bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from The Folate Pathway Inhibitor Pemetrexed Pleiotropically Enhances Effects of Cancer Immunotherapy
Authors: Schaer David A. , Geeganage Sandaruwan , Amaladas Nelusha , Lu Zhao Hai , Rasmussen Erik R. , Sonyi Andreas , Chin Darin , Capen Andrew , Li Yanxia , Meyer Catalina M. , Jones Bonita D. , Huang Xiaodong , Luo Shuang , Carpenito Carmine , Roth Kenneth D. , Nikolayev Alexander , Tan Bo , Brahmachary Manisha , Chodavarapu Krishna , Dorsey Frank C. , Manro Jason R. , Doman Thompson N. , Donoho Gregory P. , Surguladze David , Hall Gerald E. , Kalos Michael , Novosiadly Ruslan D.
Published: April 2023

AbstractPurpose:
Combination strategies leveraging chemotherapeutic agents and immunotherapy have held the promise as a method to improve benefit for patients with cancer. However, most chemotherapies have detrimental effects on immune homeostasis and differ in their ability to induce immunogenic cell death (ICD). The approval of pemetrexed and carboplatin with anti-PD-1 (pembrolizumab) for treatment of non–small cell lung cancer represents the first approved chemotherapy and immunotherapy combination. Although the clinical data suggest a positive interaction between pemetrexed-based chemotherapy and immunotherapy, the underlying mechanism remains unknown.
Experimental Design:
Mouse tumor models (MC38, Colon26) and high-content biomarker studies (flow cytometry, Quantigene Plex, and nCounter gene expression analysis) were deployed to obtain insights into the mechanistic rationale behind the efficacy observed with pemetrexed/anti-PD-L1 combination. ICD in tumor cell lines was assessed by calreticulin and HMGB-1 immunoassays, and metabolic function of primary T cells was evaluated by Seahorse analysis.
Results:
Pemetrexed treatment alone increased T-cell activation in mouse tumors in vivo, robustly induced ICD in mouse tumor cells and exerted T-cell–intrinsic effects exemplified by augmented mitochondrial function and enhanced T-cell activation in vitro. Increased antitumor efficacy and pronounced inflamed/immune activation were observed when pemetrexed was combined with anti-PD-L1.
Conclusions:
Pemetrexed augments systemic intratumor immune responses through tumor intrinsic mechanisms including immunogenic cell death, T-cell–intrinsic mechanisms enhancing mitochondrial biogenesis leading to increased T-cell infiltration/activation along with modulation of innate immune pathways, which are significantly enhanced in combination with PD-1 pathway blockade.

See related commentary by Buque et al., p. 6890
Data from Circulating Tumor Cell Clusters in Patients with Metastatic Breast Cancer: a SWOG S0500 Translational Medicine Study
Authors: Paoletti Costanza , Miao Jieling , Dolce Emily M. , Darga Elizabeth P. , Repollet Madeline I. , Doyle Gerald V. , Gralow Julie R. , Hortobagyi Gabriel N. , Smerage Jeffrey B. , Barlow William E. , Hayes Daniel F.
Published: April 2023

AbstractPurpose:
Metastasis requires malignant cell circulation from the primary to a distant tissue. Elevated levels of circulating tumor cells (CTC) portend a poor prognosis in breast and other cancers. Recent studies have suggested that CTC clusters may be a factor in the metastatic process. We conducted a prospective retrospective study of the SWOG0500 clinical trial to test whether CTC clusters are associated with poorer prognosis.
Experimental Design:
CTC CellSearch galleries from SWOG0500 trial were reread using prespecified criteria for CTC clusters, doublets, and enumeration. Survival analysis methods include Kaplan–Meier plots and log-rank tests.
Results:
Patients were classified into three prognostic subgroups based on baseline CTC/7.5 mL whole blood (WB): Arm A: <5CTC; Arm B/C: ≥5CTC and then B (<5CTC) and C (≥5CTC)/7.5 mL WB at first follow-up. At baseline, 19% of patients had CTC doublets or clusters, which were more likely in Arm B/C versus Arm A (38% vs. 1.4%; P < 0.0001). Furthermore, doublets or clusters were significantly more common in patients who were ultimately assigned to Arm C versus B (54% vs. 25%; P < 0.0001). In Arm C, doublets and clusters were associated with worse overall survival than only doublets, clusters, or no doublets nor clusters at baseline (P = 0.008) and first follow-up (P = 0.010). When compared with enumeration alone, doublets, clusters, or both were not prognostic in patients who had 5–19 or ≥20 CTC/7.5 mL WB.
Conclusions:
In patients with metastatic breast cancer starting first-line chemotherapy, mortality is independent of the presence of CTC clusters, but rather depends on the number of CTC/7.5 mL WB.
Data from Nomogram to Predict the Benefit of Intensive Treatment for Locoregionally Advanced Head and Neck Cancer
Authors: Mell Loren K. , Shen Hanjie , Nguyen-Tân Phuc Felix , Rosenthal David I. , Zakeri Kaveh , Vitzthum Lucas K. , Frank Steven J. , Schiff Peter B. , Trotti Andy M. , Bonner James A. , Jones Christopher U. , Yom Sue S. , Thorstad Wade L. , Wong Stuart J. , Shenouda George , Ridge John A. , Zhang Qiang E. , Le Quynh-Thu
Published: April 2023

