Study aims to perfect molecules with anti-tumor action

Approximately 150 compounds were synthesized based on substances found in nature.

By Karina Toledo

Agência FAPESP – Inspired by natural compounds with anti-tumor action described in scientific literature, scientists from the Institute of Chemistry at Universidade Estadual de Campinas (IQ-Unicamp) have synthesized approximately 150 molecules that are being tested in cancer cell lines and animal models.

The researchers’ objectives are to uncover the mechanism of action of these molecules, with the aim of selecting the most promising candidates, and to discover what types of structural modifications can be made to increase their medicinal effects and minimize possible toxicity.

The study is being developed as part of a Thematic Project coordinated by Professor Ronaldo Aloise Pilli, of IQ-Unicamp. Some of the results were presented during the São Paulo School of Advanced Science on Bioorganic Chemistry, a FAPESP-funded event held in Araraquara at the beginning of July.

According to Pilli, the vast majority of the synthesized compounds in the library at Unicamp were inspired by natural substances. One such natural substance, piplartine, is isolated from Piper tuberculatum, which can be found throughout northeast Brazil. Another natural substance, goniothalamin, can be extracted from the roots and stems of Goniothalamus macrophyllus, a shrub found in Asia.

“There are groups conducting tests with natural versions of these compounds, but the major advantage of chemical synthesis is that we can make the compounds even more efficient through structural modifications. Our objective is to discover the principles that will be the starting point for creating new medications,” affirmed Pilli.

The functionalities of these molecules are being tested on breast, ovarian, lung, kidney, skin (melanoma), brain (glioma), colon (intestine), pancreas and prostate cancer cell lines. To date, according to Pilli, six synthetic compounds have shown promising effects against breast, pancreas and prostate tumors. These results led the team to begin tests in animals, although such testing was not originally planned.

In an experiment described in Bioorganic & Medicinal Chemistry, the researchers implanted an extremely aggressive and rapidly growing breast adenocarcinoma, known as the Ehrlich tumor, into animals.

Some of the animals were treated with goniothalamin, a chemical produced in the lab, while the others were treated with 5-fluorouracil, which is used in clinical chemotherapy. After fifteen days, growth inhibition of the malignant cells and reduction of the tumor mass was comparable in the two groups, without any evidence of acute toxicity in the animals treated with the synthetic product.

In another experimental model described in the same article, the researchers induced edema in mice feet with a substance called carrageenan. In this case, the synthetic version of goniothalamin helped to reduce inflammation.

“We are attempting to discover the mechanism by which these substances inhibit the growth tumor cell colonies. We want to determine the stage of cellular division that these compounds interfere with, and we want to discover if they interact with a membrane protein or whether they enter the cell and alter some cytoplasmic or nuclear structure. This knowledge could help us to perfect the molecules,” explained Pilli.

Other targets

The Thematic Project also seeks to investigate the efficacy of synthetic compounds for the inhibition of two classes of proteins that participate in the regulation of several cell functions: phosphatases and chaperones.

“For a protein to perform its role in an organism, it needs to be in the correct form. A protein’s function is associated with its three-dimensional form,” explained Pilli.

The chaperones, added the researcher, help other proteins to fold, or rather mold their amino acid chains to achieve a functional form. Phosphatases, on the other hand, are enzymes that dephosphorylate other proteins; in other words, they remove a phosphate group from a molecule. The opposite role is performed by kinases, which are enzymes that add a phosphate group to another protein (phosphorylation).

“Some diseases are triggered because one or more proteins remain phosphorylated for longer than they should. Other diseases arise because the proteins were dephosphorylated when they should not have been. That’s why it would be interesting to identify molecules capable of modulating the action of phosphatases. PTB-1B is highly expressed in several malignant tumors, such as those found in the breast and ovary, whereas CDC-25b is associated with deviations in cell cycle checkpoints and genetic stability. Overexpression of LMW-PTP was observed in several neoplasms and could be related to the malignancy and aggressiveness of tumors or other metastatic processes,” explained Pilli.

Through in vitro experiments, the researchers examined 150 compounds and selected those that had the highest capacity to inhibit the expression of chaperones and phosphatases.

“The next step is to attempt co-crystallization of the compound and the protein, or rather, to unite the two together such that they can be examined by X-ray crystallography. This will allow us to see how our molecules fit into proteins and what types of modifications we can make to create even better inhibitors,” said Pilli.

In a study published in PLoS One, the researchers studied the involvement of LMW-PT in the resistance of chronic myeloid leukemia cells to multiple drugs.

According to Pilli, silencing the enzyme restored the sensitivity of malignant cells to two chemotherapy drugs: vincristine and imatinib mesylate. “The results suggest that LMW-PTP inhibition could be a therapeutic strategy worth exploring,” commented Pilli.

The thematic group is formed by nine researchers with expertise in several areas, including chemistry, biochemistry, pharmacology and structural biology. The group also includes thirty undergraduate and graduate students and five post-doctoral scholars. The studies conducted to date have resulted in the publication of nearly one hundred articles in scientific magazines.

“The range of activities is expanding as the studies are developed further. We have begun forming international partnerships, and we are partnering with the Research Center on Diabetes, Obesity and Comorbidities (one of the new Research, Innovation and Dissemination Centers - CEPIDs). The project is helping to train future generations of researchers who will know how to be active in the interface between organic synthesis, cellular biology and structural biology,” explained Pilli.

The article Effect of goniothalamin on the development of Ehrlich solid tumor in mice, can be accessed at www.sigmaaldrich.com/catalog/papers/20729093.

The article Knocking Down Low Molecular Weight Protein Tyrosine Phosphatase (LMW-PTP) Reverts Chemoresistance through Inactivation of Src and Bcr-Abl Proteins (doi: 10.1371/journal.pone.0044312), can be accessed at www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0044312.