Guidelines for Diagnosis and Treatment of Chronic Lymphocytic Leukemia. Krohem B-Cll 2017

Recent developments in the diagnosis and treatment of chronic lymphocytic leukemia (B-CLL) have led to change of approach in clinical practice. New treatments have been approved based on the results of randomized multicenter trials for fi rst line and for salvage therapy, and the results of numerous ongoing clinical trials are permanently providing new answers and further refi ning of therapeutic strategies. Th is is paralleled by substantial increase in understanding the disease genetics due to major advances in the next generation sequencing (NGS) technology. We defi ne current position of the Croatian Cooperative Group for Hematologic Disease on diagnosis and treatment of CLL in the transition from chemo-immunotherapy paradigm into a new one that is based on new diagnostic stratifi cation and unprecedented therapeutic results of B-cell receptor inhibitors (BRI) and Bcl-2 antagonists. Th is is a rapidly evolving fi eld as a great number of ongoing clinical trials constantly accumulate and provide new knowledge. We believe that novel therapy research including genomic diagnosis is likely to off er new options that will eventually lead to time limited therapies without chemotherapy and more eff ective clinical care for B-CLL based on individualized precision medicine.

## Introduction and defi nition
In recent years, dramatic change in therapeutic landscape led to unprecedented therapeutic results that were translated to clinical practice, and after years of slow to modest progress in the fi eld of B-cell chronic lymphocytic leukemia (B-CLL), the American Society of Clinical Oncology (ASCO) named transformation of CLL treatment Cancer Advance of the Year for 2015 [bib_ref] Clinical Cancer Advances 2015: Annual Report on Progress against Cancer from the..., Masters [/bib_ref]. New treatments were approved based on the results of randomized multicenter trials for fi rst line and for salvage therapy, and the results of numerous ongoing clinical trials are permanently providing new Steps 1-4 are made in single visit. Each step is diff erent with respect to aim, decision criteria, and the extent of work-up. Last column describes classifi cation categories proposed. Only about 10% of patients have indication for therapy at diagnosis. Others are followed-up repeatedly until the criteria for therapy are reached . Th e scope of work-up is diff erent in each step, pretreatment evaluation being most complete, aiming to provide all the necessary elements for patient stratifi cation and defi nition of therapeutic goal. Th e overall goal of diagnostic process is to enable individualization of therapy, defi nition of therapeutic aim and strategy, by implementing general principles to each individual case.
answers and further refi ning of therapeutic strategies. Th is is paralleled by substantial increase in understanding the disease genetics owing to major advances in the next generation sequencing (NGS) technology 2,3 . All these have led to change of the until now predominant paradigm based on chemo-immunotherapy (CIT) to a new one [bib_ref] Chronic lymphocytic leukaemia, Kipps [/bib_ref]. Defi nition. B-cell chronic lymphocytic leukemia and related disorders (monoclonal B-lymphocytosis (MBL) and small lymphocytic lymphoma (SLL)) are defi ned by the presence of clonal mature B-lymphocytes with typical immunophenotype in peripheral blood, bone marrow and lymphoid organs (WHO, iw-CLL) [bib_ref] Guidelines for the diagnosis and treatment of chronic lymphocytic leukemia: a report..., Hallek [/bib_ref] [bib_ref] Th e 2016 revision of the World Health Organization classifi cation of..., Swerdlow [/bib_ref] representing one nosologic entity. Today, it is considered that these entities are diff erent manifestations of the same disease. MBL is most prevalent and is considered as an early stage of malignancy progressing to CLL/SLL in 1%-2% of cases per year. SLL accounts for less than 10% of overt malignancy, and for this reason B-CLL is most commonly used to represent both variants (CLL/SLL) [bib_ref] Guidelines for the diagnosis and treatment of chronic lymphocytic leukemia: a report..., Hallek [/bib_ref] [bib_ref] Reassessment of small lymphocytic lymphoma in the era of monoclonal B-cell lymphocytosis, Gibson [/bib_ref] [bib_ref] Th e 2008 WHO classifi cation of lymphoid neoplasms and beyond: evolving..., Campo [/bib_ref] [bib_ref] Monoclonal B-cell lymphocytosis and chronic lymphocytic leukemia, Rawstron [/bib_ref] [bib_ref] National Cancer Institute-sponsored Working Group guidelines for chronic lymphocytic leukemia: revised guidelines..., Cheson [/bib_ref] [bib_ref] Th e natural history of CLL: historical perspective, Marti [/bib_ref] [bib_ref] Diagnostic criteria for monoclonal B-cell lymphocytosis, Marti [/bib_ref] [bib_ref] Clinical progression and outcome of patients with monoclonal B-cell lymphocytosis, Fung [/bib_ref] [bib_ref] Guidelines on the diagnosis, investigation and management of chronic lymphocytic leukaemia, Oscier [/bib_ref].
Epidemiology. B-CLL is the most common type of leukemia in Western countries. Th e incidence is estimated to more than 6 per 100,000 people annually. Th e median age at diagnosis is growing globally, so that now exceeds 70 years. It should be noted that the age at treatment initiation is several years older than the age at diagnosis, depending on the duration of observation without treatment. Th e disease is nearly twice as common in men [bib_ref] Th e roles of age and sex in the prognosis of chronic..., Jaksic [/bib_ref] [bib_ref] Chronic lymphocytic leukaemia: case control epidemiological study in Yorkshire, Cartwright [/bib_ref] [bib_ref] Incidence and mortality trends of leukemia and lymphoma in Croatia, Novak [/bib_ref]. Th e incidence and prevalence of MBL is much higher, depends on sensitivity of the methods used, and is estimated to involve up to 12% of the population aged over 40 years [bib_ref] Increased frequency (12%) of circulating chronic lymphocytic leukemia-like B-cell clones in healthy..., Nieto [/bib_ref] [bib_ref] Monoclonal B-cell lymphocytosis and early-stage chronic lymphocytic leukemia: diagnosis, natural history, and..., Strati [/bib_ref].

## Diagnostic procedure
Th e diagnostic process can be conditionally divided into several sections (steps, phases) with respect to different objectives to be achieved. In [fig_ref] Table 1: Steps and aims in diagnostic process that leads to defi nition of... [/fig_ref] , the basis for decision is specifi ed, along with the main criteria for classifi cation and the possible categories to which the classifi cation in this section should lead.

## Setting suspicion and patient referral to hematologist
Th e most common fi nding leading to suspicion of B-CLL is absolute lymphocytosis in routine blood ex-amination (70%-80%), and less frequently (20%-30%) the fi nding of organomegaly (swollen lymph nodes and/or spleen) or symptoms associated with CLL.

## Diagnosis and diff erential diagnosis
It is mandatory to make the diagnosis of typical B-CLL on the basis of morphology and fl ow cytometry in peripheral blood sample and to distinguish it from other entities in the CLL syndrome. For the diagnosis of B-CLL phenotype (typical phenotype), the following is required: restriction of sIg light-chain expression of low intensity, CD5 + , CD19 + , CD20 low , CD23 +25 . Basic hematologic clinical fi ndings and blood count allow for quantifi cation of the tumor mass in peripheral blood and lymphoid organs, which enables classifi cation of entities that meet the diagnostic criteria for:
(1) B-CLL (presence in the blood of more than 5x10 9 /L clonal cells), or (2) SLL (less than 5x10 9 /L clonal cells in the blood and clonal lymphadenopathy greater than 1.5 cm), and (3) MBL (less than 5x10 9 /L clonal cells in the blood and no clonal lymphadenopathy or symptoms). So, a diff erent size of the tumor mass between B-CLL and MBL is critical and tumor distribution is critical for distinction between B-CLL and SLL.
It is evident that for the diagnosis of disease type, a very small number of tests could be considered sufficient because if the result is positive in peripheral blood (PB), it is not necessary to analyze bone marrow (BM) or lymph nodes (LN), although these tests have their place in further diagnostic work-up. For the diagnosis of SLL, it is recommended to make lymph node biopsy to establish the diagnosis, and radiological assessment (ultrasound (US) or computed tomography (CT)) of the neck, thorax, abdomen and pelvis can be useful to distinguish between MBL and SLL.
Other B-cell malignancies that can be accompanied by increased lymphoid cell counts in the blood should be taken in consideration in diff erential diagnosis. Distinction to CLL is usually made by fl ow cytometry assessed immune phenotype. Th e majority are B-cell malignancies such as follicular lymphoma (FL), mantle cell lymphoma (MCL), marginal zone lymphoma (MZL), prolymphocytic leukemia (PLL) and hairy cell leukemia (HCL), which all together represent less than 15% of all lymphoid appearing leukocytoses, and less than 5% express T-cell lineage marker variants.
It should be remembered that this type of disorder classifi cation based only on the immune phenotype is not using morphological, cytogenetic, molecular, or other characteristics of the diseases that today show signifi cant association with prognosis.

