Search

PURPOSE Triple-negative breast cancer (TNBC) is defined by the lack of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER-2) expression. In this study, we compared response to neoadjuvant chemotherapy and survival between patients with TNBC and non-TNBC. PATIENTS AND METHODS Analysis of a prospectively collected clinical database was performed. We included 1,118 patients who received neoadjuvant chemotherapy at M.D. Anderson Cancer Center for stage I-III breast cancer from 1985 to 2004 and for whom complete receptor information were available. Clinical and pathologic parameters, pathologic complete response rates (pCR), survival measurements, and organ-specific relapse rates were compared between patients with TNBC and non-TNBC. RESULTS Two hundred fifty-five patients (23%) had TNBC. Patients with TNBC compared with non-TNBC had significantly higher pCR rates (22% v 11%; P = .034), but decreased 3-year progression-free survival rates ( P < .0001) and 3-year overall survival (OS) rates ( P < .0001). TNBC was associated with increased risk for visceral metastases ( P = .0005), lower risk for bone recurrence ( P = .027), and shorter postrecurrence survival ( P < .0001). Recurrence and death rates were higher for TNBC only in the first 3 years. If pCR was achieved, patients with TNBC and non-TNBC had similar survival ( P = .24). In contrast, patients with residual disease (RD) had worse OS if they had TNBC compared with non-TNBC ( P < .0001). CONCLUSION Patients with TNBC have increased pCR rates compared with non-TNBC, and those with pCR have excellent survival. However, patients with RD after neoadjuvant chemotherapy have significantly worse survival if they have TNBC compared with non-TNBC, particularly in the first 3 years.

A novel, convergent synthesis of aminotriazoloquinazolines is reported. These heterocycles are reliably prepared via a "click-like" reaction between readily available aryl carbodiimides and acyl or aryl hydrazides. Such products are of particular interest with respect to their inhibitory activity against the A

PTPN11 encodes the SHP2 protein tyrosine phosphatase that activates the mitogen-activated protein kinase (MAPK) pathway upstream of KRAS and MEK. PTPN11/Shp2 somatic mutations occur frequently in Juvenile myelomonocytic leukaemia (JMML); however, the role of mutated PTPN11 in lung cancer tumourigenesis and its utility as a therapeutic target has not been fully addressed. We applied mass-spectrometry-based genotyping to DNA extracted from the tumour and matched the normal tissue of 356 NSCLC patients (98 adenocarcinomas (LUAD) and 258 squamous cell carcinomas (LUSC)). Further, PTPN11 mutation cases were identified in additional cohorts, including TCGA, Broad, and MD Anderson datasets and the COSMIC database. PTPN11 constructs harbouring PTPN11 E76A, A72D and C459S mutations were stably expressed in IL-3 dependent BaF3 cells and NSCLC cell lines (NCI-H1703, NCI-H157, NCI-H1299). The MAPK and PI3K pathway activation was evaluated using Western blotting. PTPN11/Shp2 phosphatase activity was measured in whole-cell protein lysates using an Shp2 assay kit. The Shp2 inhibitor (SHPi) was assessed both in vitro and in vivo in a PTPN11-mutated cell line for improved responses to MAPK and PI3K targeting therapies. Somatic PTPN11 hotspot mutations occurred in 4/98 (4.1%) adenocarcinomas and 7/258 (2.7%) squamous cells of 356 NSCLC patients. Additional 26 PTPN11 hotspot mutations occurred in 23 and 3 adenocarcinomas and squamous cell carcinoma, respectively, across the additional cohorts. Mutant PTPN11 significantly increased the IL-3 independent survival of Ba/F3 cells compared to wildtype PTPN11 (p < 0.0001). Ba/F3, NCI-H1703, and NCI-H157 cells expressing mutant PTPN11 exhibited increased PTPN11/Shp2 phosphatase activity and phospho-ERK1/2 levels compared to cells expressing wildtype PTPN11. The transduction of the PTPN11 inactivating mutation C459S into NSCLC cell lines led to decreased phospho-ERK, as well as decreased phospho-AKT in the PTPN11-mutated NCI-H661 cell line. NCI-H661 cells (PTPN11-mutated, KRAS-wild type) were significantly more sensitive to growth inhibition by the PI3K inhibitor copanlisib (IC50: 13.9 ± 4.7 nM) compared to NCI-H1703 (PTPN11/KRAS-wild type) cells (IC50: >10,000 nM). The SHP2 inhibitor, in combination with the PI3K targeting therapy copanlisib, showed no significant difference in tumour development in vivo; however, this significantly prevented MAPK pathway induction in vitro (p < 0.0001). PTPN11/Shp2 demonstrated the in vitro features of a driver oncogene and could potentially sensitize NSCLC cells to PI3K inhibition and inhibit MAPK pathway activation following PI3K pathway targeting.

