Antiestrogenic and anticancer activities of peptides derived from the active site of alpha-fetoprotein

Cyclo[EKTOVNOGN] (AFPep), a cyclic 9-amino acid peptide derived from the active site of alpha-fetoprotein, has been shown to prevent carcinogen-induced mammary cancer in rats and inhibit the growth of ER+ human breast cancer xenografts in mice. Recently, studies using replica exchange molecular dynamics predicted that the TOVN region of AFPep might form a dynamically stable putative Type I beta-turn, and thus be biologically active without additional amino acids. The studies presented in this paper were performed to determine whether TOVN and other small analogs of AFPep would inhibit estrogen-stimulated cancer growth and exhibit a broad effective-dose range. These peptides contained nine or fewer amino acids, and were designed to bracket or include the putative pharmacophoric region (TOVN) of AFPep. Biological activities of these peptides were evaluated using an immature mouse uterine growth inhibition assay, a T47D breast cancer cell proliferation assay, and an MCF-7 breast cancer xenograft assay. TOVN had very weak antiestrogenic activity in comparison to AFPep's activity, whereas TOVNO had antiestrogenic and anticancer activities similar to AFPep. OVNO, which does not form a putative Type I beta-turn, had virtually no antiestrogenic and anticancer activities. A putative proteolytic cleavage product of AFPep, TOVNOGNEK, significantly inhibited E2-stimulated growth in vivo and in vitro over a wider dose range than AFPep or TOVNO. We conclude that TOVNO has anticancer potential, that TOVNOGNEK is as effective as AFPep in suppressing growth of human breast cancer cells, and that it does so over a broader effective-dose range.

Screening of Indian Plants for Biological Activity: Part II

Three hundred botanically identified plant materials have been extracted with 50% ethanol and the extracts put through a wide biological screen. These include tests for antibacterial, anticancer, antifertility, antifungal, anthelminthic, antiprotozoal, antiviral and pharmacological activities. Biological activities have been confirmed in fractions of fifty-six of these extracts.

Screening of Indian Plants for Biological Activity: Part III

Ethanol extracts (50%) of 295 botanically Identified plant materials have been tested for a wide variety of biological activities including anticancer, chemotherapeutic and pharmaceutical activities. Biological activities been confirmed in fractions of 34 of these extracts.

Screening of Indian Plants for Biological Activity: Part IV

Ethanol extracts (50%) of 295 botanically Identified plant materials have been tested for a wide variety of biological activities including anticancer, chemotherapeutic and pharmaceutical activities. Biological activities been confirmed in fractions of 22 of these extracts.

Screening of Indian Plants for Biological Activity : Part V

Two hundred and eighty nine botanically identified plant materials have been tested as 50% ethanol extracts, for a wide variety of bloloalcal activities including including anticancer, chemotherapeutic and pharmaceutical activities. Biological activities been confirmed in fractions of 47 of these extracts.

Screening of Indian Plants for Biological Activity: Part VI

Two hundred and ninety eight botanically identified plant materials have been tested as 50% ethanol extracts, for a wide variety of biological activities including antifertility, anticancer, chemotherapeutic and pharmacological activities. Biological activities been confirmed in fractions of 84 of these extracts.

Screening of Indian Plants for Biological Activity : Part IX

Two hundred and ninety three botanically identified plant materials have been tested as 50% ethanol extracts, for a wide variety of biological activities including antifertility, anticancer, chemotherapeutic and pharmaceutical activities. Biological activity has been confirmed in fractions of 40 of these extracts.

Screening of Indian Plants for Biological Activity: Part X

Two hundred eightythree botanically identified plant species have been tested as 50% ethanol extracts, for a wide variety of biological activities including antifertility, anticancer, chemotherapeutic and pharmaceutical activities. Biological activity has been observed in 143 of these extracts. The biological Activity has been confirmed in the fraction of 35 extracts and anticancer activity in the extracts of 23 plant species.

Screeningof Indian Plants for Biological Activity :Part XII

Alcoholic extracts of 295 botanically identified plant materials from 267 plant species have been tested for a wide variety of biological activities including chemotherapeutic and pharmacological screenings. Biological activities have been confirmed in fractions of 64 of these extracts. Follow-up studies have been carried out in some plants with confirmed activity. The active principles and results of these studies are reported.

Screening of Indian Plants for Biological Activity : Part XIII

Alcoholic extracts of 300 botanically identified plant materials from 287 plant species have been tested for various biological activities including chemotherapeutic and pharmacological screenings. Biological activities have been confirmed in 51 fractions of the extracts. Follow-up studies have been carried out in some plants with confirmed activity. The active principles and results of these studies are reported.

Screening of Indian plants for biological activity : Part XIV

Alcoholic extracts of 300 botanically identified plant materials from 275 plant species have been tested for various biological activities including chemotherapeutic and pharmacological screenings. Biological activities have been observed in 111 extracts. Follow-up studies have been carried out in some plants with confirmed activity. The active principles and results of these studies are reported.

