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Old 09-28-2013, 08:21 PM   #1
'lizbeth
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HER2 overexpression renders human breast cancers sensitive to PARP inhibition

HER2 overexpression renders human breast cancers sensitive to PARP inhibition independently of any defect in homologous recombination DNA repair

Somaira Nowsheen, Tiffiny Cooper, [...], and Eddy S. Yang
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Abstract

HER2 overexpression in breast cancer confers increased tumor aggressiveness. Although anti-HER2 therapies against have improved patient outcome, resistance ultimately occurs. Poly(ADP-Ribose) polymerase (PARP) inhibitors target homologous recombination (HR)-deficient tumors, such as the BRCA-associated breast and ovarian cancers. In this study, we show that HER2+ breast cancers are susceptible to PARP inhibition independent of a HR deficiency. HER2 overexpression in HER2 negative breast cancer cells was sufficient to render cells susceptible to the PARP inhibitors ABT-888 and AZD-2281 both in vitro and in vivo, which was abrogated by HER2 reduction. In addition, ABT-888 significantly inhibited NFκB (p65/RelA) transcriptional activity in HER2+ but not HER2 negative breast cancer cells. This corresponded with a reduction in phosphorylated p65 and total IKKα levels, with a concomitant increase in IκBα. Overexpression of p65 abrogated cellular sensitivity to ABT-888, while IκBα overexpression reduced cell viability to a similar extent as ABT-888. Therefore, susceptibility of HER2+ breast cancer cells to PARP inhibition may be due to inhibition of NF-kB signaling driven by HER2. Our findings indicate that PARP inhibitors may be a novel therapeutic strategy for sporadic HER2 positive breast cancer patients.

Keywords: breast cancer, HER2, PARP inhibitors, NFκB, homologous recombination repair

Introduction

The human epidermal growth factor receptor 2 (HER2) is a proto oncogene that belongs to a family of four transmembrane receptor tyrosine kinases that mediate the growth, differentiation, and survival of cells. Overexpression of the HER2 protein, amplification of the HER2 gene, or both, occur in approximately a third of breast cancers and are associated with aggressive behavior in the tumor (1). This may be, in part, due to activation of the NFκB signaling pathway by HER2, which enhances cell proliferation, invasion, and resistance to therapies (24). HER2 activation of NFκB requires IKKα, and this activation leads to an increase in cytokine and chemokine expression, as well as an increase in invasive phenotype (3). Additionally, activation of NFκB depends on poly(ADP-ribose) polymerase (PARP) (5, 6). Targeted therapy against HER2 has been shown to benefit patients with HER2-positive breast cancer (1). However, a significant number of patients either do not respond or quickly relapse and exhibit resistance to therapy. Thus, novel therapeutic strategies are needed.
PARP inhibitors have shown initial promise in clinical trials for their single-agent activity in BRCA-associated breast and ovarian cancers, based on synthetic lethal interactions between PARP inhibition and homologous recombination (HR) repair defects (79). This is due to the conversion of single strand DNA breaks into double strand breaks in PARP inhibited cells, which is normally repaired by HR. However, because BRCA-associated tumors are deficient in HR, the double strand break persists and is lethal to the tumor cell. Normal tissues still possess intact HR, which explains why minimal side effects have been observed in these patients. However, very few studies report the efficacy of PARP inhibition alone in sporadic cancers, presumably due to their intact HR repair. Instead, numerous trials incorporate PARP inhibitors because of their ability to enhance the action of other DNA damaging agents. Thus, the use of PARP inhibitors as part of novel therapeutic combinations is currently under extensive investigation.
In this study, we unexpectedly observed exquisite susceptibility of HER2+ breast cancer cells to PARP inhibitors alone independent of an inherent HR deficiency. HER2 overexpression itself was sufficient to render breast tumor cells susceptible to PARP inhibition. The mechanistic insight of this intriguing sensitivity is described in this report and involves attenuation of NFκB signaling driven by HER2. Our results suggest that PARP inhibitors may be useful for sporadic HER2+ breast cancer patients.