Background Glioblastoma (GBM) is an aggressive brain cancer that typically results in death in the first 15 months after diagnosis. There have been limited advances in finding new treatments for GBM. In this study, we investigated molecular differences between patients with extremely short (≤ 9 months, Short term survivors, STS) and long survival (≥ 36 months, Long term survivors, LTS). Methods Patients were selected from an in-house cohort (GLIOTRAIN-cohort), using defined inclusion criteria (Karnofsky score > 70; age Results Transcriptomic analysis of tumour samples identified cilium gene signatures as enriched in LTS. Moreover, Immunohistochemical analysis confirmed the presence of cilia in the tumours of LTS. Notably, reverse phase protein array analysis (RPPA) demonstrated increased phosphorylated GAB1 (Y627), SRC (Y527), BCL2 (S70) and RAF (S338) protein expression in STS compared to LTS. Next, we identified 25 unique master regulators (MR) and 13 transcription factors (TFs) belonging to ontologies of integrin signalling and cell cycle to be upregulated in STS. Conclusion Overall, comparison of STS and LTS GBM patients, identifies novel biomarkers and potential actionable therapeutic targets for the management of GBM. Graphical abstract

mp3 file (7.5 MB). In the July edition of the Cancer Discovery podcast, Executive Editor Judy Quong talks with Lydia W.T. Cheung about her paper on the characterization of genetic mutations found in endometrial cancer.

Supplementary Tables 1-3 from High Frequency of PIK3R1 and PIK3R2 Mutations in Endometrial Cancer Elucidates a Novel Mechanism for Regulation of PTEN Protein Stability

Supplementary Figures 1-6 from High Frequency of PIK3R1 and PIK3R2 Mutations in Endometrial Cancer Elucidates a Novel Mechanism for Regulation of PTEN Protein Stability

Supplementary Figure Legends 1-6 from High Frequency of PIK3R1 and PIK3R2 Mutations in Endometrial Cancer Elucidates a Novel Mechanism for Regulation of PTEN Protein Stability

PDF file, 94K, Fig. S1. No induction of ATP7A/ATP7B in cDDPR vs cDDPS cells by TM as determined by RPA (A) and Western blotting (B).

Supplementary Materials and Methods from High Frequency of PIK3R1 and PIK3R2 Mutations in Endometrial Cancer Elucidates a Novel Mechanism for Regulation of PTEN Protein Stability

