Paige Baldwin

PARP inhibitors: A new era of targeted therapy.

Personalized medicine seeks to utilize targeted therapies with increased selectivity and efficacy in preselected patient cohorts. One such molecularly targeted therapy is enabled by inhibiting the enzyme poly(ADP-ribose) polymerase (PARP) by small molecule inhibitors in tumors which have a defect in the homologous DNA recombination pathway, most characteristically due to BRCA mutations. Olaparib, a highly potent PARP inhibitor, has recently been the approved for ovarian cancer therapy by the FDA and European commission in patients with platinum-sensitive, recurrent, high-grade serous ovarian cancer with BRCA1 or BRCA2 mutations. Currently, clinical trials with several PARP inhibitors are being conducted to assess the toxicities, the efficacies and the benefit of the drugs as monotherapies or combined with radiation or other chemotherapeutic agents, in ovarian, breast, prostate, rectal, lung, pancreatic, peritoneal, head and neck, brain, squamous cell carcinomas and sarcomas, to list a few. In this review, our focus is to outline the emerging molecular mechanisms, preclinical evidence and clinical applications of PARP inhibitors especially in nonBRCA cancers, and review the combination strategies compatible with PARP inhibitor therapy.

Nanoformulation of Olaparib amplifies PARP inhibition and sensitizes PTEN/TP53-deficient prostate cancer to radiation

The use of PARP inhibitors in combination with radiotherapy is a promising strategy to locally enhance DNA damage in tumors. Here we show that radiation-resistant cells and tumors derived from a Pten/Trp53-deficient mouse model of advanced prostate cancer are rendered radiation sensitive following treatment with NanoOlaparib, a lipid-based injectable nanoformulation of olaparib. This enhancement in radiosensitivity is accompanied by radiation dose-dependent changes in γ-H2AX expression and is specific to NanoOlaparib alone. In animals, twice-weekly intravenous administration of NanoOlaparib results in significant tumor growth inhibition, whereas previous studies of oral olaparib as monotherapy have shown no therapeutic efficacy. When NanoOlaparib is administered prior to radiation, a single dose of radiation is sufficient to triple the median mouse survival time compared to radiation only controls. Half of mice treated with NanoOlaparib + radiation achieved a complete response over the 13-week study duration. Using ferumoxytol as a surrogate nanoparticle, MRI studies revealed that NanoOlaparib enhances the intratumoral accumulation of systemically administered nanoparticles. NanoOlaparib-treated tumors showed up to 19-fold higher nanoparticle accumulation compared to untreated and radiation-only controls, suggesting that the in vivo efficacy of NanoOlaparib may be potentiated by its ability to enhance its own accumulation. Together, these data suggest that NanoOlaparib may be a promising new strategy for enhancing the radiosensitivity of radiation-resistant tumors lacking BRCA mutations, such as those with PTEN and TP53 deletions. Mol Cancer Ther; 16(7); 1279–89. ©2017 AACR.

Abstract B48: Prostate cancer pre-treatment with nanoformulated Olaparib overcomes radiation resistance

Prostate cancers with PTEN deletions are promising candidates for DNA repair inhibitors such as olaparib and talazoparib. Here we show that radiation-resistant cells and tumors derived from Ptenpc-/-;Trp53pc-/- mice are rendered radiation-sensitive following pre-treatment with liposomal nanoOlaparib. This enhancement in radiosensitivity is accompanied by radiation dose-dependent changes in γ-H2AX expression and is specific to nanoformulated Olaparib alone. In animals, twice-weekly intravenous administration of nanoOlaparib alone results in significant tumor growth inhibition. When nanoOlaparib is administered prior to radiation, we find that a single dose of radiation is sufficient to increase mouse survival time by as much as 10 weeks (study duration = 13 weeks). Using ferumoxytol as a surrogate nanoparticle, magnetic resonance imaging (MRI) studies revealed that nanoOlaparib administration enhances the ability of nanoparticles to accumulate in tumors. Compared to untreated and radiation-only controls, nanoOlaparib-treated tumors showed 18-fold higher nanoparticle accumulation, suggesting that the in vivo efficacy of nanoOlaparib may be potentiated by its ability to enhance its own accumulation in tumors. Citation Format: Anne L. van de Ven, Shifalika Tangutoori, Paige Baldwin, Ju Qiao, Codi Gharagouzloo, Nina Seitzer, John Clohessy, Houari Korideck, G. Mike Makrigiorgos, Robert Cormack, Pier Paolo Pandolfi, Srinivas Sridhar. Prostate cancer pre-treatment with nanoformulated Olaparib overcomes radiation resistance. [abstract]. In: Proceedings of the AACR Special Conference on Engineering and Physical Sciences in Oncology; 2016 Jun 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2017;77(2 Suppl):Abstract nr B48.