## Evaluation of the stage/extent of the disease
After the diagnosis, the next step is evaluation of the disease stage or extent. It is based on clinical and [fig_ref] Table 2: Clinical stages and TTM absolute number of lymphocytes [/fig_ref]. Clinical stages represent a simple tool for clinical assessment of the disease extent. Th e basis of these systems is the assumption that the disease is gradually progressing and expanding. Th erefore, the patients who have advanced disease have a higher tumor load and more extensive disease [bib_ref] Clinical staging of chronic lymphocytic leukemia, Rai [/bib_ref] [bib_ref] A new prognostic classifi cation of chronic lymphocytic leukemia derived from a..., Binet [/bib_ref] [bib_ref] Total tumour mass score (TTM): a new parameter in chronic lymphocyte leukaemia, Jaksic [/bib_ref]. Th e clinical stages according to Rai and Binet assess tumor size by simple parameters, which estimate the size of the tumor mass (without precise quantifi cation of the aff ected compartments), along with the parameters for the assessment of bone marrow failure. In doing so, the greatest prognostic power contribution has the failure of bone marrow. Note that it is not a direct but an indirect sign of tumor size. Th eir prognostic power is relatively weak if the failure of the bone marrow is excluded.
Th e estimation of the size of the Total Tumor Mass (TTM) is diff erent in that it quantitatively evaluates tumor mass in 3 major cell compartments, regardless of bone marrow failure [fig_ref] Table 2: Clinical stages and TTM absolute number of lymphocytes [/fig_ref]. Quantitative character enables unbiased monitoring of disease progression and is a very convenient tool for the assessment of therapeutic response (see later). In addition, in most patients who have both peripheral blood and lymphoid organs aff ected, it is possible to determine the type of distribution of the tumor mass by comparing leukemic (TM 1 ) and lymphoid organ (TM 2 +TM 3 ) compartments [bib_ref] Clinical tumor cell distribution pattern is a prognostically relevant parameter in patients..., Jaksic [/bib_ref]. Th is feature of TTM system enables to estimate the dynamics of disease progression and response to therapy and redistribution (see later) [bib_ref] An old and simple solution for a new problem -more on clinical..., Jaksic [/bib_ref].

## Prognostic factors at diagnosis and predictors of response
Analyses of prognostic factors performed in the era when therapy had little eff ect on clinical course identifi ed a number of clinical or laboratory factors 31 , describing broadly the natural course of the disease. Prognostic factors that can be determined immediately at diagnosis are primarily related to the additional characterization of the tumor itself. Some of them do not change during evolution of the disease, such as the mutational status of IgVH genes that generally discriminate between more benign and more malignant disease [bib_ref] Ig V gene mutation status and CD38 expression as novel prognostic indicators..., Damle [/bib_ref] [bib_ref] Unmutated Ig V(H) genes are associated with a more aggressive form of..., Hamblin [/bib_ref] [bib_ref] CD38 expression and immunoglobulin variable region mutations are independent prognostic variables in..., Hamblin [/bib_ref] , and it is currently suggested to become minimal standard initial evaluation [bib_ref] Prognostic factors and risk stratifi cation in chronic lymphocytic leukemia, Parikh [/bib_ref]. Until now, these were rarely used in routine clinical practice and other tests showing high correlation with mutational status are used instead, such as CD38 and ZAP-70 by fl ow cytometry and immunohistochemistry, with the higher value found to correlate with poorer prognosis [bib_ref] CD38 on B-cell chronic lymphocytic leukemia cells has higher expression in lymph..., Jaksic [/bib_ref] [bib_ref] ZAP-70 has higher expression in lymph nodes and bone marrow compared to..., Jaksic [/bib_ref] [bib_ref] Immunohistochemical analysis of ZAP-70 expression in chronic lymphocytic leukemia, Korac [/bib_ref] [bib_ref] ZAP-70 expression as a surrogate for immunoglobulin-variable-region mutations in chronic lymphocytic leukemia, Crespo [/bib_ref] [bib_ref] Evaluation of ZAP-70 expression by fl ow cytometry in chronic lymphocytic leukemia:..., Letestu [/bib_ref] [bib_ref] Relative value of ZAP-70, CD38, and immunoglobulin mutation status in predicting aggressive..., Rassenti [/bib_ref] [bib_ref] Prognostic factors in chronic lymphocytic leukemia, Hallek [/bib_ref] [bib_ref] Prospective evaluation of prognostic parameters in early stage chronic lymphocytic leukemia (CLL):..., Bergmann [/bib_ref] [bib_ref] CD38 and ZAP-70 are functionally linked and mark CLL cells with high..., Deaglio [/bib_ref]. Th e usefulness of these surrogate factors is still controversial. In contrast, cytogenetic and molecular tests (FISH) to determine del (11q22-23), +12; del (13q14), del (17p13) [bib_ref] Genomic aberrations and survival in chronic lymphocytic leukemia, Dohner [/bib_ref] showed a strong independent prognostic eff ect and usually change (progress) over time. Also, fi nding certain mutations (including TP53, SF3B1, NOTCH1, etc.) [bib_ref] Th e genomic landscape of chronic lymphocytic leukaemia: biological and clinical implications, Streff Ord [/bib_ref] [bib_ref] Functional impact of NOTCH1 mutations in chronic lymphocytic leukemia, Arruga [/bib_ref] can complement these fi ndings. Th ese tests are good predictors of response in the chemo-immunotherapy era and have proven to be useful to stratify patients into groups that react diff erently to certain drugs [bib_ref] Chronic lymphocytic leukaemia, Kipps [/bib_ref] [bib_ref] Prognostication of chronic lymphocytic leukemia in the era of new agents, Eichhorst [/bib_ref]. Since they may change during the course of disease, testing must be performed just before each new line of therapy. All these tests that further characterize neoplastic clone cells are technically demanding and expensive, but relatively inexpensive compared to drugs, and for those validated the cost and eff ort are justifi ed. Predictive factors seem to change or even lose their predictive power with novel treatments. Likewise, failure to new treatments will require discovery of new predictors.
On the other hand, a number of additional factors more related to patient's state also have a strong prognostic power, so a number of diff erent parameters is often used, and multivariate analyses are performed to study their impact on prognosis [bib_ref] Validation of a new prognostic index for patients with chronic lymphocytic leukemia, Shanafelt [/bib_ref].
Because of the above, we distinguish three types of factors, given the causal relationship with B-CLL. First, those that are associated with B-CLL clonal neoplastic diseases (size distribution, growth rate of tumors, mutational status, CD38, ZAP-70, FISH) may be used as predictors of response to therapy, etc. [bib_ref] Total tumour mass score (TTM): a new parameter in chronic lymphocyte leukaemia, Jaksic [/bib_ref] [bib_ref] Ig V gene mutation status and CD38 expression as novel prognostic indicators..., Damle [/bib_ref] [bib_ref] Unmutated Ig V(H) genes are associated with a more aggressive form of..., Hamblin [/bib_ref] [bib_ref] Genomic aberrations and survival in chronic lymphocytic leukemia, Dohner [/bib_ref] [bib_ref] Prognostic indices in chronic lymphocytic leukaemia: where do we stand how do..., Baliakas [/bib_ref] [bib_ref] Prognostic value of the lymphocyte doubling time in chronic lymphocytic leukemia, Molica [/bib_ref]. Also, numerous other characteristics such as blood chemistry (serum beta 2 microglobulin, sCD23 and serum thymidine kinase [bib_ref] Serum beta(2)-microglobulin and serum thymidine kinase are independent predictors of progression-free survival..., Hallek [/bib_ref] [bib_ref] Tumor mass assessment in CLL. Distinction between sCD23 and beta-2 microglobulin, Jaksic [/bib_ref] , clonogenic, kinetic, and computer assisted cell imaging have been shown to be related to prognosis [bib_ref] New parameters of diploid histogram of image DNA cytometry and newly characterized..., Kardum-Skelin [/bib_ref] [bib_ref] Multimodal image analysis of chronic leukemic lymphoproliferative disorders and the hypothesis of..., Kardum-Skelin [/bib_ref] ; second, those that are associated with organ failure (mixed groups, they may be due to both very underlying neoplastic diseases and to consequences of associated diseases, such as anemia, thrombocytopenia, the etiology of which should be carefully evaluated); and third, those factors that are associated with the patient but not directly with the neoplasm (age, gender, performance status, comorbidity), that correlate with fi tness and the ability to tolerate aggressive treatments [bib_ref] Relative value of ZAP-70, CD38, and immunoglobulin mutation status in predicting aggressive..., Rassenti [/bib_ref] [bib_ref] Physiologic evaluation in the elderly prior to treatment with chemotherapy, Hallek [/bib_ref] [bib_ref] Comorbidity and functional status are independent in older cancer patients, Extermann [/bib_ref] [bib_ref] Prognostic factors in chronic lymphocytic leukemia -what do we need to know?, Cramer [/bib_ref] [bib_ref] Prognostic markers in chronic lymphocytic leukemia: a comprehensive review, Van Bockstaele [/bib_ref].
On the basis of strong independent predictors, composite prognostic indices are calculated [bib_ref] Characteristics associated with important clinical end points in patients with chronic lymphocytic..., Wierda [/bib_ref] [bib_ref] A new prognostic score for CLL, Tam [/bib_ref] [bib_ref] Development of a comprehensive prognostic index for patients with chronic lymphocytic leukemia, Pfl Ug [/bib_ref] , most recently the CLL International Prognostic Index (IPI) [bib_ref] An international prognostic index for patients with chronic lymphocytic leukaemia (CLL-IPI): a..., Cllipiwg [/bib_ref]. Th ey show very good prognostic power with respect to the length of survival in the chemo-immunotherapy (CIT) era, but much less so with new treatments. Also, one should be careful using them to stratify patients for treatment that is adapted to risk because a composite index usually consists of the factors that belong to each of the three above mentioned groups.
With new, more eff ective therapies, the prognostic landscape changes. Most of the important predictors in the CIT era lose their power, and new ones are yet to be identifi ed [bib_ref] Chronic lymphocytic leukaemia, Kipps [/bib_ref] [bib_ref] Prognostication of chronic lymphocytic leukemia in the era of new agents, Eichhorst [/bib_ref].

