P.W. Dykes

Limitations to the killing of tumours using radiolabelled antibodies

We have calculated the minimum requirements for effective therapy using intravenously administered, tumour-directed antibodies labelled with either iodine 131 or yttrium 90. A lethally large amount of either radionuclide would be required to achieve tumour destruction. At least a 10-fold increase in tumour uptake is necessary to combine tumour destruction with a survivable whole-body dose. The required improvement in specific uptake can be substantially reduced by accelerating the excretion of radioactivity outside the tumour. For all situations studied, yttrium 90 is superior to iodine 131 as a cytotoxic label.

MEASUREMENT OF WHOLE-BODY NITROGEN BY NEUTRON-ACTIVATION ANALYSIS

Abstract Whole-body nitrogen was measured in man using a new technique of neutron-activation analysis. Patients were irradiated with a small dose of fast neutrons and the capture γ rays arising from neutron interactions with nitrogen nuclei were recorded. Sixty-five patients with various diseases have been studied on 190 occasions. The technique has proved to be highly reproducible and offers a new experimental approach to the study of metabolic disorders in clinical medicine.

Prompt gamma neutron activation analysis to measure whole body nitrogen absolutely: Its application to studies of in-vivo changes in body composition in health and disease

Whole body nitrogen has been measured absolutely in male volunteers and patients by in-vivo neutron activation analysis using whole body hydrogen as an internal standard. The 10.8 MeV and 2.2 MeV prompt gamma rays from nitrogen and hydrogen respectively give a result reproducible to 4% for a dose of 100 mRem. Whole body potassium measured by whole body counting natural40K and whole body nitrogen have been correlated in normal adult males and patients. In the normals the correlation coefficient was 0.96 with coefficient of variation 4%. In the patients these parameters were 0.92 and 8% in 140 measurements. The ratio of N/K increased significantly as the degree of clinical wasting progressed.

Effect of Acetazolamide on Exercise at Altitude

The life-giving layer of air around the earth is so thin that we stumble to its edge within hours on trains, roads, cable cars and even walking. One percent of the worlds population reside above 3300rn where atmospheric pressure is reduced by one-third. By 5500m, pressure is halved and long term survival is impossible. Nonetheless, climbers are able to ascend and briefly survive on Mt Everest at 8848m where pressures are less than onethird of normal. These ascents are possible because of short term acclimatisation which increases the oxygen delivery to the tissues. The response depends upon the rate and duration of the ascent and the altitude reached, but if inadequate, mountain sickness develops. Fortunately, the introduction of acetazolamide as a therapeutic agent has altered this situation and assisted in understanding the processes involved. In this leading article we outline the mechanism of normal acclimatisation, describe mountain sickness and then discuss the mechanism of action and role of acetazolamide for mountain travellers.

Computer Analysis of Antibody Scans

Abstract A variety of tumours can be detected with radiolabelled antibodies and a gamma camera. Unfortunately, Low count rates and poor contrast combine to produce images that are often difficult to interpret. Subtraction, using a second isotope, may induce further artifacts. We have used automatic techniques to avoid observer bias, estimate statistical significance, and enhance resolution. Three separate techniques have been used: Firstly, the variance in the subtraction image was calculated and two standard deviations removed for each pixel and only remaining counts were regarded as significant. Secondly, noise was removed by processing with a Wiener filter and the result expressed in contour plots, each stepped in units of standard deviation. Thirdly, a non-linear deconvolution technique (maximum entropy) was used to remove both noise and camera blurring. Subtraction can then be effected using a local normalising factor, and lesions assessed by their statistical significance and shape. These techniques allow objective assessment of scans as well as improving resolution and sensitivity.

Characteristics of First and Second Antibodies for Tumour Imaging

Abstract Radiolabelled anti-tumour antibodies (1-Ab) can be cleared from non-tumour areas using a 2nd antibody (2-Ab) to give enhanced tumour:normal tissue ratios and improved tumour detection. Rabbit, mouse and pig IgGs showed higher affinities for human Fc receptors, and were considered potential 2nd antibodies, since immune complexes containing them are rapidly catabolised. In contrast, sheep and chicken antibodies showed no Fc receptor uptake and were considered useful locating antibodies, but of no value as second antibodies. 5 patients were given radiolabelled anti CEA and pig anti sheep IgG 24h later, resulting in a two-fold fall in blood counts, without a large fall in tumour counts. In contrast, patients given mouse IgM and sheep 2-Ab showed no clearance, as predicted by Fc receptor studies.

Clinical evaluation of anti-alpha-fetoprotein radioimmunodetection

Anti-alpha-fetoprotein (AFP) radioimmunodetection was performed in response to clinical requests in 16 patients. In two patients, assessment of a known tumour was required; the anti-AFP scans were accurate and provided useful clinical information in both cases. In the remaining 14 patients the request was for localisation of suspected recurrent tumour. Accurate information was provided in four of these patients. In this latter group, various conventional methods of investigation had failed to disclose the site of recurrence. However, of a total of 21 sites reported as positive in these 14 patients, eight proved to be false positives. Two false negative results also occurred in this group and nine could not be evaluated. Although occasionally patients were usefully scanned, improvements are necessary before consistently reliable information can be obtained using this technique.

Objective Interpretation of Antibody Scans

Abstract Subtraction techniques are often used in radioimmunodetection, seeking increased contrast in the gamma camera images. To reduce subjectivity in interpretation, we routinely threshold subtraction pictures so as to display significant counts, two standard deviations above background. Scans have been processed by non-linear deconvolution to improve the signal to noise ratio but there was little effect upon a series of scans with 131-1 labelled anti-insulin. Results were displayed as contour plots, stepped in standard deviations of background, and contour shape analysis demonstrated some areas of significant uptake, subsequently identified as tumour. This process may be of more value for the larger point spread functions of more sensitive (wider-holed) collimators.

Preparation of 111-Indium Labelled Antibodies

Abstract DTPA was covalently coupled to sheep anti CEA using a mixed anhydride. This was formed by mixing DTPA and a trace amount of 14-C-DTPA with triethylamine to form the triethylammonium salt, then isobutyl chloroformate to form the anhydride. The anti CEA was then covalently linked to the anhydride in 0.1M bicarbonate buffer, pH 9.5, and non-covalently bound DTPA removed by gel filtration on Sephadex G25 in 0.01M acetate buffer, pH 6. Polyvalent metal ions were removed from all buffers by extraction with dithizone. 74MBq of 111-In chloride was neutralised with acetate (pH 6.5), mixed with 0.8 mg of chelated antibody and left at 4°C overnight. Unbound 111-In was removed by gel filtration. Labelling efficiency was 50-75%, giving a specific activity of up to 74 MBq/mg protein. The chelated antibody formed stable complexes with indium. Ill-indium labelled antibodies have proved superior to iodinated antibodies for in vivo tumour imaging purposes.

Limitations in Localizing and Killing Tumors Using Radiolabelled Antibodies

Radiolabelled antibodies against tumours are being studied intensely for their use in tumour detection and tumour destruction. Although a host of different tumour types can be localised, results at present are no better than other scanning techniques. In addition no tumour has been eradicated using labelled antibodies. Dramatic improvements will depend upon a fuller understanding of tumour cell biology and optimising all the parameters involved.