Materials and Methods

Cell culture

The human breast cancer cell line MDA-MB-361 was obtained courtesy of Dr. Andra Frost (University of Alabama at Birmingham, AL) while SKBR3 and HCC 1954 were obtained courtesy of Dr. Donald Buchsbaum (University of Alabama at Birmingham, AL). The human breast cancer cell lines BT-474 (HTB-20) and MCF7 (HTB-22) and T47D (HTB-133) were obtained from ATCC. HER2 overexpressing MCF7 cells (MCF7 HER2) and its isogenic control (MCF7 NEO) were obtained courtesy of Drs. Rachel Schiff and Kent Osborne (Baylor College of Medicine, TX) (10, 11). MCF7 DRGFP cells were a kind gift from Dr. Fen Xia (Ohio State University, OH) (12). The human breast cancer cell lines MDA-MB-361, SKBR3, and HCC 1954 were maintained in RPMI-1640 (Invitrogen) supplemented with 10% fetal bovine serum (FBS) (Atlanta Biologicals). The human breast cancer cell line BT-474 (HTB-20) was cultured in RPMI-1640 supplemented with 10% FBS, 0.1% insulin (Sigma) and 4 mM L-glutamine (Invitrogen). MCF7 (HTB-22) was maintained in DMEM (Invitrogen) supplemented with 10% fetal bovine serum. MCF7 HER2 and MCF7 NEO were maintained in DMEM supplemented with 10% FBS, 0.4% G418 (Cellgro) and 15 μg/ml insulin. MCF7 DRGFP was cultured in DMEM supplemented with 10% FBS and 2 μg/mL puromycin (Sigma). T47D (HTB-133) was maintained in RPMI (Invitrogen) supplemented with 10% FBS. All cells were obtained in August 2010. The genetic background, including expression and function of key DNA repair and NF-κB signaling proteins, such as BRCA1, p53, p21, HER2, PAR, PARP, EGFR, p65, IκBα, and IKKα, as well as the growth characteristics and their response to genotoxic agents, was tested most recently on February 2012 using western blot analysis, immunohistochemistry, and colony formation assays. All experiments were performed within 10 passages, and no cell lines were kept in culture for more than 3 months after receipt or resuscitation. Cells obtained from the ATCC were also initially tested by ATCC via cytogenetic analysis and STR analysis.

Clonogenic survival assay

Cell survival was evaluated by the colony formation assay in the breast cancer cell lines as previously described (13, 14). Refer to SI Materials and Methods for details.

Cell Viability

Cell viability was measured using the ATP Lite 1 step luminescence assay (Perkin Elmer) following the manufacturer’s directions. Details are provided in SI Materials and Methods.

Immunofluorescence

To assay HR-mediated DSB repair capacity in breast cancer cell lines immunohistochemistry for radiation-induced rad51 foci was evaluated as previously described (13, 14). Immunofluorescence for HER2 expression in MCF7 HER2 tumor xenografts and H&E staining was performed as previously described (15).

Drugs, plasmids, and transfection

ABT-888 was obtained from Enzo Life Sciences (catalog # ALX-270-444) while Lapatinib (catalog # L-4804), Iniparib (catalog # I-5432) and Olaparib (catalog # O-9201) were obtained from LC Laboratories. The P65-DsRed and IκBα GFP plasmids were obtained courtesy of Dr. Markus Bredel (University of Alabama at Birmingham, AL). pDsRed and peGFP controls were obtained from Clontech. DR-GFP to measure chromosomal HR repair capacity, ISce-1 and the empty vector were gifts from Dr. Fen Xia (Ohio State University, OH) and has been described previously (12). All transfections were performed using Lipofectamine according to the manufacturer’s recommendations (Invitrogen).

Chromosomal homologous recombination mediated repair analysis

MDA-MB-361 cells were transfected with DRGFP substrate and stable integrant were selected with 2 μg/mL of puromycin (Sigma) for 3 weeks. Puromycin-resistant colonies were isolated and expanded. Chromosomal HR-mediated repair capacity was determined as described previously (12). Cells were transfected with either an empty vector, ISce-1 expression vector to measure HR-mediated repair capacity, or a GFP expression vector to measure transfection efficiency. % GFP positive cells were detected by flow cytometry. HR relative to total transfected cells was determined by division of the % GFP + cells from each ISce-1 transfection by the % GFP + cells from a parallel GFP transfection. Details are provided in SI Materials and Methods.