## Indications for treatment/the criteria for therapy initiation
Th e criteria for active disease that warrant initiation of treatment are based on iwCLL criteria, and are amended with TTM [fig_ref] Table 3: Criteria for active [/fig_ref]. Th e decision should be based solely on the assessment of parameters that are associated with neoplastic B-CLL clone [bib_ref] Guidelines for the diagnosis and treatment of chronic lymphocytic leukemia: a report..., Hallek [/bib_ref]. For parameters that may be unrelated to the neoplasm, it is necessary to carefully evaluate the extent to which the parameters are associated with neoplastic clone (e.g., fever, anemia, etc.). Criterion parameters can be classifi ed into three distinct types:
1. Quantitative parameters for which threshold consensus is defi ned, which is considered to justify treatment initiation, such as anemia, defi ned by a certain level of hemoglobin, thrombocytopenia defi ned by platelet count, organomegaly defi ned by the size of spleen or lymph nodes.
2. Monitoring data to enable assessment of trends, for example, progressive cytopenia, progressive lymphadenopathy and/or splenomegaly, increased leukocyte count or TTM values (see the previous section). It should be emphasized that only measurement of dynamic parameters can directly evaluate progression of the disease, as opposed to the a priori estimate of a possible evolution described in the previous section (see Prognostic features at diagnosis section). Here, however, we should point out certain diffi culties and ambiguities in quantitative measurement of the dynamic parameters, especially in the early stages of the disease. Th is imprecision in a priori defi nition of the criteria for progressiveness, despite a very attractive concept, often leads to decision delay until the moment when it reaches the absolute value of the threshold that is set up as described above.
3. Qualitative criteria of the occurrence of symptoms threatening organ damage and the like, considered to be the result of neoplastic disease activity.
Today, we seek to combine threshold criteria with dynamic criteria (see [fig_ref] Table 7: Treatment of relapsed/refractory CLL [/fig_ref]. Note that a number of criteria, the criteria of threshold, dynamics and qualitative changes may be used. Although it is suffi cient to indicate treatment by the presence of only one of the stipulated criteria, it is important that this criterion is compelling, and the presence of multiple criteria makes decision certainly easier. It especially holds for dynamic criteria, so it is good to compare the growth trend of the tumor mass with the trends of deepening anemia and/or thrombocytopenia. Th e indication for treatment (according to KROHEM guidance) should be documented in patient records. [fig_ref] Table 4: General schedule of investigations before, during and after therapy *Monitoring protocol frequency... [/fig_ref] shows the general scheme of tests that are used in pretreatment work-up, in therapy monitoring, and after therapy. Of the newly diagnosed CLL patients, about 10% require therapy immediately, so the pretreatment work-up is done immediately at diagnosis and visits 1-3 are completed without delay. Other patients (90% of all diagnosed) are observed after diagnosis at periodic visits until meeting the criteria for initiation of treatment. At that point, complete pretreatment work-up is performed. About 30% of all diagnosed patients never reach the criteria for initiation of treatment, while about 60% of all diagnosed patients reach the criteria, but in diff erent periods, from several months to more than 10 years.

## Th e schedule of activities and assessments
It is possible to distinguish several specifi c clinical questions that need to be answered in stipulated visits and require diff erent extent of investigations. Th ese questions are answered in the following visits: (1) diagnosis, diff erential diagnosis, evaluation of the extent of disease, and preferably but not mandatory, assessment of prognostic groups that require investigations (a-f ). At this time, it is recommended to evaluate general health profi le (j); (2) repetitive visits monitoring clinical and hematologic parameters that serve as criteria for initiation of treatment, based on simple pa- Hypogammaglobulinemia, monoclonal or oligoclonal paraproteinemia, or absolute lymphocyte count do not by themselves constitute an indication for therapy. It is out of 8 groups of criteria theoretically possible to identify 11 individual indications based on exceeding a threshold, 3 dynamic evaluation of continuous quantitative parameters, where individual trends can be compared and thereby gain additional derived criteria, and 4 qualitative assessments. Although in principle the presence of at least one indication is suffi cient, we should avoid making decisions on an isolated indication. It is clear that a larger number of indications further reinforces the decision to begin treatment. It is possible to decide that the patient needs to document the presence of at least two or more of the above indications for active (progressive/symptomatic) disease. Th e indication for treatment (according to KROHEM guidance) should be documented in patient records! rameters (investigations a, b, c, j). Th e time between the visits may vary from weeks to months; (3) immediate pre-therapeutic work-up provides defi nite classifi cation of the disease, defi nitive assessment of prognostic parameters, as well as the general state of the patient, and any associated illnesses including infection status. For this reason, this visit should be most complete, enabling stratifi cation and providing baseline parameters for treatment; (4) these repetitive visits to monitor the course of therapy should be tailored according to the needs of respective therapy; and (5) end of therapy (EOT) visit should enable evaluation of the response achieved. After that, for CIT that is time limited, monitoring visits without therapy (2') are repeated again and in case of the need for a new line of therapy, repeat visits 3', 4' and 5' are scheduled. Th e situation is diff erent for novel agents since this treatment is at present unlimited and visits 4 should enable detection of treatment failure.

## Th erapy individualization -defi ning goals and strategies, and stratifying patients
At the end of diagnostic procedure described above, there is a need to make decision on therapy, which will refl ect individualization of the respective patient. Decision is based on the integration of the factors related to the neoplasm on the one hand and the factors related to the patient on the other hand. According to new circumstances and new emerging paradigm because of the lack of curative treatment, initial therapy should maximize effi cacy while minimizing overall toxicity. Th e overall approach is essentially based on clinical judgment and the expertise.
In general, we are comparing two risks: the risk of disease and the risk of treatment. It is clear that the risk of treatment should be reasonably lower than the risk of disease.
Th e risk of disease. Some treatments have been shown to be ineffi cient in some disease subsets. However, novel treatments are also active in those subsets, which will diminish the importance of stratifi cation according to FISH and TP53, which was necessary until now. Nevertheless, since CIT has proved very effective in some subsets of patients and contraindicated in others, the classifi cation according to risk should remain until head-to-head comparisons resolve pending questions.
Th e risk of therapy, i.e. tolerance (or acute treatment toxicity) is highly dependent on the general condition of the patient and the presence of associated diseases, which is often associated (though not exclusively) with the patient's age. However, treatments with new B-cell receptor inhibitors (BRI) have much better toxicity pro- fi le that diminishes the importance of stratifi cation according to fi tness, which was necessary in the chemoimmunotherapy based paradigm. Nevertheless, for the same reason as described above, this stratifi cation should remain for CIT. Since CIT is not indicated in patients with del(17p)/TP53 mutation, the distinction between fi t and unfi t becomes irrelevant.
A patient is classifi ed as capable (fi t, Go Go) when there is a low comorbidity score (e.g., CIRS-G <6). Although age does not enter in the calculation of CIRS-G, it is known that age is a very important factor and it should be taken into account. It is common to impose an age limit for stratifi cation in therapeutic groups. Today, the limit is set at 65 years. Patients aged over 65 can be considered capable for receiving aggressive therapy if in excellent health, without substantial comorbidity.
On the basis of the two above-mentioned principles, patients are today stratifi ed into three strata with respect to antineoplastic therapy: (1) without the p53 gene deletion with good general condition (capable for aggressive, fl udarabin based CIT therapy) -(LOW RISK + FIT), (2) without the p53 gene deletions with poor general condition (incapable for aggressive therapy) -(LOW RISK + UNFIT), and (3) deletion of (17p)/TP 53 mutation (HIGH RISK + FIT/UNFIT). For those patients, no further division according to fi tness is necessary, since the new approved treatments are well tolerated [bib_ref] Guidelines on the diagnosis, investigation and management of chronic lymphocytic leukaemia, Oscier [/bib_ref] [bib_ref] State-of-the-art treatment of chronic lymphocytic leukemia, Hallek [/bib_ref] [bib_ref] How I treat CLL up front, Gribben [/bib_ref]. In other words, these strata may in principle represent the combination of risk (high correlation with TP53 abnormality) and age (high correlation with fi tness). However, for the reasons stipulated above, the chronologic age cut-off should not be rigid, so as to allow elderly patients in good health to enjoy the benefi t of more aggressive treatment, and vice versa, to spare younger patients with comorbidity of unwarranted therapy associated risks.
Th e stratifi cation is used for guidelines and refl ects general principles, but for each patient, treatment plan should be individualized and set up after careful clinical evaluation, also taking into account patients' preferences.

