**1. Introduction**

560 Selected Topics in DNA Repair

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analysis of base damage clustering associated with a site-specific radiation-induced

Here this presentation embraces dietary supplement compositions containing resveratrol material, carotenoid material, nicotinamide material, DMAE material, zinc source material, and qjuinic acid-containing material , where no other known bioactive nutrient agents having competing modes of action to these specified agents are intentionally excluded from mixtures containing at least two of these DNA repair enhancing ingredients. The compositions may be embodied in formulations for oral administration, or alternatively, in formulations for peritoneal administration.

The combined composition may be selected from the group consisting of resveratrol (3, 5, 4' trihydroxy-stilbene or an equivalent polyphenol in pure chemical form); the carotenoid material may be alpha carotene, beta carotene, canthaxanthin, lycopene and mixtures thereof; the nicotinamide material may be selected from the group consisting of nicotinamide, niacin, and mixtures thereof; the DMAE material selected from a group consisting of other choline analogs that pass the blood brain barrier; the zinc source material may be one or more zinc salts; and the quinic acid-containing material selected from a group consisting of quinic acid compounds that can enhance DNA repair by enhancing the uptake of tryptophan and nicotinamide ingredients (Pero et al 2009b; Pero and Lund 2011).

For human administration, the resveratrol material, carotenoid material, nicotinamide material, zinc source material, DMAE material, and quinic acid material may be present in proportions effective, in combination, to improve resistance to DNA damage, enhance DNA repair capacity, and stimulate immune function in a human subject to whom the composition is administered as a daily dosage (Pero et al 2009b; Pero and Lund 2011).

This formulation named Nutra-Reservatrol (Pero and Garret 2010) also contemplates the provision of a method of treating a human or other animal subject, consisting of administering resveratrol material, carotenoid material, nicotinamide material, DMAE material, zinc source material and quinic acid material to the subject to selectively supplement the subject's dietary intake thereof (i.e. without supplementing the dietary intake of any other active nutrient agents having competing modes of action) and repeating the administration on a substantially daily basis.

Enhancing DNA Repair by Combining only Dietary Supplement

additional nutrient or natural product supplements.

example, an inhibited one.

**2. Scientific history** 

2008).

learned from are as follows:

Ingredients that do not Metabolically Compete in Order to Achieve Synergism 563

combination above dietary levels, but not when co-administered together with other

The design of this previous study to prove the discovery was based on combining substances with known properties to prevent cancer and stimulate immune function, but with differing mechanisms of action; e.g. carotenoids = electrophilic scavenger of radicals produced endogenously by cells or exogenously by the environment, nicotinamide = amplified source of energy via increased production of NAD or ATP, and zinc = an essential cofactor to antioxidant, replicative and DNA repair enzymes in cells. The hypothesis was that since none of these substances have produced consistent effects in humans as a single administered agent, this shortcoming could be overcome when administered in combination because these substances might produce a consistently additive or synergistic chemopreventive biological response because of non-competitive modes of action instead of, for

In 2010 there was an extensive review article published entitled "Historic development of Uncaria preparations and their related bioactive components" (Pero 2010b). This overview has specific relevance to the current review because it was the first recognition of the concept that ingredients could in fact convey properties of enhancement of DNA repair. Before this time, there were no clear cut examples where the process of DNA repair could be shown to be stimulated to higher levels of activity by exogenous nutrients or supplements in our environment. Previously it was believed that DNA repair which regulated our genetic integrity could not afford to be anything less than perfect to satisfy the requirements of orderly evolutionary change. Now it is quite accepted that even genes need to have nutritive

Uncaria sp. is a well known herbal medicine used for generations by the Ashinka Indians native to the Amazon basin. There have been two sets of bioactive ingredients for which Uncaria extracts have been developed and standardized. The first are oxindole alkaloids, initially studied and described in 1967; the second are a set of molecules know as Carboxy Alkyl Esters (CAEs ™) first identified and described in 1997 as the bioactive ingredients in AC-11® (Reviewed in Pero 2010b). More recently in 2005 (Sheng et al 2005) it was shown that one of the acid moieties of CAEs is quinic acid, and it is now documented to be one of the more effective DNA repair ingredients found in Uncaria spp, or brightly-colored berry extracts that also contain quinic acid (Stoner et al 2008, Pero 2006), and in turn can enhance DNA repair. It is important to remember that quinic acid is a natural-occurring alpha hydroxy organic acid quite ubiquitous in berries, and also the metabolic source of all other aromatic compound production in plants via the shikimate pathway (Pero 2010a); e.g. the bioactive agents in berries such as hydroxy organic acids (hydroxy benzoic, hydroxy cinnamonic and caffeic acids), flavinoids, and ellagic acid (Stoner et al

The progression of events that established Cat's Claw extracts as the most consistent and potent DNA repair enhancer of anti-aging effects has become obvious, and signaled why some elements are built of the knowledge of the later events that have happened, to paint a more complete picture of how DNA repair regulates aging. A chronology of events that remain unbroken and additive of each other, that Pero and colleagues have in turn built and

treatment, functionality repaired and developed to maturity during life.

