“The presentations impressed upon me how much witchcraft and how little science has gone into the study of cryoprotectant agents (CPAs). This might be understandable in light of the fact that most cryobiologists are, in fact, biologists. I suspect that a great deal could be accomplished by a thorough study of the physics of the chemistry of CPAs.”
Such an observation could equally apply to the study of neuroprotectants in cerebral ischemia. There has been a growing literature investigating the potential of numerous molecules for the treatment of stroke and cardiac arrest. Although some approaches have been more successful than others, systematic reviews of the chemical and physical characteristics of effective drugs are lacking and discussion of the topic is often confined to isolated remarks.
A number of examples:
“It is not surprising that all the agents which are effective in shock carry a negative charge. This applies both to heparin, which possesses a very strong negative charge, and to hypertonic glucose solution. The same may be said about a substance now in wide use – dextran – which has small, negatively charged molecules, and also about the glucocorticoids 21, 17, and 11, which also have a negative charge.” – Professor Laborit in: Acute problems in resuscitation and hypothermia; proceedings of a symposium on the application of deep hypothermia in terminal states, September 15-19, 1964. Edited by V. A. Negovskii.
“We report that estrogen and estrogen derivatives within the hydroxyl group in the C3 position on the A ring of the steroid molecule can also act as powerful neuroprotectants in an estrogen-receptor-independent short term manner because of to their antioxidative capacity.” Christian Behl et al. Neuroprotection against Oxidative Stress by Estrogens: Structure-Activity Relationship. Molecular Pharmacology 51:535-541 (1997).
“Minocycline’s direct radical scavenging property is consistent with its chemical structure, which includes a multiply substituted phenol ring similar to alpha-tocopherol (Vitamin E)” – Kraus RL et al. Antioxidant properties of minocycline: neuroprotection in an oxidative stress assay and direct radical-scavenging activity. Journal of Neurochemistry. 2005 Aug;94(3):819-27.
“It is notable, however, that NAD+ and minocycline share a carboxamide and aromatic ring structure. A common structural feature of competitive PARP inhibitors is a carboxamide group attached to an aromatic ring or the carbamoyl group built in a polyaromatic heterocyclic skeleton. This structure is also present in each of the tetracycline derivatives with demonstrated PARP-1 inhibitory activity. Alano CC et al. Minocycline inhibits poly(ADP-ribose) polymerase-1 at nanomolar concentrations. Proceedings of the National Academy of Sciences of the United States of America. 2006 Jun 20;103(25):9685-90.
Systematic study of structure-activity relationship of neuroprotectants would not only contribute towards the development of a general theory of neuroprotection in cerebral ischemia, it would also contribute to the design of multi-functional neuroprotectants. Although it is now increasingly accepted that combination therapy offers more potential for successful treatment of stroke and cardiac arrest than mono-agents, parallel or sequential administration of multiple drugs present non-trivial challenges in research design and clinical application. Such problems may be better addressed by designing molecules with different mechanisms of action in the same structure, an approach that is currently recognized and investigated by forward-looking biomedical researchers.
Although the field of cerebral resuscitation has known some notable researchers like Vladimir Aleksandrovich Negovskii and Peter Safar, who devoted their lives to a thorough study of the mechanisms of cerebral ischemia and its treatment, the field as a whole shows a never ending stream of trial and error publications to investigate yet another drug (before moving on to other areas in neuroscience and medicine). Although there is an increased interest in meta-analysis of all these experiments, meta-analysis that places its findings in a broader biochemical and pharmacological context is rare.
The emphasis on theory and research design can be taken too far. As Nassim Nicholas Taleb recently argued, the role of design in biotechnology is overestimated at the expense of chance observations and unexpected directions. But in the area of cerebral resuscitation the risk of too much theory and systematization is low at this point. As evidenced by the successful development of vitrification agents with low toxicity in cryobiology, a committed long-term and systematical effort to find solutions to human medical needs can pay off.
Despite sensational news items about “zombie dogs,” biomedical researchers and clinicians have known for a long time that interruptions in consciousness and blood circulation can be reversed without neurological deficits, provided such events do not produce ischemic injury. There are even species who can enter a state of reversible metabolic arrest such as tardigrades (water bears). Naturally, researchers have recognized the opportunities that depressed metabolism holds for stabilizing trauma victims. Although the jury is still out on the question if therapeutic hypometabolism can be induced in large animals by chemical means, there are no doubts that lower temperatures reduce metabolism, allowing patients to tolerate longer periods of circulatory arrest.
The published record for reversible hypothermic circulatory arrest is 3 hours in a canine model. A recent study by Wu, Drabek, Tisherman et al. (2008) documented resuscitation from 3 hours of exsanguination cardiac arrest (2.5 hours of no flow) after rapid induction of profound hypothermia using cardiopulmonary bypass. These results are quite impressive in light of the fact that in the latter study cardiac arrest was induced at normal body temperature by exsanguination, and the organ preservation solution to replace the blood consisted of just chilled saline plus dissolved oxygen and/or glucose. The authors attribute their success in extending satisfactory neurological recovery from 2 to 3 hours of exsanguination cardiac arrest to the addition of energy substrates, and oxygen in particular, during induction of profound hypothermia.
Research of this nature benefits cryonics in a number of ways. If hypothermic circulatory arrest will become routine in emergency medicine and military medicine, the general public will get increased exposure to the fact that circulatory arrest does not equal death. Research of this nature also demonstrates that induction of ultra-profound hypothermic arrest in humans may be reversible and therefore the initial stages of cryonics stabilization procedures as well. The more practical application is that it offers the prospect of extending the period the brain can be kept viable after pronouncement of legal death during remote transport of cryonics patients. Last but not least, this specific study provides optimism that viability of the brain can be maintained if hypothermia is induced after circulatory arrest, provided metabolic support is given and cooling rates are fast enough to avoid irreversible injury to the brain.
Unfortunately, the technical capability to reverse 3 hours of asanguineous hypothermic arrest falls short of what is needed for cryonics patients who are stabilized in remote locations. Transport times between the location of cardiac arrest and the cryonics facility often exceed 24 hours. Although loss of viability of the brain does not constitute information-theoretic death, it would be desirable if cryonics organizations would be able to routinely secure viability of the brain between pronouncement of legal death and start of cryoprotectant perfusion.
Such advances will require substantial investments into the development and implementation of improved organ preservation solutions, perfusion techniques, and resuscitation protocols. Potential directions for such research include addition of effective neuroprotective compounds and “hibernation mimetics” to the organ preservation solution and low flow or intermittent perfusion during patient transport.
In 2005, when asked to comment on the prospects of using hypothermic circulatory arrest to treat trauma victims, Dr. Thomas Scalea, physician-in-chief at the R. Adams Cowley Shock Trauma Center at the University of Maryland Medical Center, was reported saying:
“As potentially crazy as this might sound, you’re comparing it against essentially certain death, so it’s hard to see how we can do any worse….all of us are incredibly energized by the thought of being able to do better.”
Such reasoning should equally apply to the practice of human cryopreservation, which employs even lower temperatures to protect people against “essentially certain death.”