An overview of clinical trials and its various phases

Today new drug development has reached a dynamic stage. The developed new drug must ensure its safety, efficacy and produce the desired therapeutic effect. Preclinical studies are performed before the clinical study to ensure the drug safety and efficacy in animal model. Entry of investigational drug into a human being is done by clinical trials. Clinical trials are performed in various phases to ensure the drug safety and efficacy of the desired therapeutic effect and to provide a beneficial effect to the human beings. Thorough knowledge of the clinical trial study with the defined study protocol will help the researchers to improve the properties of the upcoming new drug molecule coming to the market for various disease treatments.

Modern clinical trials have been evolved during 18th century. Clinical trial is a study to evaluate the comparative effect of the new drug (interventional group) against a control in human volunteers. Clinical trial is a specialized form of biological assay to measure the therapeutic effectiveness of the new drug. In clinical trial , one or more intervention techniques may be adopted. Clinical trials can be carried out in various ways such as treatment trial, prevention trial, diagnostic trial, screening trial and to assess the quality of life of the human beings. Steps involved in the developing a new drug are pre-discovery (understand the disease), target identification (choose a molecule to target with a drug), target validation, preclinical research, Investigational New Drug (IND) Application, clinical trials, New Drug Application (NDA) and drug approval. This article reviews the clinical trials and its various phases.

Clinical trials
A clinical trial is a study to assess the efficacy of a new drug on a patient. Based on the animal pharmacology and toxicology results, a clinical trial study is initiated in human beings. In a clinical trial, a new drug substance is administered into a person or group of people and is observed for its efficacy against a particular disease.

In all clinical trial phases, the data are collected at different time intervals as per the protocol and assess the efficacy of the new drug substance. The number of persons involved in the clinical trial is based on the nature of the disease or treatment process.

The first step in a clinical trial is determination of how large a dose can be given to the volunteers until it produces an unexpected toxicity. To determine the maximum tolerable dose, the investigator starts with lower doses and increasing it until it produces or observes an unexpected or pre-specified toxicity level.

Non-clinical pharmacological evaluation
A non-clinical pharmacological study is carried out before clinical trials. In a non-clinical pharmacological evaluation, an appropriate animal model is selected based on the requirement as well as other factors like availability, validity, selectivity, predictability and reproducibility. These study results provide the mechanism of drug action, efficacy, dose escalation schemes, start dose selection and pharmacodynamic properties.

In vitro studies in non-clinical pharmacology evaluation
In vitro studies in non-clinical pharmacology evaluation are performed by utilizing the cell lines and cell free system. Bioavailability and distribution of drugs are predicted based on the cellular uptake and membrane transport. P450 isoenzyme inhibition or induction and is used to predict the drug metabolism.

Other in vitro methodologies
Generally various tissues or metabolizing enzymes like cytochrome P450 from liver is used for assessment of metabolism. For glucuronidation, sulfation, acetylation, glutathione conjugation and other enzymatic reactions non-cytochrome P450 enzymes are used. Other tissues such as gastrointestinal mucosa, kidney, intestines, brain, placenta and skin are also used for specific purposes.

In vivo studies for nonclinical pharmacology evaluation
In vivo nonclinical pharmacological evaluation is carried out in animal models to assess the efficacy of the drug substance against the specific disease, to determine the therapeutic index, fixing the dose and selection of schedule.

Toxicity studies
In animal model, toxicity studies are carried out to determine the adverse effects of the drug substances. Different toxicity studies like acute toxicity, subacute or subchronic toxicity, chronic toxicity, carcinogencity, special toxicity, reproductive toxicity, genotoxicity and toxicokinetic studies are carried out to determine the toxicity of the new drug substance.

Acute toxicity: Acute toxicity study is carried out in single or multiple doses during a period not exceeding 24 hours to assess the adverse effects in short term.

Subacute or subchronic toxicity: Subacute or subchronic toxicity are carried out over a period of 14 to 90 days to assess the toxic potential and pathological effects.

Chronic toxicity: Chronic toxicity is carried out over a period of 180 days. Potential risk in relation to the anticipated dose and period of drug treatment, potential targets of toxicity, reversibility of any observed toxicities and no-observed adverse effect levels are observed in this study.

Reproductive toxicity: Reproductive toxicity is carried out to assess the effect of drug candidate or its metabolite on reproduction in the mammalian by evaluating the male and female fertility, embryo and fetal death, parturition and the newborn, lactation process and potential teratogenicity. This reproductive toxicological study is carried out in three segments.

Carcinogenicity: Carcinogenicity study is carried to assess any tumor induction in animal which can be relevant for humans.

Genotoxicity: Genotoxicity study is carried to asses any genomic damage which could create cancer and heritable defects.

Investigational new drug
Once the non-clinical pharmacological evaluation and toxicity studies are complete, then pharmaceutical company can file an Investigational New Drug Application (IND) with the FDA.

The FDA reviews the IND application and if the non-clinical pharmacological evaluation and toxicity study results are satisfactory, then FDA gives the approval within 30 days to the pharmaceutical company to start the clinical study in human beings.