Immunoblotting

Immunoblotting was performed as described previously (13). Antibodies used are described in SI Materials and Methods.

HER2 knockdown

Endogenous HER2 was knocked down using SignalSilence HER2/ErbB2 siRNA I (catalog # 6283, Cell Signaling Technology). Scrambled siRNA (catalog # 6568, Cell Signaling Technology) was used as a control.

Luciferase reporter assay

NFκB transcriptional activity was measured using the NFκB Secreted Luciferase Reporter System (catalog # 631728, Clontech) according to the manufacturer’s instructions.

Tumor growth delay

3–4 week old female NOD.CB17-Prkdcscid/J mice were obtained from Jackson Labs and allowed a 1 week acclimatization period. Mice were anesthetized and supplemented with 0.72 mg 17β estradiol pellets from Innovative Research. Following recovery from pellet implantation, 2.5 × 106 cells were injected into the mammary fat pad. Once the tumors were palpable (~5 mm) mice were randomized into groups (n=8): control and ABT-888 (100 mg/kg). ABT-888 was administered twice daily by oral gavage for 34 consecutive days. Tumor size was measured using digital calipers on alternate days and tumor volume was calculated using the following formula: ½ × length × width2. Tumor size was measured for 35 days following which mice were sacrificed. All animal procedures were approved by the University of Alabama at Birmingham Institutional Animal Care and Use Committee (Animal protocol no. 101109241).

Statistical analysis

The data were analyzed via analysis of variance (ANOVA) followed by a Bonferroni post-test using GraphPad Prism version 4.02 (GraphPad Software). Data presented as average +/− standard error of mean.


Results

HER2 positive breast cancers are susceptible to PARP inhibitors alone independent of steroid receptor or p53 status

We recently reported that cetuximab, which inhibits the EGFR (HER1) signaling pathway, can generate a DNA repair defect in head and neck cancer cells and subsequently induce a contextual synthetic lethality with the PARP inhibitor ABT-888 (Veliparib) (13). We thus hypothesized that lapatinib, a dual tyrosine kinase inhibitor which interrupts the HER1/HER2 growth receptor pathways, would generate a similar DNA repair deficit and induce susceptibility to ABT-888 in human HER2+ breast cancer cells. Consistent with our hypothesis, lapatinib indeed significantly reduced HR-mediated repair capacity in the well characterized BT-474 human HER2+ breast cancer cells (Supplemental Figure 1A) (16). However, unexpectedly, ABT-888 alone caused similar levels of cytotoxicity as the combination treatment of lapatinib and ABT-888 (Figure 1A). A sub-therapeutic dose of lapatinib (10 nM) alone, which was chosen to test for synergy with PARP inhibition, produced a 30% reduction in the survival fraction of these cells.
Figure 1
HER2 overexpressing breast cancer cells are susceptible to PARP inhibition alone


