Quantifying the carbon footprint of clinical trials: guidance development and case studies.

BACKGROUND: The urgency of the climate crisis requires attention from biomedical research, not least clinical trials which can involve significant greenhouse gas emissions. The Low Carbon Clinical Trials Working Group set out a strategy to reduce the emissions of clinical trials, starting with the development of a method to measure their carbon footprint (CO2e). METHODS: As a first step, we developed a process map defining clinical trial core activities. Corresponding emission factors were sourced to convert activity data into greenhouse gas emissions. The subsequent method was applied to two Cancer Research UK (CRUK)-funded trials (the international randomised sarcoma trial CASPS (ISRCTN63733470) and the UK cohort-based breast cancer trial PRIMETIME (ISRCTN41579286)). A guidance document defining the scope, method and assumptions was written to allow application to any publicly funded/investigator initiated clinical trial. RESULTS: Trial specific activities over and above routine care were grouped into 10 modules covering trial set up, conduct and closure. We identified emission factors for all trial activities within both trials and used them to estimate their total carbon footprint. The carbon footprint of CASPS, an international phase 2 trial of an investigational medicinal product with 47 participants, was 72 tonnes CO2e, largely attributable to clinical trials unit emissions and staff travel. PRIMETIME, a UK-based phase 3 non-investigational medicinal product trial with 1962 patients, produced 89 tonnes CO2e, largely attributable to trial-specific in-person participant assessments. CONCLUSION: We have developed a method and guidance that trialists can use to determine the carbon footprint of clinical trials. The guidance can be used to identify carbon hotspots where alternative approaches to trial design and conduct could reduce a trial footprint, and where methodology research is required to investigate the potential impact of interventions taken to reduce carbon emissions. We will continue to refine the guidance to increase the potential application and improve usability.

Equality, diversity, and inclusion in oncology clinical trials: an audit of essential documents and data collection against INCLUDE under-served groups in a UK academic trial setting.

BACKGROUND: Clinical trials should be as inclusive as possible to facilitate equitable access to research and better reflect the population towards which any intervention is aimed. Informed by the UK's National Institute for Health and Care Research (NIHR) Innovations in Clinical Trial Design and Delivery for the Under-served (INCLUDE) guidance, we audited oncology trials conducted by the Clinical Trials and Statistics Unit at The Institute of Cancer Research, London (ICR-CTSU) to identify whether essential documents were overtly excluding any groups and whether sufficient data were collected to assess diversity of trial participants from groups suggested by INCLUDE as under-served by research in the UK. METHODS: Thirty cancer clinical trials managed by ICR-CTSU and approved between 2011-2021 were audited. The first ethics approved version of each trial's protocol, patient information sheet, and patient completed questionnaire, together with the first case report forms (CRFs) version were reviewed. A range of items aligned with the INCLUDE under-served groups were assessed, including age, sex and gender, socio-economic and health factors. The scope did not cover trial processes in participating hospitals. RESULTS: Data relating to participants' age, ethnic group and health status were well collected and no upper age limit was specified in any trials' eligibility criteria. 23/30 (77%) information sheets used at least one gendered term to address patients. Most CRFs did not specify whether they were collecting sex or gender and only included male or female categories. The median reading age for information sheets was 15-16 years (IQR: 14-15 - 16-17). Socio-economic factors were not routinely collected and not commonly mentioned in trial protocols. CONCLUSIONS: No systemic issues were identified in protocols which would explicitly prevent any under-served group from participating. Areas for improvement include reducing use of gendered words and improving readability of patient information. The challenge of fully assessing adequate inclusion of under-served populations remains, as socio-economic factors are not routinely collected because they fall beyond the data generally required for protocol-specified trial endpoint assessments. This audit has highlighted the need to agree and standardise demographic data collection to permit adequate monitoring of the under-served groups identified by the NIHR.

Identifying treatment options for BRAFV600 wild-type metastatic melanoma: A SU2C/MRA genomics-enabled clinical trial.