AbstractPurpose:
Previous studies indicate that the benefit of therapy depends on patients' risk for cancer recurrence relative to noncancer mortality (ω ratio). We sought to test the hypothesis that patients with head and neck cancer (HNC) with a higher ω ratio selectively benefit from intensive therapy.
Experimental Design:
We analyzed 2,688 patients with stage III–IVB HNC undergoing primary radiotherapy (RT) with or without systemic therapy on three phase III trials (RTOG 9003, RTOG 0129, and RTOG 0522). We used generalized competing event regression to stratify patients according to ω ratio and compared the effectiveness of intensive therapy as a function of predicted ω ratio (i.e., ω score). Intensive therapy was defined as treatment on an experimental arm with altered fractionation and/or multiagent concurrent systemic therapy. A nomogram was developed to predict patients' ω score on the basis of tumor, demographic, and health factors. Analysis was by intention to treat.
Results:
Decreasing age, improved performance status, higher body mass index, node-positive status, P16-negative status, and oral cavity primary predicted a higher ω ratio. Patients with ω score ≥0.80 were more likely to benefit from intensive treatment [5-year overall survival (OS), 70.0% vs. 56.6%; HR of 0.73, 95% confidence interval (CI): 0.57–0.94; P = 0.016] than those with ω score <0.80 (5-year OS, 46.7% vs. 45.3%; HR of 1.02, 95% CI: 0.92-1.14; P = 0.69; P = 0.019 for interaction). In contrast, the effectiveness of intensive therapy did not depend on risk of progression.
Conclusions:
Patients with HNC with a higher ω score selectively benefit from intensive treatment. A nomogram was developed to help select patients for intensive therapy.
Data from Circulating Cytokines Predict Immune-Related Toxicity in Melanoma Patients Receiving Anti-PD-1–Based Immunotherapy
Authors: Lim Su Y. , Lee Jenny H. , Gide Tuba N. , Menzies Alexander M. , Guminski Alexander , Carlino Matteo S. , Breen Edmond J. , Yang Jean Y.H. , Ghazanfar Shila , Kefford Richard F. , Scolyer Richard A. , Long Georgina V. , Rizos Helen
Published: April 2023

AbstractPurpose:
Combination PD-1 and CTLA-4 inhibitor therapy has dramatically improved the survival of patients with advanced melanoma but is also associated with significant immune-related toxicities. This study sought to identify circulating cytokine biomarkers of treatment response and immune-related toxicity.
Experimental Design:
The expression of 65 cytokines was profiled longitudinally in 98 patients with melanoma treated with PD-1 inhibitors, alone or in combination with anti-CTLA-4, and in an independent validation cohort of 49 patients treated with combination anti-PD-1 and anti-CTLA-4. Cytokine expression was correlated with RECIST response and immune-related toxicity, defined as toxicity that warranted permanent discontinuation of treatment and administration of high-dose steroids.
Results:
Eleven cytokines were significantly upregulated in patients with severe immune-related toxicities at baseline (PRE) and early during treatment (EDT). The expression of these 11 cytokines was integrated into a single toxicity score, the CYTOX (cytokine toxicity) score, and the predictive utility of this score was confirmed in the discovery and validation cohorts. The AUC for the CYTOX score in the validation cohort was 0.68 at PRE [95% confidence interval (CI), 0.51–0.84; P = 0.037] and 0.70 at EDT (95% CI, 0.55–0.85; P = 0.017) using ROC analysis.
Conclusions:
The CYTOX score is predictive of severe immune-related toxicity in patients with melanoma treated with combination anti-CTLA-4 and anti-PD-1 immunotherapy. This score, which includes proinflammatory cytokines such as IL1a, IL2, and IFNα2, may help in the early management of severe, potentially life-threatening immune-related toxicity.

See related commentary by Johnson and Balko, p. 1452
Data from Predicted Prognosis of Patients with Pancreatic Cancer by Machine Learning
Authors: Yokoyama Seiya , Hamada Taiji , Higashi Michiyo , Matsuo Kei , Maemura Kosei , Kurahara Hiroshi , Horinouchi Michiko , Hiraki Tsubasa , Sugimoto Tomoyuki , Akahane Toshiaki , Yonezawa Suguru , Kornmann Marko , Batra Surinder K. , Hollingsworth Michael A. , Tanimoto Akihide
Published: April 2023

AbstractPurpose:
Pancreatic cancer remains a disease of high mortality despite advanced diagnostic techniques. Mucins (MUC) play crucial roles in carcinogenesis and tumor invasion in pancreatic cancers. MUC1 and MUC4 expression are related to the aggressive behavior of human neoplasms and a poor patient outcome. In contrast, MUC2 is a tumor suppressor, and we have previously reported that MUC2 is a favorable prognostic factor in pancreatic neoplasia. This study investigates whether the methylation status of three mucin genes from postoperative tissue specimens from patients with pancreatic neoplasms could serve as a predictive biomarker for outcome after surgery.
Experimental Design:
We evaluated the methylation status of MUC1, MUC2, and MUC4 promoter regions in pancreatic tissue samples from 191 patients with various pancreatic lesions using methylation-specific electrophoresis. Then, integrating these results and clinicopathologic features, we used support vector machine-, neural network-, and multinomial-based methods to develop a prognostic classifier.
Results:
Significant differences were identified between the positive- and negative-prediction classifiers of patients in 5-year overall survival (OS) in the cross-validation test. Multivariate analysis revealed that these prognostic classifiers were independent prognostic factors analyzed by not only neoplastic tissues but also nonneoplastic tissues. These classifiers had higher predictive accuracy for OS than tumor size, lymph node metastasis, distant metastasis, and age and can complement the prognostic value of the TNM staging system.
Conclusions:
Analysis of epigenetic changes in mucin genes may be of diagnostic utility and one of the prognostic predictors for patients with pancreatic ductal adenocarcinoma.
Data from CD47-Targeted Near-Infrared Photoimmunotherapy for Human Bladder Cancer
Authors: Kiss Bernhard , van den Berg Nynke S. , Ertsey Robert , McKenna Kelly , Mach Kathleen E. , Zhang Chiyuan Amy , Volkmer Jens-Peter , Weissman Irving L. , Rosenthal Eben L. , Liao Joseph C.
Published: April 2023