Background We tested the hypothesis that inhibitor of apoptosis family (IAP) proteins may be altered in BRCA1-mutated ovarian cancers and that could affect the sensitivity to IAP inhibitors. Methods The levels of IAP proteins were evaluated in human cancers and cell lines. Cell lines were used to determine the effects of IAP inhibitors. The in vivo effects of treatments were evaluated in PDX mouse models. Results Expression of X-linked inhibitor of apoptosis (XIAP) is increased in BRCA1-mutated cancers and high levels are associated with improved patient outcomes after platinum chemotherapy. XIAP overexpression is mediated by NF-kB activation and is associated with an optimisation of PARP. BRCA1-mutated cell lines are particularly sensitive to IAP inhibitors due to an inhibitory effect on PARP. Both a BRCA1-mutated cell line with acquired resistance to PARP inhibitors and one with restored BRCA1 remain sensitive to IAP inhibitors. Treatment with IAP inhibitors restores the efficacy of PARP inhibition in these cell lines. The IAP inhibitor LCL161 alone and in combination with a PARP inhibitor, exhibited antitumour effects in PDX mouse models of resistant BRCA2 and 1-mutated ovarian cancer, respectively. Conclusion A clinical trial may be justified to further investigate the utility of IAP inhibitors.

Background: De novo and acquired resistance to HER2 directed therapy is frequently encountered. Upregulation of the phosphatidylinositol-3-kinase (PI3K) pathway is an important mediator of treatment resistance. This can occur through an activating mutation of the PIK3CA gene or PTEN loss. PIK3CA mutations are present in approximately 20% of HER2 positive breast cancers and as such, the PI3K pathway has emerged as an attractive target for restoring sensitivity to HER2 directed therapy. Methods: We performed a single arm, multicentre, open label Phase Ib/II trial. Patients (pts) with advanced HER2-positive breast cancer whose disease had progressed on at least 1 line of Trastuzumab/T-DM1 based treatment in the metastatic setting were eligible if they met following criteria: ECOG PS ≤ 2 and adequate organ function. Pts with treated, controlled brain metastases were permitted to enrol. Exclusion criteria included uncontrolled hypertension or diabetes mellitus. Pts on Phase IB were treated according to a 6+6 study design with a dose escalation schedule of Copanlisib IV (level 1 = 45mg, level 2 = 60mg) on Day 1, 8 and 15 of a 28-day cycle along with a fixed dose of Trastuzumab 2mg/kg weekly. Phase II treatment was the MTD (maximum tolerated dose) of Copanlisib in combination with Trastuzumab. Archival tumour tissue, voluntary biopsies and serial plasma samples were collected for genomic sequencing. Primary endpoints were MTD (Phase I) and clinical benefit rate (CBR) which was defined as complete response (CR) or partial response (PR) at any time point; or stable disease (SD) lasting at least 24 weeks (Phase II). Secondary endpoints included safety and tolerability, tumor response rate, duration of response, time to treatment failure (TTF) and progression free (PFS) and overall survival (OS). Results: Twelve pts were enrolled in Phase IB. No dose limiting toxicity was observed. The MTD was established as Copanlisib 60mg and Trastuzumab 2mg/kg. Fourteen pts were enrolled in Phase II (6 pts treated at the MTD in Phase IB were included in the final Phase II analysis resulting in a total of 20 pts). The median number of lines of prior treatment in the metastatic setting was 3 (1-8). The most common grade 3-4 toxicities encountered in the Phase Ib/II cohorts included hypertension (n=7, 27%), hyperglycaemia (n=2, 8%) and vomiting (n=2, 8%). Three pts discontinued treatment due to toxicity. The median follow-up for the Phase II cohort was 7.5 months (95% CI 6.0-14.5). PR was observed in 4 pts (20%) and SD (at any time point) was seen in 8 pts (40%). The CBR was 30% (n=6). The duration of response was 15.0 weeks (95% CI 4.9 - 16.1). The median TTF was 11.9 weeks (95% CI 7.5 - 21.1). The median PFS was 3.0 mo (95% CI 0.2 - 5.8) and OS was 14.0 mo (95% CI 5.2-22.8). At the time of analysis, 9 of 20 patients were alive. PIK3CA mutations were detectable in the archival tissue of 11 of 26 pts (42%). PIK3CA hotspot mutations (H1047R, E542K and E545K) were detectable in the plasma of all 26 pts at various points throughout treatment. Pre and post treatment biopsies of 2 pts in the Phase IB trial revealed somatic mutations in DNAH3 and TRRAP, the latter of which encodes a PI3K-like protein kinase. Targeted next generation sequencing was performed on the circulating tumour DNA of 20 pts in the Phase II cohort taken before, during and after treatment to further validate these findings and to assess for other mechanisms of response or resistance. The final translational results will be presented at the meeting. Conclusions: The combination of Copanlisib and Trastuzumab is a safe and tolerable regimen and is associated with clinical efficacy in a heavily pre-treated metastatic HER2-positive breast cancer population. Translational studies may have identified novel resistance biomarkers in this pt cohort. Citation Format: Lisa Prior, Niamh M Keegan, Simon J Furney, Janice M Walshe, Giuseppe Gullo, John Crown, M John Kennedy, Diarmuid Smith, John McCaffrey, Catherine M Kelly, Keith Egan, Jennifer Kerr, Mark Given, Niall Sheehy, Peter O'Donovan, Andres Hernando, Ausra Teiserskiene, Imelda Parker, Elaine Kay, Ray McDermott, Maccon M Keane, Seamus O'Reilly, Liam Grogan, Oscar Breathnach, Patrick G Morris, Sinead Toomey, Bryan T Hennessy. Phase Ib/II trial evaluating safety and efficacy of copanlisib (PI3K inhibitor) and trastuzumab in pre-treated advanced HER2-positive breast cancer: Results from the PantHER study [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-16-08.