Although combination BRAF and MEK inhibitors are highly effective for the 40-50% of cutaneous metastatic melanomas harboring BRAFV600 mutations, targeted agents have been ineffective for BRAFV600wild-type (wt) metastatic melanomas. The SU2C Genomics-Enabled Medicine for Melanoma Trial utilized a Simon two-stage optimal design to assess whether comprehensive genomic profiling improves selection of molecular-based therapies for BRAFV600wt metastatic melanoma patients who had progressed on standard-of-care therapy, which may include immunotherapy. Of the response-evaluable patients, binimetinib was selected for 20 patients randomized to the genomics-enabled arm, and nine were treated on the alternate treatment arm. Response rates for 27 patients treated with targeted recommendations included one (4%) partial response, 18 (67%) with stable disease, and eight (30%) with progressive disease. Post-trial genomic and protein pathway activation mapping identified additional drug classes that may be considered for future studies. Our results highlight the complexity and heterogeneity of metastatic melanomas, as well as how the lack of response in this trial may be associated with limitations including monotherapy drug selection and the dearth of available single and combination molecularly-driven therapies to treat BRAFV600wt metastatic melanomas.

Cediranib in patients with alveolar soft-part sarcoma (CASPS): a double-blind, placebo-controlled, randomised, phase 2 trial.

BACKGROUND: Alveolar soft-part sarcoma (ASPS) is a rare soft-tissue sarcoma that is unresponsive to chemotherapy. Cediranib, a tyrosine-kinase inhibitor, has shown substantial activity in ASPS in non-randomised studies. The Cediranib in Alveolar Soft Part Sarcoma (CASPS) study was designed to discriminate the effect of cediranib from the intrinsically indolent nature of ASPS. METHODS: In this double-blind, placebo-controlled, randomised, phase 2 trial, we recruited participants from 12 hospitals in the UK (n=7), Spain (n=3), and Australia (n=2). Patients were eligible if they were aged 16 years or older; metastatic ASPS that had progressed in the previous 6 months; had an ECOG performance status of 0-1; life expectancy of more than 12 weeks; and adequate bone marrow, hepatic, and renal function. Participants had to have no anti-cancer treatment within 4 weeks before trial entry, with exception of palliative radiotherapy. Participants were randomly assigned (2:1), with allocation by use of computer-generated random permuted blocks of six, to either cediranib (30 mg orally, once daily) or matching placebo tablets for 24 weeks. Treatment was supplied in number-coded bottles, masking participants and clinicians to assignment. Participants were unblinded at week 24 or sooner if they had progression defined by Response Evaluation Criteria in Solid Tumors (version 1.1); those on placebo crossed over to cediranib and all participants continued on treatment until progression or death. The primary endpoint was percentage change in sum of target marker lesion diameters between baseline and week 24 or progression if sooner, assessed in the evaluable population (all randomly assigned participants who had a scan at week 24 [or sooner if they progressed] with target marker lesions measured). Safety was assessed in all participants who received at least one dose of study drug. This study is registered with ClinicalTrials.gov, number NCT01337401; the European Clinical Trials database, number EudraCT2010-021163-33; and the ISRCTN registry, number ISRCTN63733470 recruitment is complete and follow-up is ongoing. FINDINGS: Between July 15, 2011, and July 29, 2016, of 48 participants recruited, all were randomly assigned to cediranib (n=32) or placebo (n=16). 23 (48%) were female and the median age was 31 years (IQR 27-45). Median follow-up was 34·3 months (IQR 23·7-55·6) at the time of data cutoff for these analyses (April 11, 2018). Four participants in the cediranib group were not evaluable for the primary endpoint (one did not start treatment, and three did not have their scan at 24 weeks). Median percentage change in sum of target marker lesion diameters for the evaluable population was -8·3% (IQR -26·5 to 5·9) with cediranib versus 13·4% (IQR 1·1 to 21·3) with placebo (one-sided p=0·0010). The most common grade 3 adverse events on (blinded) cediranib were hypertension (six [19%] of 31) and diarrhoea (two [6%]). 15 serious adverse reactions in 12 patients were reported; 12 of these reactions occurred on open-label cediranib, and the most common symptoms were dehydration (n=2), vomiting (n=2), and proteinuria (n=2). One probable treatment-related death (intracranial haemorrhage) occurred 41 days after starting open-label cediranib in a patient who was assigned to placebo in the masked phase. INTERPRETATION: Given the high incidence of metastatic disease and poor long-term prognosis of ASPS, together with the lack of efficacy of conventional chemotherapy, our finding of significant clinical activity with cediranib in this disease is an important step towards the goal of long-term disease control for these young patients. Future clinical trials in ASPS are also likely to involve immune checkpoint inhibitors. FUNDING: Cancer Research UK and AstraZeneca.

