JS-K Molecule

Nitric oxide (NO) is a gaseous free radical molecule comprised of an atom of nitrogen linked to an oxygen atom with a resulting unpaired electron (1). In the atmosphere, NO is a common air pollutant. However, NO is also produced by the human body as a regulatory molecule that transmits signals within the organism (1).  When NO is produced by one cell, it penetrates through membranes and regulates the function of another cell.  NO is made by the lining of arteries (endothelium) and regulates blood flow by dilating the artery to allow more blood to perfuse the affected organ. When produced by infection fighting cells (macrophages), NO fights bacterial invasion and kills cancer cells. When produced by brain cells (neurons), NO serves as a signal between brain cells (as a neurotransmitter). When produced within all cells, NO can serve as a switch to turn enzymes on and off and to regulate the ability of genes to be expressed and their protein products made.

The biological significance of NO was first discovered in 1977 by Dr. Ferid Murad, who analyzed how nitroglycerin (used to treat angina) and sodium nitroprusside (used to treat high blood pressure) act and found that they release NO, which relaxes the smooth muscle cells in arteries. In 1998, Dr. Murad and two other scientists, Drs. Robert Furchgott and Louis Ignarro, were awarded the Nobel Prize in Medicine for the discovery of NO effects. NO has been found to regulate a plethora of normal physiologic and biochemical functions. In particular, NO has been generally agreed by experts to show great promise for treating cancer (2). Nevertheless, up to this point, no one has been able to develop a NO-based drug for the treatment of cancer.

The discovery that macrophages kill tumor cells by producing NO was made in 1987 by Dr. John Hibbs at the University of Utah (3).  Dr. Paul Shami, Co-founder and Chief Medical Officer of JSK Therapeutics™ (JSKT), began studying this phenomenon in the early 1990s while at Duke University. He found that the NO-producing vasodilator sodium nitroprusside (SNP) was effective at killing leukemia cells in culture (4). SNP cannot be used for this purpose in animals or humans because it releases too much NO directly into the blood stream, producing excessively low blood pressure. The solution to this problem was to deliver NO as part of a pro-drug that might be more selectively activated by cancer cells compared to normal cells.

 In collaboration with Dr. Larry Keefer at the National Cancer Institute, Dr. Shami has developed such a drug using compounds that release NO upon interaction with the cell antioxidant compound glutathione (GSH) in a reaction catalyzed by the cell enzymes Glutathione S-Transferases (GST) (5). This design exploits the fact that GST is present at higher levels in malignant compared to normal cells. After extensive research to determine the best drug candidate, the Shami lab has identified O2-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl) piperazin-1-yl]diazen-1-ium-1,2-diolate, or JS-K, as the most active compound of this family.

 JS-K has been through the Rapid Access to Intervention Development (RAID) program at NCI. JS-K is active against the “NCI 60-cell” screen with a mean 50% growth inhibitory concentration of 1.3 μM.

Despite its effectiveness, JS-K presents two major challenges. First, JS-K can react with components in the blood, releasing NO in the blood stream, where it causes lowering of blood pressure.  Second, JS-K is insoluble in physiologic solutions used to administer drugs. The problems of turning JS-K into a safe intravenous formulation was solved with the use of nanoscale Pluronic® surfactant micelles.

Surfactants are molecules made of a hydrophobic (“hates water, loves fat”) tail and a hydrophilic (“loves water, hates fat”) head (Fig 1). One of the properties of surfactants is the formation of clusters in solution known as micelles. When a certain surfactant level is reached in solution (the critical micelle concentration or CMC), micelles assemble spontaneously with surfactant hydrophobic tails at the center of the micelle and hydrophilic heads at the outer surface of the micelle. Insoluble drugs such as JS-K can be dissolved at the center of micelles.

Micelle formulations also lead to greater drug accumulation in tumors. Unlike normal blood vessels, the blood vessels supplying tumors are “leaky” and have holes ~ 60 nm in size allowing smaller micelle particles to exit and deposit into the cancer. To produce a nanoscale micelle formulation of JS-K, copolymers known as Pluronics®  which are made by BASF (Florham Park, NJ) and generally recognized by the FDA as safe (GRAS) were chosen. Pluronics® form nanoscale micelles with diameters of around 20 - 80 nm, are nontoxic and have been used for cancer drug delivery.

For the formulation of JS-K (P123/JS-K), Pluronic® P123 was used. P123/JS-K offers a number of distinct advantages.

First, when given to mice inoculated with HL-60 human leukemia cells, P123/JS-K was more active against cancer in this animal model than the same amount of naked JS-K given intravenously in a small amount of dimethylsulfoxide (DMSO). 

Second, P123/JS-K did not lower blood pressure in mice when given intravenously at similar doses as the naked drug.

Third, and of great importance, studies by Dr. Shami at the University of Utah showed that P123/JS-K is selectively toxic towards leukemia cells as opposed to normal hematopoietic cells.

 As such, P123/JS-K presents a new paradigm: P123/JS-K can effectively treat cancer without producing the bone marrow side effects that cause many patients to become ill, infected and hospitalized because of standard chemotherapy. Thus, JS-K offers potentially large impact and savings in cancer care from decreasing the cost of hospitalizations needed following infectious complications of current cytotoxic chemotherapy.