Palbociclib

The preparation of Palbociclib involves several synthetic routes and reaction conditions. One method includes the following steps :

Ring-closing reaction: 2-acetyl-2-butenoic acid methyl ester and malononitrile react under alkaline conditions to generate 1,4,5,6-tetrahydro-2-methoxyl-4-methyl-5-acetyl-6-oxy-3-pyridine carbonitrile.

Substitution reaction: The intermediate product reacts with halogenated cyclopentane under the effect of an acid-binding agent to generate N-cyclopentyl-1,4,5,6-tetrahydro-2-methoxyl-4-methyl-5-acetyl-6-oxy-3-pyridinecarbonitrile.

Condensation reaction: The intermediate product reacts with N-[5-(1-piperazinyl)-2-pyridinyl]guanidine to generate 6-acetyl-8-cyclopentyl-5,8-dihydro-5-methyl-2-[5-(1-piperazinyl)-2-pyridinyl]amino-pyrido[2,3-d]pyrimidin-7(6H)-one.

Dehydrogenation reaction: The intermediate product reacts with sodium selenate to prepare this compound.

This method is economical, environmentally friendly, and suitable for industrial production .

Palbociclib undergoes various types of chemical reactions, including substitution reactions, redox reactions, and condensation reactions . Common reagents used in these reactions include thionyl chloride, bromine, cyclopentylamine, and sodium selenate . Major products formed from these reactions include intermediates such as 1,4,5,6-tetrahydro-2-methoxyl-4-methyl-5-acetyl-6-oxy-3-pyridine carbonitrile and 6-acetyl-8-cyclopentyl-5,8-dihydro-5-methyl-2-[5-(1-piperazinyl)-2-pyridinyl]amino-pyrido[2,3-d]pyrimidin-7(6H)-one .

Breast Cancer

The primary indication for palbociclib is in combination with endocrine therapy for advanced breast cancer. Key clinical trials have demonstrated its efficacy:

• PALOMA-1 Trial : This Phase 2 study showed that the combination of this compound and letrozole significantly prolonged progression-free survival (PFS) in postmenopausal women with estrogen receptor-positive (ER+) and human epidermal growth factor receptor 2-negative (HER2-) breast cancer. The median PFS was reported at 20.2 months compared to 10.2 months for letrozole alone .
• PALOMA-2 Trial : A Phase 3 trial confirmed these findings, reporting a 44% reduction in the risk of disease progression when this compound was added to letrozole. The trial highlighted an improved median PFS exceeding one year .

Other Cancers

Beyond breast cancer, this compound is being investigated for its potential applications in various malignancies:

• Hepatocellular Carcinoma (HCC) : Preclinical studies indicate that this compound can suppress tumor growth in liver cancer models. It has shown effectiveness in promoting cell cycle arrest and improving survival rates when combined with standard treatments like sorafenib .
• Head and Neck Cancers : Ongoing clinical trials are exploring the use of this compound in treating head and neck squamous cell carcinoma, suggesting a broader application beyond breast cancer .
• Non-Small Cell Lung Cancer (NSCLC) : Research is also underway to evaluate the efficacy of this compound in NSCLC, particularly in cases where traditional therapies have failed .

Case Studies

Several case studies illustrate the real-world applications of this compound:

• Case Study: ER+ Breast Cancer
  A 62-year-old postmenopausal woman with advanced ER+ breast cancer received this compound combined with letrozole after progressing on prior endocrine therapy. The patient achieved a PFS of over 18 months, highlighting the drug's effectiveness even after previous treatments.
• Case Study: Hepatocellular Carcinoma
  In a preclinical model, mice treated with this compound alongside sorafenib exhibited a significant reduction in tumor size compared to those receiving sorafenib alone. This suggests that this compound may enhance the efficacy of existing HCC treatments .

Safety and Side Effects

While this compound has shown promising results, it is associated with certain adverse effects, primarily myelosuppression. Clinical trials reported higher rates of neutropenia among patients treated with this compound compared to those receiving placebo or other therapies . Monitoring blood counts is essential during treatment to manage these risks effectively.

Palbociclib exerts its effects by selectively inhibiting cyclin-dependent kinases 4 and 6 . These kinases are involved in the regulation of the cell cycle, particularly the transition from the G1 phase to the S phase . By inhibiting these kinases, this compound prevents the phosphorylation of the retinoblastoma protein, which is necessary for cell cycle progression . This inhibition leads to cell cycle arrest in the G1 phase, thereby preventing cancer cell proliferation .

Comparison with Similar CDK4/6 Inhibitors

Chemical and Pharmacological Properties

Structural and Binding Characteristics

• Palbociclib : Exhibits IC50 values of 9–11 nM (CDK4) and 15 nM (CDK6) .
• Ribociclib : Similar CDK4 inhibition (IC50 = 10 nM) but reduced CDK6 potency (IC50 = 39 nM) .