Background Clinical trials are often considered the gold standard in cancer care. However, patients face barriers in trial participation including distances to cancer centres and personal costs including changing employment status, cost of medications, inpatient admissions, and parking tariffs. Aim Our aim was to compare the distances patients travelled for clinical trials compared to those receiving standard systemic anticancer therapy (SACT). We also investigated the additional costs associated with this. Methods This was a retrospective review of electronic patient medical records. The distance from the patients’ home address to Beaumont was calculated as a one-way journey in kilometres. Patients attending for clinical trials were compared to those receiving standard of care SACT. Results A total of 271 patients receiving standard SACT over a 5-day period and 111 patients enrolled on 24 clinical trials were included. The median one-way distance travelled by patients enrolled in clinical trials was 41.4 km, compared to 14 km in those patients’ receiving standard of care SACT. The median estimated cost was €13 vs €4.20 for those enrolled on clinical trials compared to those receiving standard of care treatment, respectively. Conclusion Patients enrolled on clinical trials often travel more than twice as far to receive their anti-cancer treatment compared to those receiving standard of care SACT and incur an increased cost of travel expenses.

Supplementary Figure Legends 1-4 from Integrative Analysis of Proteomic Signatures, Mutations, and Drug Responsiveness in the NCI 60 Cancer Cell Line Set

All ADCC ratios from Figure 3 and trastuzumab/pertuzumab ADCC comparison

Densitometry analysis for Figure 6B

Supplementary Figures 1-4 from Integrative Analysis of Proteomic Signatures, Mutations, and Drug Responsiveness in the NCI 60 Cancer Cell Line Set