To further confirm our observations that ABT-888 alone was cytotoxic to HER2+ breast cancer cells, the effects of ABT-888 on cell viability (Figure 1B) were assayed using ATP Lite 1 step luminescence assay on four other HER2 overexpressing cell lines of various steroid receptor (estrogen and progesterone, ER/PR respectively) and p53 status (Supplemental Table 1), including MDA-MB-361, HCC1954, SKBR3, and ZR-7530 (16). Intriguingly, all HER2+ breast cancer cell lines demonstrated exquisite susceptibility to ABT-888.
Since assays of cell viability are often a measure of cellular metabolic activity and not necessarily cytotoxicity, and because PARP activity can change cellular levels of NAD+ and ATP (17), the effects of PARP inhibition in our assays may be due to alterations in metabolic processes rather than cell death itself. Thus, to validate PARP inhibitor-mediated susceptibility of HER2+ breast cancer cells, we used the gold standard cytotoxicity assay, the colony formation assay. As shown in Figure 1C, ABT-888 also significantly reduced the survival fraction of all HER2 overexpressing cells tested in a dose dependent manner but, as expected, demonstrated minimal cytotoxicity in non-HER2 overexpressing MCF7 and T47D cells (p<0.01). These results confirmed our cell viability data that HER2+ breast cancer cells are susceptible to PARP inhibition. Thus, for the remainder of our susceptibility experiments, we utilized the ATP Lite 1 step luminescence cell viability assay.
To further assess whether susceptibility of HER2+ breast cancer cells to ABT-888 can be generalized to other PARP inhibitors, we next tested the efficacy of BSI-201 (Iniparib) and AZD-2281 (Olaparib), currently used in clinical trials, in BT-474 and MDA-MB-361 HER2+ breast cancer cells. Similar to results observed with ABT-888, BT-474 (Figure 1D) and MDA-MB-361 (Figure 1E) cells were susceptible to AZD-2281 alone. However, consistent with recent reports that BSI-201 is not a bona fide PARP inhibitor, HER2+ breast cancer cells did not exhibit significant cytotoxicity following BSI-201 treatment (18, 19). These novel and intriguing results suggested that indeed, human HER2+ breast cancer cells are susceptible to PARP inhibitors alone independent of ER, PR, and p53 status.
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Old 10-08-2013, 11:42 PM   #2
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Evolutionary History Of The PARP Enzyme

The recently analyzed evolutionary history of the poly(ADP-ribose)polymerase (PARP) found these proteins in eukaryotes, a wide range of organisms - animals, plants, molds, fungi, algae and protozoa - whose cells contain complex structures enclosed within membranes. While PARP proteins can be found with any of these groups, they have been most extensively studied in mammals.

In these organisms, PARPs have key functions in DNA repair, genome integrity and epigenetic regulation. More recently it has been found that proteins within the PARP family have a broader range of functions that initially predicted.

Researchers used computers to identify 236 PARP proteins from 77 species across five of the six groups, and performed extensive phylogenetic analyses of the identified PARP regions.

PARPs are found in all eukaryotic groups for which sequence is available, but some individual lineages within groups have independently lost these genes. The PARP family can be subdivided into six branches or clades. Two of these clades were likely found in the last common eukaryotic ancestor. In addition, they have identified PARPs in organisms in which they have not previously been described.

Three main conclusions were drawn from the study:

First, the broad distribution and pattern of representation of PARP genes indicated to the researchers that the ancestor of all existing eukaryotes encoded proteins of this type.

Second, the ancestral PARP proteins had different functions and activities. One of these proteins likely functioned in DNA damage response.

Third, the diversity of the PARP family is larger than previously documented, suggesting as more eukaryotic genomes become available, this gene family will grow in both number and type.

Source: The study, "Evolutionary history of the poly(ADP-ribose) polymerase gene family in eukaryotes," was authored by Rebecca S. Lamb, PhD, an assistant professor of Molecular Genetics and Ohio State University colleagues Matteo Citarelli and Sachin Teotia and appeared in an issue of the journal BMC Evolutionary Biology. The work was supported by a grant from the Ohio Plant Biotechnology Consortium and by funds from the Ohio State University.
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Old 10-08-2013, 11:44 PM   #3
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PARP Inhibitiors in Breast Cancer

Some cell-based assay labs have explored the biology of PARP inhibitors, alone and in combination, in actual human tumor primary culture micro-speheroids (microclusters), in breast, ovarian and other cancers. In these investigations, the lab applies the functional profiling platform to understand how PARP inhibitors enhance the effects of drugs and drug combinations.

As seen with PARP inhibitors, mutations work with other proteins. Genes do not operate alone within the cell but in an intricate network of interactions.

To date, one lab has observed good activity for the PARP inhibitors as single agents in BRCA1 positive patients and in some triple negative patients. Work is ongoing with these BRCA1 positive patients as wells as other tumor types where the PARP inhibitors may prove useful in the future. The PARP inhibitors are turning out to be very useful.

On April 3rd, Dr. Robert Nagourney, medical director at Rational Therapeutics and instructor in Pharmacology at the University of California, Irvine School of Medicine, will have a Poster Session at the 102nd Annual Meeting of the American Association for Cancer Research (AACR) in Orlando, Florida on the most recent findings on novel compounds that target two parallel pathways in cancer cells.

