exposures: for constant dose-rates it is found that, when late-effect parameter values are used in the LQ model, there is a correspondence between the models' predictions. There is no correspondence between models when acute-effect parameter values are used in the LQ model. In the case of exponentially decaying dose-rates the predictions of the CRE and LQ models appear more divergent, although again the use of late rather than acute-effect parameter values in the LQ model gives a closer match to the CRE. 2/ Fractionated TBI is predicted to be preferable to low dose-rate TBI treatment. Although theoretically the methods can be equivalent, low dose-rate treatments would have to be over impractically long treatment times. 3/ In the case of external beam TBI, fractionated low dose-rate treatments do not appear to offer a significant improvement over fractionated high dose-rate treatments. This is because in order to achieve a significant increase in dose or reduction in toxicity impractically long exposure times are required. It is expected that this finding will be true in general for external beam radiotherapy, not just in the case of TBI. 4/ Optimal fractionation schedules for the treatment of leukaemia/lymphoma and neuroblastoma by TBI are predicted to be accelerated and hyperfractionated. It is suggested that a two fraction per day schedule of 10 fractions of 1.3-1.5 Gy is a suitable candidate for clinical evaluation. 5/ It is concluded that knowledge of ratios for tumours and normal tissues is, by itself, insufficient information to enable prediction of optimal schedules. 6/ In the case of BTR, dose-rate effects are predicted to be important for late-responding tissues. Tolerance doses may be greater or less than those for fractionated radiotherapy depending on the effective radionuclide half-life. 7/ When injected activities of targeted radionuclide are restricted by haemopoietic tolerance, curative therapy is unlikely. 131-I appears to be a better radionuclide warhead for therapy of micrometastases than 90-Y. 8/ The use of bone marrow rescue in conjunction with BTR seems to offer curative potential, however reasons are presented why a combined strategy using BTR, TBI and marrow rescue is likely to be preferable. 9/ For optimal scheduling of BTR, TBI and marrow rescue, the main characteristics of BTR which determine curative potential are its specificity and sensitivity. Specificity is defined here as the ratio of initial dose-rate at the tumour cells to that in the dose-limiting tissue. Sensitivity is inversely related to the proportion of tumour cells which escape targeting. Where biological targeting is highly specific but some tumour cells escape, a phenomenon of "overkill" will largely determine the optimal schedules. It is predicted that these are likely to consist of combinations of BTR and external beam TBI with the TBI component being the greatest in terms of radiation dose to the whole body

Multiple novel prostate cancer susceptibility signals identified by fine-mapping of known risk loci among Europeans

Genome-wide association studies (GWAS) have identified numerous common prostate cancer (PrCa) susceptibility loci. We have fine-mapped 64 GWAS regions known at the conclusion of the iCOGS study using large-scale genotyping and imputation in 25 723 PrCa cases and 26 274 controls of European ancestry. We detected evidence for multiple independent signals at 16 regions, 12 of which contained additional newly identified significant associations. A single signal comprising a spectrum of correlated variation was observed at 39 regions; 35 of which are now described by a novel more significantly associated lead SNP, while the originally reported variant remained as the lead SNP only in 4 regions. We also confirmed two association signals in Europeans that had been previously reported only in East-Asian GWAS. Based on statistical evidence and linkage disequilibrium (LD) structure, we have curated and narrowed down the list of the most likely candidate causal variants for each region. Functional annotation using data from ENCODE filtered for PrCa cell lines and eQTL analysis demonstrated significant enrichment for overlap with bio-features within this set. By incorporating the novel risk variants identified here alongside the refined data for existing association signals, we estimate that these loci now explain ∼38.9% of the familial relative risk of PrCa, an 8.9% improvement over the previously reported GWAS tag SNPs. This suggests that a significant fraction of the heritability of PrCa may have been hidden during the discovery phase of GWAS, in particular due to the presence of multiple independent signals within the same regio

The impact of immediate breast reconstruction on the time to delivery of adjuvant therapy: the iBRA-2 study

