Dr Bence Jones and the history of free light chains

From Wikilite

Figure 2.1. Henry Bence Jones. (A) Albumen print by Maull c.1850s (B) Charcoal and chalk on paper by Richmond 1865 (reproduced with permission from The Royal Institution, London and The Bridgeman Art Library).

Although immunoglobulin FLCs are synonymous with Bence Jones proteins, history might have been more generous to others involved in their discovery ^{[1][2][3][4][5]}.

On Friday, October 30^th 1845, the 53 year old Dr William MacIntyre, physician to the Western General Dispensary, St. Marylebone, London, left his rooms in Harley Street. He had been called to see Mr Thomas Alexander McBean, a 45 year-old, highly respectable grocer, who had severe bone pain and fractures. He had been under the care of his general practitioner, Dr Thomas Watson, for several months. Upon examination of the patient, William MacIntyre noted the presence of oedema. Considering the possibility of nephrosis, he tested the urine for albumin. To his consternation, the albuminous protein precipitate found on warming the urine, uncharacteristically, redissolved when heated to 75°C.

Both Dr MacIntyre and Dr Watson then sent urine samples to the chemical pathologist at St.George's Hospital. A note accompanying the urine sent by Dr Watson read as follows:-

"Dear Dr Bence Jones, The tube contains urine of very high specific gravity. When boiled it becomes highly opaque. On the addition of nitric acid, it effervesces, assumes a reddish hue, and becomes quite clear; but as it cools assumes the consistence and appearance which you see. Heat reliquifies it. What is it?"

Over the next two months, the patient deteriorated, became emaciated, weak, and was racked with pain. He eventually died on January 1st 1846, in full possession of his mental faculties. Dr MacIntyre subsequently published the post-mortem examination and the description of the peculiar urine in 1850 ^[6]. Unfortunately for him, Henry Bence Jones ^[7][8] (Figure 2.1) had already described the patient’s urinary findings in two, single author articles, one of which was published in The Lancet, in 1847. He considered the protein to be an “hydrated deutoxide of albumen”. He wisely commented:

“I need hardly remark on the importance of seeking for this oxide of albumen in other cases of mollities ossium”.

Bence Jones’s reputation was assured, while the contributions of his colleagues were consigned to the footnotes of history.

For all the apparent injustice to his colleague,William MacIntyre, Henry Bence Jones achieved much else in his career. He published over 40 papers and became rich and famous based on his clinical practice, lecturing and original observations and was elected to fellowship of The Royal Society at the tender age of 33. Florence Nightingale once described him as “the best chemical doctor in London.” Surprisingly, there was no mention of Bence Jones protein in his obituary and the eponym (and the hyphen in his name) was not used until after his death ^[2].

By 1909, over 40 cases of Bence Jones proteinuria had been reported ^[9], and the protein was thought to originate in bone marrow plasma cells that were first identified by Waldeyer in 1875. In 1922, Bayne-Jones and Wilson characterised two types of Bence Jones protein by observing precipitation reactions using antisera made by immunising rabbits with the urine of several patients. The proteins were classified as group I and group II types. However, it was not until 1956 that Korngold and Lapiri, using the Ouchterlony technique, showed that antisera raised against the different groups also reacted with myeloma proteins. As a tribute to their observations the two types of Bence Jones protein were designated kappa and lambda (κ and λ). Edelman and Gally, in 1962, subsequently showed that FLCs prepared from IgG monoclonal proteins were the same as Bence Jones protein. It had taken 117 years from the original observation for the function of Bence Jones protein to be finally determined. Remarkably, the format of the urine test had remained unchanged for a similar period. The following is a protocol from Levinson and MacFate's “Clinical Laboratory Diagnosis”, a standard textbook published in 1946.

In parallel with the clinical and scientific observations of the role of Bence Jones protein, electrophoretic techniques for protein separation were entering clinical laboratories. Longsworth et al., in 1939, recognised tall, narrow-based, “church spire” peaks in the sera of patients with MM, using moving boundary protein electrophoresis. Electrophoresis was subsequently improved by using paper as a substrate followed by cellulose acetate then agarose in the 1950's and 1960's. Finally, immunofixation electrophoresis became established in the 1980's ^[10].

Clear identification of κ and λ molecules was possible with the use of antibodies specific for each type of protein. Immunodiffusion was initially used ^[11], followed by immunoelectrophoresis in 1953 ^[12], radial immunodiffusion and ultimately nephelometry and turbidimetry. However, serum assays for Bence Jones protein (serum free light chains - sFLCs) remained unattainable because the antibodies could not distinguish between sFLCs and the overwhelming amounts of light chains bound in intact immunoglobulin molecules.

The first successful attempt to measure FLCs in serum was in 1975. Size-separation column chromatography ^[13][14][15], was used to isolate them from intact immunoglobulins, prior to analysis. Although the results were accurate and showed the potential use of serum analyses, these assays were clearly impractical for routine use.

Subsequent assays focussed on the use of antibodies directed against “hidden” epitopes on FLC molecules. These are located at the interface between the light and heavy chains of intact immunoglobulins and become detectable when the FLCs are unbound. Radioimmunoassays and enzyme immunoassays using polyclonal antisera against FLCs were used to analyse urine samples but specificity remained inadequate for serum measurements ^[16][17] and variations in FLC polymerisation caused measurement errors ^[18][19].

The use of monoclonal antibodies was an obvious approach to improving specificity but satisfactory reagents were difficult to develop ^{[20][21][22][23]} and their use was restricted to radio-immunoassays and enzyme immunoassays. Attempts were made to develop turbidimetric ^[24][25] and latex-enhanced nephelometric assays ^[26], using polyclonal antibodies but they could not detect normal sFLC concentrations, and cross-reactions with intact immunoglobulins were unacceptable. In 2001, immunoassays, based on polyclonal antibodies, were finally developed that could measure FLCs at normal serum concentrations ^[27]. Their utility was quickly made apparent when monoclonal FLCs were detected in the sera of most patients classified as having “nonsecretory myeloma” ^[28]. Furthermore, as described in The Lancet, all of 224 patients with LCMM who had Bence Jones proteinuria, also had elevated sFLC concentrations ^[29]. The serum tests were also better at detecting residual disease than urinalysis. Further studies have shown that sFLC assays can be used for screening symptomatic patients (Chapter 23) , are more sensitive than urine tests (Chapter 24) and are markers for progression in MGUS (Chapter 19).

These results and many others, herald the widespread use of sFLC immunoassays. The enduring story of Bence Jones protein and Bence Jones proteinuria may be entering its final chapter, 160 years after it began.

References