Zyflamend®, a Unique Herbal Preparation With Nonselective OX Inhibitory Activity, Induces Apoptosis of Prostate Cancer Cells That Lack COX-2 Expression

Abstract

Cyclooxygenase (COX) inhibitors have suppressive effects on several types of cancer cells including prostate cancer. In this study, we considered the potential COX-inhibitory activity of a unique anti-inflammatory herbal preparation (Zyflamend; New Chapter, Inc., Brattleboro, VT) and analyzed its effects on the human prostate cancer cell line LNCaP. COX inhibitory activity of Zyflamend was determined by a spectrophotometric-based assay using purified ovine COX-1 and COX-2 enzymes. Effects of Zyflamend on LNCaP cell growth and apoptosis in vitro were assessed by cell counting, Western blot detection of poly ADP-ribose polymerase (PARP) cleavage, and measurement of caspase-3 activity in treated and control cell extracts. Western blotting techniques were conducted to determine the effects of this herbal preparation on the expression of the cell signaling proteins, p21, androgen receptor (AR), phospho-protein kinase C (pPKC)(alpha/beta), and phospho (p)Stat3. The phospohorylation status of several signal transduction phosphoproteins was profiled using a high-throughput phosphoprotein screening assay in treated cells and compared to controls. Zyflamend dramatically decreased COX-1 and COX-2 enzymatic activity. Elevated p21 expression coincided with attenuated cell growth following treatment of LNCaP cells with Zyflamend. PARP cleavage fragments were evident, and caspase-3 activity was upregulated over the control indicating the ability of Zyflamend to induce apoptosis of these cells. Androgen receptor expression levels declined by 40%, and decreases were observed in the active forms of Stat3 and PKC(alpha/beta) in Zyflamend-treated LNCaP cells. Zyflamend inhibited both COX-1 and COX-2 enzymatic activities, suppressed cell growth, and induced apoptosis in LNCaP cells. However, our data suggests that the effects are likely due to COX-independent mechanisms potentially involving enhanced expression of p21 and reduced expression of AR, pStat3, and pPKC(alpha/beta).

Zyflamend®-mediated inhibition of human prostate cancer PC3 cell proliferation: Effects on 12-LOX and Rb protein phosphorylation

“…6,23 Sandur et al 24 found that Zyflamend inhibited activation of NFkB and consequently NFkBregulated gene products involved in inhibition of apoptosis as well as genes associated with angiogenesis. Further work by this group has shown the in vitro effects of Zyflamend in suppressing osteoclastogenesis and inhibiting tumor cell invasion.…”

“…In an examination of the possible mechanisms of action of Zyflamend, Bemis et al convincingly showed that although Zyflamend could inhibit COX-2 activity and thus lower PGE 2 production, the growth-inhibitory activity against human prostate cancer LNCaP cells was not due to suppression of COX-2 activity, since these cells lacked COX-2 enzyme expression. 6 To fully appreciate the antiproliferative effect of Zyflamend on hormone-refractory PC3 cells, we examined its effect on eicosanoid metabolism. Similar to Bemis et al, 6 we found that Zyflamend was inhibitory to cloned COX-1 and COX-2 enzyme activities; in addition, an even more potent inhibition of cloned 5-LOX activity was noted (data not shown).…”

“…6 To fully appreciate the antiproliferative effect of Zyflamend on hormone-refractory PC3 cells, we examined its effect on eicosanoid metabolism. Similar to Bemis et al, 6 we found that Zyflamend was inhibitory to cloned COX-1 and COX-2 enzyme activities; in addition, an even more potent inhibition of cloned 5-LOX activity was noted (data not shown). Notably, Zyflamend showed much stronger inhibition of the formation of 12-HETE in addition to the reduction of PGE 2 and 5-HETE levels in PC3 cells.…”

“…While specific dietary components such as lycopene from cooked tomatoes, resveratrol from grapes and red wine, and isothiocyanates from cruciferous vegetables have been shown to provide at least limited protection against prostate cancer development, 2-5 the use of whole herbs and herbal extracts for the prevention and treatment of prostate cancer is also of great interest. As pointed out by Bemis et al, 6 the anti-inflammatory activity of many plants available as intact products or as extracts has long been considered beneficial for prevention of prostate cancer. Their recent research has indicated that Zyflamend ® , a unique herbal preparation consisting of ten concentrated herbal extracts (rosemary, tumeric, ginger, holy basil, green tea, hu zhang, Chinese goldthread, barberry, oregano and Scutellaria baicalensis [skullcap]), contains ingredients that significantly inhibit both cyclooxygenase (COX)-1 and COX-2 activities and is associated with induction of apoptosis in LNCap cells.…”

“…This study, however, suggested that COX-independent mechanisms such as enhanced expression of p21 and reduced expression of androgen receptor, pStat3 and pKBC a/b also contributed to the observed inhibition of prostate tumor cell proliferation in this hormonally sensitive tumor cell model. 6 Many bioactive lipids are involved in inflammation associated with prostatitis and early stage prostate cancer. These include the well-documented ability of PGE 2 to stimulate tumor cell proliferation.…”