All three inhibitors contain protonatable amine groups with pKa >8, leading to lysosomal trapping. This property reduces cytoplasmic/nuclear drug availability but can be mitigated by lysosomotropic agents like chloroquine, enhancing efficacy .

Lysosomal Trapping and Synergism

• This compound : Lysosomal accumulation reduces cytoplasmic availability. Co-administration with chloroquine or NH4Cl increases nuclear drug levels, enhancing CDK4/6 inhibition .
• Ribociclib and Abemaciclib : Similar lysosomotropic behavior, but abemaciclib shows stronger senescence induction, while ribociclib is fluorescent, enabling intracellular tracking .

Head-to-Head Comparisons

• This compound vs. However, ribociclib trended toward longer PFS/OS in patients with treatment-free intervals (TFI) <12 months .
• This compound vs. Abemaciclib : Network meta-analysis (2023) revealed comparable PFS for this compound/fulvestrant and abemaciclib/fulvestrant. This compound showed superior clinical benefit rates (CBR) .

Emerging CDK4/6 Inhibitors and Derivatives

Novel Compounds from In Silico Studies

• ZINC585292724 and ZINC585291674 : Identified via pharmacophore-based screening, these compounds exhibit stable hydrogen bonding with CDK4/6 and lower binding free energy (∆Gbind) than this compound, suggesting higher potency .
• Four Chiral Compounds: Demonstrated similar ADMET profiles to this compound but higher hepatotoxicity and carcinogenicity risks .

Dual-Target Inhibitors

Bone Microenvironment Modulation

• Osteoclasts (OCs) : All three inhibitors similarly suppress OC differentiation.
• Osteoblasts (OBs) : this compound and abemaciclib are cytotoxic to OBs at high doses, whereas ribociclib preserves bone integrity .

Senescence and Viral Replication

• This compound induces pre-senescence in hepatoma cells, enhancing hepatitis C virus replication by blocking autophagosome-lysosome fusion—a mechanism shared with chloroquine .

Data Tables

Table 1: CDK4/6 Inhibitor Pharmacological Profiles

Table 2: Clinical Trial Outcomes

Palbociclib, a selective inhibitor of cyclin-dependent kinases 4 and 6 (CDK4/6), has emerged as a significant therapeutic agent in the treatment of hormone receptor-positive (HR+) breast cancer. This article provides a comprehensive overview of its biological activity, including detailed research findings, case studies, and data tables.

This compound functions by inhibiting CDK4 and CDK6, which play crucial roles in cell cycle regulation. By blocking these kinases, this compound prevents the phosphorylation of retinoblastoma protein (RB1), leading to cell cycle arrest in the G1 phase. This action is particularly beneficial in HR+ breast cancer, where dysregulation of the cell cycle is common.

In Vitro Studies

Recent studies have demonstrated that this compound effectively reduces phosphorylated RB1 levels in various breast cancer cell lines. For instance, a comparative study showed that treatment with this compound at doses of 100 nM and 500 nM resulted in significant decreases in p-RB1 levels after 72 hours of exposure. The cytotoxic effects were also assessed using senescence-associated β-galactosidase (SA-β-gal) staining, indicating that both this compound and ribociclib induce cellular senescence at higher concentrations .

Gene Expression Changes

Gene expression profiling revealed that this compound significantly alters the expression of genes associated with various PAM50 intrinsic subtypes. Notably, it increased the Luminal A and Normal-like signatures while decreasing Basal-like and HER2-enriched signatures. This suggests that this compound not only inhibits cell proliferation but also modifies tumor biology towards a less aggressive phenotype .

Clinical Efficacy

This compound's clinical efficacy has been substantiated through pivotal trials such as PALOMA-1 and PALOMA-2. In these studies, this compound combined with letrozole demonstrated a marked improvement in progression-free survival (PFS) compared to letrozole alone. Real-world data further support these findings, showing high clinical benefit rates among patients treated with this compound .

Case Studies

A real-world study involving 162 patients treated with this compound plus letrozole reported a 94% six-month PFS rate. The objective response rate (ORR) was noted at 65%, indicating substantial effectiveness in routine clinical practice .

Safety Profile

While this compound is generally well-tolerated, side effects such as neutropenia are common. Monitoring hematologic parameters is essential during treatment to manage potential adverse effects effectively.

Basic Research Questions

Q. How is palbociclib’s mechanism of action validated in preclinical models of hormone receptor-positive (HR+) breast cancer?

• Methodological Answer : Use in vitro assays (e.g., cell cycle arrest via flow cytometry) and in vivo xenograft models to confirm CDK4/6 inhibition. Measure phosphorylation of retinoblastoma (Rb) protein and downstream E2F targets via immunoblotting. Preclinical studies in hepatocellular carcinoma (HCC) models demonstrated reversible G1 arrest in RB1-proficient cell lines, supporting mechanism validation strategies .
• Key Evidence : this compound induces senescence in RB1-proficient tumors, with resistance linked to RB1 loss .