Purpose: Apoptosis is essential for chemotherapy responses. In this discovery and validation study, we evaluated the suitability of a mathematical model of apoptosis execution (APOPTO-CELL) as a stand-alone signature and as a constituent of further refined prognostic stratification tools.Experimental Design: Apoptosis competency of primary tumor samples from patients with stage III colorectal cancer (n = 120) was calculated by APOPTO-CELL from measured protein concentrations of Procaspase-3, Procaspase-9, SMAC, and XIAP. An enriched APOPTO-CELL signature (APOPTO-CELL-PC3) was synthesized to capture apoptosome-independent effects of Caspase-3. Furthermore, a machine learning Random Forest approach was applied to APOPTO-CELL-PC3 and available molecular and clinicopathologic data to identify a further enhanced signature. Association of the signature with prognosis was evaluated in an independent colon adenocarcinoma cohort (TCGA COAD, n = 136).Results: We identified 3 prognostic biomarkers (P = 0.04, P = 0.006, and P = 0.0004 for APOPTO-CELL, APOPTO-CELL-PC3, and Random Forest signatures, respectively) with increasing stratification accuracy for patients with stage III colorectal cancer.The APOPTO-CELL-PC3 signature ranked highest among all features. The prognostic value of the signatures was independently validated in stage III TCGA COAD patients (P = 0.01, P = 0.04, and P = 0.02 for APOPTO-CELL, APOPTO-CELL-PC3, and Random Forest signatures, respectively). The signatures provided further stratification for patients with CMS1-3 molecular subtype.Conclusions: The integration of a systems-biology–based biomarker for apoptosis competency with machine learning approaches is an appealing and innovative strategy toward refined patient stratification. The prognostic value of apoptosis competency is independent of other available clinicopathologic and molecular factors, with tangible potential of being introduced in the clinical management of patients with stage III colorectal cancer. Clin Cancer Res; 23(5); 1200–12. ©2016 AACR.

Kaplan-Meier estimates for disease-free and overall survival for n=120 stage III patients of the discovery cohort categorized based on TN staging, tumor location and lymphovascular invasion. Additional exploratory analyses investigated the prognostic value of the APOPTO-CELL-PC3 signature within each sub-group identified by the clinical features.

Purpose: We studied the expression levels of cyclins B1, D1, and E1 and the implications of cyclin overexpression for patient outcomes in distinct breast cancer subtypes defined by clinical variables and transcriptional profiling.Experimental Design: The expression levels of cyclins B1, D1, and E1 were quantified in 779 breast tumors and 53 cell lines using reverse phase protein arrays and/or transcriptional profiling.Results: Whereas cyclin E1 overexpression was a specific marker of triple-negative and basal-like tumors, cyclin B1 overexpression occurred in poor prognosis hormone receptor–positive, luminal B and basal-like breast cancers. Cyclin D1 overexpression occurred in luminal and normal-like cancers. Breast cancer subgroups defined by integrated expression of cyclins B1, D1, and E1 correlated significantly (P < 0.000001) with tumor subtypes defined by transcriptional profiling and clinical criteria. Across three hormone receptor–positive data sets, cyclin B1 was the dominant cyclin associated with poor prognosis in univariate and multivariate analyses. Although CCNE1 was present in significantly higher copy numbers in basal-like versus other subtypes (ANOVA P < 0.001), CCNB1 gene copy number did not show gain in breast cancer. Instead, cyclin B1 expression was increased in tumors with co-occurrence of TP53 mutations and MYC amplification, a combination that seems to characterize basal-like and luminal B tumors. CCNB1 gene expression was significantly correlated with PLK, CENPE, and AURKB gene expression.Conclusion: Cyclins B1, D1, and E1 have distinct expressions in different breast cancer subtypes. Novel PLK, CENPE, and AURKB inhibitors should be assessed for therapeutic utility in poor prognosis cyclin B1–overexpressing breast cancers.

Related Article from A Systems Approach to Analysis of Molecular Complexity in Breast Cancer

Unadjusted and multivariate Cox proportional hazards analyses to examine the association of APOPTO-CELL and APOPTO-CELL-PC3 signatures with DFS in n=157 stage III patients of the expansion cohort.