## Th erapeutic procedure

## Th erapeutic recommendations based on evidence/clinical trials
Th e treatment for B-CLL consists of antineoplastic therapy and supportive measures. Antineoplastic measures consist traditionally of chemotherapy, therapeutic antibodies, radiotherapy, stem cell transplantation methods, and recently novel agents that include B-cell receptor signal transduction inhibitors (BRI) and bcl-2 antagonists. In these guidelines, the recommendations are based on phase 3 clinical trials, in some cases on evidence from earlier phase trials, and on the approved agents and therapies in Croatia. Combination therapy is generally more effi cient than monotherapy.

## Choice of antineoplastic treatment options -new therapeutic paradigm
Because this disease is generally not curable, occurs in elderly population, and often progresses slowly, it is most often treated in a conservative fashion. In asymptomatic patients, treatment may be deferred until the patient becomes symptomatic as the disease progresses [bib_ref] Chemotherapeutic options in chronic lymphocytic leukemia: a meta-analysis of the randomized trials...., Cll Trialists&apos; Collaborative [/bib_ref]. Th is concept is still valid and the treatment outside clinical trials is recommended only for patients that fulfi ll the indications described. Since the rate of progression may vary from patient to patient, with long periods of stability and sometimes spontaneous regressions, frequent and careful observation is required to monitor the clinical course. At present, about 30% live without symptoms and never need therapy. Others will progress sooner or later, and will meet the criteria for therapy introduction.
Th e new, very eff ective treatments have recently dramatically changed the therapeutic landscape and led to change in the current CIT based paradigm into a new one. In comparison to therapies available to date, the new treatments show markedly improved effi cacy and considerably better tolerance. Th is has an impact on all aspects of patient management and choice of treatment options. Th is also goes for deferring therapy, but at present there are no data to support early treatment. Current and future clinical trials that include novel agents in this setting may change this current concept. For patients in whom the criteria for therapy initiation are met, the general principle in the new paradigm is that (because no curative therapy has yet been found) initial therapy should maximize efficacy with improvement of overall survival (OS), while introducing the least overall toxicity, both short term and long term. Standard chemotherapeutic agents induce not only cytopenias and sometimes fatal infections (acute treatment toxicity) but also mutational Th is chart shows diagnostics based decision steps (orange rectangles) and their sequence (orange arrows), currently approved therapies by EMA (in 2/2017) (blue rectangles), as well as the sequence for fi rst-line treatment (green arrows) and salvage treatment lines (blue arrows). Th e minority of patients (about 10%) present with indication for treatment at diagnosis, while the majority are observed until the criteria for treatment are met. Th is part (framed) did not change. When the indication is present, B-CLL patients are eligible for fi rst-line treatment. In this part, major changes have occurred because of recent approval of new options. Ibrutinib monotherapy is approved as continuous treatment of undetermined duration or until progression or unmanageable toxicity for all patient strata because of favorable effi cacy/ tolerance ratio in disease control. Th is is a new approved option, so that ibrutinib could be used to start the new path. In case of progression or toxicity, patients qualify for second-line treatment (approved option is venetoclax). If this fails, the patient is eligible for experimental treatments (combinations of novel agents with immunotherapy, allogeneic stem cell transplantation, CAR-T cell therapy, and the like). Th eoretically, all of these could be done without further diagnostics and stratifi cation, while avoiding chemotherapy. However, head-tohead comparison data between novel agents and chemo-immunotherapy (CIT) are still lacking. CIT, although associated with higher short-and long-term toxicities, has proved to be highly eff ective in achieving long, durable remissions and perhaps even a cure in some patient subsets. It was therefore in the CIT era essential to identify those who would respond, and a number of predicting factors have emerged in this setting. Th e NGS has revealed that intratumoral heterogeneity and genomic changes can be used for better CIT response prediction. Most important for CIT clinical use are two predictors, del(17p)/TP53mut and IGHV mutation status. Th e former can identify patient subset in which chemotherapy is ineff ective and even contraindicated because of inducing adverse clonal evolution, and the latter can identify disease type, where patients with unmutated IGHV poorly respond to CIT and even if they respond, the response is short and clones that are more resistant emerge. Both predictors are considered standard minimum for stratifi cation. If adverse features are present, the patients should be treated with ibrutinib or idelalisib in fi rst line. Others may continue towards CIT that is tailored according to age and comorbidities. Fit patients qualify for FCR, unfi t for Clb+Obi or like, and patients 'in-between' for BR. If they relapse late, the CIT may be repeated, tailored to current fi tness, while early relapsed/refractory patients qualify for BRI or venetoclax salvage. At present, baseline stratifi cation based on genetically defi ned risk, as well as on age and comorbidities to tailor treatment intensity is still needed for CIT, although fi tness is currently not important for novel agents. Th e current CIT based paradigm (shown horizontally) is losing importance and the new paradigm (shown vertically) is likely to take over. However, it will require identifi cation of new important predictors along the new path, since the majority of predictors identifi ed for CIT lose their power in the new setting. As data accumulate, new predictors will emerge for this setting. High throughput NGS has begun to identify new predictors for targeted therapy response, as well as new predictors of failure at molecular level, as treatment proceeds. All this may eventually lead to a new upfront stratifi cation for risk adapted precision medicine therapy in B-CLL. Th e ongoing trials and head-to-head comparison of novel agents and their combinations with immunotherapy versus CIT are under way. Th ey will hopefully resolve current dilemmas. Novel therapy research including genomic diagnostics is likely to off er new options that will eventually lead to time limited therapies, without chemotherapy. Dg = diagnosis; WW(I) = watch and wait (investigate); Ind = indications for treatment; PF = predictive factors; Late R = late relapse; Early R/R = early relapsing or refractory; IBR = ibrutinib; IDELA = idelalisib; VEN = venetoclax; Exp = experimental treatment; CIT = chemo-immunotherapy damage to the genome that can manifest as more aggressive and refractory phenotypes upon relapse and can induce second malignancies. For this reason, avoiding chemotherapeutic agents upfront, when possible, is a new paradigm of sequencing therapy for CLL. Major changes are explained in [fig_ref] Figure 1: Flow chart describing B-CLL diagnosis and treatment 2017 paradigm shift [/fig_ref].
However, in the absence of head-to-head randomized trials to assess effi cacy/tolerance ratio between novel therapies and best CIT for fi t patients, in this subset CIT should not be abandoned. Also, the access and availability of new treatments will need some time, and the current cost of novel treatment may be out of reach for insurers.

## Treatment options and current labels for approved antineoplastic medications
Observation. For patients who do not fulfi ll the criteria for starting therapy, the treatment is observation. Although it is in contrast to general oncologic tendency to treat patients with neoplasms as soon as possible, no data exist to suggest any harm in deferring therapy in those patients. Since the rate of progression may vary, frequent and careful observation is required to monitor the clinical course.
Ibrutinib. Ibrutinib is a selective irreversible inhibitor of Bruton tyrosine kinase, a signaling molecule located upstream in the B-cell receptor-signaling cascade. Label: IMBRUVICA as a single agent is indicated for the treatment of adult patients with previously untreated CLL; IMBRUVICA as a single agent or in combination with bendamustine and rituximab (BR) is indicated for the treatment of adult patients with CLL who have received at least one prior therapy. Th ose indications have been approved based on the randomized phase 3 studies RESONATE-2 (PCYC-1115-CA), RESONATE and HELIOS [bib_ref] Ibrutinib as initial therapy for patients with chronic lymphocytic leukemia, Burger [/bib_ref] [bib_ref] Ibrutinib versus ofatumumab in previously treated chronic lymphoid leukemia, Byrd [/bib_ref] [bib_ref] Ibrutinib combined with bendamustine and rituximab compared with placebo, bendamustine, and rituximab..., Chanan-Khan [/bib_ref].
Idelalisib. Idelalisib is an oral inhibitor of the delta isoform of phosphatidylinositol 3-kinase, which is located in the B-cell receptor-signaling cascade. Label: ZYDELIG is indicated in combination with an anti-CD20 monoclonal antibody (rituximab or ofatumumab) for the treatment of adult patients with CLL who have received at least one prior therapy, or as fi rst-line treatment in the presence of 17p deletion or TP53 mutation in patients who are not eligible for any other therapies. Approval is based on randomized, doubleblind, phase 3 study [bib_ref] Idelalisib and rituximab in relapsed chronic lymphocytic leukemia, Furman [/bib_ref] and phase 2 study 76 .