Dr. Nagourney will report the results of functional analysis with the mTOR/P13K and MEK/ERK inhibitors, BEZ235 and AZD6244, alone and in combination in human tumor primary culture micro-spheroids (microclusters): Exploration of horizontal pathway targeting. While the profiles of each drug alone are of interest, the profiles of the drugs in combination are better still.

The phenomenon of cross-talk defines an escape mechanism whereby cancer cells blocked from one passage, find a second. When clinical therapists have the capacity to block more than one pathway, the cancer cell is trapped and often dies.

This is what has been observed with these duel inhibitor combinations.

What is interesting is the fact that the activities cut across tumor types. Melanomas, colon cancers and lung cancers seem to have similar propensities to drive along these paths. Once again, we find that cancer biology is non-linear.

Moreover, cancers share pathways across tumor types, pathways that might not intuitively seem related. This is the beauty of cell-based functional profiling platform. It allows the exploration of drugs and combinations that most oncologists wouldn’t think of.

It is these counterintuitive explorations that will likely lead to meaningful advances.

Functional profiling measures biological signals rather than DNA indicators, which plays an important role in cancer drug selection and is demonstrably greater and more compelling data currently generated from DNA analyses.

The results of their investigation support the clinical relevance of targeting the MEK/ERK and PI3K/mTOR pathways and more importantly, suggest "dual" pathway inhibition (horizontal) to be a productive strategy for further clinical development. Disease specific profiles and sequence dependence are being explored and will be reported.

Most solid tumors reveal complex interactions between signal pathways that cross talk at points of commonality. To examine the clinical potential of BEZ235 and AZD6244 - inhibitors of PI3K and MEK/ERK pathways - they applied cell function analysis of programmed cell death to tumor micro-spheroids (microclusters) isolated from 24 patients. Drugs were tested alone and in combination.

According to researcher, Professor Alan Ashworth, director of the Breaktrhough Breast Cancer Research Centre in London, the BRCA1 and BRCA2 genes are involved in a repair pathway for double-strand DNA breaks that occur very close to each other. An elaborate mechanism called homologous recombination fixes some of these double-strand breaks, and BRCA2 and BRCA1 are critical for homologous recombination.

PARP is a very active enzyme involved in the repair of single-strand breaks in DNA or modified bases. It binds to DNA damage and adds multiple sugar molecules to the DNA that act as a beacon to recruit other components of DNA repair.

Emerging work on assays (PARP levels correlating with response to PARP inhibitors) have shown pretty good response with PARP inhibitors as single agents in BRCA1 positive patients and in some triple negative patients. There has been some results combining the PARP inhbitors with mustard alkylators, platins and drug combinations to optimize PARP inhibitor combinations.

These molecules have also been the subject of investigation using functional analysis in the laboratory of Dr. Nagourney. As will be reported in the Proceedings of the American Society of Clinical Oncology, Dr. Nagourney found activity for Olaparnib and Iniparib, in patients with BRCA mutation and in some triple negative breast cancer patients. This is a fertile area of investigation and a highly informative application of human tumor microspheroid analyses.

Source: Robert A. Nagourney, Paula Bernard, Federico Francisco, Ryan Wexler, Steve Evans, Rational Therapeutics, Long Beach, CA. Proceedings of AACR - Volume 52 - April 2011.
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Old 10-08-2013, 11:45 PM   #4
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Can the PARP Inhibitors be tested with the Functional Profiling Platform?

Poly ADP ribose polymerase (PARP) is a nuclear enzyme associated with response to DNA damage. Following single strand DNA breaks, the enzyme attaches a backbone of ADP and ribose that serves to initiate DNA repair. Certain classes of chemotherapeutics, specifically alkylating agents, can induce injury that results in extensive poly ADP ribosylation resulting in the exhaustion of intercellular pools of NAD and ATP ultimately leading to cell death.

Although PARP inhibitors have recently entered the clinical cancer literature mostly relating to the treatment of BRCA+ and triple negative patients, neither PARP nor PARP inhibitors are new to the cancer researcher community, according to Dr. Robert Nagourney, medical director at Rational Therapeutics, one of the pioneers of the functional profiling technique.