Carboplatin in BRCA1/2-mutated and triple-negative breast cancer BRCAness subgroups: the TNT Trial.

Germline mutations in BRCA1/2 predispose individuals to breast cancer (termed germline-mutated BRCA1/2 breast cancer, gBRCA-BC) by impairing homologous recombination (HR) and causing genomic instability. HR also repairs DNA lesions caused by platinum agents and PARP inhibitors. Triple-negative breast cancers (TNBCs) harbor subpopulations with BRCA1/2 mutations, hypothesized to be especially platinum-sensitive. Cancers in putative 'BRCAness' subgroups-tumors with BRCA1 methylation; low levels of BRCA1 mRNA (BRCA1 mRNA-low); or mutational signatures for HR deficiency and those with basal phenotypes-may also be sensitive to platinum. We assessed the efficacy of carboplatin and another mechanistically distinct therapy, docetaxel, in a phase 3 trial in subjects with unselected advanced TNBC. A prespecified protocol enabled biomarker-treatment interaction analyses in gBRCA-BC and BRCAness subgroups. The primary endpoint was objective response rate (ORR). In the unselected population (376 subjects; 188 carboplatin, 188 docetaxel), carboplatin was not more active than docetaxel (ORR, 31.4% versus 34.0%, respectively; P = 0.66). In contrast, in subjects with gBRCA-BC, carboplatin had double the ORR of docetaxel (68% versus 33%, respectively; biomarker, treatment interaction P = 0.01). Such benefit was not observed for subjects with BRCA1 methylation, BRCA1 mRNA-low tumors or a high score in a Myriad HRD assay. Significant interaction between treatment and the basal-like subtype was driven by high docetaxel response in the nonbasal subgroup. We conclude that patients with advanced TNBC benefit from characterization of BRCA1/2 mutations, but not BRCA1 methylation or Myriad HRD analyses, to inform choices on platinum-based chemotherapy. Additionally, gene expression analysis of basal-like cancers may also influence treatment selection.

Conducting non-commercial international clinical trials: the ICR-CTSU experience.

BACKGROUND: Academic clinical trials play a fundamental role in the development of new treatments, the repurposing of existing treatments and in addressing areas of unmet clinical need. With cancer treatments increasingly targeted at molecular subtypes, and with priority placed on developing new treatments for rare tumour types, the need for international trial participation to access sufficient patient numbers for successful trial conduct is growing. However, lack of harmonisation of international legal, ethical and financial systems can make this challenging and the cost and effort of conducting trials internationally can be considered prohibitive, particularly where the sample size is comparatively small. METHODS: The Institute of Cancer Research - Clinical Trials and Statistics Unit (ICR-CTSU) is a UK-based academic clinical trials unit that specialises in the design, conduct and analysis of clinical trials of cancer treatments with an expanding portfolio of trials in molecular subtypes of breast and urological cancers and in other rare cancer types. Implementing appropriate mechanisms to enable international participation has therefore been imperative. In this article, we explain how we have approached the challenges involved and describe examples of successful international trial conduct, achieved through robust collaborations with academic and industry partners. CONCLUSION: Conducting academic trials internationally is challenging but can and should be achieved through appropriate governance mechanisms and strong collaborations.