Background: Immediate breast reconstruction (IBR) is routinely offered to improve quality-of-life for women requiring mastectomy, but there are concerns that more complex surgery may delay adjuvant oncological treatments and compromise long-term outcomes. High-quality evidence is lacking. The iBRA-2 study aimed to investigate the impact of IBR on time to adjuvant therapy. Methods: Consecutive women undergoing mastectomy ± IBR for breast cancer July–December, 2016 were included. Patient demographics, operative, oncological and complication data were collected. Time from last definitive cancer surgery to first adjuvant treatment for patients undergoing mastectomy ± IBR were compared and risk factors associated with delays explored. Results: A total of 2540 patients were recruited from 76 centres; 1008 (39.7%) underwent IBR (implant-only [n = 675, 26.6%]; pedicled flaps [n = 105,4.1%] and free-flaps [n = 228, 8.9%]). Complications requiring re-admission or re-operation were significantly more common in patients undergoing IBR than those receiving mastectomy. Adjuvant chemotherapy or radiotherapy was required by 1235 (48.6%) patients. No clinically significant differences were seen in time to adjuvant therapy between patient groups but major complications irrespective of surgery received were significantly associated with treatment delays. Conclusions: IBR does not result in clinically significant delays to adjuvant therapy, but post-operative complications are associated with treatment delays. Strategies to minimise complications, including careful patient selection, are required to improve outcomes for patients

Destructin-1 is a collagen-degrading endopeptidase secreted by Pseudogymnoascus destructans, the causative agent of white-nose syndrome.

Pseudogymnoascus destructans is the causative agent of white-nose syndrome, a disease that has caused the deaths of millions of bats in North America. This psychrophilic fungus proliferates at low temperatures and targets hibernating bats, resulting in their premature arousal from stupor with catastrophic consequences. Despite the impact of white-nose syndrome, little is known about the fungus itself or how it infects its mammalian host. P. destructans is not amenable to genetic manipulation, and therefore understanding the proteins involved in infection requires alternative approaches. Here, we identify hydrolytic enzymes secreted by P. destructans, and use a novel and unbiased substrate profiling technique to define active peptidases. These experiments revealed that endopeptidases are the major proteolytic activities secreted by P. destructans, and that collagen, the major structural protein in mammals, is actively degraded by the secretome. A serine endopeptidase, hereby-named Destructin-1, was subsequently identified, and a recombinant form overexpressed and purified. Biochemical analysis of Destructin-1 showed that it mediated collagen degradation, and a potent inhibitor of peptidase activity was identified. Treatment of P. destructans-conditioned media with this antagonist blocked collagen degradation and facilitated the detection of additional secreted proteolytic activities, including aminopeptidases and carboxypeptidases. These results provide molecular insights into the secretome of P. destructans, and identify serine endopeptidases that have the clear potential to facilitate tissue invasion and pathogenesis in the mammalian host

Large library docking for novel SARS‐CoV‐2 main protease non‐covalent and covalent inhibitors

Antiviral therapeutics to treat SARS-CoV-2 are needed to diminish the morbidity of the ongoing COVID-19 pandemic. A well-precedented drug target is the main viral protease (MPro ), which is targeted by an approved drug and by several investigational drugs. Emerging viral resistance has made new inhibitor chemotypes more pressing. Adopting a structure-based approach, we docked 1.2 billion non-covalent lead-like molecules and a new library of 6.5 million electrophiles against the enzyme structure. From these, 29 non-covalent and 11 covalent inhibitors were identified in 37 series, the most potent having an IC50 of 29 and 20 μM, respectively. Several series were optimized, resulting in low micromolar inhibitors. Subsequent crystallography confirmed the docking predicted binding modes and may template further optimization. While the new chemotypes may aid further optimization of MPro inhibitors for SARS-CoV-2, the modest success rate also reveals weaknesses in our approach for challenging targets like MPro versus other targets where it has been more successful, and versus other structure-based techniques against MPro itself

Global identification of peptidase specificity by multiplex substrate profiling

A simple and rapid multiplex substrate profiling method has been developed to reveal the substrate specificity of any endo- or exo-peptidase using LC-MS/MS sequencing. A physicochemically diverse library of peptides was generated by incorporating all combinations of neighbor and near-neighbor amino acid pairs into decapeptide sequences that are flanked by unique dipeptides at each terminus. Addition of a panel of evolutionarily diverse peptidases to a mixture of these tetradecapeptides generated prime and non-prime site information and substrate specificity matched or expanded upon previous substrate motifs. This method biochemically confirmed the activity of the klassevirus 3C gene responsible for polypeptide processing and allowed Granzyme B substrates to be ranked by enzymatic turnover efficiency using label-free quantitation of precursor ion abundance. Furthermore, the proteolytic secretions from a parasitic flatworm larvae and a pancreatic cancer cell line were deconvoluted in a subtractive strategy using class-specific peptidase inhibitors