His group first became interested following a 1988 study by Distelhorst from Case Western Reserve (Distelhorst CW, Blood 1988 Oct;72(4):1305-09) that described a mechanism of cell death that correlated with their work in childhood leukemia. Previously, investigators at Scripps Clinic had described PARP’s role in response to 2CDA (Seto, S., et al. J Clin. Invest. 1985 Feb;75(2):377-83). His group has studied small molecule inhibitors of PARP for many years, and more recently, they have expanded these investigations to include BSI201 (iniparib) and AZD2281 (olaparib). Both of which are undergoing clinical investigations. Nagourney will be reporting their findings with these PARP inhibitors at the 2011 ASCO meeting (Nagourney, R., et al Proceedings Amer Soc Clin Oncol. 2011).

PARP inhibitors are easily studied and provide interesting signals in the tissue studied. They have seen activity in BRCA+ patients and some triple negative breast cancers. They have also identified synergy with other classes of drugs. The compounds are a welcome addition to the cancer therapy armamentarium and continue to be actively studied in the cell-based functional profiling platform.

Of interest is the recent failure of the iniparib plus Carboplatin & gemcitabine Phase III trial to meet progression-free and overall survival goals in triple negative breast cancer patients (Zacks Investment Research on January 31, 2011). This failure may reflect the need to apply predictive methodologies to select candidates for these drugs, similar to Nagourney's successful work with other classes of compounds.
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Old 10-08-2013, 11:47 PM   #5
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New mechanism of action for PARP

NIH researchers discover new mechanism of action for class of chemotherapy drugs

Bethesda, Maryland

Monday, November 05, 2012

The National Institutes of Health (NIH) researchers have discovered a significant new mechanism of action for a class of chemotherapy drugs known as poly (ADP-ribose) polymerase inhibitors, or PARP inhibitors. They have also identified differences in the toxic capabilities of three drugs in this class which are currently being tested in clinical trials. The study was conducted at the National Cancer Institute (NCI), part of NIH.

In recent years, drugs classified as PARP inhibitors have been shown to be promising anticancer agents for breast and ovarian cancer. Members of the PARP family of proteins are involved in a number of critical cellular processes, including DNA damage repair and programmed cell death. Prior to this study, PARP inhibitors were thought to work primarily by blocking PARP enzyme activity, thus preventing the repair of DNA damage and ultimately causing cell death.

In this study, scientists established that PARP inhibitors have an additional mode of action: localizing PARP proteins at sites of DNA damage, which has relevance to their anti-tumour activity. The trapped PARP protein–DNA complexes are highly toxic to cells because they block DNA replication. When the researchers tested three PARP inhibitors for their differential ability to trap PARP proteins on damaged DNA, they found that the trapping potency of the inhibitors varied widely.

"Critical to our research is that, while PARP inhibitors had been assumed to be of equivalent potency based on the degree to which they elicit PARP inhibition, we now know that they are not equivalent with respect to their potency to trap PARP," said Yves Pommier, MD, PhD., NCI Centre for Cancer Research. "Our findings suggest that PARP inhibitors should be categorized according to their potency to trap PARP, in addition to their enzyme inhibition abilities."

The PARP family of proteins in humans includes PARP1 and PARP2, which are DNA binding and repair proteins. When activated by DNA damage, these proteins recruit other proteins that do the actual work of repairing DNA. Under normal conditions, PARP1 and PARP2 are released from DNA once the repair process is underway. However, as this study shows, when they are bound to PARP inhibitors, PARP1 and PARP2 become trapped on DNA. The researchers showed that trapped PARP–DNA complexes are more toxic to cells than the unrepaired single-strand DNA breaks that accumulate in the absence of PARP activity, indicating that PARP inhibitors act as PARP poisons.

In collaboration with James Doroshow, MD, deputy director for clinical and translational research at NCI, the investigators used PARP assays (ways of measuring PARP activity in cells and tissues) to compare three PARP inhibitor compounds that are currently in clinical testing: MK-4827, olaparib, and veliparib.