Outcomes of preterm infants receiving positive pressure support at term corrected gestation.

AIM: To establish the mortality rate to hospital discharge of very preterm infants who remain on positive pressure support (PPS) at term corrected gestation and describe factors that are associated with increased mortality. METHODS: Infants born <30 weeks' gestation between 1 January 2001 and 31 December 2009 who were receiving PPS at 40 weeks' postmenstrual age (PMA) were identified from our database, and their medical records reviewed. The fraction of inspired oxygen (FiO2 ), mean airway pressure and partial pressure of carbon dioxide (PaCO2 ) at 40 weeks' PMA were recorded. Receiver operating characteristic curves for mortality before discharge were generated. RESULTS: One thousand three hundred fifty-nine of 1572 infants survived to term. Forty-nine infants were receiving PPS at 40 weeks' PMA. Of these, 15 (31%) infants died before hospital discharge. All three infants who were ventilated via an endotracheal tube died. Increased oxygen requirement at term was associated with increased risk of death before discharge (area under curve (AUC) 0.75). FiO2 > 0.5 was associated with an 80% risk of death. PaCO2 was not predictive of death (AUC 0.49). CONCLUSIONS: Two thirds of very preterm infants who remained on PPS at 40 weeks' PMA survived to hospital discharge. Risk of death rises with increasing oxygen requirements. All five infants with FiO2 > 0.70 died.

Expression of CD1c enhances human invariant NKT cell activation by α-GalCer.

Invariant natural killer T (iNKT) cells are innate T lymphocytes that specifically recognize α-linked glycosphingolipids (α-GSLs) as antigens presented by CD1d molecules. Activating iNKT cells by administering α-GSLs improves disease outcomes in murine cancer models and, thus, there is great interest in the clinical potential of these lipids for treating human cancers. However, humans possess several other CD1 isoforms that are not present in mice and it is not clear whether these CD1 molecules, which also bind lipids, affect human iNKT cell responses. We demonstrate here that CD1c, which is co-expressed with CD1d on blood dendritic cells and on a fraction of B cells, is able to present α-galactosylceramide (α-GalCer) as a weak agonist to human iNKT cells, and that the presence of CD1c synergistically enhances α-GalCerdependent activation of iNKT cells by CD1d. Primary human B cells expressing CD1c induced stronger iNKT cell responses to α-GalCer than the CD1c- subset, and an antibody against CD1c inhibited iNKT cell cytokine secretion. These results suggest that therapeutic activation of human iNKT cells by α-GSLs will be driven preferentially by CD1c+ cell types. Thus, B cell neoplasias that co-express CD1c and CD1d may be particularly susceptible to α-GSL therapy, and cancer vaccines using α-GSLs as adjuvants may be most effective when presented by CD1c+ antigen-presenting cells.

Validation of membrane vesicle-based breast cancer resistance protein and multidrug resistance protein 2 assays to assess drug transport and the potential for drug-drug interaction to support regulatory submissions.

