Combining External Beam Radiation and Radionuclide Therapies: Rationale, Radiobiology, Results and Roadblocks.

The emergence of effective radionuclide therapeutics, such as radium-223 dichloride, [177Lu]Lu-DOTA-TATE and [177Lu]Lu-PSMA ligands, over the last 10 years is driving a rapid expansion in molecular radiotherapy (MRT) research. Clinical trials that are underway will help to define optimal dosing protocols and identify groups of patients who are likely to benefit from this form of treatment. Clinical investigations are also being conducted to combine new MRT agents with other anticancer drugs, with particular emphasis on DNA repair inhibitors and immunotherapeutics. In this review, the case is presented for combining MRT with external beam radiotherapy (EBRT). The technical and dosimetric challenges of combining two radiotherapeutic modalities have impeded progress in the past. However, the need for research into the specific radiobiological effects of radionuclide therapy, which has lagged behind that for EBRT, has been recognised. This, together with innovations in imaging technology, MRT dosimetry tools and EBRT hardware, will facilitate the future use of this important combination of treatments.

Proton vs photon: A model-based approach to patient selection for reduction of cardiac toxicity in locally advanced lung cancer.

PURPOSE/OBJECTIVE: To use a model-based approach to identify a sub-group of patients with locally advanced lung cancer who would benefit from proton therapy compared to photon therapy for reduction of cardiac toxicity. MATERIAL/METHODS: Volumetric modulated arc photon therapy (VMAT) and robust-optimised intensity modulated proton therapy (IMPT) plans were generated for twenty patients with locally advanced lung cancer to give a dose of 70 Gy (relative biological effectiveness (RBE)) in 35 fractions. Cases were selected to represent a range of anatomical locations of disease. Contouring, treatment planning and organs-at-risk constraints followed RTOG-1308 protocol. Whole heart and ub-structure doses were compared. Risk estimates of grade⩾3 cardiac toxicity were calculated based on normal tissue complication probability (NTCP) models which incorporated dose metrics and patients baseline risk-factors (pre-existing heart disease (HD)). RESULTS: There was no statistically significant difference in target coverage between VMAT and IMPT. IMPT delivered lower doses to the heart and cardiac substructures (mean, heart V5 and V30, P < .05). In VMAT plans, there were statistically significant positive correlations between heart dose and the thoracic vertebral level that corresponded to the most inferior limit of the disease. The median level at which the superior aspect of the heart contour began was the T7 vertebrae. There was a statistically significant difference in dose (mean, V5 and V30) to the heart and all substructures (except mean dose to left coronary artery and V30 to sino-atrial node) when disease overlapped with or was inferior to the T7 vertebrae. In the presence of pre-existing HD and disease overlapping with or inferior to the T7 vertebrae, the mean estimated relative risk reduction of grade⩾3 toxicities was 24-59%. CONCLUSION: IMPT is expected to reduce cardiac toxicity compared to VMAT by reducing dose to the heart and substructures. Patients with both pre-existing heart disease and tumour and nodal spread overlapping with or inferior to the T7 vertebrae are likely to benefit most from proton over photon therapy.

International Variation in Criteria for Internal Mammary Chain Radiotherapy.

AIMS: Evidence has emerged that internal mammary chain (IMC) radiotherapy reduces breast cancer mortality, leading to changes in treatment guidelines. This study investigated current IMC radiotherapy criteria and the percentages of patients irradiated for breast cancer in England who fulfilled them. MATERIALS AND METHODS: A systematic search was undertaken for national guidelines published in English during 2013-2018 presenting criteria for 'consideration of' or 'recommendation for' IMC radiotherapy. Patient and tumour variables were collected for patients who received breast cancer radiotherapy in England during 2012-2016. The percentages of patients fulfilling criteria stipulated in each set of guidelines were calculated. RESULTS: In total, 111 729 women were recorded as receiving adjuvant breast cancer radiotherapy in England during 2012-2016 and full data were available on 48 095 of them. Percentages of patients fulfilling IMC radiotherapy criteria in various national guidelines were: UK Royal College of Radiologists 13% (6035/48 095), UK National Institute for Health and Care Excellence 18% (8816/48 095), Germany 32% (15 646/48 095), Ireland 56% (26 846/48 095) and USA 59% (28 373/48 095). Differences between countries occurred because in Ireland and the USA, treatment may be considered in some node-negative patients, whereas in the UK, treatment is considered if at least four axillary nodes are involved or for high-risk patients with one to three positive nodes. In Germany, treatment may be considered for all node-positive patients. CONCLUSIONS: There is substantial variability between countries in criteria for consideration of IMC radiotherapy, despite guidelines being based on the same evidence. This will probably lead to large variations in practice and resource needs worldwide.