Abstract 3900: Sustained release of PARP inhibitor Talazoparib and chemotherapeutics from biodegradable implants for treatment of breast and prostate cancer

Sustained localized delivery of cancer therapeutics is a safe and effective unique option for non-metastatic cancers. Here we report a novel biodegradable implant with the capability to encapsulate therapeutics, molecular agents, or nanoparticles for local intratumoral delivery. We have successfully demonstrated in vivo the delivery of PARP inhibitor Talazoparib to treat Brca1-mutated cancers and Docetaxel to treat localized or recurring prostate cancers. This one-time intratumoral injection provides a safe vehicle for the sustained release of PARP inhibitor Talazoparib and chemotherapeutic Docetaxel in contrast to low bioavailability and toxicity associated with oral or systemic delivery. Methods: Biodegradable implants of 1-2mm length and 0.8mm diameter were loaded with ∼50μg Talazoparib (BMN) for BRCA1-mutated breast cancer (BCa) studies and ∼500μg Docetaxel (DTX) for prostate cancer (PCa) studies. Implants were characterized using SEM and HPLC, and release studies were carried out in pH 6.0 PBS buffer at 37°C. The IC509s were determined using an MTS assay in cell lines W0069 and W780 (BCa) and PC3 (PCa). In vivo studies were carried out in Brca1 Co/Co;MMTV-Cre; p53+/− spontaneous tumored mice for BCa studies. Subcutaneous PC3 tumors were xenografted in nude mice. PCa studies were done with and without radiation. Implants were injected once intratumorally using an 18G brachytherapy needle. Results: The release profile of the drug from the implant in buffer showed a highly sustained release for multiple weeks at therapeutically relevant doses for both DTX and BMN loaded implants. BCa cell lines W0069 and W780 were highly sensitive to BMN, most likely due to Brca1 mutation. Following a one-time intratumoral implantation of BMN, tumors reduced in size by an average of 50%, while untreated tumors increased ∼5X in size. BMN dosing appeared to be well tolerated by the mice. DTX implants proved to be an effective method for PCa treatment in vivo with no weight loss observed. The local DTX group showed sustained tumor inhibition compared to empty implants and an equivalent DTX dose given systemically. At 40 days 89% survival was observed for mice treated with localized DTX implants compared with 0% in all other treatment groups. Histology samples were taken from sacrificed mice and immunohistochemistry is currently underway. Conclusions: Sustained local release of therapeutically relevant doses of BMN and DTX were observed in vitro and in vivo. Therapeutics loaded in implants represent a novel delivery modality that is well-tolerated. Sustained release of BMN appears to amplify the therapeutic efficacy of PARP inhibition in BRCA1 mutated breast cancers and sustained release of DTX is an effective chemotherapy option alone or in combination with radiation therapy. These results lay a strong foundation for the use of localized biodegradable implants for the treatment of breast and prostate cancer. Citation Format: Jodi Belz, Noelle Castilla Ojo, Paige Baldwin, Rajiv Kumar, Anne van de Ven, Karen Liby, Robert Cormack, Mike Makrigiorgos, Srinivas Sridhar. Sustained release of PARP inhibitor Talazoparib and chemotherapeutics from biodegradable implants for treatment of breast and prostate cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3900.