Thus, in a particular sense, this dietary supplement combination contemplates the provision of a method of treating a human subject consisting of selectively administering to the subject resveratrol material, carotenoid material, nicotinamide material, DMAE material, zinc source and quinic acid material in daily dosage amounts effective, in combination, to improve resistance to DNA damage, enhance DNA repair capacity, and stimulate immune function. In a specific example of currently preferred dosage range for humans, about 100- 500 mg resveratol material, about 100 mg of carotenoid material, about 100 mg of nicotinamide material, about 100-200 mg DMAE material, about 10 mg of zinc source material and between 250-700 mg quinic acid source material are administered daily in this method (Pero and Garret 2010). So far as the author is aware, this particular combination of ingredients devoid of other bioactives has heretofore already been recognized as being synergistic or even effective (Pero 2000, Sheng et al 1998).

The discovery that natural products should not be combined into a natural medicine unless one tests whether each ingredient is additive to the overall desired biological effect, and that one way to accomplish this endpoint is to not combine natural products that have similar modes of action and thus competitive routes of absorption and excretion without first testing the combination for additive effects. That is to say, the present invention avoids inhibited uptake and absorption of natural products, thereby obtaining additive biological effects, by combining only natural products having well defined different, and thus potentially non-competitive modes of action which is, for example, the case with the exclusive combination of carotenoids + nicotinamide + zinc (Pero 2000, Sheng et al 1998).

Diet supplementation of humans or animals for example, by the oral, intraperitoneal, intravenous, subcutaneous or intramuscular routes of administration with the combination of carotenoids + nicotinamide/niacin + an appropriate zinc salt at a dose of this combination that exceeds a normal dietary levels was effective. This practice showed that dietary supplementation containing this combination together with simultaneous supplementation of other nutrients and/or natural products cannot enhance immune function (Payette, H. et al 1990; Zhang et al 1995), but when daily doses of carotenoids (as lycopene at 20 mg and Vitamin E 36 IU), niacin (120 mg) zinc salt (12 mg), reservatol (300 mg) and a quinic acid material (400 mg) were administered in the absence of other known DNA repair enhancers, and above dietary levels, the resistance to oxidative cellular DNA damage, and enhancement of DNA repair and immune function were observed in the clinic (Pero and Lund 2011). These data were taken as proof of concept for this dietary supplement to avoid metabolic competition and synergize DNA repair.

A clinical evaluation already published (Pero 2000, Sheng et al 1998) was also determined by comparing each individual's biological response before and after supplementation. In such a manner, each individual became his own control; e.g. the male subjects were given baseline measurements of resistance to cellular DNA damage, enhancement of DNA repair and stimulation of immune function once a week for 4 weeks, and then they were supplemented daily and the same measurements repeated once a week for the last 5 weeks of a 7 week intervention period. The before measurements (i.e. n = 4) were the baseline biological response parameters to be compared to the after measurements (i.e. n = 5). One individual was not supplemented to provide a control for the supplemented individuals. The data from this experimental design has taught that resistance to cellular DNA damage, enhancement of DNA repair and stimulation of immune function were all significantly modulated by a combination of carotenoids + nicotinamide + zinc when administered as an exclusive drug combination above dietary levels, but not when co-administered together with other additional nutrient or natural product supplements.

The design of this previous study to prove the discovery was based on combining substances with known properties to prevent cancer and stimulate immune function, but with differing mechanisms of action; e.g. carotenoids = electrophilic scavenger of radicals produced endogenously by cells or exogenously by the environment, nicotinamide = amplified source of energy via increased production of NAD or ATP, and zinc = an essential cofactor to antioxidant, replicative and DNA repair enzymes in cells. The hypothesis was that since none of these substances have produced consistent effects in humans as a single administered agent, this shortcoming could be overcome when administered in combination because these substances might produce a consistently additive or synergistic chemopreventive biological response because of non-competitive modes of action instead of, for example, an inhibited one.