AbstractPurpose:
Near-infrared photoimmunotherapy (NIR-PIT) is a localized molecular cancer therapy combining a photosensitizer-conjugated mAb and light energy. CD47 is an innate immune checkpoint widely expressed on bladder cancer cells, but absent from luminal normal urothelium. Targeting CD47 for NIR-PIT has the potential to selectively induce cancer cell death and minimize damage to normal urothelium.
Experimental Design:
The cytotoxic effect of NIR-PIT with anti-CD47-IR700 was investigated in human bladder cancer cell lines and primary human bladder cancer cells derived from fresh surgical samples. Phagocytosis assays were performed to evaluate macrophage activity after NIR-PIT. Anti-CD47-IR700 was administered to murine xenograft tumor models of human bladder cancer for in vivo molecular imaging and NIR-PIT.
Results:
Cytotoxicity in cell lines and primary bladder cancer cells significantly increased in a light-dose–dependent manner with CD47-targeted NIR-PIT. Phagocytosis of cancer cells significantly increased with NIR-PIT compared with antibody alone (P = 0.0002). In vivo fluorescence intensity of anti-CD47-IR700 in tumors reached a peak 24-hour postinjection and was detectable for at least 14 days. After a single round of CD47-targeted NIR-PIT, treated animals showed significantly slower tumor growth compared with controls (P < 0.0001). Repeated CD47-targeted NIR-PIT treatment further slowed tumor growth (P = 0.0104) and improved survival compared with controls.
Conclusions:
CD47-targeted NIR-PIT increased direct cancer cell death and phagocytosis resulting in inhibited tumor growth and improved survival in a murine xenograft model of human bladder cancer.
Data from Genetic Characterization and Prognostic Relevance of Acquired Uniparental Disomies in Cytogenetically Normal Acute Myeloid Leukemia
Authors: Walker Christopher J. , Kohlschmidt Jessica , Eisfeld Ann-Kathrin , Mrózek Krzysztof , Liyanarachchi Sandya , Song Chi , Nicolet Deedra , Blachly James S. , Bill Marius , Papaioannou Dimitrios , Oakes Christopher C. , Giacopelli Brian , Genutis Luke K. , Maharry Sophia E. , Orwick Shelley , Archer Kellie J. , Powell Bayard L. , Kolitz Jonathan E. , Uy Geoffrey L. , Wang Eunice S. , Carroll Andrew J. , Stone Richard M. , Byrd John C. , de la Chapelle Albert , Bloomfield Clara D.
Published: April 2023

AbstractPurpose:
Uniparental disomy (UPD) is a way cancer cells duplicate a mutated gene, causing loss of heterozygosity (LOH). Patients with cytogenetically normal acute myeloid leukemia (CN-AML) do not have microscopically detectable chromosome abnormalities, but can harbor UPDs. We examined the prognostic significance of UPDs and frequency of LOH in patients with CN-AML.

Experimental Design: We examined the frequency and prognostic significance of UPDs in a set of 425 adult patients with de novo CN-AML who were previously sequenced for 81 genes typically mutated in cancer. Associations of UPDs with outcome were analyzed in the 315 patients with CN-AML younger than 60 years.
Results:
We detected 127 UPDs in 109 patients. Most UPDs were large and typically encompassed all or most of the affected chromosome arm. The most common UPDs occurred on chromosome arms 13q (7.5% of patients), 6p (2.8%), and 11p (2.8%). Many UPDs significantly cooccurred with mutations in genes they encompassed, including 13q UPD with FLT3-internal tandem duplication (FLT3-ITD; P < 0.001), and 11p UPD with WT1 mutations (P = 0.02). Among patients younger than 60 years, UPD of 11p was associated with longer overall survival (OS) and 13q UPD with shorter disease-free survival (DFS) and OS. In multivariable models that accounted for known prognostic markers, including FLT3-ITD and WT1 mutations, UPD of 13q maintained association with shorter DFS, and UPD of 11p maintained association with longer OS.
Conclusions:
LOH mediated by UPD is a recurrent feature of CN-AML. Detection of UPDs of 13q and 11p might be useful for genetic risk stratification of patients with CN-AML.
Data from High-Plex Predictive Marker Discovery for Melanoma Immunotherapy–Treated Patients Using Digital Spatial Profiling
Authors: Toki Maria I. , Merritt Christopher R. , Wong Pok Fai , Smithy James W. , Kluger Harriet M. , Syrigos Konstantinos N. , Ong Giang T. , Warren Sarah E. , Beechem Joseph M. , Rimm David L.
Published: April 2023

AbstractPurpose:
Protein expression in formalin-fixed, paraffin-embedded tissue is routinely measured by IHC or quantitative fluorescence (QIF) on a handful of markers on a single section. Digital spatial profiling (DSP) allows spatially informed simultaneous assessment of multiple biomarkers. Here we demonstrate the DSP technology using a 44-plex antibody cocktail to find protein expression that could potentially be used to predict response to immune therapy in melanoma.

Experimental Design: The NanoString GeoMx DSP technology is compared with automated QIF (AQUA) for immune marker compartment-specific measurement and prognostic value in non–small cell lung cancer (NSCLC). Then we use this tool to search for novel predictive markers in a cohort of 60 patients with immunotherapy-treated melanoma on a tissue microarray using a 44-plex immune marker panel measured in three compartments (macrophage, leukocyte, and melanocyte) generating 132 quantitative variables.
Results:
The spatially informed variable assessment by DSP validates by both regression and variable prognostication compared with QIF for stromal CD3, CD4, CD8, CD20, and PD-L1 in NSCLC. From the 132 variables, 11 and 15 immune markers were associated with prolonged progression-free survival (PFS) and overall survival (OS). Notably, we find PD-L1 expression in CD68-positive cells (macrophages) and not in tumor cells was a predictive marker for PFS, OS, and response.
Conclusions:
DSP technology shows high concordance with QIF and validates based on both regression and outcome assessment. Using the high-plex capacity, we found a series of expression patterns associated with outcome, including that the expression of PD-L1 in macrophages is associated with response.
Data from A Novel Mechanism Driving Poor-Prognosis Prostate Cancer: Overexpression of the DNA Repair Gene, Ribonucleotide Reductase Small Subunit M2 (RRM2)
Authors: Mazzu Ying Z. , Armenia Joshua , Chakraborty Goutam , Yoshikawa Yuki , Coggins Si'Ana A. , Nandakumar Subhiksha , Gerke Travis A. , Pomerantz Mark M. , Qiu Xintao , Zhao Huiyong , Atiq Mohammad , Khan Nabeela , Komura Kazumasa , Lee Gwo-Shu Mary , Fine Samson W. , Bell Connor , O'Connor Edward , Long Henry W. , Freedman Matthew L. , Kim Baek , Kantoff Philip W.
Published: April 2023