Abstract 4335: Nanoformulations of PARP inhibitors Olaparib and Talazoparib for targeted cancer therapy

Introduction: Poly(ADP-ribose) Polymerase (PARP) plays an important role in a number of DNA repair pathways. PARP inhibitors (PARPi) such as Olaparib and Talazoparib exploit the concept of synthetic lethality by selectively targeting cancer cells with defective DNA repair pathways. These drugs are currently only available in oral form which results in limited bioavailability, poor tumor accumulation, and systemic toxicity. Here we report the development of novel nanoformulations of Olaparib and Talazoparib to allow intravenous or intraperitoneal delivery, providing greater bioavailability and tumor accumulation, while limiting systemic toxicities. Methods: Nanoparticle formulations of Olaparib and Talazoparib were synthesized and tested in vitro and in vivo. Short-and long-term dose response with a panel of ovarian cancer cell lines were conducted. These cell lines include KURAMOCHI, SKOV3, OVSAHO, JHOS2, PA1, COV318, 403 and 404, derived from BRCA2-/-, PTEN-/-, TP53mut mice, and 4306 and 4412, developed from conditional LSL-K-rasG12D/+, PTENloxP/loxP mice. Radiosensitization with NanoOlaparib was tested in the radiation resistant prostate cancer cell line FK01, derived from Ptenpc-/-;Trp53pc-/- mice. In vivo, NanoOlaparib was tested in an IP spread model using 404 cells. Animals were treated IP with NanoOlaparib alone, and in combination with cisplatin. Radiosensitization with NanoOlaparib in vivo was tested in a xenograft model using FK01 cells to mimic castration resistant prostate cancer. Animals were treated biweekly with NanoOlaparib before and after radiation treatment. Results: The murine cell lines 403 and 404 were highly sensitive to this treatment due to the mutations in BRCA2, PTEN, and TP53. 4412 and 4306 showed comparable sensitivity, suggesting that a PTEN deletion confers similar sensitivity to PARP inhibitors as a BRCA2 deletion. PA1 demonstrated high sensitivity to NanoOlaparib which may be attributed to genetic instability. NanoTalazoparib is more potent than NanoOlaparib, resulting in a similar relationship in cell line sensitivity with overall lower IC50’s. Strong synergistic radiosensitization was observed in FK01 cells with NanoOlaparib. Bioluminescence imaging illustrated that NanoOlaparib administered IP daily resulted in a greater inhibition of tumor growth than those treated with oral Olaparib daily. The FK01 xenografts are highly radioresistant with little difference between untreated and radiation only animals. NanoOlaparib delays tumor growth, while the combination of radiation and NanoOlaparib clearly shrinks tumors. Conclusions: Robust nanoparticle formulations of NanoTalazoparib and NanoOlaparib have been successfully developed for in vitro and in vivo studies. These results show that NanoOlaparib and NanoTalazoparib amplify the therapeutic efficacy of PARP inhibition and imply a very promising role for the nanoformulation in ovarian and prostate cancers. Citation Format: Paige Baldwin, Anders Ohman, Jeremy Thong, Shifalika Tangutoori, Anne van de Ven, Rajiv Kumar, Daniela Dinulescu, Srinivas Sridhar. Nanoformulations of PARP inhibitors Olaparib and Talazoparib for targeted cancer therapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4335.

In vitro analysis of PARP inhibitor nanoformulations

PARP-l is a DNA repair protein that plays a role in a number of repair pathways and also helps in transcriptional regulation; thus PARP inhibitors (PARPi), such as olaparib and BMN-673, act by inhibiting DNA damage repair. This leads to an accumulation of deleterious mutations leading to genetic instability as a result of a number of cell replications. Currently, olaparib is only available in an oral form and has poor bioavailability, consequently leading to poor accumulation in the tumor due to first-pass metabolism. Therefore, in the present study, an injectable nanoparticle formulation of olaparib was created that offers a delivery route in which the drug would be fully bioavailable in the vasculature, suggesting greater tumor accumulation. Our results illustrated that injectable nanoformulations of olaparib and BMN-673, a next generation PARPi, could be developed, and an efficacy test indicated that BMN-673 is a much more potent PARPi than olaparib. The success of these molecular inhibitors as a monotherapy in inhibiting colony formation suggests enhanced efficacy of these treatments in combination with other therapies, even in tumors which have developed resistance.