Spatial distribution of Auger electrons emitted from internalised radionuclides in cancer cells: the photoresist autoradiography (PAR) method.

Microdosimetric evaluation of Auger electron-emitting radionuclides involves a detailed evaluation of energy deposition at a nanometre scale. To perform Monte Carlo modelling of such energy deposition, accurate information regarding the spatial distribution of the radionuclide is required. A recent addition to the methods for determining the spatial distribution of cellular internalised radionuclides is based on detection in a polymer photoresist (e.g. polymethyl methacralate), followed by atomic force microscopy analysis of the resultant 3D pattern. In comparison with present practice, the method offers greater spatial resolution and improved quantification. The volume of the pattern is proportional to the total dose, thereby permitting assessment of variability of accumulated activity, while the variation in depth across the pattern reflects the lateral spatial distribution in the local fluence per unit area. An added advantage is the similarity in response to ionising radiation of an organic polymer compared to that of biological material. A pattern in the resist from radiation emitted by a radionuclide treated cell gives additional spatial information about the energy deposited in the resist.

Can γH2AX be used to personalise cancer treatment?

Many cancer therapeutics, including radiation therapy, damage DNA eliciting the DNA damage response (DDR). Clinical assays that characterise the DDR could be used to personalise cancer treatment by indicating the extent of damage to tumour and normal tissues and the nature of the cellular response to that damage. The phosphorylated histone γH2AX is generated early in the response to DNA double-strand breaks, the most deleterious form of DNA damage. Translational researchers are developing tissue sampling and assay strategies to apply the measurement of γH2AX to a range of clinical questions, including that of tumour response. The presence of γH2AX is also associated with other cell states including replication stress, hypoxia and apoptosis, which could influence the relationship between γH2AX and clinical endpoints. This review aims to assess the potential of γH2AX as a practical and clinically useful biomarker of tumour and normal tissue responses to therapy.

Advances in anticancer radiopharmaceuticals.

This review highlights recent progress in the development of anticancer radiopharmaceuticals. Molecularly targeted radiotherapy refers to the selective delivery of radionuclides that emit charged particles, such as α particles, ß or Auger electrons, to cancer cells via a targeting vector. The discovery of new molecular targets through systems biology and other approaches has widened the scope for radiopharmaceutical development. Innovations in antibody engineering and humanisation, recombinant DNA technology, conjugation chemistry and, increasingly, nanotechnology have provided new approaches to the delivery of radionuclides to cancer cells. The increased availability of radioisotopes that have not traditionally been considered for therapy, such as α particle emitters, has also broadened the indications for targeted radiotherapy.

Targeting ATR in vivo using the novel inhibitor VE-822 results in selective sensitization of pancreatic tumors to radiation.