## Rituximab.
Rituximab is a murine anti-CD20 monoclonal antibody. Label: MABTHERA is indicated in combination with chemotherapy for the treatment of patients with previously untreated and relapsed/refractory CLL. Approval was based on randomized phase 3 studies with fl udarabine and cyclophosphamide (FCR) [bib_ref] Addition of rituximab to fl udarabine and cyclophosphamide in patients with chronic..., Hallek [/bib_ref] [bib_ref] Rituximab plus fl udarabine and cyclophosphamide prolongs progression-free survival compared with fl..., Robak [/bib_ref] , bendamustine (BR) [bib_ref] Bendamustine in combination with rituximab for previously untreated patients with chronic lymphocytic..., Fischer [/bib_ref] [bib_ref] Bendamustine combined with rituximab in patients with relapsed and/or refractory chronic lymphocytic..., Fischer [/bib_ref] , and chlorambucil [bib_ref] Obinutuzumab plus chlorambucil in patients with CLL and coexisting conditions, Goede [/bib_ref] [bib_ref] Rituximab plus chlorambucil as fi rst-line treatment for chronic lymphocytic leukemia: fi..., Hillmen [/bib_ref].
Obinutuzumab. Obinutuzumab is a human anti-CD20 monoclonal antibody. Label: GAZYVARO in combination with chlorambucil is indicated for the treatment of adult patients with previously untreated CLL and with comorbidities making them unsuitable for full-dose fl udarabine based therapy. Approval is based on a randomized phase 3 study [bib_ref] Obinutuzumab plus chlorambucil in patients with CLL and coexisting conditions, Goede [/bib_ref].
Ofatumumab. Ofatumumab is a human anti-CD20 monoclonal antibody. Label: ARZERRA is used in previously untreated patients who cannot be treated with fl udarabine; in these patients, it is used together with chlorambucil or bendamustine (other cancer medicines); in patients whose disease has not responded to previous treatment (known as refractory disease) with fl udarabine and a medicine called alemtuzumab; and in patients whose disease has come back after previous treatment (known as relapsed disease). In these patients, ARZERRA is used together with fl udarabine and cyclophosphamide. Approval is based on a randomized phase 3 study in combination with chlorambucil COMPLEMENT-1 [bib_ref] Chlorambucil plus ofatumumab versus chlorambucil alone in previously untreated patients with chronic..., Hillmen [/bib_ref].
Alemtuzumab. Alemtuzumab, a monoclonal antibody directed at CD52. Th e drug was withdrawn for CLL indication by the producer because of commercial reasons. However, the company can off er the drug on request for compassionate use free of charge. Label: (EMA EPAR 2011 authorization withdrawn). MAB-CAMPATH is used to treat patients with B-cell CLL for whom treatment combinations including fl udarabine are not appropriate. Approval was based on randomized clinical trials and showed activity in TP53 mutation [bib_ref] Fludarabine plus alemtuzumab versus fl udarabine alone in patients with previously treated..., Elter [/bib_ref] [bib_ref] Alemtuzumab compared with chlorambucil as fi rstline therapy for chronic lymphocytic leukemia, Hillmen [/bib_ref] [bib_ref] Alemtuzumab in combination with methylprednisolone is a highly eff ective induction regimen..., Pettitt [/bib_ref].
Venetoclax. Venetoclax is a highly selective inhibitor of Bd2. Label: VENCLYXTO monotherapy is indicated for the treatment of CLL in the presence of 17p deletion or TP53 mutation in adult patients who are unsuitable for or have failed a B-cell receptor pathway inhibitor. Venclyxto monotherapy is indicated for the treatment of CLL in the absence of 17p deletion or TP53 mutation in adult patients who have failed both chemo-immunotherapy and B-cell receptor pathway inhibitor. Approval is based on a phase 1 escalation study [bib_ref] Targeting BCL2 with venetoclax in relapsed chronic lymphocytic leukemia, Roberts [/bib_ref] and phase 2 study [bib_ref] Venetoclax in relapsed or refractory chronic lymphocytic leukaemia with 17p deletion: a..., Stilgenbauer [/bib_ref].
Oral alkylating agents with or without corticosteroids. Chlorambucil was used for the treatment of CLL for more than 60 years. Label: LEUKERAN is indicated for the treatment of CLL. Although the role of chlorambucil has considerably diminished, regulators paradoxically still accept drugs for registration on the basis of phase 3 trials, which use chlorambucil in very low doses as comparator [bib_ref] Alemtuzumab compared with chlorambucil as fi rstline therapy for chronic lymphocytic leukemia, Hillmen [/bib_ref] [bib_ref] Fludarabine compared with chlorambucil as primary therapy for chronic lymphocytic leukemia, Rai [/bib_ref] [bib_ref] bendamustine versus chlorambucil in treatment-naive patients with b-cell chronic lymphocytic leukemia (B-CLL):..., Knauf [/bib_ref] , while claiming at the same time that the therapeutic success of such therapies is extremely modest. Th us, low doses are still considered standard therapy, although it was shown that medium and especially high doses had significantly greater eff ectiveness [bib_ref] High dose continuous chlorambucil vs intermittent chlorambucil plus prednisone for treatment of..., Jaksic [/bib_ref] [bib_ref] High dose chlorambucil versus Binet's modifi ed cyclophosphamide, doxorubicin, vincristine, and prednisone..., Jaksic [/bib_ref] [bib_ref] Randomized phase II study in untreated B-cell chronic lymphocytic leukemia (B-CLL) comparing..., Jaksic [/bib_ref] [bib_ref] Fludarabine plus cyclophosphamide improves response and progression-free survival in chronic lymphocytic leukemia...., Catovsky [/bib_ref]. A meta-analysis of six trials of immediate versus deferred therapy with chlorambucil showed no diff erence in OS at 10 years 69 .
Purine analogs. Fludarabine is a purine analog, one of a group of chemotherapy drugs known as anti metabolites. Th ey stop cells making and repairing DNA. Cancer cells need to make and repair DNA in order to grow and multiply. Label: Fludarabine (gen) is used in the treatment of B-cell CLL in patients with suffi cient healthy blood cell production. First treatment for CLL with this medicine should only be started in patients with advanced disease having disease related symptoms or evidence of disease progression. Approval is based on a phase 3 randomized study [bib_ref] Fludarabine compared with chlorambucil as primary therapy for chronic lymphocytic leukemia, Rai [/bib_ref]. Th is drug is also used in combination therapies (see below).
Bendamustine. Bendamustine is a cytotoxic agent with bifunctional properties of an alkylator and a purine analog. Label: bendamustine (gen) is used as monotherapy or in combinations with other drugs for treatment of CLL in cases when combination chemotherapy containing fl udarabine is not appropriate. Approval is based on a randomized phase 3 trial for monotherapy [bib_ref] Phase III randomized study of bendamustine compared with chlorambucil in previously untreated..., Knauf [/bib_ref] and for combinations on phase 2 trials in previously treated [bib_ref] Bendamustine in combination with rituximab for previously untreated patients with chronic lymphocytic..., Fischer [/bib_ref] and untreated patients [bib_ref] Bendamustine combined with rituximab in patients with relapsed and/or refractory chronic lymphocytic..., Fischer [/bib_ref].
Combination therapy Fludarabine based combinations include FCR, FCOfa, FR, and FC. Fludarabine plus cyclophosphamide plus rituximab has proved very eff ective in those patients who can tolerate the treatment. For this reason, in the last 10 years, FCR has become the gold standard of CIT for fi t patients [bib_ref] Addition of rituximab to fl udarabine and cyclophosphamide in patients with chronic..., Hallek [/bib_ref]. Long-term results have confi rmed overall effi cacy and a subset of long-term responders defi ned by genomic risk groups has emerged [bib_ref] Long-term remissions after FCR chemoimmunotherapy in previously untreated patients with CLL: updated..., Fischer [/bib_ref] [bib_ref] Targeted therapies in CLL: mechanisms of resistance and strategies for management, Woyach [/bib_ref] [bib_ref] Fludarabine, cyclophosphamide, and rituximab treatment achieves long-term disease-free survival in IGHV-mutated chronic..., Th Ompson [/bib_ref]. Although no head-to-head comparisons have yet been completed in line with the new paradigm, the indication is narrowed to a subset of fi t patients with hypermutated IVGH in whom the likelihood of very long remission may outweigh the concerns of chemotherapy toxicity.
Bendamustine combinations are used in those patients in whom fl udarabine cannot be tolerated. In head-to-head comparison to FCR, BR combination was found inferior [bib_ref] First-line chemoimmunotherapy with bendamustine and rituximab versus fl udarabine, cyclophosphamide, and rituximab..., Eichhorst [/bib_ref].
Combination therapy without anti-CD20 monoclonal antibodies. For FC, CVP and CHOP, a metaanalysis of ten trials compared combination chemotherapy (before the availability of rituximab) with chlorambucil alone and showed no diff erence in OS at 5 years [bib_ref] Chemotherapeutic options in chronic lymphocytic leukemia: a meta-analysis of the randomized trials...., Cll Trialists&apos; Collaborative [/bib_ref].
Combination with novel agents. It is likely that combinations of BRI and Bcl2 inhibitors with anti-CD20 antibodies will be the basis in the era of the new emerging paradigm. At present, only ibrutinib + bendamustine + rituximab and idelalisib + anti-CD20 monoclonal antibodies are approved.
Bone marrow and peripheral stem cell transplantations. Although this modality is considered the only option for cure, it is still under clinical evaluation, especially in the context of novel agents and new emerging paradigm [bib_ref] Sequencing of chronic lymphocytic leukemia therapies, Barrientos [/bib_ref] [bib_ref] Managing high-risk CLL during transition to a new treatment era: stem cell..., Dreger [/bib_ref].
Th e overall therapeutic eff ect is a consequence of total therapeutic interventions, including antineoplastic and supportive measures that are particularly important. [fig_ref] Table 5: Defi nition of response to treatment [/fig_ref] shows the criteria for assessment of therapeutic eff ect. Th e criteria generally used the same grounds, based on the estimation of the tumor mass parameters in diff erent compartments on the one hand, and the parameters for the assessment of myelopoiesis on the other hand. Th e criteria are somewhat diff erent in the current NCI/iwCLL criteria 6,13 and the criteria described below (IGCI, EORTC) [bib_ref] High dose continuous chlorambucil vs intermittent chlorambucil plus prednisone for treatment of..., Jaksic [/bib_ref] [bib_ref] High dose chlorambucil versus Binet's modifi ed cyclophosphamide, doxorubicin, vincristine, and prednisone..., Jaksic [/bib_ref] [bib_ref] Randomized phase II study in untreated B-cell chronic lymphocytic leukemia (B-CLL) comparing..., Jaksic [/bib_ref] [bib_ref] More intensive treatment signifi cantly improves survival in young patients with advanced..., Jaksic [/bib_ref] [bib_ref] A randomized phase II trial of high-dose chlorambucil vs. fl udarabine in..., Eortc [/bib_ref].