Sustained release Talazoparib implants for localized treatment of Brca1-deficient breast cancer

Talazoparib, a potent PARP inhibitor, has shown promising clinical and pre-clinical activity by inducing synthetic lethality in cancers with germline Brca1/2 mutations. Conventional oral delivery of Talazoparib is associated with significant off-target effects, therefore we sought to develop new delivery systems in the form of an implant loaded with Talazoparib for localized, slow and sustained release of the drug at the tumor site in Brca1-deficient breast cancer. Poly(lactic-co-glycolic acid) (PLGA) implants (0.8 mm diameter) loaded with subclinical dose (25 or 50 µg) Talazoparib were fabricated and characterized. In vitro studies with Brca1-deficient W780 and W0069 breast cancer cells were conducted to test sensitivity to PARP inhibition. The in vivo therapeutic efficacy of Talazoparib implants was assessed following a one-time intratumoral injection in Brca1Co/Co;MMTV-Cre;p53+/- mice and compared to drug-free implants and oral gavage. Immunohistochemistry studies were performed on tumor sections using PCNA and γ-H2AX staining. Sustained release of Talazoparib was observed over 28 days in vitro. Mice treated with Talazoparib implants showed statistically significant tumor growth inhibition compared to those receiving drug-free implants or free Talazoparib orally. Talazoparib implants were well-tolerated at both drug doses and resulted in less weight loss than oral gavage. PARP inhibition in mice treated with Talazoparib implants significantly increased double-stranded DNA damage and decreased tumor cell proliferation as shown by PCNA and γ-H2AX staining as compared to controls. These results demonstrate that localized and sustained delivery of Talazoparib via implants has potential to provide superior treatment outcomes at sub-clinical doses with minimal toxicity in patients with BRCA1 deficient tumors.

Abstract B30: Sustained release of PARP inhibitor Talazoparib and chemotherapeutic Docetaxel from modified brachytherapy spacers for treatment of breast and prostate cancer

Sustained localized delivery of cancer therapeutics is a safe and effective unique option for local control of tumors. Here we report a novel biodegradable implant with the capability to encapsulate different therapeutics, molecular agents, or nanoparticles for local intratumoral delivery. We have successfully demonstrated in vivo the delivery of PARP inhibitor Talazoparib to treat Brca1-mutated cancers and Docetaxel to treat localized or recurring prostate cancers. This one-time intratumoral injection provides a safe vehicle for the sustained release of PARP inhibitor Talazoparib and chemotherapeutic Docetaxel in contrast to low bioavailability and toxicity associated with oral or systemic delivery. Methods: Biodegradable implants of 1-2mm length and 0.8mm diameter were loaded with ~50μg Talazoparib for BRCA1-mutated breast cancer studies and ~500μg Docetaxel (DTX) for prostate cancer studies. The implants were characterized in vitro using SEM and HPLC, and the release kinetic studies were carried out in PBS buffer (pH 6.0) at 37°C. The IC509s were determined using an MTS assay in breast cancer cell lines derived from Brca1 Co/Co; MMTV-Cre; p53+/−mice, W0069 and W780, and human-derived prostate cancer, PC3. In vivo studies were carried out in Brca1 Co/Co; MMTV-Cre; p53+/− spontaneous tumored mice for breast cancer studies. Subcutaneous PC3 tumors were xenografted in nude mice. Prostate cancer studies were done with and without radiation. Drug-loaded implants were injected once intratumorally using an 18G brachytherapy needle. Results: The release profile of the drug from the implant in buffer showed a highly sustained release for multiple weeks at therapeutically relevant doses for both docetaxel and Talazoparib loading implants. Breast cancer cell lines W0069 and W780 were highly sensitive to Talazoparib, most likely due to Brca1 mutation. Following a one-time intratumoral implantation of Talazoparib, tumors reduced in size by an average of 50%, while untreated tumors increased ~5X in size. Talazoparib dosing appeared to be well tolerated by the mice. Docetaxel implants proved to be an effective method for prostate cancer in vivo with no significant weight loss observed. The local docetaxel spacer group showed sustained tumor inhibition compared to empty implants and an equivalent DTX dose given systemically. At 40 days 89% survival was observed for mice treated with localized DTX implants compared with 0% in all other treatment groups. Histology samples were taken from sacrificed mice and immunohistochemistry are currently underway. Conclusions: Sustained local release of therapeutically relevant doses of Talazoparib and Docetaxel were observed in vitro and in vivo. Therapeutics-loaded implants represent a novel delivery route that are well-tolerated. Sustained release of Talazoparib appears to amplify the therapeutic efficacy of PARP inhibition in BRCA1 mutated breast cancers and sustained release of Docetaxel is an effective chemotherapy option alone or in combination with radiation therapy. These results laid a strong foundation for the use of localized biodegradable implants for the treatment of breast and prostate cancer. This work was supported by the Army- W81XWH-14-1-0092, Breast Cancer Research Foundation and Northeastern University–Dana Farber Cancer Institute collaborative grant. Citation Format: Jodi Belz, Noelle Castilla Ojo, Paige Baldwin, Rajiv Kumar, Anne van de Ven, Karen Liby, Robert Cormack, Mike Makrigiorgos, Srinivas Sridhar4. Sustained release of PARP inhibitor Talazoparib and chemotherapeutic Docetaxel from modified brachytherapy spacers for treatment of breast and prostate cancer. [abstract]. In: Proceedings of the AACR Special Conference on Engineering and Physical Sciences in Oncology; 2016 Jun 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2017;77(2 Suppl):Abstract nr B30.