Q. What statistical methods are recommended for analyzing progression-free survival (PFS) in phase III trials like PALOMA-3?

• Methodological Answer : Employ Cox proportional hazards models with stratification by metastatic site (visceral/nonvisceral), endocrine sensitivity, and menopausal status. Preplanned interim analyses should use independent data monitoring committees to assess efficacy and safety. In PALOMA-3, hazard ratios (HR) were calculated with 95% confidence intervals, and log-rank tests determined significance (P<0.001) .
• Key Evidence : Median PFS improved from 3.8 months (placebo + fulvestrant) to 9.2 months (this compound + fulvestrant) in PALOMA-3 .

Q. How are patient cohorts stratified in trials evaluating this compound combinations?

• Methodological Answer : Stratify by endocrine therapy sensitivity (primary vs. secondary resistance), menopausal status, and presence of visceral metastases. Subgroup analyses in PALOMA-3 revealed stronger overall survival (OS) benefits in endocrine-sensitive populations (HR=0.72; 10-month OS difference) .
• Key Evidence : Patients with prior endocrine sensitivity had a median OS of 39.7 months vs. 29.7 months in the placebo group .

Advanced Research Questions

Q. How can exposure-response analysis optimize this compound dosing in heterogeneous populations?

• Methodological Answer : Use pharmacokinetic-pharmacodynamic (PK-PD) modeling with average steady-state concentration (Cavg,t) to correlate drug exposure with PFS. In PALOMA-2, patients with Cavg,t below the median had shorter PFS, suggesting exposure-dependent efficacy. Adjust dosing in renal/hepatic impairment using population PK models .
• Key Evidence : Dose reductions did not significantly impact efficacy if Cavg,t remained above threshold levels .

Q. What experimental approaches identify mechanisms of this compound resistance in HR+ breast cancer?

• Methodological Answer : Perform genomic profiling (e.g., whole-exome sequencing) to detect RB1 loss or cyclin E1 amplification. Use in vitro models with long-term this compound exposure to mimic acquired resistance. In HCC models, RB1 loss conferred intrinsic resistance, observed in 30% of tumors .
• Key Evidence : RB1-deficient tumors show no response to CDK4/6 inhibition .

Q. How do real-world studies address limitations of randomized controlled trials (RCTs) for this compound?

• Methodological Answer : Apply propensity score matching to balance baseline characteristics (e.g., prior therapies, age) between real-world and RCT cohorts. Retrospective analyses, such as the POLARIS study, validate this compound’s PFS benefit (median 12.4 months in real-world vs. 9.2–24.8 months in trials) despite broader eligibility criteria .
• Key Evidence : Real-world data confirm neutropenia (80% incidence) as the primary toxicity, consistent with RCTs .

Q. What methodologies assess synergism between this compound and PI3K/AKT/mTOR inhibitors?

• Methodological Answer : Use combination index (CI) assays (e.g., Chou-Talalay method) in vitro. In the INAVO120 trial, inavolisib (PI3K inhibitor) + this compound + fulvestrant improved PFS vs. This compound + fulvestrant alone (15.0 vs. 7.3 months; HR=0.43) .
• Key Evidence : Synergistic effects are linked to dual pathway inhibition, delaying compensatory resistance .

Q. How are biomarkers like Rb phosphorylation or Ki-67 integrated into early-phase trial designs?

• Methodological Answer : Include exploratory endpoints using immunohistochemistry (IHC) or RNA sequencing to quantify Rb pathway activity. In PALOMA-1, Rb-positive tumors showed higher response rates to this compound + letrozole .
• Key Evidence : Rb loss or cyclin D1 amplification predicts poor response to CDK4/6 inhibitors .

Q. What pharmacodynamic endpoints are critical for this compound combination trials?

• Methodological Answer : Measure cell cycle arrest (G1 phase fraction) via flow cytometry and senescence markers (e.g., SA-β-galactosidase). In breast cancer models, this compound + EGFR inhibitors increased apoptosis (e.g., 40% apoptotic cells in HCT15 lines) .
• Key Evidence : Senescence-associated secretory phenotype (SASP) markers (e.g., IL-6, IL-8) are elevated in resistant tumors .

Q. How do ethnic and sex-specific factors influence this compound pharmacokinetics and efficacy?

• Methodological Answer : Conduct subgroup analyses stratified by ethnicity (e.g., Asian vs. non-Asian) or sex. In Japanese patients, this compound exposure correlated with body weight, necessitating dose adjustments. Male patients showed similar safety profiles to females in real-world studies .
• Key Evidence : FDA approval for male patients was based on real-world data and PK consistency across sexes .