## Criteria for evaluation of response to therapy
To monitor the dynamics of the disease (both progression and response to therapy), TTM score (described in clinical stage) is very convenient because it is the only clinical system that is based on a continuous, quantitative parameter that is easy to apply and NCI based criteria, but TTM shows the advantage in assessing partial remission (PR) by comparing total tumor mass before and after treatment, so it is possible to set a minimum threshold for minimal remission (MR), e.g., reduction by >25%, partial remission (PR) >50%, and very good PR >75% and more. Likewise, the TTM based criteria are more accurate and without bias on estimating stable disease (SD) and progressive disease (PD). Th e continuous quantitative character of TTM size allows for comparison of trends between the criterion group A (TTM) and group B (function of residual normal hematopoiesis). It is possible to evaluate the benefi cial antineoplastic eff ect of therapy independently of the toxic eff ect on hematopoiesis. validated in thousands of patients in various international clinical trials. To estimate the doubling time (DT), it is more reliable than just the number of lymphocytes because it can compensate for changes in the distribution of the tumor mass occurrence after the administration of corticosteroids or TKIs, when there may be an increase in the number of leukocytes while reducing the nodes or spleen. For this reason, the NCI/ iwCLL criteria have recently been amended, but are still suboptimal for monitoring disease response [bib_ref] Increased frequency (12%) of circulating chronic lymphocytic leukemia-like B-cell clones in healthy..., Nieto [/bib_ref] [bib_ref] Novel targeted agents and the need to refi ne clinical end points..., Cheson [/bib_ref] , while the TTM scoring system is much better for measuring redistribution of clonal cells among compartments [bib_ref] An old and simple solution for a new problem -more on clinical..., Jaksic [/bib_ref] [bib_ref] Redistribution pattern and assessment of early response to ibrutinib in CLL by..., Jaksic [/bib_ref].
When assessing the response to therapy, complete remission (CR) is assessed equally in the TTM and Group A criteria defi ne tumor mass, group B criteria defi ne hematopoietic system (or bone marrow) function. 1 CR (complete remission): all criteria must be present, and patients must be without general symptoms associated with CLL; PR (partial remission): at least two criteria in group A plus one in group B must be present; SD (stable disease) is the absence of progressive disease (PD) if at least PR is not reached; PD (progressive disease): at least one criterion from group A or B must be present. [bib_ref] Th e clinical implications of gene mutations in chronic lymphocytic leukaemia, Rossi [/bib_ref] Th e sum of the products of multiple lymph nodes (as evaluated by CT in clinical trials or by physical examination in general practice). [bib_ref] Presence of multiple recurrent mutations confers poor trial outcome of relapsed/refractory CLL, Guieze [/bib_ref] Th ese parameters are irrelevant for certain types of responses.
Minimal residual disease (MRD) is an important end point, and should refl ect no measurable disease in the body. However, MRD testing should be standardized and the sample source well defi ned. By doing this, we should avoid reports of MRD negative patients with persistent signifi cant organomegaly. Th ose cases in fact have 'clean' blood but sometimes may not fulfi ll common criteria for partial remission, and the term MRD without specifi cation is misleading.

## Antineoplastic therapy -current treatment options
Th ere are several treatment options. Th erapeutic recommendations summary for fi rst-line treatment and for salvage treatment in major therapeutic stratifi cation groups are shown in respective tables.

## Management of patients with no accepted criteria for therapy
If the patient is not showing any signs of active/ progressive/symptomatic disease, the antineoplastic therapy is not recommended, but the patient is monitored and reviewed without therapy. Th us 'Watch and Wait' should be transformed to 'Watch and Investigate' (W&I). Th is view is based on the evidence collected in randomized trials during the 1980s, when it was shown that chlorambucil based treatment did not contribute to longer survival, moreover, despite the relative ease of controlling symptoms and achieving clinical remission, overall survival was marginally worse [bib_ref] High dose continuous chlorambucil vs intermittent chlorambucil plus prednisone for treatment of..., Jaksic [/bib_ref] [bib_ref] More intensive treatment signifi cantly improves survival in young patients with advanced..., Jaksic [/bib_ref] [bib_ref] Chlorambucil in indolent chronic lymphocytic leukemia, Dighiero [/bib_ref]. Until now, there are no data indicating that harm is due to deferring therapy in asymptomatic, stable disease. However, trials that are under way, which include novel agents and/or combinations, may change this concept, but it will take time since those trials require prolonged follow up.
While for these patients antineoplastic medication is not recommended, standard care should include infection prophylaxis such as annual fl u and pneumococcal every 5 years, and in case of infection early treatment.

## Initial treatment (fi rst-line treatment)
Th e fi rst-line treatment relates to previously untreated patients. All patients in standard care must have clinical indication for treatment initiation (i.e. must fulfi ll the criteria stipulated above). Recommendation depends on the risk associated with B-CLL (High or Low) and patient general condition (Fit or Unfi t) [fig_ref] Table 6: First-line treatment of CLL [/fig_ref] , [fig_ref] Figure 1: Flow chart describing B-CLL diagnosis and treatment 2017 paradigm shift [/fig_ref]. Each stratum will be discussed separately.

## Initial treatment for patients with no del(17p/tp53 mutation that are fi t (low risk + fit)
As a rule, the patients in this stratum (about onethird of fi rst-line treatment) are younger than 65. Th erapeutic goal is to be set high, to achieve complete, durable remission, prolong survival, and perhaps even off er a possible cure.
FCR (fl udarabine, cyclophosphamide, rituximab) is recommended as a standard initial therapy for previously untreated fi t patients outside clinical trials 78 . According to the DCLLSG CLL8 protocol, 6 cycles at intervals of 28 days if the patients tolerate the treatment well, and after EOT, no further treatment is anticipated, only follow up visits. Long-term follow up identifi ed a subset of patients in whom long and durable response was achieved [bib_ref] Targeted therapies in CLL: mechanisms of resistance and strategies for management, Woyach [/bib_ref] [bib_ref] Fludarabine, cyclophosphamide, and rituximab treatment achieves long-term disease-free survival in IGHV-mutated chronic..., Th Ompson [/bib_ref] [bib_ref] Long-term remissions after FCR chemoimmunotherapy in previously untreated patients with CLL: updated..., Fischer [/bib_ref]. Th ose patients had mutated IGHV, no 11q and were aged <65. Other patients may respond, but they tend to relapse soon. For this reason, for fi t patients aged <65, mutated IGHV and no 11q (nor del(17p)/TP mutation) who fulfi ll the criteria for treatment, FCR is the treatment of choice. In others, according to the new paradigm, it is suggested to avoid chemotherapy, especially FC.
In patients who are unsuitable for fl udarabine therapy, bendamustine + rituximab (BR) can be tried. It seems to be both less toxic and less eff ective.
Ibrutinib is also approved for this indication. According to the new paradigm, BRI may replace CIT but at present, there are no data of randomized trials to support it. Head-to-head comparison of FCR or BR with ibrutinib is under way and is likely to resolve this question.