Abstract B29: Nanoformulations and sustained delivery systems for the PARP inhibitors Olaparib and Talazoparib

Introduction: Poly(ADP-ribose) Polymerase (PARP) plays an important role in a number of DNA repair pathways. PARP inhibitors (PARPi) such as Olaparib and Talazoparib exploit the concept of synthetic lethality by selectively targeting cancer cells with defective DNA repair pathways. These drugs are currently only available in oral form which results in limited bioavailability, poor tumor accumulation, and systemic toxicity, specifically when combined with other DNA damaging agents. Here we report the development of two different delivery techniques including nanoformulations of Olaparib and Talazoparib and a biodegradable implant for localized delivery of Talazoparib. The nanoformulations allow for intravenous or intraperitoneal delivery, providing greater bioavailability and tumor accumulation, while limiting systemic toxicities and the implant provides a sustained release for intratumoral delivery to enhance the dose at the tumor site thereby limiting systemic toxicity. Methods: Nanoparticle formulations of Olaparib and Talazoparib were synthesized and characterized via TEM, DLS, and HPLC to elucidate size, loading, and release before testing in vivo. In vitro experiments for testing nanoformulations include dose response with a panel of ovarian cancer cell lines. In vivo, NanoOlaparib was tested in an IP spread model using 404 cells derived from BRCA2-/-, TP53-/-, PTEN-/- genetically engineered mouse models. Animals were treated IP with NanoOlaparib alone, and in combination with cisplatin. The 1-2 mm long Talazoparib implant was characterized with SEM and HPLC for structure, loading, and release. A spontaneous tumor forming breast cancer model was used to test the Talazoparib implant. Implants were directly injected into the tumor of Brca1Co/Co;MMTV-Cre;p53+/- mice for a one time treatment. Results: PA1 demonstrated high sensitivity to NanoOlaparib which may be attributed to genetic instability at 11/13 polymorphic loci. Both PTEN and BRCA deficiencies resulted in comparable IC509s likely due to synthetic lethality attributed to dysfunctional homologous recombination pathways. NanoTalazoparib is more potent than NanoOlaparib, resulting in a similar relationship in cell line sensitivity with overall lower IC509s. Bioluminescence images illustrated that NanoOlaparib administered IP daily in the ovarian cancer model resulted in a greater inhibition of tumor growth than those treated with oral Olaparib daily. The Talazoparib implant released therapeutically relevant doses of Talazoparib over multiple weeks. In vivo the implant reduced tumors 50% following the one time treatment, while untreated tumors increased significantly in size. Conclusions: Robust nanoparticle formulations of NanoTalazoparib and NanoOlaparib have been successfully developed and characterized. These results show that NanoOlaparib and NanoTalazoparib amplify the efficacy of PARP inhibitors in vivo. The Talazoparib implant provides a safe strategy for sustained local delivery of Talazoparib directly to the tumor. These strategies provide opportunities to increase the efficacy of PARP inhibitors via tailored delivery to the type of tumor. This work was supported by the DOD Ovarian Cancer Research Program under Army- W81XWH-14-1-0092 and IGERT grant NSF-DGE- 0965843. Citation Format: Paige Baldwin, Anders Ohman, Jodi Belz, Jeremy Thong, Noelle Castilla Ojo, Karen Liby, Daniela Dinulescu, Srinivas Sridhar. Nanoformulations and sustained delivery systems for the PARP inhibitors Olaparib and Talazoparib. [abstract]. In: Proceedings of the AACR Special Conference on Engineering and Physical Sciences in Oncology; 2016 Jun 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2017;77(2 Suppl):Abstract nr B29.