The scientists found that the three PARP inhibitors differed in their ability to inhibit PARP enzyme activity, with olaparib being the most potent inhibitor, followed by veliparib and then MK-4827. However, in terms of toxicity, MK-4827 was the most potent, followed by olaparib and then veliparib. Moreover, PARP1 complexes with MK-4827 and olaparib were shown to be more tightly bound to DNA than complexes with veliparib.

These findings suggest that there may be two classes of PARP inhibitors, catalytic inhibitors that act mainly to inhibit PARP enzyme activity and do not trap PARP proteins on DNA, and dual inhibitors that both block PARP enzyme activity and act as PARP poison.

"Our findings suggest that clinicians who use PARP inhibitors in clinical trials should carefully choose their drug, because we now suspect results may differ, depending upon the PARP inhibitor used," said Junko Murai, MD, PhD., NCI Centre for Cancer Research. "As a next step, we are working to categorize other leading PARP inhibitors based upon both PARP trapping and PARP inhibition."

This work was supported by the Intramural Programme of NCI and by the Japan Society for the Promotion of Science (JSPS) Core-to-Core Programme. First author Junko Murai is a JSPS fellow working at NCI Centre for Cancer Research. Funding was provided by NCI grant Z01 BC 006150-19LMP.

The National Cancer Institute (NCI) leads the National Cancer Programme and the NIH effort to dramatically reduce the burden of cancer and improve the lives of cancer patients and their families, through research into prevention and cancer biology, the development of new interventions, and the training and mentoring of new researchers.

NIH, the nation's medical research agency, and is a component of the US Department of Health and Human Services. NIH is the primary federal agency conducting and supporting basic, clinical, and translational medical research, and is investigating the causes, treatments, and cures for both common and rare diseases.
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Old 10-11-2013, 11:31 AM   #6
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Re: HER2 overexpression renders human breast cancers sensitive to PARP inhibition

GDP,

Thanks for the postings.

I am watching the PARP inhibitors with interest. My friend - who has BRCA1 ovarian cancer was in a Stanford trial. In the beginning she had dramatic results, then the cancer took off like it was on steroids.

So again, the metabolomics trumped genetics, and the cancer triumphed in survival of the fittest.

I had mentioned that she had her initial tumor tested with Rational Therapeutics and Estrogen played a key role. When she developed resistance to the initial treatment (platinum salt and Tamoxifen I think) she thought that blocking the estrogen does not work.

In the clinical trial, I do not believe estrogen receptors were addressed. It was like Lani's example - this time the windows were shut, but the front door was left wide open. It only took a few months for the puppy (cancer) to figure out it could get out! If she had also been taking Tamoxifen, or an Aromatase Inhibitor – I wonder if the resistance would have been delayed or overcome. If I remember correctly it was a monotherapy, but I wasn’t privy to all the details, so I’m not sure.

I’m not knocking functional profiling. I’m with you – I want to see the Phase III Battle of the Bands. I’m sure I already know which band is going to come out on top.

I see that Susan G. Komen gave a significant amount to explore the Her2/Parp issue.

http://www.news-medical.net/news/201...nhibitors.aspx

I wonder what it takes to get these charities interested in supporting the Battle of the Band? I think one of the issues is that it is testing, and it is easier to envision the benefits of new treatments.

Do you have knowledge of who is supporting funding? Is it Vanguard Cancer Foundation? We need bigger guns to get the Phase III funded. If we could get one of the top charities interested supporting the Battle of the Bands – then personalized medicine could really get somewhere. It would potentially save many, many patients from needlessly suffering through ineffective & toxic treatments.

I mean think of it, in the doctor’s office: Well, Mrs. Smith you can go into this clinical trial and get randomized into a standard of care treatment that may or may not work for you, or you can get the arm that personalizes your treatment into the one that works best for you.

Of course, you could avoid the placebo effect by not telling the patient which arm they are in.
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Old 10-11-2013, 01:11 PM   #7
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A couple of years ago, Nagourney was explaining what happened to the PARP inhibitors in breast cancer.

It turned out that iniparib, a member of the benzamine chemical family, at physiological concentrations achievable in humans is not a PARP inhibitor. This, in retrospect, should have been obvious because a full-dose PARP inhibitor, plus a potent combination of carboplatin plus gemcitabine would not likely be tolerable if PARP inhibition were achieved.

