Selective delivery of CB300638, a cyclopenta[g]quinazoline-based thymidylate synthase inhibitor into human tumor cell lines overexpressing the alpha-isoform of the folate receptor

The alpha-isoform of the glycosylphosphatidylinositol cell membrane tethered folate receptor (alpha-FR) is overexpressed in some carcinomas (notably ovarian carcinomas) relative to normal tissues. The nonpolyglutamatable folate-based thymidylate synthase (TS) inhibitor, CB300638 (TS K(i) = 0.24 nM) displayed an IC(50) of 0.0028 microM for the inhibition of the growth of human A431-FBP cells transfected with the alpha-FR. In contrast, the IC(50) for the neotransfected A431 cells was 0.81 microM (300-fold higher). Similarly, this compound inhibited the growth of human KB cells that constitutively overexpress the alpha-FR with an IC(50) of 0.0036 microM. These data were derived from cells grown in a physiological concentration of folate (20 nM R,S-leucovorin). Incubation of KB cells with a 1 microM excess of folic acid (FA), to selectively block uptake via the alpha-FR, increased the CB300638 IC(50) to 0.39 microM. The relatively low potency of CB300638 under these conditions, or in cell lines not expressing the alpha-FR, is ascribed to its low affinity for the ubiquitously expressed folate transporter, the reduced-folate carrier (K(i) for inhibition of [(3)H]methotrexate transport >100 microM). The high potency of CB300638 in alpha-FR-overexpressing cell lines is attributable to high affinity of the alpha-FR (53% of FA) and efficient endosomal trafficking mediated by the alpha-FR. Sixteen-h exposure to CB300638 inhibited the rate of (3)H(2)O release from 5-[(3)H]dUrd (in situ TS assay) in A431, A431-FBP, and KB cells with IC(50) values of 0.1 microM, 0.005 microM, and 0.002 microM, respectively. The coaddition of 1 micro M FA increased the IC(50)s for A431-FBP and KB cells to approximately 0.1 microM consistent with alpha-FR-mediated transport of CB300638. In conclusion, alpha-FR-mediated uptake of CB300638 leads to TS and growth inhibition that is highly selective for alpha-FR overexpressing tumor cell lines. The low expression of the alpha-FR in normal tissues, particularly those sensitive to TS inhibitors, together with the low affinity of CB300638 for the reduced-folate carrier, suggests that the compound may have potential as an antitumor agent with a high therapeutic index

IMPORT Trial Management Group. Clinical impact of IMPORT HIGH trial (CRUK/06/003) on breast radiotherapy practices in the United Kingdom.

Objective: IMPORT HIGH is a multicentre randomized UK trial testing dose-escalated intensity-modulated radiotherapy (IMRT) after tumour excision in females with early breast cancer and higher than average local recurrence risk. A survey was carried out to investigate the impact of this trial on the adoption of advanced breast radiotherapy (RT) techniques in the UK. Methods: A questionnaire was sent to all 26 IMPORT HIGH recruiting RT centres to determine whether the trial has influenced non-trial breast RT techniques in terms of volume delineation, dosimetry, treatment delivery and verification. In order to compare the clinical practice of breast RT between IMPORT HIGH and non-IMPORT HIGH centres, parts of the Royal College of Radiologists (RCR) breast RT audit result were used in this study. Results: 26/26 participating centres completed the questionnaire. After joining the trial, the number of centres routinely using tumour bed clips to guide whole-breast RT rose from 5 (19%) to 21 (81%). 20/26 (77%) centres now contour target volumes and organs at risk (OARs) in some or all patients compared with 14 (54%) before the trial. 14/26 (54%) centres offer inverse-planned IMRT for selected non-trial patients with breast cancer, and 10/14 (71%) have adopted the IMPORT HIGH trial protocol for target volume and OARs dose constraints. Only 2/26 (8%) centres used clip information routinely for breast treatment verification prior to IMPORT HIGH, a minority that has since risen to 7/26 (27%). Data on 1386 patients was included from the RCR audit. This suggested that more cases from IMPORT HIGH centres had surgical clips implanted (83 vs 67%), were treated using CT guided planning with full three-dimensional dose compensation (100 vs 75%), and were treated with photon boost RT (30 vs 8%). Conclusion: The study suggests that participation in the IMPORT HIGH trial has played an important part in providing the guidance and support networks needed for the safe integration of advanced RT techniques, where appropriate, as a standard of care for breast cancer patients treated at participating cancer centres. Advances in knowledge: We investigated the impact of the IMPORT HIGH trial on the adoption of advanced breast RT techniques in the UK and the trial has influenced non-trial breast RT techniques in terms of volume delineation, dosimetry, treatment delivery and verification

A multicentre observational study evaluating image-guided radiotherapy for more accurate partial-breast intensity-modulated radiotherapy: comparison with standard imaging technique