Combined radiochemotherapy is the currently used therapy for locally advanced pancreatic ductal adenocarcinoma (PDAC), but normal tissue toxicity limits its application. Here we test the hypothesis that inhibition of ATR (ATM-Rad3-related) could increase the sensitivity of the cancer cells to radiation or chemotherapy without affecting normal cells. We tested VE-822, an ATR inhibitor, for in vitro and in vivo radiosensitization. Chk1 phosphorylation was used to indicate ATR activity, γH2AX and 53BP1 foci as evidence of DNA damage and Rad51 foci for homologous recombination activity. Sensitivity to radiation (XRT) and gemcitabine was measured with clonogenic assays in vitro and tumor growth delay in vivo. Murine intestinal damage was evaluated after abdominal XRT. VE-822 inhibited ATR in vitro and in vivo. VE-822 decreased maintenance of cell-cycle checkpoints, increased persistent DNA damage and decreased homologous recombination in irradiated cancer cells. VE-822 decreased survival of pancreatic cancer cells but not normal cells in response to XRT or gemcitabine. VE-822 markedly prolonged growth delay of pancreatic cancer xenografts after XRT and gemcitabine-based chemoradiation without augmenting normal cell or tissue toxicity. These findings support ATR inhibition as a promising new approach to improve the therapeutic ration of radiochemotherapy for patients with PDAC.

Locoregional treatments for triple-negative breast cancer.

The absence of drug-targetable receptors in triple-negative breast cancer (TNBC) makes the use of targeted systemic therapy inappropriate for this breast cancer subgroup. Although patients with TNBC show sensitivity to some chemotherapy regimens, in early-stage disease greater emphasis is placed on locoregional treatments, based on surgery and radiation therapy (RT). Ongoing improvements in both screening and surgical techniques have reduced the need for radical surgical intervention in all breast cancers, and breast-conserving surgery (BCS) followed by RT is now increasingly common for all tumour types. However, while evidence has clearly established the importance of post-surgical RT for favourable long-term outcomes in breast cancer, it is less well-established as to where and under which conditions more radical surgeries than BCS, such as modified radical mastectomy (MRM), may be indicated for TNBC. A high proportion of TNBC tumours are BRCA1-mutated and therefore patients with this type of tumour are at a potentially elevated risk of ipsilateral or contralateral recurrence. In addition, while some studies indicate that post-BCS locoregional TNBC relapse rates generally appear similar to other tumour types, some evidence suggests that distant relapse rates may be higher. There is evidence that some subtypes of TNBC may require MRM rather than BCS for optimal long-term outcomes. More research is needed to establish whether TNBC-specific approaches to locoregional treatment may be required.

DNA repair capacity as a possible biomarker of breast cancer risk in female BRCA1 mutation carriers.

The BRCA1 gene product helps to maintain genomic integrity through its participation in the cellular response to DNA damage: specifically, the repair of double-stranded DNA breaks. An impaired cellular response to DNA damage is a plausible mechanism whereby BRCA1 mutation carriers are at increased risk of breast cancer. Hence, an individual's capacity to repair DNA may serve as a useful biomarker of breast cancer risk. The overall aim of the current study was to identify a biomarker of DNA repair capacity that could distinguish between BRCA1 mutation carriers and non-carriers. DNA repair capacity was assessed using three validated assays: the single-cell alkaline gel electrophoresis (comet) assay, the micronucleus test, and the enumeration of gamma-H2AX nuclear foci. DNA repair capacity of peripheral blood lymphocytes from 25 cancer-free female heterozygous BRCA1 mutation carriers and 25 non-carrier controls was assessed at baseline and following cell exposure to gamma-irradiation (2 Gy). We found no significant differences in the mean tail moment, in the number of micronuclei or in the number of gamma-H2AX nuclear foci between the carriers and non-carriers at baseline, and following gamma-irradiation. These data suggest that these assays are not likely to be useful in the identification of women at a high risk for breast cancer.

A phase I study of 99mTc-hR3 (DiaCIM), a humanized immunoconjugate directed towards the epidermal growth factor receptor.