Purpose:Antibody-dependent cell-mediated cytotoxicity (ADCC) is one mechanism of action of the monoclonal antibody (mAb) therapies trastuzumab and pertuzumab. Tyrosine kinase inhibitors (TKIs), like lapatinib, may have added therapeutic value in combination with mAbs through enhanced ADCC activity. Using clinical data, we examined the impact of lapatinib on HER2/EGFR expression levels and natural killer (NK) cell gene signatures. We investigated the ability of three TKIs (lapatinib, afatinib, and neratinib) to alter HER2/immune-related protein levels in preclinical models of HER2-positive (HER2+) and HER2-low breast cancer, and the subsequent effects on trastuzumab/pertuzumab-mediated ADCC.Experimental Design:Preclinical studies (proliferation assays, Western blotting, high content analysis, and flow cytometry) employed HER2+ (SKBR3 and HCC1954) and HER2-low (MCF-7, T47D, CAMA-1, and CAL-51) breast cancer cell lines. NCT00524303 provided reverse phase protein array–determined protein levels of HER2/pHER2/EGFR/pEGFR. RNA-based NK cell gene signatures (CIBERSORT/MCP-counter) post-neoadjuvant anti-HER2 therapy were assessed (NCT00769470/NCT01485926). ADCC assays utilized flow cytometry–based protocols.Results:Lapatinib significantly increased membrane HER2 levels, while afatinib and neratinib significantly decreased levels in all preclinical models. Single-agent lapatinib increased HER2 or EGFR levels in 10 of 11 (91%) tumor samples. NK cell signatures increased posttherapy (P = 0.03) and associated with trastuzumab response (P = 0.01). TKI treatment altered mAb-induced NK cell–mediated ADCC in vitro, but it did not consistently correlate with HER2 expression in HER2+ or HER2-low models. The ADCC response to trastuzumab and pertuzumab combined did not exceed either mAb alone.Conclusions:TKIs differentially alter tumor cell phenotype which can impact NK cell–mediated response to coadministered antibody therapies. mAb-induced ADCC response is relevant when rationalizing combinations for clinical investigation.

In this issue of Clinical Cancer Research, Andre et al. apply high-resolution arrays to elucidate copy number anomalies in breast cancer. They identify distinct copy number anomaly patterns in different breast cancer subtypes that implicate a number of genes as potential therapeutic targets and as potential markers of therapy responsiveness.

Supplementary Tables 1-4 from Integrative Analysis of Proteomic Signatures, Mutations, and Drug Responsiveness in the NCI 60 Cancer Cell Line Set

Densitometry for Figure 1D

Detailed description of the measurements used as inputs for the apoptosis-based signatures in the discovery, expansion and validation cohorts.

Aberrations in oncogenes and tumor suppressors frequently affect the activity of critical signal transduction pathways. To analyze systematically the relationship between the activation status of protein networks and other characteristics of cancer cells, we did reverse phase protein array (RPPA) profiling of the NCI60 cell lines for total protein expression and activation-specific markers of critical signaling pathways. To extend the scope of the study, we merged those data with previously published RPPA results for the NCI60. Integrative analysis of the expanded RPPA data set revealed five major clusters of cell lines and five principal proteomic signatures. Comparison of mutations in the NCI60 cell lines with patterns of protein expression showed significant associations for PTEN, PIK3CA, BRAF, and APC mutations with proteomic clusters. PIK3CA and PTEN mutation enrichment were not cell lineage-specific but were associated with dominant yet distinct groups of proteins. The five RPPA-defined clusters were strongly associated with sensitivity to standard anticancer agents. RPPA analysis identified 27 protein features significantly associated with sensitivity to paclitaxel. The functional status of those proteins was interrogated in a paclitaxel whole genome small interfering RNA (siRNA) library synthetic lethality screen and confirmed the predicted associations with drug sensitivity. These studies expand our understanding of the activation status of protein networks in the NCI60 cancer cell lines, demonstrate the importance of the direct study of protein expression and activation, and provide a basis for further studies integrating the information with other molecular and pharmacological characteristics of cancer. Mol Cancer Ther; 9(2); 257–67

Supplementary Data from Integrative Analysis of Cyclin Protein Levels Identifies Cyclin B1 as a Classifier and Predictor of Outcomes in Breast Cancer