Background: Whole-breast radiotherapy (WBRT) is the standard treatment for breast cancer following breast-conserving surgery. Evidence shows that tumour recurrences occur near the original cancer: the tumour bed. New treatment developments include increasing dose to the tumour bed during WBRT (synchronous integrated boost) and irradiating only the region around the tumour bed, for patients at high and low risk of tumour recurrence, respectively. Currently, standard imaging uses bony anatomy to ensure accurate delivery of WBRT. It is debatable whether or not more targeted treatments such as synchronous integrated boost and partial-breast radiotherapy require image-guided radiotherapy (IGRT) focusing on implanted tumour bed clips (clip-based IGRT). Objectives: Primary – to compare accuracy of patient set-up using standard imaging compared with clip-based IGRT. Secondary – comparison of imaging techniques using (1) tumour bed radiotherapy safety margins, (2) volume of breast tissue irradiated around tumour bed, (3) estimated breast toxicity following development of a normal tissue control probability model and (4) time taken. Design: Multicentre observational study embedded within a national randomised trial: IMPORT-HIGH (Intensity Modulated and Partial Organ Radiotherapy – HIGHer-risk patient group) testing synchronous integrated boost and using clip-based IGRT. Setting: Five radiotherapy departments, participating in IMPORT-HIGH. Participants: Two-hundred and eighteen patients receiving breast radiotherapy within IMPORT-HIGH. Interventions: There was no direct intervention in patients’ treatment. Experimental and control intervention were clip-based IGRT and standard imaging, respectively. IMPORT-HIGH patients received clip-based IGRT as routine; standard imaging data were obtained from clip-based IGRT images. Main outcome measures: Difference in (1) set-up errors, (2) safety margins, (3) volume of breast tissue irradiated, (4) breast toxicity and (5) time, between clip-based IGRT and standard imaging. Results: The primary outcome of overall mean difference in clip-based IGRT and standard imaging using daily set-up errors was 2–2.6 mm (p < 0.001). Heterogeneity testing between centres found a statistically significant difference in set-up errors at one centre. For four centres (179 patients), clip-based IGRT gave a mean decrease in the systematic set-up error of between 1 mm and 2 mm compared with standard imaging. Secondary outcomes were as follows: clip-based IGRT and standard imaging safety margins were less than 5 mm and 8 mm, respectively. Using clip-based IGRT, the median volume of tissue receiving 95% of prescribed boost dose decreased by 29 cm3 (range 11–193 cm3) compared with standard imaging. Difference in median time required to perform clip-based IGRT compared with standard imaging was X-ray imaging technique dependent (range 8–76 seconds). It was not possible to estimate differences in breast toxicity, the normal tissue control probability model indicated that for breast fibrosis maximum radiotherapy dose is more important than volume of tissue irradiated. Conclusions and implications for clinical practice: Margins of less than 8 mm cannot be used safely without clip-based IGRT for patients receiving concomitant tumour bed boost, as there is a risk of geographical miss of the tumour bed being treated. In principle, smaller but accurately placed margins may influence local control and toxicity rates, but this needs to be evaluated from mature clinical trial data in the future. Funding: This project was funded by the Efficacy and Mechanism Evaluation (EME) programme, a MRC and NIHR partnership

Dose-escalated simultaneous integrated boost radiotherapy in early breast cancer (IMPORT HIGH): a multicentre, phase 3, non-inferiority, open-label, randomised controlled trial.

BACKGROUND: A tumour-bed boost delivered after whole-breast radiotherapy increases local cancer-control rates but requires more patient visits and can increase breast hardness. IMPORT HIGH tested simultaneous integrated boost against sequential boost with the aim of reducing treatment duration while maintaining excellent local control and similar or reduced toxicity. METHODS: IMPORT HIGH is a phase 3, non-inferiority, open-label, randomised controlled trial that recruited women after breast-conserving surgery for pT1-3pN0-3aM0 invasive carcinoma from radiotherapy and referral centres in the UK. Patients were randomly allocated to receive one of three treatments in a 1:1:1 ratio, with computer-generated random permuted blocks used to stratify patients by centre. The control group received 40 Gy in 15 fractions to the whole breast and 16 Gy in 8 fractions sequential photon tumour-bed boost. Test group 1 received 36 Gy in 15 fractions to the whole breast, 40 Gy in 15 fractions to the partial breast, and 48 Gy in 15 fractions concomitant photon boost to the tumour-bed volume. Test group 2 received 36 Gy in 15 fractions to the whole breast, 40 Gy in 15 fractions to the partial breast, and 53 Gy in 15 fractions concomitant photon boost to the tumour-bed volume. The boost clinical target volume was the clip-defined tumour bed. Patients and clinicians were not masked to treatment allocation. The primary endpoint was ipsilateral breast tumour relapse (IBTR) analysed by intention to treat; assuming 5% 5-year incidence with the control group, non-inferiority was predefined as 3% or less absolute excess in the test groups (upper limit of two-sided 95% CI). Adverse events were assessed by clinicians, patients, and photographs. This trial is registered with the ISRCTN registry, ISRCTN47437448, and is closed to new participants. FINDINGS: Between March 4, 2009, and Sept 16, 2015, 2617 patients were recruited. 871 individuals were assigned to the control group, 874 to test group 1, and 872 to test group 2. Median boost clinical target volume was 13 cm3 (IQR 7 to 22). At a median follow-up of 74 months there were 76 IBTR events (20 for the control group, 21 for test group 1, and 35 for test group 2). 5-year IBTR incidence was 1·9% (95% CI 1·2 to 3·1) for the control group, 2·0% (1·2 to 3·2) for test group 1, and 3·2% (2·2 to 4·7) for test group 2. The estimated absolute differences versus the control group were 0·1% (-0·8 to 1·7) for test group 1 and 1·4% (0·03 to 3·8) for test group 2. The upper confidence limit for test group 1 versus the control group indicated non-inferiority for 48 Gy. Cumulative 5-year incidence of clinician-reported moderate or marked breast induration was 11·5% for the control group, 10·6% for test group 1 (p=0·40 vs control group), and 15·5% for test group 2 (p=0·015 vs control group). INTERPRETATION: In all groups 5-year IBTR incidence was lower than the 5% originally expected regardless of boost sequencing. Dose-escalation is not advantageous. 5-year moderate or marked adverse event rates were low using small boost volumes. Simultaneous integrated boost in IMPORT HIGH was safe and reduced patient visits. FUNDING: Cancer Research UK