Glycosylated oleic acid/Vitamin D-binding protein suppresses HER2 oncogene expression in human breast cancer

Marco Ruggiero, Jacopo Branca, David Noakes, Massimo Gulisano, Gabriele Morucci, Lynda Thyer and Stefania Pacini 2014 

Department of Experimental and Clinical Biomedical Sciences University of Firenze Florence Italy.
Imuno Biotech Ltd.  Correspondence: jacopo.branca@libero.it

Published atx the International Institute of Anticancer Research

Introduction
The healthy properties of Oleic acid (OA) in breast cancer have been known for centuries and recent evidence suggests that these properties are amplified by association of OA with proteins such as alpha-lactalbumin and lactoferrins. These proteins form OA protein complexes that exhibit highly selective anti-tumour activity in vitro and in vivo. 

We recently demonstrated that also a serum protein with the capability to bind OA shows anti-cancer effects: this is the glycosylated vitamin D binding protein also known as GC-protein derived macrophage activating factor or GcMAF. This protein binds both OA and vitamin D and exerts its immune-stimulating and anti-cancer effects through crosstalk with the vitamin D receptor. Here we report a clinical observation suggesting that OA GcMAF, that is GcMAF complexed with OA, suppresses the expression of a major oncogene involved in human breast cancer that is human epidermal growth factor receptor 2 (HER 2)

Results
Amplification or over expression of HER 2 plays an important role in the development and progression of breast-cancer and has become an important biomarker and target of therapy since it is strongly associated with increased disease recurrence and a poor prognosis [8]. Consistent with the aggressive nature of the cancer in this patient, pre-operative biopsy on four specimens collected under ultrasound guidance showed significant positivity for HER 2 assessed by polyclonal antibody A 0485 with more than 10% of positivity and a score of 2+ (Figure 1). After three weeks of OA-GcMAF treatment, and subsequent mastectomy, analysis of the surgical specimen showed no positivity for HER 2 expression (negative score Zero figure 1) this indicating complete suppression of oncogene expression.

Study of the expression of progesterone receptor (PGR clone 1E2) was consistent with such a reversal of the neoplastic phenotype.  PGR expression in the biopsy was low, less than one %, a finding consistent with poor differentiation and aggressiveness. However in the surgical specimen taken after the  3 weeks of treatment with OA-GcMAF PGR expression was significantly increased to 20% (Fig 1). The selectivity of these effects was confirmed by study of the expression of Ki67 and oestrogen receptor (30% and 90% respectively) that did not show any change following OA-GcMAF treatment.  (Fig 1)

Patient and Methods
No drug was administered or was programmed in the three weeks preceeding surgery, nor had the patient received any treatment for the previous eight years. The analyses on HER 2 and other gene expression on the biopsy and surgical specimens were performed by the Laboratory of the University Hospital of Careggi of the Italian Public Health Service in Florence Italy. Analyses were performed according to the European standards of quality (UNI EN ISO 9001:2008), and were examined and countersigned by four different professionals.  The original documents are conserved in the archives of the Department of Biomedicine of the Careggi Hospital (Molecular diagnostic and pathologic histology).s

The patient gave informed consent to the treatment as well as to this description of her results. Surgery was performed at the division of General Surgery n. 2 of the University Hospital of Careggi

Discussion
These results demonstrate that OA-GcMAF administered by subcutaneous injections aerosol or in a functional food products supressed the expression of HER 2, an oncogene which plays a key role in the aetiology, invasive progression and metastases in breast cancer. This effect was paralleled by increase of PGR expression thus indicating that OA-GcMAF treatment induced healthy differentiation of cancer cells. We hypothesise that these multifaceted effects on the regulation of gene expression in human breast cancer are due to the peculiar association of OA with GcMAF that is an association between two molecules endowed with anti-cancer properties.

In fact OA has been shown to down regulate HER2 expression in cancer cell lines and we demonstrate that GcMAF inhibits human breast cancer cell proliferation and reverts their malignant phenotype.We hypotheise that OA-GCMAF, but not OA or GcMAF taken singularly   interacts with the HER 2 protein through hydrophobic interaction between the amino-terminal of GcMAF and the extracellular region of HER 2 and between the OA binding region of GcMAF and the plasma membrane [4]. In fact the stretch of amino acids between position 17-46 of OA-GcMAF, and position 243-273 of HER 2, there is a high density of hydrophobic amino acids that may favour selective binding. Whatever the case, these results indicate that the effects of OA-GcMAF in  cancer are due to multiplicity of actions that involves suppression of oncogene expression.