Screening of Indian plants for biological activity: Part XV

Alcoholics extracts of 266 botanically identified plant materials from 222 plant species have been tested for various biological activities including chemotherapeutic and pharmacological screenings. Biological activities have been observed in 89 extracts. Follow-up studies have been carried out in some plants with confirmed activity. The active principles and results of these studies are reported.

Screening of Indian plants for biological activity: Part XVI

Alcoholic extracts of 288 of plant materials from 199 plant species have been tested for various biological activities including chemotherapeutic and pharmacological screening. Biological activities, ranging from moderate to good degree, have been observed in 61 plants extracts. Follow up studies have been carried out in these extracts and some of them have shown moderate degree of activities at this Institute. However, none of the extracts was found to be good enough for further development. Results of the present studies, alongwith chemical investigations on different species of similar genera which were screened earlier, are also discussed.

The nonsteroidal anti-inflammatory drug indomethacin induces heterogeneity in lipid membranes: potential implication for its diverse biological action.

BACKGROUND: The nonsteroidal anti-inflammatory drug (NSAID), indomethacin (Indo), has a large number of divergent biological effects, the molecular mechanism(s) for which have yet to be fully elucidated. Interestingly, Indo is highly amphiphilic and associates strongly with lipid membranes, which influence localization, structure and function of membrane-associating proteins and actively regulate cell signaling events. Thus, it is possible that Indo regulates diverse cell functions by altering micro-environments within the membrane. Here we explored the effect of Indo on the nature of the segregated domains in a mixed model membrane composed of dipalmitoyl phosphatidyl-choline (di16:0 PC, or DPPC) and dioleoyl phosphatidyl-choline (di18:1 PC or DOPC) and cholesterol that mimics biomembranes. METHODOLOGY/PRINCIPAL FINDINGS: Using a series of fluorescent probes in a fluorescence resonance energy transfer (FRET) study, we found that Indo induced separation between gel domains and fluid domains in the mixed model membrane, possibly by enhancing the formation of gel-phase domains. This effect originated from the ability of Indo to specifically target the ordered domains in the mixed membrane. These findings were further confirmed by measuring the ability of Indo to affect the fluidity-dependent fluorescence quenching and the level of detergent resistance of membranes. CONCLUSION/SIGNIFICANCE: Because the tested lipids are the main lipid constituents in cell membranes, the observed formation of gel phase domains induced by Indo potentially occurs in biomembranes. This marked Indo-induced change in phase behavior potentially alters membrane protein functions, which contribute to the wide variety of biological activities of Indo and other NSAIDs.

STUDIES ON THE BIOLOGICAL ACTIVITY OF MACROMOLECULES MICROINJECTED INTO MAMMALIAN SOMATIC CELLS

Red Blood cell mediated and glass needle mediated microinjection technology was used to introduce macromolecules into mammalian somatic cells. The biological activities of DNA synthesis inducing factor(s) (Chapter 1), mitotic factor(s) (Chapter 2), and DNA coding for ovalbumin and thymidine kinase (Chapter 3) were studied following injection into mammalian somatic cells.^ Chapter 1. A cell undergoing DNA replication (S phase) contains a factor(s) that induces DNA synthesis prematurely in a G(,1) nucleus when an S phase cell is fused to a G(,1) cell. An assay for the active factor(s) was developed in which a mixture of s phase extract loaded red blood cells (RBC) and synchronous G(,1) HeLa cells was centrifuged onto Concanavalin A (Con A) treated coverslips and fused by PEG. This technique is called "Centrifusion". The synchronous G(,1) HeLa cells injected with S phase extract initiated DNA synthesis earlier than the control G(,1) cells mock injected with RBC loaded with buffer.^ Chapter 2. It has been demonstrated that fusion between a mitotic and an interphase cell usually leads to breakdown of the interphase nucleus, followed by condensation of the interphase chromatin into discrete chromosomes, a process termed premature chromosome condensation. I wanted to develop an assay for the mitotic factor(s) that induces premature chromosome condensation. Experiments were performed utilizing glass needle mediated microinjection of HeLa cell mitotic extract into interphase somatic mammalian cells in an attempt to induce premature chromosome condensation. However, I was not able to induce premature chromosome condensation in the interphase cells, probably because of an inability to introduce sufficient mitotic factor(s) into the cells.^ Chapter 3. A recombinant plasmid containing the chicken ovalbumin gene and three copies of the Herpes thymidine Kinase gene (pOV12-TK) was introduced into mouse LMTK('-) cell nuclei using glass needle mediated gene transfer resulting in LMTK('+) clones that were selected for in HAT medium. Restriction enzyme analysis of the high molecular weight DNA from 6 HAT medium survivor cell clones revealed the presence of one or at best only a few copies of the 12kb ovalbumin gene per mouse genome. Further analysis showed the ovalbumin DNA was not rearranged and was associated with high molecular weight mouse cell DNA. Each of the analyzed cell clones produced ovalbumin demonstrating that the biological activity of the microinjected ovalbumin was retained. ^