Mark R. Burns

Combination therapy with 2-difluoromethylornithine and a polyamine transport inhibitor against murine squamous cell carcinoma

Using a recently developed autochthonous mouse model of squamous cell carcinoma (SCC), a combination therapy targeting polyamine metabolism was evaluated. The therapy combined 2‐difluoromethylornithine (DFMO), an inhibitor of ornithine decarboxylase (ODC), and MQT 1426, a polyamine transport inhibitor. In 1 trial lasting 4 weeks, combination therapy with 0.5% DFMO (orally, in the drinking water) and MQT 1426 (50 mg/kg i.p., bid) was significantly more effective than with either single agent alone when complete tumor response was the endpoint. In the combination group, 72% of SCCs responded completely vs. 21 and 0% for DFMO and MQT 1426, respectively. A second trial involved a 4‐week treatment period followed by 6 weeks off‐treatment. With apparent cures as an endpoint, combination therapy was again more effective than either agent alone: a 50% apparent cure rate was observed in the combination group vs. 7.7% in the DFMO group. MQT 1426 had no inhibitory effect on SCC ODC activity nor did it enhance the inhibition by DFMO, but SCC polyamine levels declined more rapidly when treated with combination therapy vs. DFMO alone. The apoptotic index in SCCs was transiently increased by combination therapy but not by DFMO alone. Thus, targeting both polyamine biosynthesis and polyamine transport from the tumor microenvironment enhances the efficacy of polyamine‐based therapy in this mouse model of SCC.

AMXT‐1501, a novel polyamine transport inhibitor, synergizes with DFMO in inhibiting neuroblastoma cell proliferation by targeting both ornithine decarboxylase and polyamine transport

Neuroblastoma (NB) is associated with MYCN oncogene amplification occurring in approximately 30% of NBs and is associated with poor prognosis. MYCN is linked to a number of genes including ornithine decarboxylase (ODC), the rate‐limiting enzyme in polyamine biosynthesis. ODC expression is elevated in many forms of cancer including NB. Alpha‐difluoromethylornithine (DFMO), an ODC inhibitor, is currently being used in a Phase I clinical trial for treatment of NB. However, cancer cells treated with DFMO may overcome their polyamine depletion by the uptake of polyamines from extracellular sources. A novel polyamine transport inhibitor, AMXT‐1501, has not yet been tested in NB. We propose that inhibiting ODC with DFMO, coupled with polyamine transport inhibition by AMXT‐1501 will result in enhanced NB growth inhibition. Single and combination drug treatments were conducted on three NB cell lines. DFMO IC50 values ranged from 20.76 to 33.3 mM, and AMXT‐1501 IC50 values ranged from 14.13 to 17.72 µM in NB. The combination treatment resulted in hypophosphorylation of retinoblastoma protein (Rb), suggesting growth inhibition via G1 cell cycle arrest. Increased expression of cleaved PARP and cleaved caspase 3 in combination‐treated cells starting at 48 hr suggested apoptosis. The combination treatment depleted intracellular polyamine pools and decreased intracellular ATP, further verifying growth inhibition. Given the current lack of effective therapies for patients with relapsed/refractory NB and the preclinical effectiveness of DFMO with AMXT‐1501, this combination treatment provides promising preclinical results. DFMO and AMXT‐1501 may be a potential new therapy for children with NB.

Synthesis of bis-spermine dimers that are potent polyamine transport inhibitors

A series of novel spermine dimer analogues was synthesized and assessed for their ability to inhibit spermidine transport into MDA-MB-231 breast carcinoma cells. Two spermine molecules were tethered via their N(1) primary amines with naphthalenedisulfonic acid, adamantanedicarboxylic acid and a series of aliphatic dicarboxylic acids. The linked spermine analogues were potent polyamine transport inhibitors and inhibited cell growth cytostatically in combination with a polyamine synthesis inhibitor. Variation in the linker length did not alter polyamine transport inhibition. The amount of charge on the molecule may influence the molecular interaction with the transporter since the most potent spermidine transport inhibitors contained 5-6 positive charges.

Induction of apoptosis by aryl-substituted diamines: role of aromatic group substituents and distance between nitrogens.

A series of aromatic substituted diamines was synthesized and characterized for their cytotoxic profiles against human breast and prostate tumor cell lines. Following a structure function analysis of the effects of changes of the benzyl substituents and the distance between amino groups the most potent analogues were analyzed biologically and were shown to induce apoptosis. These compounds do not induce the enzyme SSAT or deplete intracellular polyamine levels, mechanisms demonstrated by other cytotoxic polyamine analogues.