## Initial treatment for patients with no del(17p)/tp53 abnormality that are unfi t (low risk+unfit)
Th e majority of patients (more than 60% of treated in fi rst line) belong to this group. As a rule, the patients are older than 65, with comorbidity, and therefore not capable to tolerate aggressive CIT therapeutic approach; thus, it is necessary to modify therapeutic goal this subset, since the TP53 mutated clone is not responding and CIT may enhance unfavorable clonal selection and is therefore harmful. We recommend for both fi t and unfi t the induction with ibrutinib or idelalisib plus rituximab. Ibrutinib appears superior to idelalisib in all settings as fi rst choice BRI [bib_ref] Indirect treatment comparisons of ibrutinib versus physician's choice and idelalisib plus ofatumumab..., Sorensen [/bib_ref] [bib_ref] Outcomes of CLL patients treated with sequential kinase inhibitor therapy: a real..., Mato [/bib_ref]. In selected cases, this therapy can be followed by elective AlloSCT. HDMP plus rituximab [bib_ref] Rituximab in combination with high-dose methylprednisolone for the treatment of chronic lymphocytic..., Castro [/bib_ref] [bib_ref] High dose methylprednisolone and rituximab is an eff ective therapy in advanced..., Dungarwalla [/bib_ref] or alemtuzumab 122 should be used if BCI therapy is unavailable.

## Treatment for relapsed/refractory cll
Th is relates to previously treated patients. Again, they should fulfi ll the criteria for retreatment, essentially the same as described above. For this reason, time to progression (measured as PFS) is distinct from time to next treatment (TTNT). Th e situation in this setting is much more complex, since in addition to four major therapeutic strata, special attention should be paid to previous treatment(s) (type of treatment, num-and choose remission or stabilization of disease with a well-preserved quality of life.
We recommend chlorambucil plus one of the anti-CD20 antibodies as a standard [bib_ref] Obinutuzumab plus chlorambucil in patients with CLL and coexisting conditions, Goede [/bib_ref] [bib_ref] Rituximab plus chlorambucil as fi rst-line treatment for chronic lymphocytic leukemia: fi..., Hillmen [/bib_ref] [bib_ref] Chlorambucil plus ofatumumab versus chlorambucil alone in previously untreated patients with chronic..., Hillmen [/bib_ref] [bib_ref] Rituximab plus continuous or intermittent high-dose chlorambucil for previously untreated CLL: dose,..., Basic-Kinda [/bib_ref]. Best results have been published with chlorambucil + obinutuzumab.
Also, BR is an option of chemo-immunotherapy for patients with appropriate fi tness.
Ibrutinib is also approved for this indication because of excellent results in this patient subset. According to the new paradigm, BRI may replace CIT but at present, there are no direct data to support it. Head-to-head comparison of ibrutinib + obinutuzumab with chlorambucil + obinotuzumab is under way and is likely to resolve this question.

## Initial treatment for fi t/unfi t patients with del(17p) /tp53 mutation (high risk + fit/unfit)
In this stratum, we expect less than 7% of all patients treated in fi rst line. CIT is contraindicated in ber of treatment lines, the period that has elapsed from previous treatment, etc.). In principle, with the exception of very late relapses, the patients require more therapy to achieve less response. Since the vast majority (>90%) of all CLL fi rst-line treated patients have no TP53 abnormality, their treatment allocation was essentially dependent on their general condition (Fit or Unfi t). Th us, the fi t patients receive more aggressive treatment aiming at MRD negativity (hopefully eradication of the disease), while the unfi t patients receive less aggressive treatment that is less likely to achieve MRD negativity and consequently the therapeutic aim is less ambitious.
Th e relapse is, therefore, primarily linked to the fi rst-line therapeutic stratum. It is an indicator of respective therapy failure. In principle, the longer the period to relapse, the more eff ective fi rst-line treatment was.
General principles of therapeutic strategies in relapsing/refractory patients are shown in [fig_ref] Table 7: Treatment of relapsed/refractory CLL [/fig_ref] and [fig_ref] Figure 1: Flow chart describing B-CLL diagnosis and treatment 2017 paradigm shift [/fig_ref]. Th e relapsing patient in early relapse is considered refractory and requires change of therapy. If the treatment results in remission of long duration, it is reasonable to try the same treatment that has proved eff ective again in the relapse. For this reason, relapses will be described as a function of fi rst-line stratifi cation therapeutic failures. However, in all patients in relapse, TP53 status should be checked (before each new line of therapy) to assess whether the risk grade has changed in comparison to front-line stratifi cation. Most of the patients currently relapsing early were in fi rst line treated by CIT adjusted to fi tness status. Occasionally, some patients in fi rst relapse were in fi rst line treated by chemotherapy (chlorambucil, fl uradabine alone or in combination with cyclophosphamide).

## Patients relapsing from the low risk + fit stratum
Patients relapsing early are considered refractory and should be treated with ibrutinib, or idelalisib plus rituximab.
If the anti BCR drugs cannot be provided, the current options include BR, HDMP+R, FA, other chemo-immunotherapy, ofatumumab and alloSCT.
Patients relapsing late who have not acquired TP53 abnormality remain fi t enough for fl udarabine-based treatment, and those with a clinical indication for treatment may receive FCR, provided that they have mutated IGHV, no TP53 or 11q [bib_ref] Sequencing of chronic lymphocytic leukemia therapies, Barrientos [/bib_ref] [bib_ref] fl udarabine, alemtuzumab, and rituximab as salvage therapy for heavily pretreated patients..., Badoux [/bib_ref]. If the patient at the time of relapse changes to unfi t stratum, the relapse treatment described in respective section applies.

## Patients relapsing from the low risk + unfit stratum
Over 50% of all treated patients belong to this group. In this group, less aggressive treatment was applied in front line because these patients are not likely to tolerate FCR. Th e response obtained is less likely MRD negative and relapses are expected in a wide range from early (less than 24 months) or late (more than 24 months).
In case of early relapse, patient is considered refractory to given treatment and ibrutinib or idelalisib + rituximab is recommended. Less benefi t can be expected from BR or chlorambucil + antiCD20 or HDMP + R. In selected responsive cases, ofatumumab maintenance is applied.
In case of late relapse, the patients relapsing after chlorambucil can be retreated with chlorambucil + anti-CD20 monoclonal antibody [bib_ref] Rituximab plus continuous or intermittent high-dose chlorambucil for previously untreated CLL: dose,..., Basic-Kinda [/bib_ref]. Th is therapy may be repeated several times (in function of achieved du-ration) until the duration of remission is shortened to 2 years, after which it is justifi ed to go to the secondline therapy (ibrutinib or idelalisib+R).

## Patients relapsing from the high risk + fit/unfit stratum
Th ese patients are at a particularly high risk. If the patients did not receive BRI treatment, those drugs are recommended. If the patients relapse on BRI drugs, venetoclax or alternative BRI is indicated. If those are not available, HDMP + R or alemtuzumab  R can be tried.
If the fi t patients were not previously allo transplanted, reinduction should be considered with diff erent combinations (including ofatumumab) and if successful, proceed to transplantation.
In selected responsive cases, ofatumumab maintenance may be considered 124 .

## Consolidation/maintenance therapy
It has been observed that MRD negative remission is associated with prolonged progression free survival both in previously untreated [bib_ref] Fludarabine, cyclophosphamide, and mitoxantrone as initial therapy of chronic lymphocytic leukemia: high..., Bosch [/bib_ref] and relapsed 126 patients. Th is has led to studies of additional treatment in patients with residual disease post therapy. In line with the new paradigm, although chlorambucil maintenance can prolong survival 127 , maintenance treatment should be chemotherapy free. Ofatumumab in selective responsive patients was shown to prolong PFS but not OS 128 , and is approved for this indication by FDA. Other anti-CD20 antibodies, although approved in some other B-cell malignancies [bib_ref] Rituximab maintenance strategy in advanced follicular lymphoma: facts and controversies, Milunovic [/bib_ref] , are not yet approved in B-CLL by regulators. Lenalidomide was also shown to improve PFS, but is not yet approved by regulators.
As described earlier, BRI therapies are given for a prolonged period to maintain the response as distinct from chemo-immunotherapy. Early (months) period of treatment is characterized by marked tumor mass redistribution, so that monitoring should be adjusted accordingly (see section on response criteria).

## Th e role of allogeneic transplantation
Th e indication for allogeneic SCT is currently changing in line with excellent results of BRI and venetoclax. Current indications include poor respond-ers to BRI and Bcl-2 antagonist, and appearance of Richter syndrome [bib_ref] Managing high-risk CLL during transition to a new treatment era: stem cell..., Dreger [/bib_ref].

## Th e role of radiotherapy
Radiotherapy can provide eff ective palliation in cases with symptomatic bulky lymphadenopathy and should be off ered to patients in which chemo-immunotherapy has been ineff ective or is contraindicated. Low doses of external beam radiotherapy (2x2 Gy) can be highly eff ective in this situation and a higher dose (30 Gy in 2-3 Gy fractions) may be required in patients with transformed aggressive disease or those known to have a TP53 abnormality 132 .