Breast cancer resistance protein (BCRP) and multidrug resistance protein 2 (MRP2) can play a role in the absorption, distribution, metabolism, and excretion of drugs, impacting on the potential for drug-drug interactions. This study has characterized insect cell- and mammalian cell-derived ABC-transporter-expressing membrane vesicle test systems and validated methodologies for evaluation of candidate drugs as substrates or inhibitors of BCRP or MRP2. Concentration-dependent uptake of BCRP ([³H]oestrone 3-sulfate, [³H]methotrexate, [³H]rosuvastatin) and MRP2 ([³H]oestradiol 17ß-glucuronide, [³H]pravastatin, carboxydichlorofluorescein) substrates, and inhibitory potencies (IC50) of BCRP (sulfasalazine, novobiocin, fumitremorgin C) and MRP2 (benzbromarone, MK-571, terfenadine) inhibitors were determined. The apparent K(m) for probes [³H]oestrone 3-sulfate and [³H]oestradiol 17ß-glucuronide was determined in insect cell vesicles to be 7.4 ± 1.7 and 105 ± 8.3 µM, respectively. All other substrates exhibited significant uptake ratios. Positive control inhibitors sulfasalazine and benzbromarone gave IC50 values of 0.74 ± 0.18 and 36 ± 6.1 µM, respectively. All other inhibitors exhibited concentration-dependent inhibition. There was no significant difference in parameters generated between test systems. On the basis of the validation results, acceptance criteria to identify substrates/inhibitors of BCRP and MRP2 were determined for insect cell vesicles. The approach builds on earlier validations to support drug registration and extends from those cell-based systems to encompass assay formats using membrane vesicles.

In vitro risk assessment of AZD9056 perpetrating a transporter-mediated drug-drug interaction with methotrexate.

Methotrexate is the most commonly used drug to treat rheumatoid arthritis. Since clinical efficacy studies in rheumatoid arthritis are conducted on a background of methotrexate therapy, it is necessary to assess the potential of rheumatoid arthritis candidate drugs to perpetrate a drug-drug interaction (DDI) with methotrexate during development. Consequently, we need to identify the regulatory in vitro studies required to facilitate this assessment. We therefore reviewed the literature to ascertain the methotrexate disposition pathways implicated with known DDIs. Experiments were conducted to confirm that methotrexate was identified as a substrate for these pathways in our laboratory. The literature indicated active renal elimination (mediated by the human transporters OAT1, OAT3, MRP2 and BCRP) to be the principal pathway for methotrexate DDI risk. With the exception of MRP2, methotrexate was confirmed as a substrate of these transporters using oocyte and membrane vesicle test systems. A rheumatoid arthritis candidate drug (AZD9056) and sulfasalazine were subsequently assessed as inhibitors of OAT1, OAT3 and BCRP to determine their DDI potential towards methotrexate. AZD9056 was neither an inhibitor of OAT1 nor OAT3 and did not inhibit their transport of methotrexate. AZD9056 was an inhibitor of BCRP and weakly inhibited BCRP-mediated transport of methotrexate (IC(50)=92µM). Sulfasalazine inhibited methotrexate transport mediated by all transporters studied (IC(50)<5µM). Subsequent assessment of the in vitro data using [I]/IC(50) ratios indicated that both AZD9056 and sulfasalazine were unlikely to cause a DDI with methotrexate in vivo. In conclusion, to support rheumatoid arthritis drug development it is proposed that regulatory in vitro studies for OAT1, OAT3 and BCRP inhibition be routinely conducted to assess the potential for a transporter-mediated DDI with methotrexate in vivo.

Determination of cellular lipids bound to human CD1d molecules.

CD1 molecules are glycoproteins that present lipid antigens at the cell surface for immunological recognition by specialized populations of T lymphocytes. Prior experimental data suggest a wide variety of lipid species can bind to CD1 molecules, but little is known about the characteristics of cellular ligands that are selected for presentation. Here we have molecularly characterized lipids bound to the human CD1d isoform. Ligands were eluted from secreted CD1d molecules and separated by normal phase HPLC, then characterized by mass spectroscopy. A total of 177 lipid species were molecularly identified, comprising glycerophospholipids and sphingolipids. The glycerophospholipids included common diacylglycerol species, reduced forms known as plasmalogens, lyso-phospholipids (monoacyl species), and cardiolipins (tetraacyl species). The sphingolipids included sphingomyelins and glycosylated forms, such as the ganglioside GM3. These results demonstrate that human CD1d molecules bind a surprising diversity of lipid structures within the secretory pathway, including compounds that have been reported to play roles in cancer, autoimmune diseases, lipid signaling, and cell death.