AbstractPurpose:
Defects in genes in the DNA repair pathways significantly contribute to prostate cancer progression. We hypothesize that overexpression of DNA repair genes may also drive poorer outcomes in prostate cancer. The ribonucleotide reductase small subunit M2 (RRM2) is essential for DNA synthesis and DNA repair by producing dNTPs. It is frequently overexpressed in cancers, but very little is known about its function in prostate cancer.
Experimental Design:
The oncogenic activity of RRM2 in prostate cancer cells was assessed by inhibiting or overexpressing RRM2. The molecular mechanisms of RRM2 function were determined. The clinical significance of RRM2 overexpression was evaluated in 11 prostate cancer clinical cohorts. The efficacy of an RRM2 inhibitor (COH29) was assessed in vitro and in vivo. Finally, the mechanism underlying the transcriptional activation of RRM2 in prostate cancer tissue and cells was determined.
Results:
Knockdown of RRM2 inhibited its oncogenic function, whereas overexpression of RRM2 promoted epithelial mesenchymal transition in prostate cancer cells. The prognostic value of RRM2 RNA levels in prostate cancer was confirmed in 11 clinical cohorts. Integrating the transcriptomic and phosphoproteomic changes induced by RRM2 unraveled multiple oncogenic pathways downstream of RRM2. Targeting RRM2 with COH29 showed excellent efficacy. Thirteen putative RRM2-targeting transcription factors were bioinformatically identified, and FOXM1 was validated to transcriptionally activate RRM2 in prostate cancer.
Conclusions:
We propose that increased expression of RRM2 is a mechanism driving poor patient outcomes in prostate cancer and that its inhibition may be of significant therapeutic value.
Data from Long-Term Survival Analysis of Adjuvant Chemotherapy with or without Trastuzumab in Patients with T1, Node-Negative HER2-Positive Breast Cancer
Authors: He Xuexin , Ji Jiali , Tian Mei , Esteva Francisco J. , Hortobagyi Gabriel N. , Yeung Sai-Ching Jim
Published: April 2023

AbstractPurpose:
Adjuvant therapy for small, node-negative HER2-positive breast cancer (HER2+ BC) is controversial. We aimed to identify the subgroup that would benefit most from adjuvant chemotherapy and trastuzumab.
Experimental Design:
We reviewed records of patients with pT1N0M0 HER2+ BC treated at our institution from January 1, 1998, through October 31, 2009. We compared three groups: A, no adjuvant chemotherapy; B, adjuvant chemotherapy only; and C, adjuvant chemotherapy with trastuzumab. We evaluated disease-free survival (DFS), overall survival (OS), distant recurrence-free survival (DRFS), and breast cancer-specific survival (BCSS) in each group.
Results:
We reviewed 587 consecutive patients with a median follow-up of 123.0 months. The 10-year DFS rate was 81.0%, 65.4%, and 97.3% in groups A, B, and C, respectively (P < 0.001). The restricted mean survival time ratio did not differ between groups A and B [ratio = 0.982; 95% confidence interval (CI), 0.930–1.036; P = 0.498). Cox regression showed that adjuvant chemotherapy with trastuzumab was associated with better DFS compared with no adjuvant chemotherapy [hazard ratio (HR), 0.071; 95% CI, 0.025–0.204; P < 0.001). Larger tumor size was associated with short DFS (HR, 2.384; 95% CI, 1.549–3.056; P < 0.001); improvements in DFS, OS, DRFS, and BCSS were observed with adjuvant chemotherapy plus trastuzumab in patients with tumors ≥0.8-cm diameter. Receiving adjuvant chemotherapy with trastuzumab was not associated with improved DFS, OS, or DRFS for tumors <0.8 cm.
Conclusions:
Adjuvant chemotherapy plus trastuzumab should be recommended for patients with pT1N0M0 HER2+ BC ≥0.8 cm in diameter; adjuvant therapy may not be necessary for tumors <0.8 cm.
Data from A Phase Ib/II Study of Ramucirumab in Combination with Emibetuzumab in Patients with Advanced Cancer
Authors: Harding James J. , Zhu Andrew X. , Bauer Todd M. , Choueiri Toni K. , Drilon Alexander , Voss Martin H. , Fuchs Charles S. , Abou-Alfa Ghassan K. , Wijayawardana Sameera R. , Wang Xuejing Aimee , Moser Brian A. , Uruñuela Arantxa , Wacheck Volker , Bendell Johanna C.
Published: April 2023

AbstractPurpose:
Inhibition of the VEGFR-2 blocks angiogenesis and attenuates tumor growth, but cancers may evade this effect through activation of the hepatocyte growth factor receptor MET. Here we report results of the phase Ib/II study of ramucirumab, a monoclonal anti-VEGFR-2 antibody, plus the anti-MET mAb emibetuzumab.
Patients and Methods:
A 3+3 dose escalation of emibetuzumab plus ramucirumab (phase Ib) was followed by tumor-specific expansion cohorts. Primary objectives were to determine the recommended phase II dose and to evaluate antitumor activity. Secondary objectives included safety, pharmacokinetics, and immunogenicity. Tumoral MET expression was explored by immunohistochemistry (IHC).
Results:
A total of 97 patients with solid tumor [6 phase Ib, 16 gastric or gastroesophageal junction adenocarcinoma, 45 hepatocellular carcinoma (HCC), 15 renal cell carcinoma, and 15 non–small lung cancer] received emibetuzumab at 750 or 2,000 mg flat dosing plus ramucirumab at 8 mg/kg every 2 weeks. No dose-limiting toxicities were observed. Common adverse events were primarily mild or moderate and included fatigue (36.1%), peripheral edema (28.9%), and nausea (14.4%). Emibetuzumab exposures were similar as in previous studies with no apparent drug–drug interactions. Five partial responses (5.2%) were observed across all tumor types. The greatest antitumor activity was noted in HCC with a 6.7% overall response rate, 60% disease control rate, and 5.42 months (95% confidence interval, 1.64–8.12) progression-free survival (PFS). HCC with high MET expression showed improved PFS with approximately 3-fold increase in PFS (8.1 vs. 2.8 months) relative to low MET expression.
Conclusions:
Ramucirumab plus emibetuzumab was safe and exhibited cytostatic antitumor activity. MET expression may help to select patients benefitting most from this combination treatment in select tumor types.
Data from NK Cell Infiltrates and HLA Class I Expression in Primary HER2+ Breast Cancer Predict and Uncouple Pathological Response and Disease-free Survival
Authors: Muntasell Aura , Rojo Federico , Servitja Sonia , Rubio-Perez Carlota , Cabo Mariona , Tamborero David , Costa-García Marcel , Martínez-Garcia María , Menéndez Sílvia , Vazquez Ivonne , Lluch Ana , Gonzalez-Perez Abel , Rovira Ana , López-Botet Miguel , Albanell Joan
Published: April 2023