Supplementary Data from Pharmacodynamic Markers of Perifosine Efficacy

Densitometry analysis for Figure 6D/E

Purpose: Using reverse phase protein array, we measured protein expression associated with response to primary chemotherapy in patients with advanced-stage, high-grade serous ovarian cancer.Experimental Design: Tumor samples were obtained from 45 patients with advanced high-grade serous cancers from the Gynecology Tumor Bank at the British Columbia Cancer Agency. Treatment consisted of platinum-based chemotherapy following debulking surgery. Protein lysates were prepared from fresh frozen tumor samples, and 80 validated proteins from signaling pathways implicated in ovarian carcinogenesis were measured by reverse phase protein array. Normalization of Ca-125 by the 3rd cycle of chemotherapy was chosen as the primary outcome measure of chemotherapy response. Logistic regression was used for multivariate analysis to identify protein predictors of Ca-125 normalization and Cox regression to test for the association between protein expression and progression-free survival. A significance level of P ≤ 0.05 was used.Results: The mean age at diagnosis was 56.8 years. epidermal growth factor receptor, YKL-40, and several transforming growth factor β (TGF-β) pathway proteins [c-jun–NH2–kinase (JNK), JNK phosphorylated at residues 183 and 185, plasminogen activator inhibitor 1, Smad3, TAZ] showed significant associations with Ca-125 normalization on univariate testing. On multivariate analysis, epidermal growth factor receptor (P < 0.02), JNK (P < 0.01), and Smad3 (P < 0.04) were significantly associated with normalization of Ca-125. Contingency table analysis of pathway-classified proteins revealed that the selection of TGF-β pathway proteins was unlikely because of false discovery (P < 0.007; Bonferroni adjusted).Conclusion: TGF-β pathway signaling likely plays an important role as a marker or mediator of chemoresistance in advanced serous ovarian cancer. On this basis, future studies to develop and validate a useful predictor of treatment failure are warranted. Clin Cancer Res; 16(10); 2852–60. ©2010 AACR.

Flow cytometry MFI and % positive cells data for fluorescently labelled trastuzumab and pertuzumab in SKBR3, HCC1954, T47D and MCF-7 treated with TKIs

Rituximab ADCC negative control

Purpose: It is critical to develop methods to quantify the early pharmacodynamic effects of targeted therapeutics in vivo to make drug development more efficient and ensure biologically relevant dosing. Furthermore, an ability to identify patients likely to respond to targeted therapeutics would decrease the size, duration, and cost of clinical trials, resulting in more efficient translation to improved patient outcomes. Recent studies suggest that perifosine inhibits the phosphatidylinositol-3′-kinase (PI3K) pathway by preventing cell membrane recruitment of the AKT pleckstrin homology domain.Experimental Design: A novel functional proteomics technology, reverse phase protein array, was used to establish and quantify pharmacodynamic markers of perifosine efficacy.Results: Perifosine selectively prevents AKT recruitment to the membrane and blocks activation of downstream effectors. Perifosine inhibited breast, ovarian, and prostate cancer models. Growth inhibition was associated with apoptosis. Activation of AKT as a consequence of genomic aberrations predicted perifosine efficacy. In cell lines and xenografts, there was a highly statistically significant correlation between the degree of antitumor efficacy of different perifosine doses and quantified down-regulation of phosphorylation of AKT and of its downstream targets, particularly S6.Conclusions: Because of a strong correlation between proportional modulation of PI3K pathway biomarkers and quantified perifosine efficacy, it is likely that early measurement of such pharmacodynamic biomarkers with reverse phase protein array will optimize selection of responding patients and guide perifosine dosing. Furthermore, PI3K pathway activation status may allow baseline selection of patients most likely to respond to perifosine alone or in combination with other therapies.

Flow cytometry control scatter plots for trastuzumab-488 and pertuzumab-550

MFI comparison at T0 and T6 for T47D/MCF-7

Supplementary Table S1 from Functional Proteomic Analysis of Advanced Serous Ovarian Cancer Using Reverse Phase Protein Array: TGF-β Pathway Signaling Indicates Response to Primary Chemotherapy