Chemotherapeutic Treatment Options for Triple Negative Breast Cancer Lauren Barney April 17, 2013

Breast Cancer Subtypes Breast cancer is classified into clinical subtypes based upon receptor expression These subtypes dictate possible therapeutic options and vary in their prognosis Luminal: derived from the luminal cells ER+, PR+ Can use hormonal therapy Less aggressive Basal: derived from myoepithelial cells ER-, PR- No specific target for therapies More aggressive HER2-enriched Luminal A Luminal B Claudin-Low HER2-enriched Basal ER: estrogen receptor PR: progesterone receptor HER2: human epidermal growth factor receptor 2

Luminal and Basal Characteristics Basement membrane Myoepithelial Cells  Basal Luminal Cells  Luminal Basal Luminal Low ER Low HER2 High CK5/6 c-KIT higher High EGFR High p53 mutation High p53 protein High cyclin E Very high vimentin High ER Higher HER2 Low CK5/6 Low c-KIT Low EGFR Low p53 mutation Low p53 protein Low cyclin E Low vimentin Cyclin E: Cyclin E binds to G1 phase Cdk2, which is required for the transition from G1 to S phase of the cell cycle that determines cell division Vimentin: type III intermediate filament protein that is expressed in mesenchymal cells. EMT marker

Triple Negative Breast Cancer 15-25% of all breast cancer, but much higher proportion of all breast cancer mortality Lack ER, PR and HER2 – no targeted therapies Much more aggressive Younger age at diagnosis, high grade, large tumor size, aggressive relapse High proliferation, poor differentiation, basal marker (cytokeratin 5/6) expression, and aggressive clinical course, with early relapse and decreased survival TN tumors have specific morphologic characteristics: elevated mitotic count, tumor necrosis, pushing margin of invasion, and stromal lymphocytic response and high nuclear-cytoplasmic ratio http://ww5.komen.org/BreastCancer/SubtypesofBreastCancer.html Not all basal are TN

TN vs Basal Subtypes The terms triple negative and basal are often used interchangeably in breast cancer subtyping. Triple negative denotes the lack of ER, PR and HER2 receptors (clinical observation) Basal describes the tumors that overexpress those genes that characterized breast basal epithelial cells based on gene expression These often overlap! Basal-like breast cancer is characterized by certain features that include the TN phenotype, but TN and basal-like are not synonymous terms. A discordance of up to 30% has been described between the two groups.

Treatments can be targeted for cancers that express hormonal receptors or HER2; TN remains a clinical challenge. Hormonal therapy: blocks estrogen activity Tamoxifen, ER antagonist Competitively binds to ER & inhibits estrogen effects HER2 targeted therapy Herceptin & others These targeted therapies work really well! There is no specific target on TN cells! Must use cytotoxic chemotherapeutics, surgery, radiation. Tamoxifen

Current Options for TNBC Standard course of treatment is very aggressive: surgery with adjuvant and neoadjuvant chemotherapy and radiation therapy Neoadjuvant: administration of a drug before a main treatment – increases rate of breast conserving therapies and helps to understand a patient’s response to drugs Adjuvant: any therapy given after primary therapy – used when there is a high risk of recurrence The search is on for specific targets! http://ww5.komen.org/BreastCancer/SubtypesofBreastCancer.html http://www.cancer.gov/cancertopics/factsheet/Therapy/adjuvant-breast

TNBC Treatment Chemotherapy typically includes combinations of taxanes (T), anthracyclines (A), and oxazophorines (C) Taxanes: disrupt microtubules & inhibit cell division Paclitaxel, docetaxel Anthracyclines: most effective chemotherapeutics! Three mechanisms: inhibit DNA and RNA synthesis, blocks transcription and replication, creates oxygen free radicals Daunorubicin, doxorubicin, epirubicin, idarubicin Oxazophorine: DNA alkylating agent Cyclophosphamide (C)