AbstractPurpose:
We investigated the value of tumor-infiltrating NK (TI-NK) cells and HLA class I tumor expression as biomarkers of response to neoadjuvant anti-HER2 antibody–based treatment in breast cancer.
Experimental Design:
TI-NK cells and HLA-I were determined by IHC in pretreatment tumor biopsies from two cohorts of patients with HER2-positive breast cancer [discovery cohort (n = 42) and validation cohort (n = 71)]. Tumor-infiltrating lymphocytes (TIL) were scored according to international guidelines. Biomarker association with pathologic complete response (pCR) and disease-free survival (DFS) was adjusted for prognostic factors. Gene set variation analysis was used for determining immune cell populations concomitant to NK-cell enrichment in HER2-positive tumors from the Cancer Genome Atlas (n = 190).
Results:
TI-NK cells were significantly associated with pCR in the discovery cohort as well as in the validation cohort (P < 0.0001), independently of clinicopathologic factors. A ≥3 TI-NK cells/50x high-power field (HPF) cutoff predicted pCR in the discovery and validation cohort [OR, 188 (11–3154); OR, 19.5 (5.3–71.8)]. Presence of TI-NK cells associated with prolonged DFS in both patient cohorts [HR, 0.07 (0.01–0.6); P = 0.01; HR, 0.3 (0.08–1.3); P = 0.1]. NK-, activated dendritic- and CD8 T-cell gene expression signatures positively correlated in HER2-positive tumors, supporting the value of NK cells as surrogates of effective antitumor immunity. Stratification of patients by tumor HLA-I expression identified patients with low and high relapse risk independently of pCR.
Conclusions:
This study identifies baseline TI-NK cells as an independent biomarker with great predictive value for pCR to anti-HER2 antibody–based treatment and points to the complementary value of tumor HLA-I status for defining patient prognosis independently of pCR.
Data from Coordinately Targeting Cell-Cycle Checkpoint Functions in Integrated Models of Pancreatic Cancer
Authors: Chung Sejin , Vail Paris , Witkiewicz Agnieszka K. , Knudsen Erik S.
Published: April 2023

AbstractPurpose:
Cancer cells often have deficiencies in cell-cycle control mechanisms and could be dependent on specific cell-cycle checkpoints to maintain viability. Because of the documented role of KRAS in driving replication stress, we targeted the checkpoint governing DNA replication using CHK1 kinase inhibitors in pancreatic ductal adenocarcinoma (PDAC) models and examined mechanisms of resistance.
Experimental Design:
Single-agent efficacy of CHK1 inhibition was investigated in established and primary PDAC lines. Drug screening was performed to identify cooperative agents. In vitro and in vivo studies were employed to interrogate combination treatment efficacy and mechanisms of resistance.
Results:
Many PDAC models evade single-agent inhibition through mechanisms that allow S-phase progression with CHK1 inhibited. Gene expression analysis revealed FOXM1 as a potential marker of CHK1 sensitivity and defined a form of pancreatic cancer with poor prognosis. Drug screen analysis identified WEE1 as a cooperative agent with CHK1 and was effective in cell culture. In vivo experiments validated the combination efficacy; however, resistance could evolve. Resistance was due to selection of a stable subclone from the original PDX tumor, which harbored high baseline replication stress. In vitro analysis revealed that gemcitabine could eliminate viability in the resistant models. The triplet regimen of gemcitabine, CHK1, and WEE1 inhibition provided strong disease control in all xenograft models interrogated.
Conclusions:
These results demonstrate the therapeutic resiliency of pancreatic cancer and indicate that coordinately targeting cell-cycle checkpoints in concert with chemotherapy could be particularly efficacious.
Data from Molecular Profiling of Cohorts of Tumor Samples to Guide Clinical Development of Pembrolizumab as Monotherapy
Authors: Ayers Mark , Nebozhyn Michael , Cristescu Razvan , McClanahan Terrill K. , Perini Rodolfo , Rubin Eric , Cheng Jonathan D. , Kaufman David R. , Loboda Andrey
Published: April 2023

AbstractPurpose:
Molecular profiling of large databases of human tumor gene expression profiles offers novel opportunities for informing decisions in clinical development programs.
Experimental Design:
Gene expression profile of programmed death ligand 1 (PD-L1) was explored in a dataset of 16,000 samples, including approximately 4,000 metastatic tumors, across >25 tumor types prevalent in the United States, looking for new indications for the programmed death 1 (PD-1) inhibitor pembrolizumab. PD-L1 expression was highly concordant with several genomic signatures indicative of immune-inflamed tumor microenvironment. Prevalence of activated immune-inflamed tumors across all tumor types was explored and used to rank tumor types for potential response to pembrolizumab monotherapy.
Results:
The analysis yielded 3 tiers of indications in which high levels of PD-L1 and immune-inflamed signatures were found in up to 40% to 60%, 20% to 40%, and 0% to 20% of tumors. Tier 1 contained novel indications known at the time of analysis to be responsive to PD-1 checkpoint blockade in the clinic (such as melanoma and non–small cell lung cancer), as well as indications not studied in the clinic previously, including microsatellite instability–high colorectal, head and neck, bladder, and triple-negative breast cancers. Complementary analysis of an Asian/Pacific cancer dataset (gastric cancer) revealed high prevalence of immune-inflamed tumors in gastric cancer. These data contributed to prioritization of these indications for clinical development of pembrolizumab as monotherapy.
Conclusions:
Data highlight the value of molecular profiling in identifying populations with high unmet needs with potentially favorable response characteristics and accelerating development of novel therapies for these patients.