A phase I trial was conducted to evaluate the safety, tumour and normal tissue localization, pharmacokinetics and radiation dosimetry of Tc-hR3, a humanized monoclonal antibody directed towards the epidermal growth factor receptor, in 12 patients with recurrent or metastatic epithelial malignancies. Patients were injected intravenously with 3.0 mg or 6.0 mg (1010 MBq) of Tc-hR3. Blood and plasma concentrations of radioactivity were measured and a complete 24 h urine collection was obtained. Whole-body images were acquired up to 24 h post-injection and normal organ uptake quantified. Radiation dosimetry was estimated using MIRDose. Safety was evaluated by clinical observation, biochemical/haematological testing and by measuring immune response to Tc-hR3. There were no adverse effects, no changes in biochemical/haematological indices and no immune response to Tc-hR3. Tc-hR3 was rapidly cleared from the blood with a distribution half-life of 10.8+/-3.8 min. The volume of distribution, and clearance, were 180+/-37 ml.kg and 14+/-3 ml.kg.min, respectively. The elimination phase could not be discerned due to increasing blood radioactivity at later times. About 19-24% was excreted in the urine. Normal tissue uptake was mainly in the liver (44-50%), spleen (3-4%) and kidneys (3%). Imaging was positive in one patient with squamous cell carcinoma of the mouth and an involved cervical lymph node. The whole-body radiation dose from Tc-hR3 was 1.34+/-0.02x10 mSv.Bq. We conclude that Tc-hR3 exhibited an excellent safety profile. Future studies to determine the sensitivity and specificity of imaging with Tc-hR3 in a larger group of patients with pre-selection for epidermal growth factor receptor positivity are planned.

Evaluation of toremifene for reversal of multidrug resistance in renal cell cancer patients treated with vinblastine.

PURPOSE: Expression of P-glycoprotein (Pgp), which confers the multidrug resistance (MDR) phenotype, is thought to contribute to the insensitivity of renal cell cancer (RCC) to chemotherapy. The development of Pgp inhibitors for clinical application has been hampered by unacceptable toxicity at doses required to achieve adequate cellular concentration. Toremifene is able to reverse MDR and sensitise RCC to vinblastine in vitro. However, in vivo toremifene is tightly bound to serum proteins, in particular the acute phase protein alpha1-acid glycoprotein (AAG), which may limit tissue availability. In this phase I-II study we assessed the tolerability of short courses of high dose toremifene in combination with vinblastine and evaluated the key determinants of MDR reversal in vivo. METHODS: Twenty-seven patients with metastatic RCC received escalating doses of oral toremifene for 3 days every 2 weeks in combination with vinblastine 6 mg/m2 i.v. on day 3 of each cycle. The serum concentration of toremifene, its metabolites and AAG were measured and the effect of patients' serum on inhibition of Pgp in vitro was determined. RESULTS: Twenty-six patients were evaluable for response. Eight patients (31%) had stable disease and 18 patients (69%) progressive disease. The mean serum concentration of toremifene at 780 mg daily for 3 days was 7.82 microM [standard deviation (SD) 2.48, range 2.50 to 14.70], which exceeds that known to reverse MDR in vitro. The serum concentration of the major metabolite of toremifene, N-demethyltoremifene, which also reverses MDR, was 5.13 microM (SD 1.78, range 1.80 to 9.00). In 60% of patients the pre-treatment AAG concentration was above that known to block the effects of toremifene in vitro. However, addition of serum from patients on toremifene to MCF-7 adr cells in vitro inhibited Pgp-mediated efflux of rhodamine 123. CONCLUSIONS: We have shown that short course, high-dose toremifene in combination with vinblastine is generally well tolerated and that the concentration of toremifene required to reverse MDR in vitro is achievable in vivo.

Expression trapping: identification of novel genes expressed in hematopoietic and endothelial lineages by gene trapping in ES cells.

