Conformational Propensity and Biological Studies of Proline Mutated LR Peptides Inhibiting Human Thymidylate Synthase and Ovarian Cancer Cell Growth

Abstract

LR and [d-Gln]LR peptides bind the monomer-monomer interface of human thymidylate synthase and inhibit cancer cell growth. Here, proline-mutated LR peptides were synthesized. Molecular dynamics calculations and circular dichroism spectra have provided a consistent picture of the conformational propensities of the [Pro ]-peptides. [Pro]LR and [Pro]LR show improved cell growth inhibition and similar intracellular protein modulation compared with LR. These represent a step forward to the identification of more rigid and metabolically stable peptides.

Cyclic Peptides Acting as Allosteric Inhibitors of Human Thymidylate Synthase and Cancer Cell Growth

“…We observed that the difference in binding sites between these peptides (allosteric) and such classical TS-active-site inhibitors as PMX and RTX goes together with a different protein set modulation by the drugs [22]. This also held true for the proline derivatives that we designed after the LR peptide (see below), a finding that points to a conserved mechanism of action in cells of LR and its proline mutants [20]. This protein set modulation was observed in different types of ovarian cancer cell models such as A2780, A2780/CP, IGROV-1, 2008, and C13* [17,20,21].…”

“…To this aim, we previously identified some small peptidic inhibitors that mime a portion of the monomer–monomer interfacial region in the h TS dimer and bind therein [17,18,19,20,21]. More precisely, from a parent peptide, C20, whose sequence came from the interface region of h TS (Figure 1A), we synthesized a series of octapeptides.…”

“…The crystal structure of the h TS/ LR complex showed that the peptide binding site has the shape of an inverted cone (highlighted in green in Figure 1C,D), narrower at the top and broader at the bottom, where the LR peptide arranges with a longer extended portion that involves residues from Leu1 to Leu5 and a helical turn that involves the last three residues at the C-terminus. To improve the activity of the LR and [ d -Gln 4 ]LR octapeptides, we sequentially mutated residues 1 to 7 to proline, and examined the conformational and functional consequences of the introduction of this rigidifying residue in three regions, namely, near the C- and the N-termini and in the central portion of the peptides [20]. According to circular dichroism (CD) spectroscopic evidence, peptides [Pro 1 ]LR , [Pro 2 ]LR , and [Pro 3 ]LR shared with LR the long-extended geometry of residues from 1 to 5 and similar tendencies to form compact helical turns that involve the residues near the C-terminus.…”

“…Overall, the seven tested proline-mutated octapeptides showed conformational propensities similar to those of the starting peptide, LR , but more compact and ordered shapes. This structural feature, possibly combined with a stabilization versus peptidase degradation, may explain the stronger inhibition of ovarian cancer cell growth shown by [Pro 3 ]LR and [Pro 4 ]LR relative to the lead peptide, LR [20].…”

Correlation of LAMA3 with onset and prognosis of ovarian cancer

“…Moreover, LAMA3 is essential for the formation and function of basement membrane and plays an additional role in regulating cell migration and mechanical signal transduction (4). This gene encodes an α-subunit and can regulate keratinocyte growth factor, epidermal growth factor and insulin-like growth factor (5,6). Currently, research on this gene mostly focuses on gastric cancer, albeit some studies have focused on breast and prostate cancer (7).…”