See related commentary by Mansfield and Jen, p. 1443
Data from Polo-like Kinase 1 Inhibition as a Therapeutic Approach to Selectively Target BRCA1-Deficient Cancer Cells by Synthetic Lethality Induction
Authors: Carbajosa Sofía , Pansa María Florencia , Paviolo Natalia S. , Castellaro Andrés M. , Andino Diego L. , Nigra Ayelén D. , García Iris Alejandra , Racca Ana C. , Rodriguez-Berdini Lucía , Angiolini Virginia , Guantay Laura , Villafañez Florencia , Federico María Belén , Rodríguez-Baili María Celeste , Caputto Beatriz L. , Drewes Gerard , Madauss Kevin P. , Gloger Israel , Fernandez Elmer , Gil Germán A. , Bocco José Luis , Gottifredi Vanesa , Soria Gastón
Published: April 2023

AbstractPurpose:
BRCA1 and BRCA2 deficiencies are widespread drivers of human cancers that await the development of targeted therapies. We aimed to identify novel synthetic lethal relationships with therapeutic potential using BRCA-deficient isogenic backgrounds.
Experimental Design:
We developed a phenotypic screening technology to simultaneously search for synthetic lethal (SL) interactions in BRCA1- and BRCA2-deficient contexts. For validation, we developed chimeric spheroids and a dual-tumor xenograft model that allowed the confirmation of SL induction with the concomitant evaluation of undesired cytotoxicity on BRCA-proficient cells. To extend our results using clinical data, we performed retrospective analysis on The Cancer Genome Atlas (TCGA) breast cancer database.
Results:
The screening of a kinase inhibitors library revealed that Polo-like kinase 1 (PLK1) inhibition triggers strong SL induction in BRCA1-deficient cells. Mechanistically, we found no connection between the SL induced by PLK1 inhibition and PARP inhibitors. Instead, we uncovered that BRCA1 downregulation and PLK1 inhibition lead to aberrant mitotic phenotypes with altered centrosomal duplication and cytokinesis, which severely reduced the clonogenic potential of these cells. The penetrance of PLK1/BRCA1 SL interaction was validated using several isogenic and nonisogenic cellular models, chimeric spheroids, and mice xenografts. Moreover, bioinformatic analysis revealed high-PLK1 expression in BRCA1-deficient tumors, a phenotype that was consistently recapitulated by inducing BRCA1 deficiency in multiple cell lines as well as in BRCA1-mutant cells.
Conclusions:
We uncovered an unforeseen addiction of BRCA1-deficient cancer cells to PLK1 expression, which provides a new means to exploit the therapeutic potential of PLK1 inhibitors in clinical trials, by generating stratification schemes that consider this molecular trait in patient cohorts.
Data from Prognostic Impact of Natural Killer Cell Count in Follicular Lymphoma and Diffuse Large B-cell Lymphoma Patients Treated with Immunochemotherapy
Authors: Klanova Magdalena , Oestergaard Mikkel Z. , Trněný Marek , Hiddemann Wolfgang , Marcus Robert , Sehn Laurie H. , Vitolo Umberto , Bazeos Alexandra , Goede Valentin , Zeuner Harald , Knapp Andrea , Sahin Deniz , Spielewoy Nathalie , Bolen Christopher R. , Cardona Andres , Klein Christian , Venstrom Jeffrey M. , Nielsen Tina , Fingerle-Rowson Günter
Published: April 2023

AbstractPurpose:
Natural killer (NK) cells are key effector cells for anti-CD20 monoclonal antibodies (mAb), such as obinutuzumab and rituximab. We assessed whether low pretreatment NK-cell count (NKCC) in peripheral blood or tumor tissue was associated with worse outcome in patients receiving antibody-based therapy.
Patients and Methods:
Baseline peripheral blood NKCC was assessed by flow cytometry (CD3−CD56+ and/or CD16+ cells) in 1,064 of 1,202 patients with follicular lymphoma treated with obinutuzumab or rituximab plus chemotherapy in the phase III GALLIUM trial (NCT01332968) and 1,287 of 1,418 patients with diffuse large B-cell lymphoma (DLBCL) treated with obinutuzumab or rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone (G-CHOP or R-CHOP) in the phase III GOYA trial (NCT01287741). The prognostic value of tumor NK-cell gene expression, as assessed by whole-transcriptome gene expression using TruSeq RNA sequencing, was also analyzed. The association of baseline variables, such as treatment arm, was evaluated using multivariate Cox regression models using a stepwise approach.
Results:
In this exploratory analysis, low baseline peripheral blood NKCC was associated with shorter progression-free survival (PFS) in both follicular lymphoma [hazard ratio (HR), 1.48; 95% confidence interval (CI), 1.02–2.14; P = 0.04] and DLBCL (HR, 1.36; 95% CI, 1.01–1.83; P = 0.04), and overall survival in follicular lymphoma (HR, 2.20; 95% CI, 1.26–3.86; P = 0.0058). Low tumor NK-cell gene expression was associated with shorter PFS in G-CHOP–treated patients with DLBCL (HR, 1.95; 95% CI, 1.22–3.15; P < 0.01).
Conclusions:
These findings indicate that the number of NK cells in peripheral blood may affect the outcome of patients with B-cell non-Hodgkin lymphoma receiving anti-CD20–based immunochemotherapy.
Data from Ibrutinib Therapy Releases Leukemic Surface IgM from Antigen Drive in Chronic Lymphocytic Leukemia Patients
Authors: Drennan Samantha , Chiodin Giorgia , D'Avola Annalisa , Tracy Ian , Johnson Peter W. , Trentin Livio , Steele Andrew J. , Packham Graham , Stevenson Freda K. , Forconi Francesco
Published: April 2023