Clinical study
Clinical trial is to determine whether the new drug has the definite or specific effect. Clinical trials and observational studies are the two types of clinical studies. Clinical study is a study intended to add medical knowledge of the drug substance by researchers and assessing its efficacy and safety in human volunteers.

Protocol content
Clinical trial is carried based on a designed protocol. The following parameters are the typical protocol content in the clinical trials. The typical protocol contents in the clinical trial includes

A. Background of the study
B. Objectives (Primary question and response variable, secondary questions and response variables, subgroup hypotheses, adverse effects)

C. Design of the study

1. Study population (Inclusion criteria and exclusion criteria)
2. Sample size assumptions and estimates
3. Enrollment of participants (Informed consent, assessment of eligibility, baseline examination, intervention allocation)
4. Intervention(s) (Description and schedule, measures of compliance)
5. Follow-up visit description and schedule
6. Ascertainment of response variables (Training, data collection, quality control)
7. Safety assessment (Type and frequency, instruments, reporting)
8. Data analysis (interim monitoring, final analysis)
9. Termination policy

D. Organization

1. Participating investigators (Statistical unit or data coordinating center, laboratories and other special units, clinical center(s))
2. Study administration (Steering committees and subcommittees, Data monitoring Committee, Funding organization)

Clinical trial phases
Phase I
Phase I includes single rising dose study, short multiple dose study and food interaction study. Phase I trial is called “clinical pharmacology studies” or “first-in-man studies” represent the first introduction of a new drug into human subjects to assess the clinical safety of the drug. In Phase I, fixing the starting dose is a critical because of the inadequate difference in the pharmacokinetic parameters between the animals and human beings. Various parameters such as animal half-life, dosing intervals and their relationship to toxicity are taken into consideration for fixing the starting dose. In general highest no-effect dose in animal toxicity studies is taken into consideration and fixing one tenth of the same will be given to human volunteers. The number of human volunteers varies based on the category of the investigations; generally 20-80 human volunteers are used in Phase I trial.

Steps followed in fixing Maximum Recommended Starting Dose (MRSD) to first in human trials are

Step 1: Determination of the No Observed Adverse Effect Level (NOAEL)

Step 2: Conversion of NOAEL to Human Equivalent Dose (HED)

Step 3: Selection of the most appropriate animal species

Step 4: Application of a safety factor to determine MRSD

Step 5: Compare MRSD with pharmacologically active dose (PAD)

Phase II
Phase II consists of a pre-pilot study, a pilot study and a dose ranging study (utilize several dose levels). Phase II trial is carried out with the patients who have particular indication and to determine the side effects and risks associated with the drug. Phase II study involves experimental design of drug against a placebo or a standard therapy to assess the safety, effectiveness and comparison among the two therapies. If the drug has no or little biological activity, then the drug is rule out in the initial stage based on the expected minimal level of activity. If the drug produces estimated minimal level of activity, more human volunteers are utilized to determine the better dose response rate curve. The number of human volunteers varies generally 100-200 human volunteers who suffer from the condition that the drug is intended to treat or diagnose. This phase can establish the foundation key aspects for Phase III study design.

The main objectives of the Phase II trial is to

1. Determine the minimum dose that is maximally effective without undue toxicity.
2. Determine whether subpopulation require different dosing regiments.
3. Provide pharmacokinetic and pharmacodynamic values.
4. Determine the drug short term side effects.

Phase III
Phase III trial is usually the third and final stage in testing, prior to submission of a New Drug Application. Phase III trial is consisting of double blind comparative studies with registered compound which have similar indications and a placebo. This phase is conducted in closely resembling of the drug would be used if the drug is approved for marketing. These trials may compare

1. Completely new treatment with the standard treatment.
2. Different doses or ways of giving a standard treatment.
3. A new way of giving radiotherapy with the standard way.

The number of human volunteers in the phase III trial between several hundred and several thousand. Phase III trial is conducted in several hospitals, clinics or other sites to collect the information about the dosage effects, efficacy, safety, benefit-risk relationship and to provide adequate information about the drug to be included in the medical leaflet for the physician. Usually Phase III trials are randomized, homogenous population is utilized to reduce the subject’s variability. Pivotal clinical study in Phase III trial may produce the data from controlled and uncontrolled trials by fulfilling the high scientific standards.

Pivotal study criteria are

1. Pivotal study must be a controlled trial.
2. Must have a blinded design when such a design is practical and ethical.
3. Must be randomized.
4. Must be adequate size.

When the Phase III is nearly completing stage, sponsor or the manufacturer may request to give permission to market the drug by submission of an INDA.

Phase IV
Phase IV study is performed after the drug has been shown some therapeutic effect and after granting of license for marketing the drugs. This study is performed under circumstances

1. To satisfy the FDA request made prior to an NDA’s approval.
2. To collect new information about the drug.
3. To develop pharmacoeconomic or cost- effectiveness.
4. To assess the efficacy and safety against a control treatment for the extended treatment and long term follow up.
5. To assess the benefit associated with new treatment.
6. To develop new formulations, dose regimens and route of administration.
7. To support the publication of its research findings in the different specialized medicinal journal and meetings.