Taxane and Anthracycline Based Therapy Typical regimens: AC-T: doxorubicin plus cyclophosphamide every 2 weeks for four cycles followed by docetaxel every 2 weeks for 4 cycles Investigating taxol before AC (not standard therapy) TAC: docetaxel, doxorubicin, and cyclophosphamide every 3 weeks for 6 cycles Different dosing regimens, frequencies can help to improve efficacy Dense dosing is better (more frequent doses are better)

CMF therapy may actually reduce recurrence of TNBC compared to anthracycline or taxane-based treatment CMF is a much older therapeutic regimen than TAC or AC-T Cyclophosphamide (alkylating agent, oxazophorine) Methotrexate (antimetabolite, stops cell growth & division) Fluorouracil (called 5FU; pyrimidine analog, antimetabolite) Many different dosing schedules possible

TN Tumors are Chemosensitive Recently, studies have shown that TNBC is more responsive to anthracycline or anthracycline/taxane chemotherapy than Luminal subtypes Patients who had a complete response to chemotherapy had good prognosis regardless of subtype Despite this, TNBC patients still have a worse distant disease free-survival and a poor prognosis Result of high likelihood of relapse in TNBC HER2+ subtype has a similar response to TNBC Bosch et al. / Cancer Treatment Reviews 36 (2010) 206–215 Basal-like and HER2+/ER subtypes are more sensitive to anthracycline-based neoadjuvant chemotherapy than luminal breast cancers. Patients that had pathologic complete response to chemotherapy had a good prognosis regardless of subtype. The poorer prognosis of basal-like and HER2+/ER breast cancers could be explained by a higher likelihood of relapse in those patients in whom pathologic complete response was not achieved.

Beyond brute force chemo: What are some potential treatment options for TNBC? Current and developing therapies Many in clinical trials Most target proliferative pathways Targets: General proliferation, surface molecules, secondary messengers

Potential Systemic Targets for TNBC

Platinum Agents Platinum agents can bind to DNA and cause cross-linking to occur  cell death Cisplatin, carboplatin and oxaplatin are approved for some types of cancers and are being studied as treatments for TNBC

PARP Inhibitors PARP: poly ADP ribose polymerase Involved in DNA repair by detecting single-strand breaks Can be activated in cells with damaged DNA Several types of cancer are more dependent on PARP, so it can be a good therapeutic target PARP inhibitors prevent breaks from being repaired, causing cell death.

Anti-EGFR EGFR is overexpressed in 45-70% of TNBC Cetuximab is an anti-EGFR antibody used to treat metastatic cancer Breast cancer patients with metastatic disease respond twice as well when Cetuximab is added Other treatments include tyrosine kinase inhibitors (erlotinib, gefitinib) Gefitinib is the only one currently approved for breast cancer, but the others are in clinical trials Inhibits an important signaling pathway and provides a specific target!

Angiogenesis in Cancer Angiogenesis: formation of new blood vessels. Tumors need blood vessels to grow and spread. Angiogenesis inhibitors prevent the formation of new blood vessels, thereby stopping or slowing the growth or spread of tumors. http://www.cancer.gov/cancertopics/factsheet/Therapy/angiogenesis-inhibitors

Anti-Angiogenesis Bevacizumab (Avastin) Monoclonal antibody to VEGF Improves survival in breast cancer patients with combined with Taxol Approved for metastatic breast cancer but benefit isn’t subtype specific – this has since been revoked because it slowed progression but didn’t extend length or quality of life and had many adverse effects Metronomic chemotherapy: repeated, low, less than toxic doses can destroy endothelial cells and prevent angiogenesis, slowing tumor growth – works in clinical trials

Androgen Receptor Nuclear receptor activated by binding testosterone or dihydrotestosterone Closely related to PR Expressed in 75% of breast cancer and 10-20% of TNBC TNBC that express AR are molecularly similar to prostate cancer and could potentially be treated similarly. Bicalutamine: anti-androgen used to treat prostate cancer 17-DMAG: semi-synthetic antibiotic derivative, has shown promise in clinical trials Enzalutamide: androgen agonist used to treat prostate cancer; is in Phase II for TNBC