AbstractPurpose:
In chronic lymphocytic leukemia (CLL), disease progression associates with surface IgM (sIgM) levels and signaling capacity. These are variably downmodulated in vivo and recover in vitro, suggesting a reversible influence of tissue-located antigen. Therapeutic targeting of sIgM function via ibrutinib, an inhibitor of Bruton tyrosine kinase (BTK), causes inhibition and tumor cell redistribution into the blood, with significant clinical benefit. Circulating CLL cells persist in an inhibited state, offering a tool to investigate the effects of drug on BTK-inhibited sIgM.
Experimental Design:
We investigated the consequences of ibrutinib therapy on levels and function of sIgM in circulating leukemic cells of patients with CLL.
Results:
At week 1, there was a significant increase of sIgM expression (64% increase from pretherapy) on CLL cells either recently released from tissue or persisting in blood. In contrast, surface IgD (sIgD) and a range of other receptors did not change. SIgM levels remained higher than pretherapy in the following 3 months despite gradual cell size reduction and ongoing autophagy and apoptotic activity. Conversely, IgD and other receptors did not increase and gradually declined. Recovered sIgM was fully N-glycosylated, another feature of escape from antigen, and expression did not increase further during culture in vitro. The sIgM was fully capable of mediating phosphorylation of SYK, which lies upstream of BTK in the B-cell receptor pathway.
Conclusions:
This specific IgM increase in patients underpins the key role of tissue-based engagement with antigen in CLL, confirms the inhibitory action of ibrutinib, and reveals dynamic adaptability of CLL cells to precision monotherapy.

See related commentary by Burger, p. 2372
Data from Investigating PSMA-Targeted Radioligand Therapy Efficacy as a Function of Cellular PSMA Levels and Intratumoral PSMA Heterogeneity
Authors: Current Kyle , Meyer Catherine , Magyar Clara E. , Mona Christine E. , Almajano Joel , Slavik Roger , Stuparu Andreea D. , Cheng Chloe , Dawson David W. , Radu Caius G. , Czernin Johannes , Lueckerath Katharina
Published: April 2023

AbstractPurpose:
Prostate-specific membrane antigen (PSMA) targeting radioligands deliver radiation to PSMA-expressing cells. However, the relationship between PSMA levels and intralesion heterogeneity of PSMA expression, and cytotoxic radiation by radioligand therapy (RLT) is unknown. Here we investigate RLT efficacy as function of PSMA levels/cell, and the fraction of PSMA+ cells in a tumor.
Experimental Design:
RM1 cells expressing different levels of PSMA (PSMA−, PSMA+, PSMA++, PSMA+++; study 1) or a mix of PSMA+ and PSMA− RM1 (study 2, 4) or PC-3/PC-3-PIP (study 3) cells at various ratios were injected into mice. Mice received 177Lu- (studies 1–3) or 225Ac- (study 4) PSMA617. Tumor growth was monitored. Two days post-RLT, tumors were resected in a subset of mice. Radioligand uptake and DNA damage were quantified.
Results:
177Lu-PSMA617 efficacy increased with increasing PSMA levels (study 1) and fractions of PSMA positive cells (studies 2, 3) in both, the RM1 and PC-3-PIP models. In tumors resected 2 days post-RLT, PSMA expression correlated with 177Lu-PSMA617 uptake and the degree of DNA damage. Compared with 177Lu-PSMA617, 225Ac-PSMA617 improved overall antitumor effectiveness and tended to enhance the differences in therapeutic efficacy between experimental groups.
Conclusions:
In the current models, both the degree of PSMA expression and the fraction of PSMA+ cells correlate with 177Lu-/225Ac-PSMA617 tumor uptake and DNA damage, and thus, RLT efficacy. Low or heterogeneous PSMA expression represents a resistance mechanism to RLT.

See related commentary by Ravi Kumar and Hofman, p. 2774
Data from Dose Optimization for Anticancer Drug Combinations: Maximizing Therapeutic Index via Clinical Exposure-Toxicity/Preclinical Exposure-Efficacy Modeling
Authors: Bottino Dean C. , Patel Mayankbhai , Kadakia Ekta , Zhou Jilai , Patel Chirag , Neuwirth Rachel , Iartchouk Natasha , Brake Rachael , Venkatakrishnan Karthik , Chakravarty Arijit
Published: April 2023

AbstractPurpose:
Recommended phase II dose (RP2D) determination for combination therapy regimens is a constrained optimization problem of maximizing antitumor activity within the constraint of clinical tolerability to provide a wide therapeutic index. A methodology for addressing this problem was developed and tested using clinical and preclinical data from combinations of the investigational drugs TAK-117, a PI3Kα inhibitor, and TAK-228, a TORC1/2 dual inhibitor.
Experimental Design:
Utilizing free fraction-corrected average concentrations, C_{117}^{avg}\ and C_{228}^{avg}, which are the primary pharmacokinetic predictors of single-agent preclinical antitumor activity, a preclinical exposure-efficacy surface was characterized, allowing for nonlinear interactions between growth rate inhibition of the agents on a MDA-MB-361 cell line xenograft model. Logistic regression was used to generate an exposure-effect surface for C_{117}^{avg}\ and C_{228}^{avg} versus clinical toxicity outcomes [experiencing a dose-limiting toxicity (DLT)] in single-agent and combination dose-escalation studies. A maximum tolerated exposure curve was defined at which DLT probability was 25%; predicted antitumor activity along this curve was used to determine optimal RP2D.
Results:
The toxicity constraint curve determined from early clinical data predicted that any clinically tolerable combination was unlikely to result in greater antitumor activity than either single-agent TAK-117 or TAK-228 administered at their respective MTDs. Similar results were obtained with 10 other cell lines, with one agent or the other predicted to outperform the combination.
Conclusions:
This methodology represents a general, principled way of evaluating and selecting optimal RP2D combinations in oncology. The methodology will be retested upon availability of clinical data from TAK-117/TAK-228 combination phase II studies.