Second, the patients receiving the drug are probably not a homogeneous population. That is, some TNBC patients may be similar to the BRCA patients, while others may not have the DNA repair deficiencies associated with PARP inhibitor response.

Finally, he originally reported the carboplatin plus gemcitabine combination in breast cancer, as a split-dose doublet in 2008 (Nagourney, Clin Breast Cancer Research, 2008). He observed, in that original clinical trial, that even a lower starting dose of gemcitabine (i.e. 800mg/ml2 vs. the O’Shaughnessy 1000 mg/m2) resulted in significant toxicity and in his concluding comments in that paper, he suggested 600mg/ml2. At 1000 mg/m2, Dr. O’Shaughnessy’s trial nearly doubled our recommended dose in this patient population.

He, like other investigators, entered into the original studies of these molecules believing iniparib to be a PARP inhibitor. To his surprise, a direct comparison of olaparib to inapaprib revealed no correlation. He described this in an abstract, “Of interest, BSI201 & AZD2281 activity did not correlate in parallel analyses (R = 0.07, P > 0.5).” The human tumor primary culture analysis scooped the other investigators.

So, what was earned? First, they learned that iniparib is not a true PARP inhibitor.
Second, they learned that the combination of platins plus gemcitabine in breast cancer is synergistic, highly active and can be toxic (particularly at the doses chosen for the trial).

Finally, they learned that TNBC, indeed all breast cancers, even more to the point, all cancers in general, are heterogeneous. That is precisely why the use of human tumor primary culture analyses are so instructive and should be incorporated into clinical trials for these and other targeted agents.

I know I'm preaching to the choir and I'm just as interested in a Battle of the Band clinical trial (if anything, in the memory of Ann). I would suspect that Vanguard has the interest, but whether they have the means. I think that something like $5 million would be minimum to start something like this. Looking for a Sugar Daddy!
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Old 10-11-2013, 01:42 PM   #8
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Re: HER2 overexpression renders human breast cancers sensitive to PARP inhibition

or a Sugar Mama! BTW . . . I believe Dr. O’Shaughnessy is a female, as in Joyce. I remember her because she said:
Quote:
TCH/pertuzumab may become the standard toward the end of the year.

She was discussing that the:

Quote:
TRYPHAENA trial showed similar pCR rates of about 60% to 70% when patients were assigned preoperatively to TCH with pertuzumab versus FEC followed by taxane/trastuzumab/pertuzumab.

Maybe we need to get the ball rolling on social media - "Battle of the Bands" sounds a lot more appealing to me than the stink of Pink!

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Old 10-11-2013, 02:06 PM   #9
gdpawel
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Re: HER2 overexpression renders human breast cancers sensitive to PARP inhibition

"Battle of the Bands" sounds a lot more appealing to me than the stink of Pink! LOL! 10-4!
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Old 10-11-2013, 02:49 PM   #10
'lizbeth
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Re: HER2 overexpression renders human breast cancers sensitive to PARP inhibition

LOL, well I'm still annoyed about the Pink vaccine. I know they mean to be helpful, but talking about FastTracking and how it could be available to the public in just a few years - that is just blatantly deceptive. It hasn't even been approved for a Phase I trial, it is only effective in mice, and if it is been tested on people who might someday have cancer, well, I would expect the clinical trial process would be minimum 10 - 15 years.

There is so much hype that we don't see what is right under our nose, and working. Sometimes it seems like the low budget, highly successful ideas get run over and left behind by the high budget, incrementally successful ideas.

What we need to do is sell the Simon Cowell, Ryan Secrest, Randy Jackson, Steven Tyler, Keith Urban, Jennifer Lopez, Mariah Carey, Paula Abdul to have an American Idol: Battle of the Bands season, and when people call in to vote for a band a portion goes to Vanguard for the "Battle of the Bands" clinical trial.

I like it.
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Old 10-28-2013, 04:52 AM   #11
aleena
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Re: HER2 overexpression renders human breast cancers sensitive to PARP inhibition

nice post, thanks for sharing
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