Neonatal hyperparathyroidism and pamidronate therapy in an extremely premature infant.

We describe the use of pamidronate to control marked hypercalcemia in an extremely premature infant with neonatal hyperparathyroidism that resulted from an inactivating mutation (R220W) of the calcium-sensing receptor. Despite improvement in bone mineralization and subsequent parathyroidectomy with normalization of the serum calcium level, the combination of chronic lung disease, osteomalacia, and poor thoracic cage growth ultimately proved fatal. Pamidronate therapy seems to be safe in the short-term and effective in helping control hypercalcemia even in the very premature infant, allowing for planned surgical intervention when it becomes feasible.

Comparison of species differences of P-glycoproteins in beagle dog, rhesus monkey, and human using Atpase activity assays.

P-glycoprotein (P-gp) is a transmembrane efflux transporter which possesses many important functions in drug absorption, disposition, metabolism, and toxicity. The ultimate goal of investigating drug interactions between P-gp and drug molecules in early drug discovery is to understand the contribution of P-gp to the pharmacokinetic and pharmacodynamic properties of drug candidates and to project drug-drug interaction (DDI) potentials in humans. Understanding species differences in P-gp activities further helps the prediction of P-gp-mediated drug disposition and DDI in humans from preclinical pharmacokinetics data. The objective of the present study is to investigate the species difference in P-gp activities, via P-gp ATPase assays, using rhesus monkey Mdr1, beagle dog Mdr1, and human MDR1 expressed insect cell membranes. Twenty-one compounds with diverse chemical structures and different P-gp binding sites were chosen for the ATPase assays. P-gp ATPase binding affinities (alphaKa) and fold increases in P-gp ATPase activities (beta) of P-gp substrates were determined. Consistent with the gene and amino acid similarity, the binding affinities of test compounds to rhesus monkey P-gp were much closer to those of human P-gp than beagle dog P-gp. This is the first study which investigates the ligand affinities of P-gp from three different species. The result of this study provides an example of how to use membrane P-gp ATPase assays to evaluate interspecies P-gp differences.

Central neurologic involvement in mycosis fungoides: ten cases, actuarial risk assessment, and predictive factors.

BACKGROUND: Neurologic involvement in mycosis fungoides is rare. Isolated case reports in the literature suggest the pattern and the natural history for such occurrences, while a literature summary can provide direction on diagnosis and management. Although case series may confirm such information, cohort data are required to establish an overall risk of occurrence and to evaluate possible predictive factors. METHODS: We presented a case of central nervous system involvement in mycosis fungoides from Haifa, Israel and tabulated a series of nine cases from Canada. To estimate the risk of neurologic involvement, a cohort of 680 consecutive patients with newly diagnosed mycosis fungoides, of which the nine cases of neurologic involvement emerged during follow up, was analyzed using the Kaplan-Meier method. The actuarial risk of developing neurologic involvement was related to the baseline tumor-node-metastasis-blood classification factors. RESULTS: The pattern of disease in these 10 additional cases confirms the overall pattern in the approximately 40 patients described in the literature. The main symptoms are fluctuating higher cognitive functions and cranial nerve dysfunction, with fairly rapid clinical onset of symptoms. Most cases of central neurologic involvement with mycosis fungoides emerge within a setting of advanced disease. In patients with newly diagnosed mycosis fungoides, the greatest risk of developing neurologic involvement is within the first several years after diagnosis and is associated with the initial stage of disease. Patients with two or more of the T3-4, N3, M1, and B1 classification factors have a one in six chance of developing central neurologic involvement, while there is about a one in a hundred chance for the corresponding control group. CONCLUSIONS: Neurologic involvement with mycosis fungoides is indeed rare, but it is associated with a more advanced stage at diagnosis and with other visceral disease that can precede it. Although the role of low-dose prophylactic cranial radiation is uncertain, overt neurologic involvement requires urgent palliative treatment.