We have developed a large-scale, expression-based gene trap strategy to perform genome-wide functional analysis of the murine hematopoietic and vascular systems. Using two different gene trap vectors, we have isolated embryonic stem (ES) cell clones containing lacZ reporter gene insertions in genes expressed in blood island and vascular cells, muscle, stromal cells, and unknown cell types. Of 79 clones demonstrating specific expression patterns, 49% and 16% were preferentially expressed in blood islands and/or the vasculature, respectively. The majority of ES clones that expressed lacZ in blood islands also expressed lacZ upon differentiation into hematopoietic cells on OP9 stromal layers. Importantly, the in vivo expression of the lacZ fusion products accurately recapitulated the observed in vitro expression patterns. Expression and sequence analysis of representative clones suggest that this approach will be useful for identifying and mutating novel genes expressed in the developing hematopoietic and vascular systems.

Menadione-resistant Chinese hamster ovary cells have an increased capacity for glutathione synthesis.

A cell line (MRc40) resistant to the model quinone compound, menadione, has been isolated from a parental Chinese hamster ovary cell line (CHO-K1). The known relationship between menadione toxicity and glutathione (GSH) depletion led us to investigate whether the mechanism of resistance of MRc40 was related to alteration in GSH homeostasis. Intracellular concentrations of GSH and cysteine (CySH) were twofold and 3.2-fold greater in MRc40 than in CHO-K1. Following exposure to menadione, GSH and CySH were depleted, but subsequent recovery of thiols was more rapid and of greater magnitude in MRc40 than in CHO-K1. Twelve hours after exposure to menadione, the concentrations of GSH and CySH were 9.7- and 4.2-fold greater in MRc40 than in CHO-K1. Using nuclear magnetic resonance (NMR) spectroscopy, we observed the in situ removal of menadione from cell suspensions of CHO-K1 and MRc40. However, only in CHO-K1 did we observe concomitant depletion of NMR-visible GSH. We conclude that the perturbation of GSH metabolism contributes to the resistant phenotype and is an important characteristic of menadione-resistant CHO cells.

External beam and intraluminal radiotherapy for locally advanced bile duct cancer: role and tolerability.

BACKGROUND AND PURPOSE: Cholangiocarcinoma is rare but carries a poor prognosis. Radiotherapy has been used either as an adjuvant treatment following surgical resection of tumour or for palliation. The purpose of this study was to assess the feasibility and morbidity of accelerated external beam radiotherapy with or without intraluminal radiotherapy in the treatment of locally advanced bile duct cancer. MATERIALS AND METHODS: Thirty eight patients were treated. Surgical procedures performed prior to radiotherapy were extended hepatectomy (3), hepaticojejunostomy with tumour resection (6), palliative biliary-enteric bypass (6), biopsy (4), Whipple's procedure (1), gastrojejunostomy (1) and cholecystectomy (1). Twenty patients received external beam radiotherapy (ERT). Six patients received one Phase of ERT and 12 received two Phases, separated by a 2-week gap. Dose per Phase was 22.5 Gy in 10 twice daily fractions. After 1989, dose per Phase was increased to 27.5 Gy. One patient received Phase I ERT (30.0 Gy) using conventional fractionation and one patient received an uninterrupted, conventionally fractionate course of treatment (50.0 Gy). Fourteen patients received both ERT and intraluminal radiotherapy (IRT) using iridium-192 (192Ir) wire passed through a percutaneous, transhepatic catheter (median dose, ERT 23.8 Gy + IRT 40.0 Gy). In addition, four patients received IRT alone (median dose 45.0 Gy at 1 cm radius). Patients were followed for at least 42 months. RESULTS: Median overall survival was 15 months. Overall survival for the whole group at 1,2 and 3 years was 59.6%, 32.5% and 16.2%. Thirty four patients died of disease. Radiotherapy caused acute toxicity in seven patients. According to RTOG/EORTC criteria toxicity was Grade 1 in four cases, Grade 2 in two cases and Grade 3 in one case. Two patients developed gastrointestinal bleeding as a late complication of radiotherapy. CONCLUSIONS: Accelerated external beam radiotherapy with or without intraluminal radiotherapy is feasible and associated with acceptable toxicity when used in the management of advance cholangiocarcinoma.

Relationship between the adaptive response to oxidants and stable menadione-resistance in Chinese hamster ovary cell lines.