Abstract 3100: Nanoformulated Talazoparib and Olaparib for enhanced delivery

Introduction: PARP inhibitors such as Talazoparib and Olaparib exploit deficiencies in DNA repair pathways, making them attractive candidates for treatment of a number of different cancers. These drugs are particularly effective when used in combination with other DNA damaging agents such as chemotherapeutics and radiation therapy. Combination trials, however, have resulted in severe toxicities, necessitating either dose reduction or delay. Dose reduction leads to suboptimal dosing and provides little therapeutic benefit compared to monotherapy. Systemically administered nanoparticles offer a more effective way to selectively accumulate drugs in tumors and bypass toxicities associated with oral delivery. We have developed nanoparticle delivery systems for both Olaparib and Talazoparib in order to improve tumor accumulation while bypassing the toxicity associated with oral administration. Methods: Lipid nanoformulations of Olaparib and Talazoparib have been developed and characterized in regard to size, surface charge, drug loading, release, and stability. NanoTalazoparib has been tested in vitro in breast cancer cell lines including W0069, W780, and HCC1937 which exhibit BRCA1 and 2 mutations, and NanoOlaparib in the lung cancer cell line Calu-6 which also has a defective FA-BRCA pathway. Mice have been treated with NanoOlaparib and NanoTalazoparib alone and in combination with radiation or temozolomide in order to evaluate toxicity. Therapeutic efficacy studies are currently underway. Results: The nanoformulations have been formulated to encapsulate a clinically relevant dose of either Talazoparib or Olaparib and release at 37°C over a period of days, while remaining stable during storage at 4°C. In vitro, both nanoformulations show the same activity as free drug with IC50s in the nanomolar range for these cell lines with varying deficiencies in the BRCA pathway. Mice have shown no appreciable weight loss during treatment with either nanoformulation alone or in combination with other treatment modalities. Conclusion: Nanoformulations of Talazoparib and Olaparib have been developed and characterized to demonstrate activity in vitro and tolerable doses in vivo. We have found that mice tolerate NanoTalazoparib at higher doses when combined with Temozolomide than when given oral Talazoparib. The sustained release from the nanoparticles allows for the nanoformulation to be administered less often than the daily administration for oral drug and the improved tolerability opens the door for combination therapy with both chemotherapeutics and radiation therapy. Therapeutic efficacy studies are underway and we expect that as a monotherapy NanoTalazoparib will be more effective at lower doses than oral Talazoparib, based on the longer circulation time and more selective accumulation in tumors. We also anticipate that combination therapy will be more effective with the nanoformulation, as the maximum tolerated dose is higher than that of the oral drug. Citation Format: Paige Baldwin, Rajiv Kumar, Edward Favours, Karen Liby, Raushan Kurmasheva, David Kozono, Srinivas Sridhar. Nanoformulated Talazoparib and Olaparib for enhanced delivery [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3100. doi:10.1158/1538-7445.AM2017-3100