RTK Inhibitors Suninitib (Sutent) Imatinib (Gleevec) Multiple-target RTK inhibitor All PDGFRs and VEGFRs KIT (CD17) which drives the majority of all GI stromal tumors & several others Imatinib (Gleevec) Prevents phosphorolation of BCR-Abl, inhibiting signaling pathways necessary for cancer cell growth BCR-Abl: Exists only in cancer cells! Worked in vitro; no effect on metastatic breast cancer patients in Phase II

Src Tyrosine Kinase Src is overexpressed in breast cancer Dasatinib: multiple tyrosine kinase inhibitor approved for CML Possible efficacy in breast cancer - small effect seen in Phase II In vitro: basal breast cancer cells were more sensitive! Several others in trials also seem to have promising preclinical activity

mTOR Cell cycle regulator and a downstream effector in the PI3K/PTEN/AKT pathway PTEN is often mutated in TNBC, leading to increased AKT and mTOR activation Everolimus and temsirolimus block mTOR function and inhibit proliferation Everolimus is approved for some types of cancers - currently in clinical trials for TNBC in combination with chemotherapy Temsirolimus is approved for renal cell carcinoma and completed a Phase II trial with promising results

Other possible therapeutic options Hsp90 (heat shock protein 90) – upregulated in response to stress signals; regulates and stabilizes many key proteins, including downstream targets of p53, PI3K, AKT and EGFR – can be recruited to ‘protect’ oncogenic proteins, leading to protein overexpression HDAC (Histone deacetylase) – can effect epigenetics and cause re-expression of epigenetically silenced genes

Other ways to sensitize cells to chemotherapy Inhibition of TGF-beta sensitizes to chemo TRAIL: Lexatumumab (monoclonal antibody in clinical trials) TRAIL controls proliferation & induces apoptosis Chk1 (checkpoint kinase 1): involved in cell cycle control. Inhibition sensitizes proliferating tumor cells to chemotherapies that damage DNA TGFbeta: controls prolif in normal cells; not cancer cells. Trail induces apoptosis

Mutations that Could be Targeted p53 (75% of TNBC) – complex, so target downstream components of pathway Myc (40% of TNBC) Loss of retinoblastoma gene (20% of TNBC) Mutation in BRCA1 or BCRA2 (15-20% of TNBC) Rare: PTEN PIK3CA Amplification of HER2 Amplification of FGFR2 http://www.cancernetwork.com/conference-reports/aacr2013/content/article/10165/2136430 RB: tumor suppressor

We need to get creative: changes in formulation EndoTAG®-1: formulation of paclitaxel combined with neutral and positive lipids Interacts with newly developing and negatively charged endothelial cells that are forming new blood vessels Attacks the activated endothelial cells as they divide Targets blood supply to tumors without affecting healthy tissue Prevents angiogenesis and inhibits tumor growth!! http://www.medigene.com/products-pipeline/development-projects/endotag-1

What’s in clinical trials now? New compounds New drug combinations or dosing regimens New formulations Interesting Current Clinical Trials Re-expression of ER in Triple Negative Breast Cancers Bevacizumab, Metronomic Chemotherapy (CM), Diet and Exercise After Preoperative Chemotherapy for Breast Cancer Laboratory-Treated T Cells After Chemotherapy in Treating Women With Stage II or Stage III Breast Cancer Undergoing Surgery Preoperative Clinical Trial of Sorafenib in Combination With Cisplatin Followed by Paclitaxel for Triple Negative (ER-, PR-, Her2-) Early Stage Breast Cancer Clinicaltrials.gov

Recent news stories March 18, 2013 - Copper depletion shows early success in triple-negative breast cancer April 8, 2013 – Paragazole (HDAC) excels in preclinical models of triple-negative breast cancer April 12, 2013 - Omega-3 Fatty Acids Slow Triple-Negative Breast Cancer Cell Proliferation April 15, 2013 - Nanodiamonds could improve effectiveness of breast cancer treatment

Outlook for now and future Need targeted therapies, new formulations to be able to treat TNBC Combination therapies will be necessary because tumors are heterogeneous and can change Also need to attack tumors from all sides Reaching complete remission and preventing recurrence are key

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