## Treatment of sll
Th e biological similarities between SLL and CLL are so close that a similar response to treatment could be expected. Th is is supported by the MDACC single centre retrospective study [bib_ref] Assessment of chronic lymphocytic leukemia and small lymphocytic lymphoma by absolute lymphocyte..., Tsimberidou [/bib_ref]. Indications for and choices of treatment are the same as for CLL. Rare patients in whom SLL is diagnosed following biopsy of an enlarged lymph node in the absence of detectable disease at any other site may be off ered local radiotherapy with curative intent.

## Risk of other diseases
Infections Th e risk of infections is related to progressive defect both in antibody-and cell-mediated immunity. In addition, therapy may worsen immune impairment, particularly fl udarabine and anti-CD20 antibodies, but also B-cell receptor inhibitors. For this reason, infections represent a frequent cause of morbidity and mortality in CLL. Infections are typically bacterial and should be treated early. For those who have repeated infection and require repeated antibiotics, immunoglobulin replacement therapy should be considered. Prophylactic vaccination is advised, but live vaccine should be avoided.

## Autoimmune complications
Autoimmune complications are common in CLL, occurring in 10%-20% of patients [bib_ref] Chronic lymphocytic leukemia and autoimmunity: a systematic review, Hodgson [/bib_ref]. Th ey almost exclusively target blood cells, most commonly red blood cells. Hemolytic anemia (AIHA) is predominant and immune thrombocytopenia (ITP) is 4-5 times less common. A bone marrow aspirate is usually required to confi rm the diagnosis of autoimmune cytopenia.
AIHA or ITP should be treated before deciding whether therapy for CLL is needed. First-line therapy is prednisolone. Second-line therapies for patients intolerant of or refractory to steroids include cyclosporine, intravenous immunoglobulin (ITP), thrombopoietin mimetic agents (ITP), CVP, low-dose cyclophosphamide, rituximab, alemtuzumab and splenectomy. CLL treatment may be initiated to control recurrent or refractory AIHA/ITP. Rituximab-containing regimens are recommended in patients who do not have a TP53 abnormality. If AIHA/ITP develops during CLL treatment, the same regimen should be used again in this patient with extreme caution and only if no eff ective alternative is available. Autoimmune neutropenia usually responds to GCSF.

## Prolymphocytic transformation
B-cell prolymphocytic transformation is diagnosed in progressive organomegaly and lymphocytosis with increase of prolymphocytes >55% and is treated as aggressive CLL. It occurs rarely in <1% of cases.

## Richter syndrome
Richter syndrome is CLL transformation to aggressive lymphoma, usually DLBCL or Hodgkin like. LN biopsy is mandatory for diagnosis, PET may be helpful. It occurs in 2%-7% of patients. While novel agents are under investigation, CIT is still the recommended approach. Depending on the histologic subtype of lymphomatous transformation, patients who are suitable for intensive therapy should receive regimens currently employed for either primary diff use large B cell lymphoma (Richter syndrome) or Hodgkin's lymphoma. Younger patients who achieve good response are candidates for allogeneic stem cell transplantation.
Check point inhibitor pembrolizumab showed activity in RS, but not in CLL [bib_ref] Th e PD-1/PD-L1 axis contributes to T-cell dysfunction in chronic lymphocytic leukemia, Brusa [/bib_ref] [bib_ref] Th e emerging role of immune checkpoint inhibition in malignant lymphoma, Hude [/bib_ref].

## Myelodysplastic syndrome and acute leukemia
Although MDS and AML are uncommon in CLL, the rate of therapy related to fl uadarabine based CIT is about 5% and much higher after autologous stem cell - Annual vaccination against infl uenza. Care must be taken of the fact that the recovery of B-cell system after anti CD20 antibody therapy lasts for about 9 months, so that the response to vaccination in this period is inadequate. - Pneumococcal vaccine every 5 years - Avoid all live vaccines, including Zoster 2
Anti-infective prophylaxis
- For patients receiving purine analogs and/or alemtuzumab, and in period after that the following prophylaxis is recommended:  Herpes viruses (acyclovir)  PCP (sulfamethoxazole/trimethoprim) - Special attention in patients receiving alemtuzumab should be paid to the problem of CMV reactivation. Although there is no common position in the literature, most reports recommend that prophylactic ganciclovir is prescribed if viremia present.
Th e viral load levels must be monitored every few weeks. - For patients receiving anti CD20 antibody and BRI and positive for HBV, HCV consult GE/infectologist for antiviral prophylaxis.
3

## Respiratory infections requiring iv antibiotics and hospitalization
- Apply appropriate antibiotic therapy.
- Determine serum IgG, and if the value is less than 5 g/L:  Apply monthly IVIG 0.3-0.5 g/kg  Adjust the dose so that the value is maintained above 5 g/L 4 Immunoglobulin replacement therapy
- Should be considered as a means of reducing the incidence of bacterial infections in patients with a low serum IgG level who have experienced previous major or recurrent minor bacterial infections despite optimal antibacterial prophylaxis. - Th e goal should be to reduce the incidence of infection and the immunoglobulin dose should be adjusted accordingly. - Patients should be reviewed regularly to evaluate the eff ectiveness of immunoglobulin replacement therapy and whether there is a continuing need for treatment. - Patients who develop serious and/or recurrent infections despite antimicrobial prophylaxis and immunoglobulin replacement should be managed in conjunction with a microbiologist, infectious disease specialist, and/or immunologist.

## Blood transfusion
- Th e use of irradiated blood products should be considered in the following situations: indefi nitely in patients treated with a purine analog, following bendamustine until more evidence emerges about the risk of transfusion-associated graft versus host disease, following alemtuzumab and for 3 months post conditioning with chemotherapy or immunotherapy (6 months after total body irradiation) for patients undergoing autologous transplantation.
6 Tumor lysis - Venetoclax can cause severe tumor lysis syndrome. Special precautions and ramp-up therapy should be followed strictly.
transplantation. Th e response to therapy is poor. Whether novel agents that do not induce genotoxic stress to stem cells reduce the incidence of this serious complication is currently under evaluation.

## Supportive therapy
Supportive therapy plays an important role in management and is carried out in accordance with the generally accepted good clinical practice [bib_ref] Guidelines on the diagnosis, investigation and management of chronic lymphocytic leukaemia, Oscier [/bib_ref]. [fig_ref] Table 8: Supportive therapy in patients with B-CLL [/fig_ref] shows the basic characteristics of supportive therapy in B-CLL. Th is covers the area of vaccination, anti-infective prophylaxis, respiratory recurrent infections requiring IV antibiotics and hospitalization, immunoglobulin replacement therapy, blood transfusions, and tumor lysis.
Extended version and updates can be found on KROHEM website: http://www.krohem.hr/Documents/AMENDMENT%20KROHEM%20CLL% 20GUIDELINES%20v1.%202016%20%20ENGL.pdf
[fig] Figure 1: Flow chart describing B-CLL diagnosis and treatment 2017 paradigm shift. [/fig]
[table] Table 1: Steps and aims in diagnostic process that leads to defi nition of therapeutic goal and strategySTEPS AND AIMS BASIS FOR DECISION CRITERIAComment: the table shows diagnostic steps. [/table]
[table] Table 2: Clinical stages and TTM absolute number of lymphocytes; TTM = Total Tumor Mass score Note: if the largest lymph node is found by imaging (US or CT), those values should be used for TM 2 Tumor Mass Distribution (TD) is calculated as quantitative parameter according to formula: Doubling Time of TTM (DT) is calculated as quantitative parameter according to formula: where TTM beg is the size of TTM at the beginning of period M: TTM end is TTM size at the end of period M; M is interval between TTM beg and TTM end in months. M should be at least 3 months. Electronic calculator available online at: http://www.krohem.hr hematologic evaluation, under the criteria shown in [/table]
[table] Table 3: Criteria for active (progressive/symptomatic) disease [/table]
[table] Table 4: General schedule of investigations before, during and after therapy *Monitoring protocol frequency varies depending on clinical condition from several weeks to several months, or even one year if the situation is stable, without change. However, in the emergence of new circumstances, it is necessary to check-up early; **these repetitive visits depend on treatment applied. For novel agents with at present unlimited duration, these visits should enable detection of treatment failure; b preferred but not required tests. [/table]
[table] Table 5: Defi nition of response to treatment(NCI updated guidelines, Blood 2008) [/table]
[table] Table 6: First-line treatment of CLL (KROHEM v1 2017) Percentages of patients with distinct general condition and molecular genetics groups refer to treated patients. Fit patients are less than 65 years of age and with CIRS score less than 6. Younger patients with CIRS score of 6 and more and patients aged 65 years or more (regardless of CIRS score) qualify as unfi t. c Standard treatments are in order of preference, all are 2A or less according to NCCN consensus, treatments with higher grade are marked (1). [/table]
[table] Table 7: Treatment of relapsed/refractory CLL (KROHEM v1 2017) A = alemtuzumab; Allo SCT = allogeneic stem cell transplantation; HDMP (high dose methylprednisolone); antiCD20 (ofatumumab or obinutuzumab or rituximab). [/table]
[table] Table 8: Supportive therapy in patients with B-CLL [/table]