Basic study design in clinical trials
Mostly the trials use the parallel design. In the parallel design, simultaneously follows the interventional group and control groups from the time of allocation to one or the other. Cross-over trial is a modification of the parallel design. In this study, each participant utilizes at least twice, at least once as a member of the control group (placebo, no treatment, usual or standard care, or a specified treatment) and at least once as a member of one or more intervention group.

Randomized control trials
Randomization is a process by which all participants or subjects or human volunteers are equally assigned to receive either the new drug or the control group.

Advantages of randomization are

1. Removes the potential bias in the participant allocation.
2. Produce comparable groups.
3. Significance of the statistical tests is guaranteed.

The disadvantage is that all clinical studies cannot be randomized.

Non-randomized concurrent control studies
In the non-randomized concurrent control study, the participants are treated without the new intervention group approximately at the same time as the control group is treated. In non-randomized concurrent control study, the results are not strictly comparable between intervention group and control group.

Randomization process
Randomization process is classified into two types.
They are

1. Fixed allocation randomization
2. Adaptive randomization procedures

Fixed allocation randomization
In fixed allocation randomization, both the intervention group and control group are usually equal and the study progress is not altered by the allocation probability.

Simple randomization: Simple randomization is the most commonly referred in practice for the comparison between the intervention and control group in the pre-determined sample size.

Blocked Randomization: It is also called as permuted block randomization, used to avoid the imbalance between the assigned participants in each group.

The advantage of blocked randomization is that balance between the number of participants in each group and the trial can be terminated before the enrollment is completed. The disadvantage is, it is a strictly theoretical one and the data analysis is complicated one.

Stratified randomization: It is a method help to achieve between group comparability study of the specific characters.

Adaptive randomization procedures
In the adaptive randomization procedure, the study progresses is altered by the allocation probability.

1. Baseline Adaptive Randomization Procedures: In the baseline adaptive randomization procedure the number of participants are balanced based on the previous assignment but does not consider the participant response between the participant in the interventional group and control.
2. Response Adaptive Randomization: In the response adaptive randomization procedure the participant response is considered when allocation of the next participant.

Types of clinical trials
Unblinded: In unblinded or open trial, both the participant and the investigator know to whom the intervention has been assigned.

Single-Blind: In a single blind study, the investigator alone knows to whom which intervention has been assigned to each participant.

Double-Blind: In a double blind study, neither the participant nor the investigators are responsible for following the participants, collecting data and assessing. Such designs are usually restricted to trials of drug or biologics.

Triple-Blind: A double-blind study in which, in addition, the identities of those enrolled in the study and control groups and/or the details about the nature of the interventions (experimental medications), are withheld from the statistician(s) who conduct the analysis of the data.

Choices of clinical trial controls
Placebo concurrent control: In this study, the subjects are randomly assigned to a test treatment or to an identical appearing treatment. Test drug will not be given in this treatment. This is a double blinded and the drug may be given at one or more fixed doses to assess the effect or tolerance.

No treatment concurrent control: In this study, the human volunteers are assigned to receive either test treatment or no study treatment randomly. This treatment is conducted only when the double blind is impossible.

Dose response concurrent control: In this study, subjects are randomized to receive one of the several fixed dose group, it is a dose-response trial. In this study initially the subjects they receive one fixed dose and then raising the dose gradually. The comparison is made between the groups on their final dose and it is a double blind study.

Active (positive) concurrent control: It is a double blind study. In this study, subjects are randomly assigned to receive either the test treatment or an active control treatment.

The objective of the study is

1. To show the efficacy of the test treatment is as good as a known effective treatment
2. To show efficacy by showing superiority of the test treatment to the active control.

External control (including historical control): In this study, a comparison is done between the subjects receiving the test treatment with a group of patient’s external to the study rather than to an internal control group consisting of patients from the same population assigned to a different treatment.

Development of bioanalytical method
Development of a sensitive and selective bioanalytical method is essential to quantify the drug substance in the biological matrix. First step is to characterize a suitable analytical technique to determine the drug substance with the range of reliable results, lower and upper limit of quantification, specificity, accuracy and precision; also the stability of the candidate in the biological matrix. Various chromatographic techniques like High Performance Chromatography, Ultra Performance Liquid Chromatography and Gas Chromatography with various detectors like UV, fluorescent, diode array, flame ionization, electron capture and mass spectrometry, capillary electrophoresis-mass spectrometry are used. For any specific quantification of any drug substance Enzyme-Linked Immunosorbent Assay (ELISA) or Radioimmunoassay (RIA) techniques are used.

Conclusion
Preclinical studies are primary study to establish the drug safety and efficacy in a suitable animal model. Based on the preclinical study results, clinical trials are conducted in human volunteers as per ICH and GCP guidelines to establish its safety and efficacy. The clinical trial study provide the information about the pharmacokinetic, pharmacodynamic properties and its side effects which may be harmful . The fruitful outcome of the clinical trial is an availability of improved new drug for a specific treatment to the patients. The post marketing surveillance about the new drug is done as Phase IV study which provides new information about the drug during extended treatment, benefit associated with new treatment and development of new formulation.