To study the genetic changes that generate resistance to oxidants in mammalian cells, we isolated cell lines that are resistant to the naphthoquinone, menadione, from a Chinese hamster ovary cell line (CHO-K1). Corss-resistance to other oxidants (H2O2 and sodium arsenite) was observed. The IC50 of menadione (measured using a clonogenic assay) was 7.8-fold greater for one menadione-resistant cell line (MRc40) than for CHO-K1. Acquisition of resistance was associated with elevations of 2- and 3.2-fold in the low molecular weight thiols, glutathione and cysteine, respectively. Further, characterization demonstrated significant changes in the expression of enzymes associated with the oxidative stress response and with protection against oxidizing agents. The expressions of glutathione S-transferase pi (GST pi), glutathione peroxidase (GPX) and heme oxygenase mRNAs were all increased. Accompanying these changes the enzyme activity of GST pi, GPX and gamma-glutamyl transpeptidase (gamma-GT) were also elevated. Interestingly, in a revertant cell line heme oxygenase overexpression approached wild-type levels. Intriguingly, similar changes in gene expression seen in the menadione-resistant cells were also observed in wild-type cells following transient oxidative stress, indicating that the observed changes in the resistant line may be due to the immortalization of a normally transient adaptive stress response.

Radiotherapy for stages I and II testicular seminoma: results and morbidity in 238 patients.

We have undertaken a retrospective analysis of 238 patients with Stages I and II seminoma of the testis treated with radiotherapy in Edinburgh between 1974 and 1989. There were five deaths from seminoma. Cause-specific survival for the whole group at 2 and 5 years was 99.2% and 98.1%, respectively. Cause-specific survival at 2 and 5 years by stage (Royal Marsden staging classification) was: Stage I, 99.5% and 98.7% and Stage II, 98.1% and 96.1%. Fourteen (5.9%) patients relapsed (one after treatment for his second testicular seminoma). Eight were given successful salvage treatment, five died of seminoma and one died of intercurrent disease. 13 (5.5%) patients developed World Health Organisation (WHO) grade 3 gastrointestinal or haematological toxicity and two developed grade 4 gastrointestinal toxicity as a result of abdominal radiotherapy. 22 patients (9.2%) developed problems ascribed to late morbidity of abdominal radiotherapy including 18 with peptic ulcer disease. Contralateral testicular tumours occurred in seven (2.9%) patients and five (2.1%) patients developed malignancies at other sites.

Relationship between the debrisoquine hydroxylase polymorphism and cancer susceptibility.

There have been a series of reports on the association of a genetic polymorphism at the cytochrome P450 CYP2D6 gene locus with cancer susceptibility. Many of these reports have remained contradictory either because of small numbers of patients studied or because of the limitations and controversy surrounding the pharmacokinetic assay used to identify affected individuals (poor metabolizers; PMs). We have recently developed a DNA-based assay that will allow the unequivocal identification of poor metabolizers and have applied this to the study of 1635 patients with different forms of cancer. Out of 361 lung cancer patients studied no statistically significant change in the proportion of PMs relative to controls was found. However, a significant increase in the proportion of poor metabolizers or heterozygotes was seen in leukaemia, bladder cancer and melanoma patients. This could be explained by a role for CYP2D6 in carcinogen detoxification or by linkage to another cancer-causing gene.

Distress associated with radiotherapy for malignant disease: a quantitative analysis based on patients perceptions.

Distress associated with attendance at a radiotherapy department was assessed in 80 consecutive patients. All patients were interviewed within 24 h of their first fraction of radiotherapy; 31 patients were also interviewed at the end of treatment. The problem identified at first interview as causing the most distress was worry about the effects of disease and its treatment upon the patient's family. At second interview the dominant complaint was of not being allowed to wash. Psychological problems, including anxiety and sleep disturbances, caused more overall distress than did physical symptoms. The method used in this study for eliciting information on the side-effects of therapy is straightforward and has yielded data that are provocative and suggest interesting avenues for further investigation.