See related commentary by Mayawala et al., p. 6564
Data from Phase I Study of Intermittent High-Dose Lapatinib Alternating with Capecitabine for HER2-Positive Breast Cancer Patients with Central Nervous System Metastases
Authors: Morikawa Aki , de Stanchina Elisa , Pentsova Elena , Kemeny Margaret M. , Li Bob T. , Tang Kendrick , Patil Sujata , Fleisher Martin , Van Poznak Catherine , Norton Larry , Seidman Andrew D.
Published: April 2023

AbstractPurpose:
Lapatinib and capecitabine cross the blood–tumor barrier in breast cancer brain metastasis but have modest clinical efficacy. Administration of high-dose tyrosine kinase inhibitor has been evaluated in brain metastases and primary brain tumors as a strategy to improve drug exposure in the central nervous system (CNS). We derived a rational drug scheduling of intermittent high-dose lapatinib alternating with capecitabine based on our preclinical data and Norton–Simon mathematical modeling. We tested this intermittent, sequential drug schedule in patients with breast cancer with CNS metastasis.
Patients and Methods:
We conducted a phase I trial using an accelerated dose escalation design in patients with HER2-positive (HER2+) breast cancer with CNS metastasis. Lapatinib was given on day 1–3 and day 15–17 with capecitabine on day 8–14 and day 22–28 on an every 28-day cycle. Lapatinib dose was escalated, and capecitabine given as a flat dose at 1,500 mg BID. Toxicity and efficacy were evaluated.
Results:
Eleven patients were enrolled: brain only (4 patients, 36%), leptomeningeal (5 patients, 45%), and intramedullary spinal cord (2 patients, 18%). Grade 3 nausea and vomiting were dose-limiting toxicities. The MTD of lapatinib was 1,500 mg BID. Three patients remained on therapy for greater than 6 months.
Conclusions:
High-dose lapatinib is tolerable when given intermittently and sequentially with capecitabine. Antitumor activity was noted in both CNS and non-CNS sites of disease. This novel administration regimen is feasible and efficacious in patients with HER2+ breast cancer with CNS metastasis and warrants further investigation.
Data from A Constitutional Activating MET Mutation Makes the Genetic Link between Malignancies and Chronic Inflammatory Diseases
Authors: El Bouchtaoui Morad , Do Cruzeiro Marcio , Leboeuf Christophe , Loisel-Ferreira Irmine , Fedronie Carèle , Ferreira Chrystophe , Ait El Far Rachida , Ziol Marianne , Espié Marc , Falgarone Géraldine , Cassinat Bruno , Kiladjian Jean-Jacques , Feugeas Jean-Paul , Janin Anne , Bousquet Guilhem
Published: April 2023

AbstractPurpose:
The genesis of all cancers results from an accumulation of mutations, constitutional and/or acquired when induced by external mutagenic factors. High-speed technologies for genome sequencing have completely changed the study of disease genetics, but with limited knowledge of the functional value of most genetic changes.
Experimental Design:
Here, we proposed an innovative individual approach by studying tissue samples from a young woman with an unusual association of breast cancer, polycythemia vera, and rheumatoid arthritis. We performed genomic analyses for copy number variations and point mutations on laser-microdissected tumor cells from the breast cancer, and on CD34+ cells sorted from bone marrow aspiration, to identify gene abnormalities common to these two types of cell populations.
Results:
Using ONCOSCAN technology, we identified a constitutional pR988C, c2962C>T mutation of MET. Using CRISPR-Cas9 technology, we established pR988C MET-mutated transgenic mice, which reproduced the autoimmune diseases and myeloproliferation found in our index-case; one of the transgenic mice spontaneously developed a skin squamous cell carcinoma. We also showed that additional mutagenic factors were required to induce cancers, including skin squamous cell carcinoma and thyroid cancer. Using an anti-MET drug, cabozantinib, we demonstrated for the first time the functional role of this mutation in the maintenance of myeloproliferation and rheumatoid arthritis, and in cancer genesis.
Conclusions:
Our study opens a considerable field of application in the domain of constitutional genetics, to establish genetic links between cancers and other very different severe diseases.
Data from Longitudinal Photoacoustic Imaging of the Pharmacodynamic Effect of Vascular Targeted Therapy on Tumors
Authors: Johnson S. Peter , Ogunlade Olumide , Lythgoe Mark F. , Beard Paul , Pedley R. Barbara
Published: April 2023

AbstractPurpose:
Photoacoustic imaging (PAI) is a novel noninvasive and nonionizing imaging technique that allows longitudinal imaging of tumor vasculature in vivo and monitoring of response to therapy, especially for vascular targeted chemotherapy agents. In this study, we used a novel high-resolution all-optical PAI scanner to observe the pharmacodynamic response to the vascular-disrupting agent OXi4503.
Experimental Design:
Two models of colorectal carcinoma (SW1222 and LS174T) that possess differing pathophysiologic vascularization were established as subcutaneous tumors in mice. Monitoring of response was performed over a 16-day “regrowth” period following treatment at 40 mg/kg, and at day 2 for a “dose response” study at 40 mg/kg, 10 mg/kg, 1 mg/kg, and sham dose.
Results:
Qualitative and quantitative changes in PA signal are observed, with an initial decrease followed by a plateau and subsequent return of signal indicating regrowth. Both tumor types exhibited a decrease in signal; however, the more vascularized SW1222 tumors show greater response to treatment. Decreasing the dose of OXi4503 led to a decrease in PA signal intensity of 60%, 52%, and 20% in SW1222 tumors and 30%, 26%, and 4% for LS174T tumors.
Conclusions:
We have shown for the first time that PAI can observe the pharmacodynamic response of tumor vasculature to drug treatment both longitudinally and at different dose levels. Assessment of differing response to treatment based on vascular pathophysiologic differences among patients has the potential to provide personalized drug therapy; we have demonstrated that PAI, which is clinically translatable, could be a powerful tool for this purpose.